key: cord-0033659-at7pq6u1
authors: nan
title: The 20th Conference of the Asian Pacific Association for the Study of the Liver Poster Presentation
date: 2010-03-11
journal: Hepatol Int
DOI: 10.1007/s12072-010-9169-3
sha: 48699491bf7a4dafd65d11a0cf2f028370544432
doc_id: 33659
cord_uid: at7pq6u1

nan

(p \ 0.05), but the numbers were not significantly different between the negative HBeAg group and the control group (p[0.05). The frequencies of mDC, pDC and CD4 + /CD8 + ratios did not vary from each other among the three groups (p [ 0.05).

The statistic analyse result of our cases HBeAg positive group (n = 5) HBeAg negative group (n = 6) Normal control group (n = 5) , in oncohematological patients-HBV-37 (3.7%), HCV-63 (6.3%), co-infection in 6 (0.9%). Of 687 patients 33% (229)-female, 67% (458)-male, the mean age was 40 (3-77). 587 solid tumor patients consisted of 103 (18%)-breast, 58 (9.9%)-lung, 57 (9.7)%gastrointestinal, 33 (5.6%)-urological, 60 (10%)-gynecological, 20 (3.4%)brain, 53 (9%)-soft tissue, 203 (34%)-other cancers. 100 oncohematological patients consisted of 53 (53%)-leukemia, 30 (30%)-lymphoma, 17 (17%)others. Conclusion: Hepatitis B and C virus prevalence is 3.8% and 6.1% accordingly in oncohematological patients, which is higher, than in solid tumor patients (1.9% and 3.1%). Hepatitis B and C virus prevalence in solid tumor patients is the same, as in general population. Hepatitis B and C virus high prevalence in oncohematological patients, its association with nozology is suggested with possible role of hepatitis viruses in the pathogenesis of lymphoproliferative diseases and with necessity of multiple transfusions in these patients. Background: To investigate clinical feature and antifungal therapeutic effect of chronic severe hepatitis (CSH) cases with invasive fungal infection (IFI) in order to improve the level of diagnosis and treatment. Methods: Clinical manifestation, hemogram, imageology, mycetology characteristic, antifungal treatment prescription and therapeutic effect of 79 CSH cases with IFI was retrospectively analyzed. Results: 13 (16.5%) patients received glucocorticoid or other immunodepressant pre-admission, (50.6%) cases were taken invasive operation and (77.2%) patients were administered 1-6 kind of broad-spectrum antibiotic. 92.4% cases had fever. Leucocytes and neutrophilic granulocyte increased in 96.2% patients. Lung (31.6%), intestinal tract (26.2%) and oral cavity (14%) infection was common. Fungus was found in 70.9% cases. Candida albicans (40.9%) and Aspergillus (21.1%) was often seen. There was relative specificity halo sign and crescent sign on lung CT in 40% patients with fungal pneumonia. Voriconazole was more effective than fluconazol (71.4 vs. 39%, P \ 0.05). 12 cases with lung aspergillus infection were administered voriconazole, 8 (66.7%) cases of which was effective, 4 (33.3%) cases were dead. Conclusion: There were high risk factors in major CSH patients with IFI. The most clinical manifestation of CSH patients with IFI is fever, leukocytosis, lung and intestinal tract infection. Candida albicans and Aspergillus infection is common. Voriconazole is more effective than fluconazol in the CSH patients with IFI, and it can increase survival rate of the CSH patients with IFI.

Five Years Follow-up Study of 220 Chronic HBV Carriers Zhong-Hua Lu 1 1 Wuxi Infectious Diseases Hospital, No. 88 Xingyuan Zhong Road, Wuxi, China Objective: To understand the turnover of hepatic tissue pathology, hepatitis B reactivation rate and serological examination in chronic HBV carriers. Methods: A 5 years dynamic observation and research on 220 chronic HBV carriers in WuXi district has been taken via viewing the clinical symptom, histology, virology and HBV immunological markers, et al. Results: 35 of 220 patients, accounting for 15.9%, showed hepatitis B reactivation. The hepatitis B reactivation rate of patients with obvious hepatic tissue inflammation (CG 2 ) was 27% (33/122) and the patients with mild hepatic tissue inflammation (G 0 -G 1 ) was 2% (2/98) , showing a significant differences (v 2 = 25.41, P \ 0.01) and reactivation rate of patients with high inflammation was clearly higher ones with mild inflammation. 27 of 35 hepatitis B reactivation cases were older than 40 years, showing the significant statistical differences between the dissimilar age group cases and hepatitis B reactivation rate (v 2 = 6.72, P \ 0.01), moreover no relation between the sex and hepatitis B reactivation rate. There were differences about inflammation grade and fibrosis stage between HBeAg positive and anti-HBe positive group cases (Kruskal-Wallis test, v 2 = 8.6822, P = 0.0032; v 2 = 6.8382, P = 0.0089), showing inflammation grade and fibrosis stage of anti-HBe positive group was higher HBeAg positive group. There were no obvious differences about inflammation grade between age \40 years old and C40 years old group cases (v 2 = 0.62, P [ 0.05), but significant statistical differences about fibrosis stage (v 2 = 7.37, P \ 0.01), showing fibrosis stage of C40 years old group cases was clearly higher \40 years old group cases. 56 cases received liver biopsy for the second time and 23 for the third time. We find that whose pathology of hepatic tissue was normal kept stable for several years while abnormal ones could hardly get recovery or occur light change. The HBsAg darkening rate per year is 1.55% and the HBeAg darkening rate per year is 5.4%. Conclusion: The hepatic tissue pathology for most chronic HBV carriers (55%) had significant abnormity (inflammation grade CG 2 ), and whether falling to hepatitis B reactivation was high relevant to the liver inflammation grade (G) and dissimilar age group.

Iran) were closer than the distances between Xinjiang and European countries.

Conclusion: This study suggested that the predominant HBV strain circulated in Uigur patients of Xinjiang was genotype D, and the dominant subgenotype was D1.

Background: HBV infection is endemic in Mongolia. So, HBV vaccination introduced as mandatory immunization in newborn since 1991. Objective: To assess the epidemiological situation and vaccination efficacy of HBV infection. Methods: We are randomly, matching by age and sex, selected 337 ''healthy'' subjects, who is living in Ulaanbaatar (1-60 ages), in May 2009. In serum of all subjects tested HBsAg (by ELISA) and HBsAb who is below 16 age. Positive serum for HBsAg also tested HBeAg and anti-HBe. The results compared with analogical data obtained in 2002.

Year 2009 There was tested positive for HBeAg is in two and positive for anti-HBe in 13 subjects from 29. Conclusions: After introduction of HBV vaccination, the epidemiological situation of HBV infection is significantly improved. But, in last 6 years, has not firmly extended improvement of HBV vaccination efficacy.

Hepatol Int (2010) 4:94-345 99

Background: Natural killer (NK) cells have capacity to mediate liver injury in viral hepatitis. This study aimed to characterize hepatic NK cells and defined their functional roles in liver pathology in patients with chronic hepatitis B virus (HBV) infection. Methods: Intrahepatic and peripheral NK cell frequency, phenotype and function were comprehensively investigated in 46 immune activation (IA) patients with chronic hepatitis B (CHB) and 24 immune tolerant (IT) carriers, as well as 21 healthy individuals. Flow cytometric, immunohistochemical, and immunofunctional assays were performed to analyze the impact of NK cell function on liver pathogenesis. Results: Hepatic NK cells in vivo were preferentially accumulated in liver necrotic regions of IA patients where they were activated and skewed towards to cytotoxicity but without an accompanied increase in interferon-c production; by contrast, they were less seen in the fibrotic than non-fibrotic region in the livers of these IA patients. Importantly, the up-regulation of hepatic NK cell cytotoxicity was preferentially dependent on the increased interleukin (IL)-12, IL-15 and IL-18 and reduced IL-10 cytokine expression seen in vivo in livers of IA patients, and was found to be closely associated with the severity of liver injury and fibrosis in these patients. In vitro, the exposure of NK cells from healthy subjects to HBV-induced IL-12, IL-15 and IL-18 leads to increased NK cell activation and CD107a degranulation.

Conclusions: Activated NK cells in vivo were enriched in the livers and skewed to cytolytic activity in IA patients. These findings therefore defined the role of hepatic NK cells in HBV-mediated liver pathogenesis, and may allow the rational development of immunotherapeutic strategies that limiting or blocking liver inflammation and fibrosis.

Benjamin Cowie 1,2,3 , Heath Kelly 1,3 1 Victorian Infectious Diseases Reference Laboratory, North Melbourne Victoria Australia, 2 Victorian Infectious Diseases Service, Royal Melbourne Hospital, Parkville Victoria Australia, 3 University of Melbourne, Parkville, Victoria, Australia

Background: The prevalence of chronic hepatitis B virus (HBV) infection in Australia is increasing, as is the burden of complications such as cirrhosis and hepatocellular carcinoma. Methods: We constructed a mathematical model of HBV infection in Australia which simulated acute and chronic HBV infections in Australia from 1951 to 2050, attendant mortality, and the impact of existing vaccination programs which are the cornerstone of our public health response. Results: The model estimates that in 2009 the number of people living with chronic HBV in Australia will reach 160,000, increasing by another 20,000 over the next decade. Fewer than 60% of these infections have been diagnosed. Lifetime attributable mortality in Australians with chronic HBV is estimated to be 22.5%, resulting in over 20,000 deaths in the first half of this century. Annual acute infections are expected to fall 75% by 2050 with current immunisation coverage, averting approximately 800 attributable deaths at the cost of vaccinating 18 million people with 65 million doses of vaccine. HBsAg prevalence in Australia in 2050 is predicted to be 0.74% with current vaccination levels, compared with 0.78% with no vaccination at all. Conclusions: This model, validated using external data, suggests a significant and growing burden of chronic HBV in Australia, a burden which no domestic vaccination program can address, and of which our surveillance system has detected less than two thirds. Suggestions for additional control measures derived from model outcomes will be presented, as will recommendations for further research and policy priorities.

disease progression.Therefore, IFN-k mightbe a predicator for HBV replication and a promising drug for HBV antiviral therapy. The age-specific incidence density (ID) was calculated and comparison was performed. For vaccinated cohort, a case-control study was conducted through data linkage of NNSS and National Immunization Informative System (NIIS). Each case of AHB reported to NNSS during 1990-2008 was residency and age matched with four controls randomly selected through household registration information. Results of maternal HBV screening at pregnancy and records of HBV vaccination in infancy were analyzed for case and control subjects. Results: The ID of AHB for unvaccinated and vaccinated cohorts was 3.62 and 0.24 persons/10 5 person-years, respectively. The estimated aged-specific ID among subjects aged 15-24 years for the two cohorts was 0.55 and 3.07 persons/10 5 person-years, with a ID ratio of 0.18 (95% CI: 0.15-0.22) in vaccines in comparison to unvaccinated cohort. For those who were born after the implementation of universal infant HBV immunization program, maternal HBV carrier status at pregnancy (odds ratio: 2.59, 95% CI: 1.54-4.34) or incomplete HBV vaccination history (odds ratio: 2.89, 95% CI: 1.29-6.40) was associated with an increased risk of AHB. Conclusion: Implementation of county wide at birth HBV immunization programs for more than 24 years has resulted in preventing the occurrence of AHB among adolescents and young adults in Taiwan.

Results: Six (15.4%) patients were treated by interferon, 25 (64.1%) were done by lamivudine, and 8 (20.5%) were done by more than two antiviral medication. The mean duration of antiviral treatment was 44.0 months. After HBsAg seroclearance, 35 patients (89.7%) had normal alanine aminotransferase levels.

Sixteen patients (41.0%) with initially HBeAg(+) had acquired HBeAg seroconversion in 14 (87.5%) at mean 41.1 months after initiation of treatment. The mean duration since HBeAg seroconversion to HBsAg seroclearance was 76.2 months. Twenty-six (66.7%) patients had undergone HBsAg seroconversion (HBsAb positive), and 13 (33.3%) had only HBsAg loss. Mean HBV viral load was 2.82 9 10 7 IU/mL before initiation of antiviral treatment. HBV-DNA showed positive results in two patients (5.1%) after HBsAg seroclearance in treated patients by lamivudine. One patient of these had a HBsAg reversion 8 months after HBsAg loss. Liver cirrhosis and hepatocellular carcinoma did not develop in any of the 26 patients without evidence of liver cirrhosis at the time of HBsAg seroclearance. Conclusion: The outcome following HBsAg seroclearance after antiviral treatment is excellent and continuous acquisition of HBsAg seroclearance shoud be monitored with HBsAb and HBV-DNA level. Background: To investigate the incidence and distribution of spontaneous decrease of HBV DNA level in chronic hepatitis B and Liver cirrhosis (CHB/LC) patients. Method: The definition of spontaneous decrease of HBV DNA level means that serum HBV DNA level of CHB/LC patients spontaneously decreases to 1% (decrease of 2log 10 ) or less than 1% of the previous level (C100,000 copies/ mL) within 12 weeks without any antiviral and immunoregulatory drug. The incidence, demographic features and distribution of spontaneous decrease of HBV DNA level in CHB/LC patients with different clinical types were studied with prospectively epidemiological methods. Results: Among all the CHB/LC patients (n = 315) with previous HBV DNA level C100,000 copies/mL, spontaneous decerase of HBV DNA level occurred in 171 (54.3%) patients within the observation period of 12 weeks, including 61 (19.4%) patients with undetectable HBV DNA level (\1,000 copies/mL), and 110 (34.9%) patients with detectable HBV DNA level, but decrease more than 2log 10 s. Stratified by age and clinical types, the incidence (25.0%, 2/8) of spontaneous decrease of HBV DNA level in juvenile patients (B19 years) was lower than that (58. 6%, 116/198) in young patients (20-39 years), with significant difference (v 2 = 2.956, P = 0.048), and the incidence (78.7%, 48/61) of spontaneous decrease of HBV DNA level in patients with hepatic failure was higher than that (48.4%, 123/254) in patients without hepatic failure (v 2 = 5.528, P = 0.002). The incidence (75.0%, 21/28) of spontaneous decrease of HBV DNA level in patients with HEV/HBV co-infection was higher than that (51.8%, 146/282) in patients with HBV infection alone (v 2 = 5.530, P = 0.019). Conclusion: Spontaneous decrease of HBV DNA level is a quite common phenomenon during the natural progressing courses of CHB/LC patients, especially in patients with hepatic failure and those with HEV/HBV coinfection. In this situation, the efficacy of antiviral therapies may be overestimated.

Loss of HBsAg Antigen in 894 HBsAg+ Patients during Long Term Follow Up Hasan Ozkan 1 , Mustafa Yakut 1 1 Ankara University Faculty of Medicine, Cebeci Ankara/Turkey

Background: Clearance of HBsAg has been associated with very low or undetectable HBV DNA. HBsAg seroclearance results in an improvement of liver biochemistry and hepatic necroinflammation. The aim of this study is to evaluate the rate of loss of HBsAg in patients with chronic hepatitis B treated with antivirals and/or interferon treatment and in the inactive HBsAg positive carrier patients without treatment during follow up. Method: We followed the ALT, HBVDNA and HBsAg levels of 894 HBsAg+ patients during 6-12 months intervals.HBVDNA levels were calculated with real time PCR method. Liver biopsy was performed to the patients who are in need of treatment.

Results: Mean age of the patients were 42 (±14.5) years old. Patients were followed 3-140 months (Mean follow up time was 51 months). Mean ALT was 86 IU/ml at the beginning. Among patients 215 (24%) of them had HBVDNA (PCR) C10 4 copy/ml and were given treatment. Of those patients who are treated, 66 of them had only interferon based regimen [52 (78.7%) of them HBeAg-] and 149 of them had nucleos(t)ide analogue alone or in combination. 679 (75.9%) patients were regarded to be inactive carriers, and were followed without treatment. HBsAg loss was seen in 51 (5.7%) patients among 894 HBsAg+ cases. HBsAg loss rate in treated patients was (22/215) 10.2%. Also HBsAg loss rate in treated patients with interferon was (14/66) 21.2%. On the other hand, HBsAg loss rate was 31/679 (4.56%) among inactive carriers. Conclusion: In this study we find out that HBsAg loss occurs so often in patients with chronic hepatitis B treated with antiviral and/or interferon treatment and even in the inactive HBsAg positive carriers. Our results show that HBsAg loss was statistically more in patients who are treated, compared to those who are not treated.

(Fibroscan, FS) and fibrotest (FT) have become popular methods to assess liver fibrosis. The aim of this study was to assess the performance of FS and FT to differentiate inactive hepatitis B carriers from HBe Ag-negative HBV disease. Patients and Methods: Sixty-four patients (mean age: 56 years; male/female ratio: 1/1) considered inactive hepatitis B carriers (permanently normal ALT and HBV DNA undetectable or\10,000 ui/ml) were evaluated in the same day by liver biopsy, FT and FS. The histological assessment was compared with the results of non-invasive methods. Results: Histological assessement confirmed inactive hepatitis B carriers state in 44 (68.6%) patients (HAI \ 3, F \ 1) and shown morphological features suggestive for chronic hepatitis in 18 (28.3%) patients and for liver cirrhossis in the remaining 2 (3.1%) patients. Median FS and FT values were significantly lower in inactive biopsy proved hepatitis B than in HBe-negative chronic HBV patients (4.6 vs.7.4 KPa, respectively, p \ 0.001 and 0.18 vs. 0.38, respectively, p \ 0.001). Conclusion: Both FS and FT are simple non-invasive and effective methods to differentiate the inactive hepatitis B carriers from HBeAg-negative chronic HBV disease and could assist therapeutic decisions.

Hong-Tao Xv 1 , Tong-Jing Xing 1 , Hao Li 1 , Jun Ye 1 1 Taizhou People's Hospital, 399 Hailing South Street, Taizhou, China

The possible role of different regulatory T cell populations in HBV persistence has been suggested, especially CD4 + CD25 + T regulatory cell. However, there is discrepancy on the percentage of CD4 + CD25 + T regulatory cell in chronic hepatitis B infection in recent reports. Lack reliable markers is one of the reasons. Three kinds of common markers of regulatory T cells were used to detect the numbers of regulatory T cells by flow cytometry in healthy controls and patients with chronic hepatitis B. No significant difference was found between gated with CD25 hi and CD25 + FoxP3 + . However, significant difference were found between gated with CD25 + CD127 low/and CD25 hi or CD25 + FoxP3 + in healthy controls and patients with chronic hepatitis B. Patients with asymptomatic HBV carriers, inactive HBsAg carrier and chronic hepatitis B showed a significantly higher percentage of Treg compared with healthy controls (8.56 ± 2.01, 8.74 ± 3.04, 10.7 ± 2.93 vs.7.42 ± 1.28; F = 20.6, P = 0.00). These results supported that (1) there is certain discrepancy by different markers to detect the CD4 + CD25 + T regulatory cells, might be affect the study and isolation of CD4 + CD25 + T regulatory cells. (2) the frequencies of T reg cells were increased in the different stages of chronic hepatitis B infection, might be association with the disease activity and contributed to prevent the extensive liver injury.

(GG + GA: OR = 0.371, , p = 0.007; Allele G: OR = 0.445, , p = 0.008) were statistically significant. But they weren't between anti-HBe positive group and normal control group (GG + GA: OR = 0.795, , p = 0.499; Allele G: OR = 0.914, 95%CI: 0.547-1.527, p = 0.731). Conclusions: Allele G on SNP rs2660 of OAS-1 gene maybe benefit patients with chronic HBV infection to achieve spontaneous HBeAg seroconversion. Genotyping on this SNP may be predicting valuable for interferon therapy for chronic HBV infection.

Mongolia is a country with high prevalence of Hepatitits B infection. 10% of the population is estimated to be positive for HBsAg. Hepatitis virus is the leading cause of cancer related death in Mongolia with HBV being responsible for half the incidence. Our study is the first of the kind that combines data on HBV viral load and other serology markers in Mongolian patients. Aim: We are committed to unveil which of the diagnostic modalities are important for clinicians to assess the current state of liver disease for chronic hepatitis B patients. Method: Statistical analysis was performed in 300 randomly selected patients who were tested for HBV viral load at Happy Veritas, Clinical Diagnostics Laboratory. Diagnostic methods by Taqman Real-Time PCR, HBV-combo rapid tests, biochemistry, CBC were used in the study. Result: 300 patients with positive HBsAg were tested for viral load. Only 40.6% of patients had detectable viral load of [500 copies/ml. Serum HBV DNA negativity rate was 58% in cirrhotic patients, whereas 51% was negative in non-cirrhotic patients with no significant difference between the two. HBeAg positivity was 5.3% among HBsAg positive patients. Though the mean age of HBV positive patients were 38, it was 22 for HBeAg(+) patients. Summary: HBV viral load is not important for estimating progression of chronic liver disease. Though viral load is important to assess response to treatment in chronic HBV patients, HBV DNA positive patients make only 40% of HBsAg positive patients. It is difficult to assume that HBV DNA negative patients are healthy carriers because they still have ALT elevations and liver fibrosis. In such cases HBsAg quantification could be of use to those patients. HBsAg is hardly used to assess treatment response, but there is immense pressure to develop sensitive, wide range reproducible assays for that purpose. Though HBeAg positivity has important role in assessing HBV infection and in decision making in western countries, it is not the case in Mongolia, like other Asian countries. HBeAg(+) patients make only a small portion of HBV infected population.

Scoring Objective: The study aimed to develop clinically useful scoring systems based on laboratory tests to predict the probability of acute hepatitis B (AHB) among HBV-related acute hepatitis patients whose prior history of HBV infection was unknown.

Methods: The independent predictors employed to construct the scoring systems were identified by multivariate analysis from laboratory variables, which were selected by univariate analysis between patients with acute selflimited hepatitis B (ASL-HB) and chronic hepatitis B (CHB) with an acute exacerbation (CHB-AE).The scores assigned to the predictors were determined empirically and arbitrarily by referring to estimated regression coefficients.

Results: Multivariate analysis identified IgM anti-HBc, HBV DNA/HBeAg-/+ and prothrombin time (PT) as independent predictors for AHB. Scoring System I was constructed by using of IgM anti-HBc and HBV DNA/HBeAg-/+, and Scoring System II by adding PT into the algorithm of the Scoring System I. The areas under of receiver operating characteristics curves of IgM anti-HBc, the Scoring Systems I and II were 0.960 (95% CI: 0.934-0.985), 0.989 (0.979-0.998), and 0.990 (0.982-0.999) (chi square = 10.28, P = 0.006), respectively. Positive likelihood ratios of the two models were 34.8 and 27.6, respectively. Applied in validation set patients, two models of the Scoring Systems had the same powers for identifying CHB, but the Scoring System I correctly identified more patients with ASL-HB than did the Scoring System II (80/81 vs. 76/81). Conclusions: The scoring systems constructed by serum IgM anti-HBc and HBV DNA/HBeAg-/+, or combination with prothrombin time were clinically useful in identifying AHB. The Scoring System I is more practicable than IgM anti-HBc alone and Scoring System II in differential diagnosis between AHB and CHB-AE. study was to evaluate the role of maternal data during perinatal period as well as the role of serological and virological cord blood data on SPB among 102 HBeAg(-) chronic HBV infected pregnant women, without several known risk factors for preterm delivery. Methods: Haematological, biochemical and serological parameters were calculated using commercially available automated techniques. Serum HBV-DNA was determined by using the Cobas Amplicor HBV Test (lower limit of quantification: 300 copies/ml) in both maternal (n = 76) and cord blood (n = 73) available samples. Results: Fifteen women exhibited SPB (14.7%). The presence of SPB was not related with any maternal parameter [age (p = 0.096), HBV-DNA positivity (p = 0.478), haemoglobin (p = 0.122), platelets (0.735), AST (p = 0.337), ALT (p = 0.178)], except of maternal absolute lymphocyte count (p = 0.007). A significant association between SPB and HBV-DNA presence in cord blood was observed (p = 0.007). HBV-DNA positivity in cord blood was significantly associated with maternal viremia (median maternal HBV-DNA was 147.543 copies/ml in women with HBV-DNA positive cord blood vs. 1.646 copies in women with HBV-DNA negative cord blood, p = 0.002). The relative risk of HBV-DNA presence in cord blood was 6.43 times higher among women with serum HBV-DNA [10.000 copies/ml and lymphocyte count \1,500 compared to those with all the other combinations of both parameters (p = 0.001).

Conclusions: The presence of HBV-DNA in cord blood is significantly associated with SPB in HBeAg(-) chronic HBV infected pregnant women. Women with HBV-DNA [10.000 copies/ml and absolute lymphocyte count \1,500 during perinatal period have the higher probability of HBV-DNA presence in their cord blood.

Backgrounds/aims: Although liver stiffness measurements (LSMs) were useful for predicting fibrosis stage, predictive accuracy was not adequate to replace liver biopsy, especially in patients with chronic hepatitis B (CHB). This study was performed to establish a noninvasive method to avoid unnecessary liver biopsy in patients with CHB. Methods: Consecutive patients with CHB who were admitted for liver biopsy were enrolled. Patients underwent LSMs and blood tests on the same day as the liver biopsy. Results: One-hundred eighty patients were randomly classified to the training set (n = 120) or validation set (n = 60). AUROC of LSM for predicting significant fibrosis was 0.83 (optimal cutoff value 7.2 kPa, sensitivity 85.9%, specificity 67.9%). Hyaluronic acid, a 2 -macroglobulin, and LSMs were significant factors for predicting significant fibrosis on multiple regression analysis. With this model, although 50% of patients in training set could avoid liver biopsy, 8.3% of patients in validation set were misdiagnosed. To establish more accurate criteria, patients in training set were classified into two groups according to their LSMs. a 2 -macroglobulin and hyaluronic acid were the only significant factors for predicting significant fibrosis in low-LSM group (\7.2 kPa) and in high-LSM group (C7.2 kPa), respectively. In low-LSM group, no patient with a 2 -macroglobulin \160 mg/dL had significant fibrosis; in high-LSM group, all patients with hyaluronic acid C27 ng/mL had significant fibrosis. With this algorithm, 51.7% in training set and 55% in validation set could avoid liver biopsy without misdiagnosis. Conclusions: Our algorithm using LSMs, a 2 -macroglobulin, and hyaluronic acid suggests an accurate criterion for avoiding unnecessary liver biopsy. Background: To investigate the HBV DNA load in peripheral blood mononuclear cells (PBMCs) during the course of treatment patients with chronic hepatitis B (CHB), and to explore which relationship with serum HBV DNA load.

Methods: According to serum HBV DNA quantitative results, 68 cases of CHB patients were divided into four groups (À C10 7 ,`\10 7 to C 10 5 ,´\10 5 to C 10 3 ,ˆ\10 3 ). HBV DNA load in PBMCs was detected by fluorescent quantitative real-time PCR. Results: HBV-DNA in PBMCs of the fourth group were lower than the detection limit, and the Logarithmic values of PBMCs HBV-DNA from the other three groups (copies/10 6 PBMCs) were 4.60 ± 0.91; 4.31 ± 0.44; 2.80 ± 0.78. There was no significant difference between the first group and that of second group (P [ 0.05). The third group was lower than the form two groups (P \ 0.01).

Conclusions: The load of serum HBV-DNA and HBV-DNA level in PBMCs was no significant correlation. HBV-DNA in PBMCs was not detected while HBV-DNA load in serum drop to below detection limit after treatment. Our data indicate the PBMCs HBV DNA load could be used as indicators to treatment efficacy in patients with CHB. Background/Aim: To explore peripheral T-lymphocyte subpopulation profile and the variation of serum interferon-c (IFN-c) and interleukin-4 (IL-4) levels in patients with chronic HBV infection. Methods: Distribution of T-lymphocyte subpopulations in peripheral blood were measured by flow cytometry in patients with chronic HBV infection. The cytokines IFN-c and IL-4 levels were examined by ELISA. HBV markers were detected with ELISA. Serum HBVDNA load was assessed with PCR. The results of peripheral T-cell subsets and serum levels of IFN-c and IL-4 were compared among groups based on HBeAg express status and the quantities of HBVDNA. Results: Chronic HBV infection patients had significantly decreased CD3 + cells, CD4 + cells and CD4 + /CD8 + ratio, and increased CD8 + cells compared with unifected controls, all with P \ 0.001. CD4 + cells and CD4 + /CD8 + ratio were significant lower (P \ 0.05), and CD8 + cells was obvious higher (P = 0.01) in chronic hepatitis B (CHB) patients than those in asymptomatic chronic HBV carriers (ASC). There was a trend of decreasing CD4 + cells and CD4 + /CD8 + ratio and of increasing CD8 + cells in the order of control and ASC and CHB. Comparing with HBVDNA/HBeAg-negative group respectively, the ASC patients with HBVDNA/HBeAg-positive had significant decreased CD4 + cells and CD4 + /CD8 + ratio and increased CD8 + cells (P \ 0.01). CHB patients with HBVDNA/HBeAg-positive had significant decreased CD4 + /CD8 + ratio and increased CD8 + cells (P \ 0.05). CHB patients had significant increased serum IFN-c level and IFN-c/IL-4 ratio than those in control group and ASC patients (P \ 0.01, respectively). There was a trend of increasing of serum IFN-c levels and IFN-c/IL-4 ratio in the order of control and ASC and CHB. No significant differences were observed for serum levels of IL-4 and IFN-c and the IFN-c/IL-4 ratio between HBVDNA(+)/HBeAg(+) and HBVDNA(-)/HBeAg(-) patients, respectively. Conclusion: The disorder of host cellular immunity in chronic HBV-infected patients would be caused by HBV infection, and the increasing HBVDNA replication and HBeAg express could make the disorder advanced and the more severe liver injury.

MS. Cellular immunfluresence and cytometry assay were employed to determine the membrane-binding property of antibodies in mice sera. Results: Total 46 autoantigens recognized by autoantibodies in mice sera were identified by MS/MS. Among the antigens identified, mitochondrium was the most frequent auto-antigen. Immunofluorescence and cytometry assay demostrated that the auto-antibodies in sera of HBsAg positive mice bound to the membrane of the liver. Using an HBsAg expression cell line, Ss1, it was found that HBsAg promoted some sequestered autoantigens identified in this study to secrete into the supernatant of cell culture and co-existed with HBsAg, which could be apt to trigger autoimmune reaction. Conclusions: The repertoire of autoantibodies in an HBsAg transgenic mice were obtained, and a possible mechanism was demostrated that expression of HBsAg was associated with the occurrence of autoantibodies.

Background/Aims: A promising risk prediction score for hepatocellular carcinoma (HCC) remained to be derived and validated in different international cohorts. This study aimed to develop and validate a novel risk scoring system for the prediction of the risk of HCC in patients with chronic hepatitis B.

Methods: A risk score to predict 3-, 5-, and 10-year HCC risk was developed using a multivariate Cox proportional hazards model from 3,584 cirrhosis-free participants in the community-based REVEAL-HBV study cohort in Taiwan. The risk score was externally validated in a composite cohort consisted of hospital-based patients from Hong Kong and Korea (N = 1, 505) . The validation was done by calculating the area under receiver operating curve (AUROC) and testing the calibration of predicted HCC risk with observed HCC risk. Results: There were 131 newly developed HCC cases in the risk score developing cohort. Five variables including gender, age, serum ALT level, HBeAg status, and serum HBV DNA level were significantly associated with HCC risk and incorporated in a risk score. The predicted 3-, 5-, and 10-year HCC risk ranged 0.006-15.2%, 0.01-32.4%, and 0.04-64.4%, respectively, for those with the lowest (score = 0) to highest (score = 16) HCC risk. In the validation cohort, 111 patients developed HCC in a mean follow-up of 5.3 (maximum 11.2) years. The risk score had AUROCs of 0.811, 0.796, and 0.769, respectively, to predict 3-, 5-, and 10-year risk of HCC in the validation cohort; and AUROCs of 0.902, 0.783, and 0.806, respectively, after excluding cirrhotic cases (N = 277). The risk score-predicted HCC risk was well calibrated with the Kaplan-Meier-observed HCC risk.

Conclusions: This study proposed and validated a simple 16-point risk score of baseline clinical variables to estimate the risk of developing HCC with specified time frames. Aims: To investigate the dynamic changes of liver stiffness measurements (LSM) by FibroScan Ò in patients with chronic hepatitis B (CHB) flare and evaluate the relationship between LSM and alanine aminotransferase (ALT) and bilirubin. Methods: Forty-nine consecutive treatment naïve patients with CHB flare [ALT [5 times upper limit of normal range (ULN)] were enrolled and divided into two groups: group Child-Turcotte-Pugh (CTP) A (n = 14) and group CTP B/C (n = 35). Fibroscan Ò was performed every 7-10 days during hospitalization and every 1-3 months for follow-up outpatients. Hematology and liver function were tested at the same time. Liver biopsy was performed during hospitalization for 11 patients. Results: Total of 167 LSM were performed. Overall, a statistically significant, positive correlation was found between LSM and ALT, LSM and bilirubin, whereas a positive partial correlation was only found between LSM and bilirubin (r = 0.468, P \ 0.001). Following with continuously declined ALT and bilirubin, LSM fell down to lower than cirrhosis cut-off 13.4 kPa in 10 patients with high initial LSM and without clinical signals or typical images of cirrhosis (4 patients proved by biopsy) (Fig. 1) . Whereas, LSM remained larger than 13.4kPa in 10 patients, including 4 biopsy proved patient (Fig. 2 ), 3 patients with clinical sign or images of cirrhosis. In patients with ALT normalization, bilirubin and CTP score were increased in patients with LSM larger than 13.4 kPa (P \ 0.001). Conclusion: Although influenced by ALT, LSM was majorly affected by bilirubin in patients with CHB flare. In CTP A patients, dynamic evaluation of LSM associated with bilirubin and ALT was essential to improve accuracy of FibroScan Ò for cirrhosis diagnosis.

replication error rate of 3 9 10 -5 (1/3.3 9 10 4 ) by the viral polymerase. Interestingly, the mutant/WT ratio in nine treatment-naïve (never been treated) HBeAg positive patients is in the range of 1/10 5 -1/10 7 . All nine individuals harbor mixtures of YIDD and YVDD mutants, some of them confirmed by sequencing of the cloned PCR products. The rt204ATA (YIDD) is the most prevalent variant in treatment naive patients, and is the only variant detected in the cultured cells. The mutant/WT ratio for the precore and BCP mutations in the same patients is more variable and often higher than that for YMDD. Conclusion: Our data suggest that the rate of YMDD mutant generation in tissue culture and in treatment naïve individuals may be dictated mostly by viral factor(s). The rate at which these mutants increase in people following introduction of antiviral agents may reflect host contributions to the process and the degree of liver disease. Since YMDD mutations confer resistance to lamivudine, the extent to which this information can be used to predict the time at which drug resistance breakthrough occurs, and other disease pathologies, will be discussed.

Nuclease Background: Identical blood samples tested using different kits can give markedly different hepatitis B virus (HBV) DNA levels, which can cause difficulty in the interpretation of viral load. A universal double-stranded DNA control or standard that can be used in all commercial HBV DNA nucleic acid amplification kits is urgently needed. Methods: By aligning all HBV genotypes (A-H), we found that the surface antigen gene and precore-core gene regions of HBV are the most conserved regions among the different HBV genotypes. We constructed a chimeric fragment by overlapping extension polymerase chain reaction and obtained a 1,349-bp HBV C+S fragment. We then packaged the fragment into lambda phages using a traditional lambda phage cloning procedure. Results: The obtained armoured DNA was resistant to DNase I digestion and was stable, noninfectious to humans, and could be easily extracted using commercial kits. More importantly, the armoured DNA can be used with all HBV DNA nucleic acid amplification kits. Conclusions: The lambda phage packaging system can be used as an excellent expression platform for armoured DNA. The obtained armoured DNA possessed all characteristics of an excellent positive control or standard. In addition, this armoured DNA is likely to be appropriate for all commercial HBV DNA nucleic acid amplification detection kits. Thus, the constructed armoured DNA can probably be used as a universal positive control or standard in HBV DNA assays. The pathogenesis of HBV-associated liver failure is very complicated and it has achieved great progress in recent years. On the recent literature, combining with the novel findings of our research team, here, we summarized the pathogenesis of HBV-associated liver failure as follows: (1) both host and viral factors are contributed to the pathogenesis of HBV-associated liver failure; (2) polymorphisms G-201A from CXCL10, -592C and -819C from IL-10 are closely related to hepatitis flares and liver failure; (3) genotype C and B, particularly B/C co-infection, are correlated with liver failure; (4) quasispecies studies during antiviral therapy by using NAs is quite important; (5) attention should be paid to missed detection in HBV DNA by real-time PCR.

Expression and Purification of a Functional Human HBV Polymerase Yu Yang 1 , Dipendra Raj Pandeya 1 , Seong-Tshool Hong 1 1 Department of Microbiology, Medical School, Chonbuk National University, Chonju Chonbuk 561-712, South Korea

Hepatitis B virus polymerase plays an essential role in the life cycle of HBV. Biochemical analysis of human HBV polymerase is hampered by difficulties in obtaining pure and active recombinant human HBV-Pol (rhHBV-Pol) . In this study, we developed a method for the efficient large-scale purification of fully active rhHBV-Pol without the simultaneous overexpression of cellular factors. Full length rhHBV-Pol tagged with a 5 0 polyhistidine tag (843 amino acids) was expressed at a high level in an Escherichia coli system. SDS lysis buffer was utilized to dissolve insoluble rhHBV-Pol, and Ni-NTA resin affinity chromatography was utilized for rhHBV-Pol purification. The majority of rhHBV-Pol was eluted with 100 mM imidazole in the presence of NP-40, which is a weak detergent that was used to keep dissolved rhHBV-Pol insoluble. A high-concentration reducing agent was utilized throughout purification steps to keep soluble rhHBV-Pol from forming disulfide bonds. Purified rhHBV-Pol showed both stable reverse transcriptase activity and DNA polymerase activity. The purified protein was highly pure and had stable reverse transcriptase activity. The ability of this purification approach to yield rhHBV-Pol in an enzymatically active form may facilitate the development of novel therapeutics for the treatment of chronic Hepatitis B.

Chien-Hung Chen 1 , Chuan-Mo Lee 1 , Chao-Hung Hung 1 , Tsung-Hui Hu 1 , Jing-Houng Wang 1 , Sheng-Nan Lu 1 , Chi-Sin Changchien 1 1 Chang Gung Memorial Hospital-Kaohsiung Medical Center, Chang Gung Memorial Hospital-Kaohsiung Medical Center, 123 Ta Pei Road, Kaohsiung, Taiwan

Background and Aims: This study investigated the risk predictors of reactivation of hepatitis B virus (HBV) following hepatitis B e antigen (HBeAg) seroconversion in patients with or without interferon-based therapy. Patients and Methods: One hundred and sixty-nine HBeAg-positive chronic hepatitis B patients were recruited for this study including 119 with spontaneous HBeAg seroconversion and 50 with interferon (n = 16) or peginterferon (n = 34) induced HBeAg seroconversion. The HBV genotypes and the sequences core promoter and precore regions were determined before HBeAg seroconversion. Results: Of the 169 patients, 111 males and 58 females with a mean age of 35.1 ± 10.7 years. Cox regression analysis revealed that older age at HBeAg serovoncersion, interferon-based therapy, HBV genotype C, ALT [200 IU/ml before HBeAg seroconversion, and A1899 mutant were significantly associated with the reactivation of HBV following HBeAg seroconversion. In patients without interferon therapy, age at HBeAg seroconversion, male and ALT [200 IU/ml before seroconversion were independent factors for reactivation of HBV following HBeAg seroconverison. In patients with interferon therapy, HBV genotype C and A1899 mutant were independent factors for reactivation of HBV. In patients without interferon therapy, age at HBeAg serovoncersion and T1762/A1764 were independent factors for developing liver cirrhosis. Moreover, T1762/A1764 was a significant risk factor for HCC development.

Conclusions: Patients with IFN-based therapy had a higher rate of reactivation of HBV following HBeAg seroconverion than those with spontaneous HBeAg seroconversion. The present of T1762/A1764 mutant before HBeAg seroconversion was a useful molecular marker for predicting the clinical outcomes in patients with spontaneous HBeAg seroconversion.

Top 10% Posters Background: Patients with genotype B HBV infection have lower serum ALT level, and earlier HBeAg seroconversion than genotype C. Whether this different manifestation is related to the HBV genomic evolution remains unknown. We thus investigated the HBV genomic evolution rate in HBeAgpositive patients during different stages of HBV infection. Methods: We enrolled 25 HBeAg-positive patients (genotype B in 13 and genotype C in 12) with a mean follow-up period of 3 years: 7 had persistently low normal serum ALT level (group I, immune tolerance phase), 9 had high normal or minimally elevated serum ALT level (group II, early immune clearance phase), and 9 developed active hepatitis B (group III, immune clearance phase). The paired dominant full-length viral genome at enrollment and last follow-up of each patient was compared according to HBV genotype and serum ALT level by DNaSP 5.0 software and Mann-Whitney test. We counted the evolution rate in full genome (Pi), nonoverlapping region as synonymous (Ks) and non-synonymous (Ka), and overlapping region as K.

Results: The evolution rate was low in group I patients (1.35 9 10 -5 ± 3.58 9 10 -5 nucleotide substitution/site/year), and significantly increased in group II (1.57 9 10 -3 ± 2.10 9 10 -3 , p \ 0.001) and III patients (2.48 9 10 -3 ± 3.27 9 10 -3 , p \ 0.001). Ka/year of genotype C was higher than genotype B in group II patients (p = 0.05), and Ks/year of genotype B was higher than genotype C (p \ 0.05). A positive correlation (R 2 = 0.5841) between Ka/year of genotype B and the area under the average ALT-time curve was found. Conclusions: The HBV genomic evolution rate in immune clearance phase was higher than that in immune tolerance phase. In immune clearance phase, the evolution rate of genotype B was significantly higher than that of genotype C. Backgrounds/Aims: The literature for association between X mutations of hepatitis B virus (HBV) and clinical status were few still now. We conducted to examine difference of clinical status according the X gene mutations in patients with HBV genotype C infection.

Methods: X gene of HBV were determined by direct sequencing in 194 patients. We compared 60 patients with chronic hepatitis B to 65 patients with cirrhosis, and 69 patients with HCC, respectively. Results: HBeAg serostatus was associated with clinical status of liver disease. G1386M, C1485T, C1653T, T1753V, A1762T and G1764A mutations were each significantly associated with clinical severity. Prevalence of the X mutations (G1386M, C1653T, T1753V, A1762T and G1764A) were commonly increased as HBV infection progressed from chronic hepatitis status to liver cirrhosis or HCC, but C1485D mutation was high prevalence in the HCC as compared to chronic hepatitis B. In the comparison of X mutation between the liver cirrhosis and HCC, only T1753V mutation was significantly associated with HCC. T1753V, A1762T and G1764A mutations were found more frequently in HBeAg negative than HBeAg positive patients. Conclusions: These findings suggest that the X gene mutations of specific types are contribute to disease progression of chronic HBV infection.

Top 10% Posters Background: To explore the biological function of HBV X deletion mutation in NH 2 -terminus or COOH-terminus and its roles in carcinogenesis of hepatocellular carcinoma. Methods: HBV X gene fragments encoding mutant HBV X protein with deletion 50 amino acid residues at NH 2 -terminus or COOH-terminus (HBxn or HBxc) were amplified from subtype adr of HBV DNA by PCR. The purified HBxn and HBxc gene fragment was inserted into GFP expression vector, pEGFP-C1, to construct recombinant expression vector pGFP/HBxn and pGFP/HBxc. HepG2 cells were transfected with these recombinant constructs by lipofectectin reagent, and HepG2 cell lines stable expressing GFP/HBxn or GFP/HBxc fusion protein were established. The expression of deletion mutant HBV X gene was demonstrated by RT-PCR analysis. Then, the growth rates and cell cycle were determined in HepG2, HepG2/GFP, HepG2/GFP-HBx, HepG2/GFP-HBxn and HepG2/GFP-HBxc cells.

Result: RT-PCR analysis showed that HBxn, HBxc expressed in HepG 2 /GFP-HBxn and HepH2/GFP-HBxc cells. MTT analysis showed that HepG2/GFP-HBx cells and HepG2/GFP/HBxn cells grow faster than HepG2/GFP and HepG2/GFP/HBxc cells. Flow cytometry analysis showed that the percentage of G0/G1 phase in HepG2/GFP-HBx (51.043±3.628)% and HepG2/GFP-HBxc cells (48.143 ± 3.721)% were significant decreased compared with HepG2 (65.767 ± 1.665)%, HepG2/GFP (65.193 ± 2.952)% (P \ 0.05). However, no significant differences was observed among HepG2/GFP-HBxn (67.950 ± 2.128)%, HepG2 and HepG2/GFP cells. Conclusions: Wild HBx and the COOH-terminal region-deleted HBx promote growth and proliferation of hepatocellular carcinoma cell via induction of cell cycle progression. It also indicates that the NH 2 -terminus of HBx is more important in presenting biologic function of HBx in carcinogenesis of hepatocellular carcinoma.

Core Internal Deletions in Spontaneous, Interferon-induced or Lamivudine Treated HBeAg Seroconverter patients Yan Cheng 1 , Peizhen Hu 1 , Bee Leng Seet 1 , Seng Gee Lim 1,2 1 Dept of Medicine, Yong Yoo Lin School of Medicine, National University of Singapore, 5 Lower Kent Ridge Road, Singapore, 2 Dept of Gastroenterology and Hepatology, National University Health System, Singapore, 5 Lower Kent Ridge Road, Singapore

Backgroud: While Hepatitis B virus core deletions have been found in asymptomatic carriers, chronic hepatitis B and hepatocellular carcinoma patients, their role in HBeAg seroconversion is unclear, consequently we aimed to examine their relationship to this event.

Method: Three groups of chronic hepatitis B HBeAg seroconverters (spontaneous, interferon and lamivudine induced) with matching non-seroconverter controls were selected. Ten patients per group and three time points per patient were included. Nested PCR of precore/core gene was performed followed by cloning and sequencing. At least 20 clones/sample were sequenced. Sequences were aligned with ClustalX and the frequencies of patient and sequences with core deletions were calculated.

Hepatol Int (2010) 4:94-345 115 Results: Spontaneous seroconverters showed significantly more core deletion mutations than untreated non-seroconverters at all time points (22 vs. 1.5% at baseline, 17 vs. 2.9% before seroconversion, 3 vs. 0% after seroconversion, p \ 0.05). Interferon induced seroconverters had more sequences with core deletions than inteferon non-responders at the time before seroconversion (22.3 vs. 2%, p \ 0.001). Lamivudine induced seroconverters, and non-seroconverters showed similar low frequency of sequences bearing core deletions at all time points. The frequency of sequences with core deletion mutations decreased drastically after seroconversion in both spontaneous and interferon induced seroconverters (17-3 and 22.3-3%, respectively). Large core deletions occured usually before seroconversion in spontaneous and interferon induced seroconverters with half being in-frame deletions. Conclusions: The core internal deletions may play a role in spontaneous and interferon induced seroconversion but not in lamivudine induced seroconversion. The association between increased core deletions, and spontaneous/ interferon induced HBeAg seroconversion suggests that this could be related to a different mechanism of seroconverion compared to lamivudine induced seroconversion.

Top 10% Posters Background and Aims: Viral infected cells express on their surface MHC-class I/viral peptide complexes (pMHC), which are recognized by specific T cell receptors of CD8+ T cells. Our aim was to produce antibodies with T cell receptor (TCR) like features able to specifically recognize HBV peptide/MHCclass I complexes and distinguish HBV infected hepatocytes from non-infected ones. These antibodies were used to analyze the distribution and quantification of pMHC on HBV infected cells and their ability to deliver compounds to infected cells. Methods: Antibody secreting B-cell hybridomas were generated from mice immunized with HBVc18-27/A201 or HBVe183-91/A201 monomers. The number of pMHC on individual cells was enumerated by flow cytometry based quantification technique, while HBV antigen was quantified by Western blot. Confocal microscopy was used to detect intracellular delivery of fluorescent dye. Results: After screening several hybridomas, five were found producing antibodies that recognize T2 cells pulsed with core 18-27 (2 clones) or env 183-91 peptides (3 clones) but not T2 cells pulsed with irrelevant HLA-A0201 binding peptide. The TCR-like antibodies recognized the pMHC expressed naturally after endogenous processing on HBV infected cells. The binding of the antibodies was not inhibited by the presence of serum HBV antigens. These antibodies were used to quantify pMHC complexes expressed on different cell types. On average, 1 9 10 5 core proteins produced one pMHC on B cells, while in hepatocytes, a single pMHC requires endogenous synthesis of approximately 1.5 9 10 6 proteins, implicating the limited antigen processing ability of hepatocytes. Furthermore, TCR-like antibodies could deliver the fluorescent dye to the endosomal compartment of HBVinfected hepatocytes. Conclusions: We have produced TCR-like antibodies recognizing HBV infected cells. These antibodies open the possibilities to obtain precise quantitative analysis of the pMHC on HBV infected hepatocytes and provide the base for new therapeutic strategies for targeted delivery of drugs to HBV infected cells. Background: As a major therapy for hepatitis B virus (HBV) infection, Interferon alpha (IFN-alpha) triggers intracellular signal transduction including JAK-STAT pathway to produce numbers of antiviral effectors. However, patients with chronic hepatitis B usually show low response to IFN-alpha treatment and the underlying mechanism remains unclear. Methods: HepG2 and HepG2.2.15 cells were used to examine the Type I IFN receptors expression, STAT1 phosphorylation and methylation. STAT1-PIAS1 interaction in cells was tested by protein co-immunoprecipitation. The potential improvement of S-adenosylmethionine in the antiviral effect of IFN-alpha was investigated by measuring the PKR and 2 0 , 5 0 -OAS mRNA levels, together with the viral gene expression in both mRNA and protein levels.

Results: Both chains of the Type I IFN receptors were expressed much higher in HepG2.2.15 cells than in HepG2 cells. HBV inhibited dramatically the methylation rather than the phosphorylation of STAT1, which was consistent with an increased STAT1-PIAS1 interaction. S-adenosylmethionine treatment effectively improved the HBV-caused STAT1 demethylation and attenuated STAT1-PIAS1 binding, followed by increased PKR and 2 0 , 5 0 -OAS mRNA expression. Combined with IFN-alpha, S-adenosylmethionine treatment significantly reduced the HBsAg and HBeAg protein levels and the HBV DNA in the supernatant of HepG2.2.15 cells. Conclusions: STAT1 demethylation and subsequent STAT1-PIAS1 interaction are involved in the mechanism of IFN-alpha antagonistic activity of HBV. By improving STAT1 methylation, S-adenosylmethionine can sensitize in a great degree the antiviral effect of IFN-alpha.

Overexpression of Toll-Like Receptor Background: HBV carriers constitute major reservoir, especially in developing countries. Generally they are recommended for regular check up and treatment is recommended when patients have liver damage. This is mainly attributable to lack of information about extent of liver damage. Aim of this study was to assess extent of liver damage in HBeAg -ve patients who were completely unaware of their HBV infection in Bangladesh. Methods: In this retrospective study, we reviewed the records of 310 HBeAgve CHB patients. They were tested for serologic markers of HBV, anti-HCV and ALT. All underwent per-cutaneous liver biopsy. Result: 77.4% (240/310) were males and 22.6% (70/310) females. Patients were between 13 and 60 years of age. ALT was raised [2 times UNL (cutoff 42 U/L) in 7.1% (22/310). 37.4% (116/310) had high HBV DNA ([10 5 copies/ml by PCR), while low DNA (\10 5 copies/ml) was seen in 62. 6% (194/310) . In those with high DNA, significant necro-inflammation (HAI-NI [ 7) was seen in 46.6% (54/116) and significant fibrosis (HAI-NI [ 3) in 30.2% (35/116). The figures were 31. 4% (61/194) and 14.4% (28/ 194) , respectively, in those having low viral load. None tested positive for HCV infection. Conclusion: Machinery should be developed to characterize undetected HBV carriers in developing countries by multi-center studies. We have shown that considerable number of these patients suffers from progressive liver damage.

Overall strategy of management of CHB should also be revisited.

Hepatol Int (2010) 4:94-345 117

Background/Aims: The mechanisms that regulate hepatitis B virus (HBV) replication within the liver are poorly understood. HBV DNA has been shown to contain CpG islands that are methylated in human tissue, which suggests a role for methylation in regulating viral protein production. Methods: This study was conducted during a 1 year follow-up to characterize the viral kinetics in HBeAg-positive chronic hepatitis B. We count the mutation rate within the period of viral kinetic changes comparing the stationary pattern and fluctuated pattern. Fifty three patients with HBeAg-positive chronic hepatitis B were enrolled. Results: We compared mutation rates of nucleotides and amino acids in different phases between stationary and fluctuated viral kinetic patterns. The fluctuated pattern had statistically significantly higher mutation rate of nucleotides than stationary pattern for the following nucleotide segments, full length genome, core gene, spacer of polymerase gene, negative regulatory element (NRE), NRE gamma domain, and basic core promoter. CpG island 1 (nt55-281) is equal to the partially location of spacer of polymerase. CpG island 2 (nt1,228-1,663 ) is equal to the location of NRE (nt1, 626) and CURS_A (nt1, 704) . CpG island 3 (nt2, 446 ) is equal to the location of terminal protein of polymerase. Conclusions: The fluctuated pattern had statistically significantly higher mutation rate of nucleotides than stationary pattern in CpG island 1 and 2 for the following nucleotide segments. This project will help us to understand the relationship between not only the possible virologic factor and viral kinetics but also the mutation sites and CpG islands methylation and provide potential model for viral kinetic in natural course patients.

HBcAg Th17 cells have been demonstrated to be crucial in host defense against viral infection. However, there has been much less information on the regulation mechanism of Hepatitis B core antigen (HBcAg)-specific Th17 cells in HBV infections. In the present study, Peripheral blood mononuclear cells (PBMC) isolated from chronic hepatitis B (CHB) patients with anti-IL-10 antibody or recombinant IL-10, respectively. The frequency of hepatitis B core antigen (HBcAg)-specific Th17 cells was characterized and produced cytokines were determined by flow cytomix. Our data indicated that HBcAg stimulation promotes the production of IL-17A, IL-22, IL-6, TGF-b and IL-10 in CHB patients, but not in normal control. Furthermore, endogenous IL-10 inhibited HBcAg-stimulated production of IL-17A, IL-22, IL-6, but not TGF-b. Treatment with IL-10 inhibited the HBcAg-stimulated activation of Th17 cells while anti-IL-10 antibody significantly increased the frequency of Th17 and Th1 cells. Hence, HBcAg induced IL-10 negatively regulates Ag-specific Th17 cells immune response in CHB patients and this could represent one example of an evasion strategy by the virus to subvert specific antiviral T cell responses of the host.

Expression and Localization of APOBEC3G in PBMC and Liver Tissue in Different Chronic HBV Infected Patients Hui Chen 1 , Lu-Wen Wang 1 , Xiao-Gang Chu 1 , Shao-Nan Yan 1 , Zuo-Jiong Gong 1 1 Renmin Hospital of Wuhan University, 238 Jiefang Road, Wuchang, Wuhan, China

Background: To study the expression level and intracellular localization of APOBEC3G in PBMC and liver tissue in different chronic HBV infected patients. Methods: By using the western-blot and confocal laser scanning microscope (CLSM), the expression level and intracellular localization of APOBEC3G in peripheral blood mononuclear cells (PBMC) and liver tissue were detected in different types of patients with chronic HBV infection (chronic hepatitis B, chronic severe hepatitis, liver cirrhosis, liver cancer). Results: The expression level of APOBEC3G in PBMC of healthy people was very low. The expression level of APOBEC3G in PBMC of patients with chronic hepatitis B, chronic severe hepatitis, liver cirrhosis, or liver cancer were all significantly higher than that of healthy people (P \ 0.01), while there was no statistical difference on expression of APOBEC3G in PBMC among these patients (P [ 0.05) and there was also no statistical difference on APOBEC3G in liver tissue between chronic hepatitis B patients and hepatocellular carcinoma patients (P [ 0.05). CLSM observations indicated that the localization of APOBEC3G protein was in cytoplasms of PBMC and liver cells and was not in the nucli. Conclusion: It is suggested that chronic HBV infection could increase the expression of APOBEC3G, but high expression of APOBEC3G may be only the satellite phenomenon of chronic HBV infection. Whether APOBEC3G is involved in the inactive immune response to chronic HBV infection is still uncertain.

Zhen-Hua Zhang 1, 2 Background: To establish the reference sequences for HBV genotypes B and C that are prevalent in China and to conduct a thorough analysis of the mutations in each site. Methods: Full-length genomic sequences of HBV isolated in different regions of China were retrieved from GenBank, genotyping and sequence comparison were carried, and reference sequences were compiled through genotyping and sequence comparison by choosing the most frequently occured nucleotide at the same position, the mutation rate of each site was calculated and classified as high, moderate and low by analyzing the mutation at each position. These DNA sequences and the deduced amino acid sequences were then compared with other reference sequences, and amino acid sequences of different coding regions likewise. Results: The reference sequences for HBV genotypes B and C that are dominant in China were successfully established and the mutation rate at each position was determined.These seuences are different from other reference sequences that have been published.The homology between the genomic sequence of Bc and those of Ba, Bj were 99.32, 95.52% respectively, Cc and those of C, Caus were 99.27, 95.01% respectively, the homology between the S gene of Bc and those of Ba, Bj were 99.71, 98.68% respectively, Cc and those of C, Caus were 99.27, 95.01% respectively, suggesting homology between Bc and Ba is higher than that between Bc and Bj,Cc and C is higher than that between Cc and Caus.The amino acid sequences of different coding regions are also different,and the distribution of the base of 1,762, 1,764, 1,858 between genotypes B and C have significant differences. Conlusion: These sequences can serve as the reference sequences for HBV genotypes B and C that are prevalent in China, and can be reliably used to guide primer design and mutation analysis at the nucleotide and/or amino acid level.

Features and Clinical Implications of HBV Genotypes and Mutations in Basal Core Promoter/Precore Region in 507 Chinese Patients with Acute and Chronic Hepatitis B Yan Liu 1 , Yan-Wei Zhong 1 , Zheng-Sheng Zou 2 , Zhi-Hui Xu 1 , Bao-Sen Li 2 , Xiao-Qiang Ren 1 , Xiao-Dong Li 1 , Pan-Yong Mao 1 , Dong-Ping Xu 1 Background: The mechanisms for chronicity of HBV infection remain clarification. The association of HBV genotypes and basal core promoter (BCP) and precore (PC) mutations with the clinical characteristics is increasingly recognized but the data is still poor in China. This study is to investigate these virologic features and clinical implications between large-size patients with acute hepatitis B (AHB) and chronic hepatitis B (CHB). Methods: One hundred and eighty-two AHB patients and 325 CHB patients were investigated. HBV genotypes and BCP/PC mutations were determined by direct PCR sequencing. Mutations at ten interested sites of the BCP/PC region were compared between the two groups of patients. Results: AHB patients had a significantly higher ratio of genotype B to C than CHB patients (37.4-62.6% vs.16.6-83.4%, P \ 0.001). The prevalence of BCP/PC wild-type virus was 60.4% in AHB patients in contrast to 28.9% in CHB patients. Significantly lower prevalence of A1762T, G1764A, G1896A, and G1899A but higher prevalence of T1758C was found in AHB patients. Interestingly, T1758C and A1762T/G1764A appeared mutual restraint. Genotype B virus had lower BCP mutation frequency and similar PC mutation frequency compared to genotype C virus. AHB patients with BCP/PC mutant virus had higher viral load, whereas CHB patients with BCP/PC mutant virus had lower viral load and elevated alanine aminotransferase, in comparison with those with the wild-type virus. Conclusion: Patients with genotype B virus, BCP/PC wild-type virus or T1758C mutant virus were more susceptible to develop AHB, whereas high prevalence of the BCP/PC mutations was associated with CHB development.

after transfection by flow cytometry. The levels of COX-2, ERK1/2 in HepG2 cells were detected 24 h after transfection by semi-quantitative RT-PCR and western blotting analysis, respectively. Results: The results from MTT showed the proliferation inhibition ratio in COX-2siRNA intervention group was higher (67.08%) than that in negative control siRNA group (2.45%) and empty liposome group (1.56%) (P \ 0.01).Flow cytometry analysis showed that cell cycle distribution in the spot G1 (DNA presynthetic phase) of HepG2 cell was in creased in COX-2siRNA intervention group (72.80%) as compared to control siRNA group (53.13%), empty liposome group (50.97%) and blank control group (50.27%), respectively (P \ 0.05). Semi-quantitative RT-PCR analysis showed the amounts of mRNA of COX-2(0.58), ERK1(0.32) and ERK2(0.48) in COX-2siRNA intervention group were significantly decreased after incubated 24 hs as compared to negative control siRNA group ( Cytoplasmic HBsAg expression were positively related to HBV DNA (r = 0.412, P \ 0.0001), ALT (r = 0.193, P = 0.002) and AST (r = 0.201, P = 0.001) levels, and negatively related to fatty liver degree (r = -0.196, P = 0.001). Membranous HBsAg expression were positively related to serum HBeAg (r = 0.317, P \ 0.0001) and HBV DNA (r = 0.347, P \ 0.0001), and negatively related to age (r = -0.132, P = 0.006), but not related to ALT or AST. Hepatocyte HBsAg was not correlated with either fibrosis stage or inflammation grade. Conclusions: Cytoplasmic staining of HBsAg may suggest a maker of active hepatocyte injury. Membranous HBsAg expression which declining with age reveals the possibility of spontaneous virus clearance.

Relationship between Local Cellular Immune in Liver and Viral Replication in Different Phases of Chronic HBV Infection Yong-Xiao Zhao 1 , Li-Ying Feng 1 , Yan-Jin Liu 1 1 The Second Hospital of Hebei Medical University, 215 Heping West Road, Shijiazhuang, China

Background: The cellular immune response is thought to play a critical role in viral clearance and disease pathogenesis during HBV infection. CD4+ and CD8+ T cells, as main subpopulations of T cell, play a key role in the viral clearance and liver cell injuries. To investigate local cellular immune in liver and viral replication in different phases of chronic HBV infection is significant. Methods: CD4+, CD8+ cells were detected in 40 cases liver biopsy with chronic HBV infection by immunohistochemical technology. Liver tissue HBV-DNA were detected by real-time fluorescence quantitative PCR.

Results: CD4+ cells in immune active phase was significantly more than it in immune tolerance and inactive phase (P \ 0.01, P \ 0.05).CD8+ cells in immune active phase was significantly more than it in immune tolerance (P \ 0.05), also more than it in inactive phase, but there was no significant difference (P [ 0.05). CD4+, CD8+ cells in immune inactive phase was more than it in immune tolerance phase, but the differences were not significant. HBV-DNA can still be detected in liver tissue in eight cases with serum HBV-DNA negative. Conclusion: HBV-DNA quantitative of liver tissue can more accurately reflect the level of viral replication than serum HBV-DNA quantitative. The local immune state in liver was different in different phases of chronic HBV infection. Result: Va24NKT cells in patients with CHB and ACLF were 0.8012 ± 0.2979 and 0.4638 ± 0.2244%, respectively, which were significantly lower than that in NC (1.1114 ± 0.3546%, P \ 0.05). Va24NKT cell levels showed significantly positive correlation with that of prothrombin time (PT) and significantly negative correlation with that of total bilirubin (TB), Va24NKT cell levels showed no significant correlations with that of alanine aminotransferase (ALT) and HBVDNA. Conclusion: Va24 NKT cells in peripheral blood in patients with chronic HBV infection are decreased and correlate with the severity of hepatic injury. these cell lines with or without tumor necrosis factor (TNF)-alpha by reporter assay. We also examined the activation of IFN-beta promoter in these cells. Results: We confirmed HBeAg secretion in cell culture fluid of HBeAg-positive HepG2 cells (COI 241 ± 47.9) . Relative expressions of IL8, IL6, TNF,  IFNB, IL12A, IFNA, IL2, IL1A, IL10, IL1B, IRF3 and NFkB mRNAs were down-regulated in HBeAg-positive HepG2, compared with HBeAg-negative cells. The higher activation of NFkB with and without TNF-alpha was seen in HBeAg negative cells by reporter assay. The higher activation of IFN-beta promoter was also detected in HBeAg negative cells. Conclusions: HBeAg may suppress cytokines and IFN gene expression and promote HBV replication through the inhibition of NFkB activation and/or IRF3 activation. Further studies are now going on.

Evaluation of Two HBV Promoters for Sustained Transgene Expression in Mouse Liver Juan Du 1 , Fang Zhao 1 , Sheng-Qiang Liang 1 , Ying-Li Wang 1 , Hu Yan 1 , Lin-Sheng Zhan 1 1 Beijing Institute of Transfusion Medicine, Taiping Road, Beijing 100850, China

Aims: To find new expression cassettes for long-term expression of therapeutic genes to the liver in the context of pDNA. Methods: we have evaluated the function and specificity of HBV two hepatic enhancers combined with HBV core and X promoters in cultured cells from hepatic and nonhepatic origin. By bioluminescence imaging and hydrodynamic gene transfer technology, we have evaluated the persistence of transgene expression containing these regulatory sequences in the liver of mice. Results: We found that either EI or EII was able to stabilize HBV core and X promoters in vivo. Among the tested constructs, HBV core promoter linked to hepatitis B enhancer I and enhancer II (EI-EII-Pcore) and X promoter linked to enhancer I and enhancer II (EI-EII-Px), can direct constant and high level of gene expression in the liver of mice, but EI-EII-Px shows less specificity than the EI-EII-Pcore sequence. Conclusions: EI-EII-Pcore and EI-EII-Px are the most potent in application in gene therapy protocols. Background: HBV infection is considered to be the primary cause of hepatocellular carcinoma (HCC), the fourth most common cancer worldwide. The present study was undertaken to evaluate the association of Th1 cytokine genes, interleukin-12B (IL-12B) and interferon-gamma (IFN-c) polymorphism with hepatitis B virus related HCC risk in Indian population. Subjects and Methods: Five groups of subjects were enrolled viz. control (n = 100), HBV-carriers (n = 60), chronic active HBV (n = 60), HBVcirrhotics (n = 57) and HBV-related HCC (n = 29). PCR-RFLP was done to study the various polymorphic forms of the above mentioned cytokine genes. Genotype distributions between categories were compared using the odds ratios (ORs) and 95% CI were calculated to express the relative risk.

Results: In case of IL-12 (+1188 3 0 UTR), presence of both AC and CC genotypes, significantly (p \ 0.001) increased the risk for HCC development among controls and HBV-cirrhotics. However, among HBV-carriers and chronic-active HBV patients, the same genotypes were associated with a decreased risk for HCC. In case of IFN-c (+874), the TA genotype, was in negative association (p \ 0.01) with HCC development among controls and HBV-cirrhotics, while it was found to be a significant risk factor (p \ 0.001) for the same, among HBV-carriers. In contrast to this, the AA genotype was in positive association (p \ 0.01) with HCC development among chronic-active HBV subjects, while was a significant protective factor (p \ 0.001) for the same among the controls.

Conclusion: Polymorphic forms of IL-12B and IFN-c share an association with HBV-HCC risk in Indian population and should be further evaluated as the candidate genes to determine individual susceptibility for the same.

The Background: The pathogenesis of the hepatitis B virus (HBV) infection is still unknown. At present, the interaction between human immune response and the virus is considered as a crucial factor.HBV exists as quasispecies. The research on the relationship between HBV quasispecies and the pathogenesis of the hepatitis B has not been reported. Methods: Three acute hepatitis B, three HBV carriers, three chronic hepatitis B and three chronic severe hepatitis B were recruited.All the patients are male, HBeAg positive.The genotype of HBV is C. HBV S gene was amplified from sera of these patients. In each patient, 20 clones of HBV fragment were randomly selected and sequenced. Results: The complexity of the severe chronic hepatitis B is significantly different from that of the acute hepatitis B (p \ 0.05), and there is no difference among the other groups. In the T cell epitope 45, 47, 85 amino acid sites of the HBV S gene, the constitution of quasispecies in the chronic hepatitis B is different from that fo HBV carriers and the acute hepatitis B (p = 0.01), and it seems to be different from that of the chronic severe hepatitis, but it is not statistically proved. Besides,in the acute hepatitis B, HBV carrier and chronic hepatitis B, the constitution of quasispecies is different from one to another in the a determinant 126 amino acid site of the HBV S gene (p \ 0.05). Conclusion: The difference of the constitution of quasispecies in some T cell epitopes and a determinant of HBV S gene may be related to the clinical outcomes of hepatitis B, of which the mechanism remains to be further investigated. Background: Oral nucleos(t)ide analogues (NUCs) is current standard for treatment of chronic hepatitis B. However, emergence of viral resistant mutations and rebound of HBV DNA results in disease progression. Empirical adding of another NUCs is recommended but this practice is not always successful. The aim of this study is to evaluate HBV polymerase mutation patterns and its role in management NUCs treatment. Methods: Chronic hepatitis B patients who had received one or more of NUCs regularly who had [1log 10 HBV DNA rebound and [2,000 IU/mL from September 2007 to 31 September 2008 were included. HBV polymerase resistance profile was performed by using line probe assay INNO-LiPA HBV DR v2 (LiPA; Innogenetics, Ghent, Belgium) according to the manufacturer's recommendation.

Result: There were 27 patients (22 males and 6 females), mean initial HBV viral load before treatment was 1.0 9 10 7 IU/ml. Twenty-four patients were first treated with lamivudine and three with adefovir. Fifteen were refractory to lamivudine alone and 12 were refractory to lamivudine plus other NUCs.

Mean HBV viral load at the time of viral breakthrough was 2.8 9 10 7 IU/ mL and biochemical breakthrough was found in 8 (29.6%). HBV Hepatol Int (2010) 4:94-345 121 polymerase mutations found were M204V/I in 24 (88.9%), M180/A181 21(77.7%), L80I 9 (33.3%), L80V 10 (37%), and L173 3 (11%), respectively. Two in 15 patients (12%) in lamivudine refractory and 5/10 (50%) of both lamivudine and adefovir refractory had resistant mutation to both drugs. Conclusion: Line probe assay (INNO-LiPA HBV DR v2) is useful for guiding the rescue treatment in chronic hepatitis B patients who failed NUCs treatment. Routine adding of adefovir in lamivudine refractory patients may result in treatment failure as many as 12%. However, current version can detect only known mutation to lamivudine and adefovir. More sensitive test with more mutation points is required for current treatment. Methods: The TCRBV gene families in 11patients with CHB and 4 normal controls (NC) were examined by the real-time PCR with DNA melting curve analysis, and we further determine sequence bias and characteristics of TCRBV CDR3 gene family by analyzing the gene melting spectral pattern (GMSP) of PCR products from 24 TCRBV gene families. Results: We found that the GMSP of most of the TCRBV CDR3 gene families in NC showed a diverse multimodal shape, however, The GMSP of specific TCRBV gene families in T cells in the livers and in the peripheral blood showed a single peak in all 11 CHB patients, furthermore, specific TCRBV families in T cells in the livers and/or in the peripheral blood in 6 patients with CHB shared the same motif ''Q Y'' in CDR3. More interesting is that a shared TCR CDR3 bsequence was found in peripheral blood and in the liver of specific TCRBV families in 1 patient with CHB. Conclusion: Some of the GMSP of TCRBV gene families in T cells in the livers and in the peripheral blood in patients with CHB has been changed compared to NC and some patients with CHB may result as a shared TCR CDR3 bsequence in peripheral blood and in the liver.

China, but the problem of NA resistance (NAr) is increasing accordingly. This study aimed to characterize the genotypic resistance in treatment-naïve and NA-treated patients with chronic HBV infection. Methods: Full-length HBV reverse transcriptase (RT) sequences were amplified and sequenced directly among 156 treatment-naïve and 170 NA-treated patients with chronic genotype C HBV infection, who were enrolled from five hospitals in four northern cities of China during January 2007 to July 2009. Deduced amino acids at 11 previously-reported NAr mutation positions (including rtL80, rtI169, rtV173, rtL180, rtA181, rtT184, rtA194, rtS202, rtM204, rtN236 and rtM250) in RT region were analyzed. Result: Among 326 HBV isolates, 17.48% (57/326) harbored NAr mutations which were all found in treated-patients (57/170, 33.53%). The most prevalent NAr mutations were rtM204V/I (40/170, 23.53%), rtL180M (25/170, 14.71%) and rtL80V/I (20/170, 11.76%), respectively, which exclusively belonged to lamivudine (LMV) resistance pathway. This was in line with the fact that 66.47% (113/170) of treated-patients were receiving or experienced LMV treatment at sampling. The prevalence of NAr mutations belonging to adefovir dipivoxil (ADV) resistance pathway was relatively lower with rtA181T/V (18/170, 10.59%) and rtN236T (7/170, 4.12%) . No mutations occurred at rtI169, rtT184, rtA194 and rtS202. Twenty-four kinds of mutation patterns were detected in 57 mutants. Single site mutation was found in 16 isolates, while double, triple, quadruple and quintuple combination mutations were found in 29, 10, 1 and 1 isolates, respectively. The most predominant mutation patterns were rtM204I+rtL80I (8/57, 14.04%) and rtM204V+rtL180M (8/57, 14.04%). Conclusion: The prevalence of NAr mutations in treatment-naïve patients was not found. However, the characterization of LMV and ADV resistance mutations in NA-treated patients provided important background information for the future add-on or switch-to antiviral therapies.

Ling-Li Yang 1 , Fumio Imazeki 1 , Makoto Arai 1 , Tatsuo Kanda 1 , Kenichi Fukai 1 , Keiichi Fujiwara 1 , Osamu Yokosuka 1 1 Department of Medicine and Clinical Oncology, Graduate School of Medicine, Chiba University, 1-8-1, Inohana, Chuou-ku, Chiba, Japan

Background: Hepatitis B virus (HBV) replicates through RNA intermediates and there are several reports about serum HBV RNA detection. We examined whether HBV RNA is useful for the pathogenesis and treatment response of hepatitis B. Methods: We studied 74 patients in four groups during different stages of chronic HBV infection and 47 patients with lamivudine treatment. HBV DNA and RNA were extracted from the serum of these patients. The nucleic acids were amplified by the real-time PCR with or without reverse transcription (RT). The amount of HBV RNA was determined by the differences between real-time PCR and RT-real-time PCR. Result: Serum HBV RNA was detectable in 63.5% of the chronically HBVinfected patients. No correlation was observed between the serum HBV RNA and ALT (n.s.). The ratio of HBV RNA in all the HBV nucleic acids (HBV RNA%) was ranged between 10 and 20% in each infection stage. HBV RNA% in HBeAg-ASC group was lower than HBeAb-ASC group (11.5 vs. 21.0%, p = 0.0186). After 1-month lamivudine treatment, more patients with HBV RNA% increase were observed in the patients with viral breakthrough (VBT) than the group without VBT (22/28 vs. 7/19, p = 0.006). Conclusion: Serum HBV RNA was detectable in each stage of chronic HBV infection. HBV RNA% at 1 month after the commencement of lamivudine treatment could be a potential predictor of VBT.

Objectives: To study the features and rules of mutation in BCP, precore and preS region of HBV genome with different status of liver disease after HBV infection, and to analyze the clinical significance of the mutation. Methods: Blood samples were retrieved from China northern patients with HBV infection. Serum HBV-DNA was acquired from samples and amplified with routine PCR. Sequences of the cloning products were got and analyzed. Results: 201 patients were enrolled in the study, which included acute hepatitis B (7 cases), asyptomatic hepatits B virus carriers (13 cases), chronic hepatitis B (89 cases), liver cirrohsis (62 cases) and HBV related carcinoma (30 cases). Nucleotide sites such as nt C1726, T1727, G1730, G1752 and G1799 in BCP and precore region of HBV genome have genotype specificity. The result shows that G1776A (OR = 10.7, 95% CI: 2.4-48.1, P = 0.002), A1846T (OR = 3.8, 95% CI: 1.2-11, 7 , P = 0.02), G1896A (OR = 3.4, 95% CI: 1.4-8.3 , P = 0.01) and mutation sites C3 (OR = 2.7, 95% CI: 1.1-6.4, P = 0.027) may be associated with HBeAg negativity. G1896A mutation may be correlated with disease progression (P = 0.004) and be essential in many mutation combinations. In mutants bearing more than three substitutions, 53% had G1896A (35/66), and only 8% (8/98) in strains with point mutations less than 3 (P \ 0.05). The experiment also hints various substitutions could coexist in one strain after long history of HBV infection. Our experiment shows HBV preS1 and preS2 region deletion could coexist in the same patient and often occur in liver cirrhosis patients. Conclusion: A1727G mutation is a common mutation in northern China. A novel G1776A mutation is identified to be statistically responsible for HBeAg negativity. G1896A mutation may be the risk factor in liver disease progression and prognosis independent of age. HBV preS gene deletion may be associated with progression of liver disease.

Application of the HBV Replication Mouse Model in the Biological Study of HBV Hong Tang Background: The lack of reliable and convenient animal models has hindered progress in hepatitis B virus (HBV) research. Recently, we have established a simple HBV replication mouse model using hydrodynamic tail vein injection of HBV replication competent plasmid (pHBV4.1) into Balb/C mice. In present study, the application of this mouse model in several aspects of HBV biology was evaluated. Methods: To determining its application in detecting the efficacy of anti-HBV agents, polyinosinic-polytidylin acid (polyIC, interferon inducer) and nucleotide analogues (adefovir and entecavir) were administrated to HBV replication mouse model with wild-type pHBV4.1 injected, and the inhibiting efficacy of these agents on HBV DNA replication was evaluated. To identify its application in biological study of HBV drug-resistance mutants, single-point mutation (rtM204I, ayw subtype) and double-point mutation (rtA181V+ rtN236T, ayw subtype) were injected respectively and their corresponding HBV DNA replication intermediates in liver was assessed. Results: Compared with mouse models without antiviral agent treatment, the HBV DNA replication intermediates in polyIC treated group was significantly decreased; and after the treatment of ETV and ADV, the level of HBV DNA replication were decreased by 75 and 35% respectively. These findings suggested that HBV DNA replication in our mouse model could be inhibited by administration of anti-viral agents. For mouse model injected with rtM204I and rtA181V+rtN236T mutant HBV, HBV DNA replication could still be detected, but the level of HBV DNA replication of mutant plasmids decreased a lot compared with the mouse model with wild-type HBV. Conclusions: The HBV replication mouse model we established was a useful tool to study the biology of HBV. It could be applied to detecting the efficacy of antiviral agents and studying the biological characteristics of HBV drug-resistance mutants. This work was supported by 973 Program of China, No. 2007CB512902 and 2006CB504302. C-terminal transactivation or coactivation domain. The role of HBx in regulating HBV transcription and replication were investigated in HepG2 in vitro. The execution of HBx function requires the participation of host cell protein factors, whose types and levels may be different between HepG2 cell lines in vitro and liver cells in vivo. The purpose of this study was to investigate the possible role of HBx in regulating HBV transcription and replication in vivo. Methods: HBV replication mouse models were established using hydrodynamic tail vein injection of wild-type or HBx-minus HBV genome (with defective X gene), and HBx expression plasmid. After injection, mice were sacrificed on day 3, and liver tissue was collected and total RNA and DNA were extracted. The levels of HBV mRNA in the liver were analyzed by Northern blot hybridization, and the levels of HBV DNA replication intermediates in the liver were detected by Southern blot hybridization. Results: The levels of 3.5-kb HBV RNA and HBV DNA replication intermediates of HBx-minus HBV mutants in liver were lower than that of wild-type HBV genome or HBx-minus HBV genome plus HBx expression plasmid. The absence of HBx expression caused the reduction of HBV transcription and replication and the decreases could be restored by expression of HBx in liver tissue. Conclusions: HBV mutant genome with defective X gene led to decreased levels of HBV DNA and HBx has augmentation effects on HBV transcription and replication in vivo of mouse. This work was supported by the National Natural Science Foundation of China, NO.30570064.

Zhi-Xin Zhao 1 , Qing-Xian Cai 1 , Huan-Huan Cao 1 , Xiao-Yan Guo 1 , Zhi-Liang Gao 1 1 The Third Affiliated Hospital of Sun Yat-Sen University, Tianhe Road, No. 600, Guangzhou, China

Aim: To explore the expression of suppressor of cytokine signaling (SOCS)-1 gene in the peripheral blood mononuclear cells (PBMCs) of patients with Hepatitis B Virus (HBV)-related liver diseases and its clinical significance. Method: 50 patients with HBV-related liver diseases including 35 cases with acute-on-chronic liver failure (ACLF) and 15 with chronic hepatitis B (CHB) were enrolled into this study.10 healthy people were taken as control. SOCS-1 mRNA levels in peripheral blood mononuclear cells (PBMC) freshly obtained from all the subjects were detected by real-time quantitative retroverse polymerase chain reaction (QRT-PCR) and their correlations with clinical data were analyzed by SPSS statistics software. Result: SOCS-1 mRNA levels in PBMC of HBV-related liver disease group were significantly higher than those of control (P \ 0.05) and those of CHB group were significantly higher than those of ACLF group (P \ 0.05). SOCS-1 mRNA levels were found negatively correlated with serum HBV DNA levels and HBeAg titers in HBV-related liver disease group (R = -0.555, p \ 0.05 and R = -0.48, p \ 0.05, respectively). Conclusion: The decline of SOCS-1 gene expression in PBMC of ACLF group may contribute to the pathogenesis of ACLF. The expression of SOCS-1 gene is influenced by HBV in vivo. Background and Aims: Antiviral resistance is one of the most important factors for treatment failure with nucleos(t)ide analogues in CHB. Pre-existing drug resistance mutations may occur in untreated patients and may affect their treatment outcomes. A few prior limited studies report high rate of HBV rt mutations in reportedly treatment-naïve patients though methods of assessing the absence of prior therapy in such studies were not well defined. Our goal is to establish the prevalence of clinically-relevant HBV rt mutations in truly treatment-naïve patients with CHB using a sensitive line probe assay. Methods: We prospectively enrolled 70 CHB patients at 2 U.S. gastroenterology clinics during 2/09-5/09 and used the INNO-LiPA HBV DR3 test (Innogenetics, Belgium) to detect relevant mutations in the HBV rt at the following positions: 80, 173, 180, 181, 204, 236, 184, 194, 202, 233, and 250 . All patients gave written consent and were interviewed using a detailed questionnaire to illicit a history of prior anti-HBV treatment (herbal or conventional Western therapy). Results: The majority of patients were Asians (97%) and foreign-born (97%) with median age being 42 . Family history of CHB and HCC were present in 54 and 17%, respectively. None had cirrhosis or liver cancer. Median ALT was 25 (7-164). Median HBV DNA was 4.5 log 10 IU/mL (2.8-8.7 ) and most had either genotype B or C (94%). HBeAg was positive in 22%. All patients were found to have viral populations that were sensitive to lamivudine, adefovir, telbivudine, and entecavir and none had any detectable HBV rt mutations.

Conclusions: Contrary to prior preliminary reports, no baseline mutations associated with antiviral resistance were detected by sensitive INNO LiPA test in our treatment-naïve CHB patients. Such viral quasispecies, if present, are probably rare in CHB patients without prior exposures to oral nucleas(t)ide anti-HBV therapy.

Circulating CD4+CD25high Background: The roles of regulatory T cells (Tregs) and PD-1 in hepatitis B have not been clearly described. Also, the role of B and T lymphocyte attenuator (BTLA), which serves as a negative regulator of T cell activation, is still unknown in hepatitis B. In this study, we analyzed the frequency of circulating Tregs in patients with chronic hepatitis B (CHB) infection as well as the expression of PD-1 and BTLA on CD4 + T cells and their relationships with HBV-related clinical markers. Methods: Flow cytometry was performed to assess the expression of PD-1 and BTLA on the CD4 + Tcells and circulating CD4 + CD25 high Tregs of 39 patients with chronic hepatitis B (CHB) and 19 healthy controls (NC). Result: We found that the frequency of CD4 + CD25 high Tregs and PD-1 expression on CD4 + T cells was significantly increased in CHB patients compared with normal controls. However, BTLA expression on CD4 + T showed no significant difference between the two groups. The frequency of Tregs was significantly higher in patients with HBV DNA titer C10 8 than that of HBV DNA titer\10 8 . Circulating CD4 + CD25 high Treg frequency and PD-1 expression on CD4 + T cells correlated positively with serum HBV DNA load in CHB patients. Coclusion: Our findings suggested that the increased frequency of CD4 + CD25 high Tregs and PD-1 expression on CD4 + T lymphocytes may inhibit cellular immune response against HBV and affect viral clearance, leading to the persistence of chronic HBV infection.

Results: Of 5, 676 blood donors, 779 (14.1%) was positive for ant-HBc with HBsAgnegative, anti-HBs-negative and high-titiered anti-HBc (above 1:128) was found in seven blood donors. HBV DNA-positive was detected in four blood donors (3 from high-titiered anti-HBc group) and the load of HBV DNA of four donors was 1.2 9 10 5 , 4 9 10 3 , 1.6 9 10 3 and 4 9 10 4 copies/ml, respectively. The sequence analysis showed that two new HBsAg variants were found. Conclusions: The risk of transfusion-transmitted HBV was high in blood donors with high-titered anti-HBc. HBsAg-negative HBV infection carriers was major low-level infection of HBV and it was possible association with HBsAg antigentic variations PP115 Background: Chronic HBV infection is an important risk factor for the development of liver cirrhosis and hepatocellular carcinoma. In chronic HBV infection cellular immune responses are weak or undetectable, which lead to a state of relative collapse of HBV-specific adaptive immunity. Recent years, a new T helper cell lineage, called Th17 cells, producing interleukin (IL)-17 family cytokines and IL-22, has been identified. To clarify whether Th17 cells are involved in HBV infection we determined the frequency of IL-17-producing Th17 cells and expression of Th17-related cytokines in patients with acute and chronic hepatitis B. Methods: Flow cytometry was performed to assess frequency of peripheral Th17 cells (CD3 + CD8 -IL-17A + T cells) and CD3 + CD8 + IL-17A + T cells in 16 patients with acute hepatitis B (AHB), 41 patients with chronic hepatitis B (CHB), 20 of asymptomatic HBV carriers (AsC), and 16 normal controls (NC). Serum levels of IL-17, IL-22, TNF-a, and IFN-c were measured simultaneously by enzyme linked immunosorbent assay. Results: We found that the percentage of Th17 cells in peripheral blood of AHB patients was significantly elevated than that in NCs. The frequency of Th17 cells showed a positive correlation with serum ALT and AST in AHBs. IL-17 and IFN-c levels in serum showed no significant difference among the four groups. However, IL-22 concentration in AHB patients was notably increased as compared with levels in NCs, CHB patients, and AsCs. This marked elevation of serum IL-22 positively correlated with serum TNF-a concentration in patients with AHB. Conclusions: Th17 cells secret IL-22 as a major cytokine to take part in the immunopathogenesis of AHB disease process and they play an important role in limitation and counteraction of severe injury of liver function in acute HBV infection via TNF-a signal transduction.

Background: The alteration of peripheral blood NK cells subsets in the course of HBV infections is not completely defined. Methods: Forty-six patients with chronic hepatitis B virus infection including 25 patients in immune-tolerance phase (HBV-DNA[10 7 copy/ml, ALTB40U/ L) and 21 patients in the immune-clearance phase (HBV-DNA positive, ALT [40 U/L) were enrolled for observation. 10 healthy volunteers were taken as control. The expression of peripheral blood NK cells subsets were assessed by flow cytometry. Serum HBV DNA load was determined by real-time fluorescent quantitative polymerase chain reaction (PCR), and serum ALT levels were examined simultaneously. Results: The ratio of peripheral blood CD56 bright NK cells in healthy control group, immune-tolerant phase group and immune-clearance phase group were 0.546 ± 0.173, 0.647 ± 0.294, 0.877 ± 0.493, respectively. Compared with healthy control group and immune-tolerant phase group, the proportion of CD56 bright NK cells in immune-clearance phase group were increased and showed statistically significant (P = 0.023, 0.04). But no significant difference found among this three groups in CD56 dim NK cells. Correlation analysis showed peripheral blood CD56 bright NK cells ratio and serum HBV DNA load in immune-tolerance phase group were negative correlation (r = -0.575, P = 0.003), but in immune-clearance phase group showed no such correlation. Conclusions: The proportion of peripheral blood CD56 bright NK cells were significantly higher in the immune-clearance phase, it suggested the CD56 bright NK cells play an important immune regulation role in the process of immuneclearance in chronic hepatitis B infection.

The Resistance Mutation Pattern of Hepatitis B Virus during Adefovir Dipivoxil Sequential Therapy after Lamivudine Resistance Zhen-Ping Wu 1 , Tao Han 1 , Ying-Tang Gao 1 , Zhi Du 1 1 The Third Central Hospital of Tianjin, Tianjin Hedong District Jintang Road No. 83, China

Objective: To investigate the resistance mutation pattern of hepatitis B virus (HBV) during adefovir dipivoxil (ADV) monotherapy or lamivudine (LAM) with ADV combination therapy after lamivudine resistance. Methods: A series of serum samples from fifteen patients with suboptimal viral response to ADV sequential therapy after LAM resistance were collected.The RT region of the polymerase gene from the serum was amplified, cloned and sequenced, mutation pattern in relation to resistance was analyzed.

Results: These ADV resistance mutation patterns of A181T + N236T, A181V, A181T were selected in ADV monotherapy group.These LAM resistance mutation patterns of M204V + L180M, M204V + L180M + L229V, M204I + L80I, M204V + L180M + V207I were detected in LAM + ADV combination therapy group. Besides, 20% of clones of three serum samples were double resistance to LAM and entecavir (ETV) in combination therapy group, which were M204I + L80I + T184I (2/10), M204V + L180M + T184S (2/10), M204V + L180M + G173L + S202G (2/10) . The mutation rates of I269L, P109S were both high in two groups, but they were higher in monotherapy group than that of combination therapy group.Whatever known resistance mutation wasn't detected in two serum samples from two groups respectively, in which the mutation rate of I269L was 100%, the mutation rate of P109S was also 100% in the one from monotherapy group. Conclusion: In LAM resistance patients with suboptimal viral response to ADV sequential therapy, ADV resistance mutation patterns of A181T + N236T, A181V, A181T were easily selected during ADV monotherapy, in LAM + ADV combination therapy group, these LAM resistance emutation patterns of M204V + L180M, M204V + L180M + L229V, M204I + L80I, M204V+ L180M + V207I were predominant, mutation patterns of M204V + L180M + G173L + S202G, M204I + L80I + T184I, M204V + L180M + T184S could be selected during combination therapy, which were both resistance to LAM and ETV. The mutation rates of I269L, P109S were both high in two groups, they may affect the response of ADV sequential therapy. The aim of this study was to characterize virological factors of HBV associated with vaccine failure in Thailand. Methods: Sera collected from 14 infected infants with vaccine failure and their respective mothers (13 HBeAg-positive and 1 HBeAg-negative) (group I) were tested for HBsAg and HBV DNA levels, HBV genotypes and mutations by direct sequencing. Sera collected from 15 HBeAg-positive (group II) and 15 HBeAg-negative (group III) mothers whose infants had been successfully vaccinated served as controls.

Results: The results showed that group I mothers had higher mean levels of HBsAg and HBV DNA than group II and III. All infected infants and their respective mothers had the same HBeAg status and HBV genotypes. DNA analysis in a pair of HBeAg-negative infant and mother revealed that both were infected with an HBV precore mutant (G1896A). Escape mutants in the 'a' determinant region were detected in 2 out of 14 (14%) infected infants. Conclusion: Instead of HBeAg status, high HBsAg and HBV DNA levels in mothers are the major contributing factors to vertical transmission of HBV. The precore mutant with high viral load could be transmitted from mothers to their offspring. Escape mutants seem to be uncommon, and might not be associated with vaccine failure among Thai populations. Background/Aims: The biphasic clearance of HBV serum viral loads by the nucleos(t)ide analogues has been reported. In this study, we used a novel mathematic model to analyze the early viral kinetics in chronic hepatitis B (CHB) patients, and compared the viral dynamic profiles of different treatments. Methods: Thirty CHB patients (HBeAg/HBsAg positive) from Queen Mary Hospital, Hong Kong, were enrolled in this study. 18 patients had received 48 weeks lamivudine treatment and 12 patients had received 48 weeks clevudine treatment. Serum samples were collected at baseline, days 4, 7, 10, 14, and weeks 3, 4, 8, 12 . HBV DNA was quantified by Abbott Realtime HBV assay. Viral kinetics were analyzed by the two first order independent secay (TFOID) model: y = a exp(-bx) + c exp(-dx) (y: percentage of viral load compared with baseline; x: duration of treatment). The time needed to achieve 50% (T 50 ), 95% (T 95 ) and 99% (T 99 ) of HBV DNA reduction were calculated by Table Curve2D . Results: All viral kinetics data of the thirty patients fitted the TFOID model well (r 2 [ 0.98). T 50 of these patients [1.76 (1.38-5.07 ) day] were significantly shorter than the terms of achieving the latter 49% viral load reduction (T 99 -T 50 ) [13.14 (3.38-34.15 ) day, P \ 0.0001]. T 95 of these patients [6.30 (3.43-19.37 ) day] had no difference with terms of achieving the latter 4% viral load reduction (T 99 -T 95 ) [7.60 (1.36-28.96 ) day, P = 0.994]. There was no significant difference between T 99 of lamivudine and clevudine therapy [19.01 (4.97-35.94 ) day vs. 9.62 (5.10-27.24 ) day, P = 0.087]. Conclusion: TFOID model is suitable to analyze early viral kinetics of HBV DNA under nucleos(t)ide analogues therapy. Efficacies of these antiviral therapies in the early phase of treatment were higher than the late phase. Viral dynamic profiles of lamivudine and clevudine therapy had no significant difference.

Background: HBV carriers constitute major reservoir, especially in developing countries. Generally they are recommended for regular check up and treatment is recommended when patients have liver damage. This is mainly attributable to lack of information about extent of liver damage. Aim of this study was to assess extent of liver damage in HBeAg -ve patients who were completely unaware of their HBV infection in Bangladesh. Methods: In this retrospective study, we reviewed the records of 310 HBeAgve CHB patients. They were tested for serologic markers of HBV, anti-HCV and ALT. All underwent per-cutaneous liver biopsy. Result: 77.4% (240/310) were males and 22.6% (70/310) females. Patients were between 13 and 60 years of age. ALT was raised [2 times UNL (cutoff 42U/L) in 7.1% (22/310). 37.4% (116/310) had high HBV DNA ([10 5 copies/ ml by PCR), while low DNA (\10 5 copies/ml) was seen in 62. 6% (194/310 Objective: To investigate the evolution characteristic of hepatitis B virus (HBV) quasispecies during sequential therapy after lamivudine resistance. Methods: A series of serum samples from seven patients of lamivudine resistance during sequential therapy were collected.The reverse-transcriptase (RT) region of the HBV polymerase gene was amplified, cloned and sequenced, sequence mutation combination with clinical drugs was analyzed, then the evolution characteristic was investigated. Results: Patient 1, 60% (6/10) of clones were M204I + L80I during LAM + ADV treatment, the new mutation L180M was detected in 57% (4/7) of clones during LdT treatment, all clones (10/10) were M204I + L180M + L80I during ETV treatment, Patient 2, 80% (8/10) of clones were M204V + L180M at LAM resistance,then with LAM + ADV treatment, at week11, 80% (8/10) of clones were M204V + L180M + G173L, at week 89, M204V + L180M + T184S and M204V + L180M were detected in 25% (2/8) and 75% (6/8) of clones respectively. Patient 3, all clones (10/10) were M204I + L80I at LAM resistance, then with LAM + ADV treatment, at week 20, all clones were wild, but at week 36 and 54, the mutations were M204I + L80I (11/11) and M204V + L180M (10/10), respectively. Patient 4, 90% (9/10) of clones were M204I at LAM resistance, at week 22 during LAM + ADV treatment, M204I + L80I + L180M and M204I + L80I were detected in 56% (5/9) and 44% (4/9) of clones, respectively. Patient 5, M204V + L180M + G173L and M204V + L180M were detected in 55% (6/11) and 45% (5/11) of clones, respectively at week 8 during LAM + ETV 0.5 mg/day treatment,then ETV 1 mg/day was given for sequential therapy, at week 16, 44 all clones were M204V + L180M + G173L (10/10, 7/7). Patient 6 and 7, all mutations were M204V + L180M + G173L (6/6, 10/10) at LAM resistance. Patient 6, double resistance mutations M204V + L180M + G173L + T184A were detected in 10% (1/10) of clones during ADV + ETV treatment. Patient 7, double resistance mutations M204V + L180M + G173L + N236D were detected in 10% (1/10) of clones during ADV monotherapy.

Conclusion: HBV existed resistance mutation is not easy to disappear, morever it is easy to select cross-resistance or multi-resistance mutation. It effects the response to sequence therapy, moreover it results in the failure of sequence therapy. Once the resistance mutation was presented, it would effect sequence therapy consecutively. So for naïve antivirus treatment of HBV infected patient, it is necessary to choose antiviral agents with low risk of resistance. Background and Aims: Binding of beta 2-glycoprotein I (b2-GPI) to hepatitis B surface antigen (HBsAg) has an effect on HBV infection. The aim is to identify the affinity of b2-GPI to HBsAg, the influence of glycosylation of b2-GPI and binding rates among chronic hepatitis B (CHB) patients. Methods: HBsAg and b2-GPI were radiolabeled with Na 125 I by chloramine-t. The affinity constant (Ka) was checked by radioimmunoassay. Nonglycosylated rb2-GPI was expressed in M15/pQE30/hb2-GPI. The precipitates of b2-GPI-125 I-HBsAg and rb2-GPI-125 I-HBsAg were by competitive conjugation method. The Ka 1 and Ka 2 were calculated by Adrion method. A total of 23 samples were collected from 9 HBeAg positive, 9 HBeAg negative patients and 5 normal controls. Serum HBsAg quantification, HBV-DNA and alanine aminotransferase (ALT) levels were monitored. The binding rates of 125 I-b2-GPI with the serum of CHB patients were calculated. Results: The specific activity of 125 I-HBsAg and 125 I-b2-GPI was (2.6-2.8) 9 10 6 and (5.8-6.0) 9 10 6 Bq/lg. The affinity of b2-GPI to HBsAg was (2.795 ± 1.846) 9 10 8 L/mol. There was no statistically significant difference between Ka 1 and Ka 2 of b2-GPI from two separate sources (P [ 0.05) ( Table  1 ). The binding rates between HBeAg positive and HBeAg negative had a significant difference and the same difference was observed between active hepatitis B and carriers among the HBeAg positive patients (P \ 0.05). (Lim et al. in Gastroenterology, 2007) , but this is unknown in lamivudine treated chronic hepatitis B patients. The aim of this study was to evaluate the evolution pattern of HBV quasispecies in lamivudine induced seroconverters. Methods: Ten seroconverters and 10 non-seroconverters during lamivudine treatment were selected. Nested PCR of the precore/core gene were performed followed by cloning and sequencing for four time-point samples per patient. Twenty clones per sample were sequenced. Sequences were aligned using ClustalX and sUPGMA pylogenetic trees were constructed using Pebble 1.0 following maximum likelihood estimates of pairwise distances under a GTR + I + G model testing the viral diversity and evolution. Results: Lamivudine induced HBeAg seroconverters had a viral diversity of 1.38 9 10 -2 (substitutions/site, median) and evolutionary rate 5.97 9 10 -4 (substitutions/site/month, median) which were 3.2-and 5.2-fold higher respectively than those of non-seroconverters. The viral diversity increased significantly after starting lamivudine treatment (from baseline 7.25 9 10 -3 to 1.93 9 10 -2 substitutions/site, p = 0.01) and reached a plateau just before and after HBeAg seroconversion (2.06 9 10 -2 and 2.07 9 10 -2 substitutions/site) in seroconverters. Non-seroconverters showed no alteration in viral diversity over time.

Seroconverters showed more complex phylogenetic trees than non-seroconverters. Positive selection was similar in seroconverters and non-seroconverters. Conclusion: High viral diversity is also a feature of lamivudine induced seroconversion supporting the hypothesis that viral diversity plays a key role in pathogenesis of HBeAg seroconversion.

secretion of TNF-a in groups of CSHB, CHB and NC was 15317.69 ± 4124.90 pg/ml, 9670.29 ± 3654.68 pg/ml (P \ 0.05) and 6547.43 ± 748.18 pg/ml (P \ 0.05); the production of IL-6 in groups CSHB, CHB and NC was 1423.78 ± 375.14, 862.68 ± 93.68 pg/ml (P \ 0.05), 567.26 ± 167.04 pg/ ml (P \ 0.05), respectively. NF-jBp50 showed significant correlations with TNF-a (r = 0.70, P \ 0.01) and IL-6 (r = 0.69, P \ 0.01) in mDC. Furthermore, the secretion of TNF-aand IL-6 in mDC of group CSHB was negatively associated with PTA (r = -0.43, P \ 0.05; r = -0.40, P \ 0.05), but not with ALT, TBil and virus loads. Conclusions: The expression level of NF-jB and its regulated cytokines in MoDC in patients with CSHB were up-regulated and associated with the deterioration in liver function. (2) There was a high positive correlation between the HBV cccDNA copies of hepatocytes and the serum HBV DNA (r = 0.665, p = 0.009); but there is no correlation with serum HBsAg (r = 0.253, p = 0.383) in HBeAg negative group.

(3) There is no difference of the HBV cccDNA copies of hepatocytes between HBeAg positive and negative group (t = 0.09, p = 0.9287). Conclusions: (1) When HBV replication actively serum HBV DNA may reflect the level of hepatocytes cccDNA, but when the HBV replication inactively the level of serum HBV DNA may not match the level of hepatocytes cccDNA. (2) The clinical significance and academic value of the quantity assay of HBV cccDNA in the hepatocytes could not be substituted by the detection of sera HBV DNA, HBsAg. (5/30) , there were significant difference between two groups (P \ 0.01). There were no significant difference on the decline of TBIL and ALT between two groups (P [ 0.05). After 12 weeks the liver function were normal in two groups, there was only one patient with Abnormal liver function in model group on the 24th and 48th week, in control group, there were three and five patients with Abnormal liver function on the 24th and 48th

week. 20 patients in model group and 12 patients in control group were alive after 48 weeks. The survival percent in model group was 67.7%, in control group was 40%, there were significant difference between two groups (P \ 0.05). Conclusion: Telbivudine has strong suppression of hepatitis B virus activity, and effect soon, which will significantly improve the liver function, improve the survival percentage and avoid repeatedly abnormal liver function. patients with decompensated liver cirrhosis who underwent nucleoside analogues treatment in our hospital were enrolled in this study. Decompensated liver cirrhosis was defined as prolongation of prothrombin time (PT) for more than three seconds or serum total bilirubin level higher than 3 mg/dl. Among them, 27 cases received lamivudine (LAM) 100 mg/day, and 10 cases received entecavir (ETV) 0.5 mg/day treatment, respectively. The survival of the two groups was compared. Results: The mean baseline Child-Pugh scores of patients underwent LAM and ETV treatment were 9.1 ± 1.6 and 8.7 ± 1.9, respectively. Thirteen (48%) cases in LAM group and three (30%) cases in ETV group were classified in Child-Pugh C stage. The mean treatment duration for LAM was 13 ± 5.4 months, and for ETV was 7.3 ± 5 months. During the follow-up period, three (11.1%) cases in LAM group and one (10%) case in ETV group died. None case developed biochemical breakthrough. The causes of death were liver failure (n = 2), hepatocellular carcinoma (n = 1), and variceal bleeding (n = 1). The 6-, 12-and 18-month survival rates were 100, 91 and 85% in LAM group, and 90, 90 and 90% in ETV group (p = 0.48). Conclusions: By analysis the short-term survival, LAM could provide equal survival benefit to ETV in patients with decompensated cirrhosis related to hepatitis B. Further study is needed to determine the long-term response of LAM treatment. Methods: We examined virologic response (HBV DNA drop drop below the level of 4 log 10 copies/ml) and viral breakthrough (HBV DNA surge more than 1 log 10 copies/ml from nadir at least two occasions after on an initial virologic response) in 77 patients who had been taking adefovir mono therapy because of LAM-resistant CHB, retrospectively. Result: The rates of virologic response at month 6 and 12 were 51.9% (40/77) and 62.3% (48/77), respectively. At base line, there was no difference in age (50.1 ± 10.9 vs. 49.8 ± 8.1), male to female ratio (66.7 vs. 82.8%), HBV DNA levels (6.8 ± 1.0 vs. 6.9 ± 1.0 log 10 copies/ml), and proportion of patients with HBeAg positive (75.0 vs. 82.8%) between the patients with and without viral response at month 12. However, the proportion of patients with acute flare prior to adefovir mono therapy was higher in responders than in non-responders (54.2 vs. 27.6%, p = 0.03). Mean duration of therapy in patients with virologic response was 31.6 ± 13.4 months. After virologic response (5-38 months) , viral breakthrough occurred in 10 patients (20.8%). Of these 10 patients, genotypic resistance to adefovir (rtN236T, rtA181V) and acute flare (ALT [120 IU/ml or bilirubin [3.0 mg/ml) were noticed in 7 (70.0%) and 3 (30.0%), respectively. Conclusion: In patients with LAM-resistant resistant CHB, adefovir mono therapy showed not only low virologic response rate but also high viral breakthrough rate. Acute flare prior to adefovir mono therapy was a factor in achieving virologic response. Among patients who showed virologic response to adefovir mono therapy, viral breakthrough without genotypic resistance to adefovir and acute flare after viral breakthrough were frequent. However, until recently there have been few commercially available assays for HBsAg quantification other than the Abbott Laboratories ARCHITECT Ò assay, which has been used in most studies. Roche Diagnostics have recently developed a prototype HBsAg quantification assay which can be run on the Elecsys instrument. Using a range of samples encompassing HBV genotypes A-D for which we had previously determined HBsAg titres using the ARCHITECT assay, we compared results to those obtained using the Elecsys system. Methods: We evaluated serum samples (n = 90) from subjects with chronic HBV infection which had previously been assayed for HBsAg and genotyped. The genotype distribution was as follows: genotype A (n = 22), B (n = 24), C (n = 24) and D (n = 20). Samples were diluted when required to fall within the dynamic range of the respective assays. Serial dilutions ranging from undiluted to 10 -6 were evaluated on the Elecsys platform. Samples in the appropriate reference range of 0-250 IU/ml, as determined using the Elecsys instrument, were re-assayed in parallel using the ARCHITECT system. Results: Data obtained using the two systems were compared using diluted serum samples with litres in the range of approximately 1-500 IU/ml. Results showed a strong correlation. Conclusions: The ARCHITECT platform has been used frequently for quantification of the HBsAg. Our results show that the prototype HBsAg quantification assay using the Elecsys platform provides a satisfactory alternative, giving comparable results. Backgroud: Here are the efficacy result in a cohort of patients who were confirmed advanced fibrosis or cirrhosis and received maximum of 9 years of monotherapy with Lamivudine. Methods: 49 Nucleoside-naïve HBeAg(+)/(-) patients who had histologically confirmed advanced fibrosis or cirrhosis (the Ishak fibrosis score or 4-6) were treated with Lamivudine for 8-9 years. The objects of this observation were to evaluation the histological improvement and the virological, ALT responses, and resistance outcomes. The progression of the diseases was also observed. Results: 1.16 (32.65%) patients had evaluable repeat biopsies. Among the total, the results of Scheuer inflammation score are as below: 6 (37.50%) patients with G0, 6 (37.50%) G1, 4 (25%) G2, and no G3 and G4 in patients were found. The Scheuer fibrosis score are as below: 4 (25%) patients with S0, 3 (18.75%) S1, 2 (12.50%) S2, 5 (31.25%) S3, 2 (12.50%) S4. It is considered that the Scheuer fibrosis score 4 is roughly equal to the Ishak fibrosis score 5-6. Background: Safety and therapeutic efficacy of a combination therapy of antiviral drug and immune modulator was assessed in IDHAS in Bangladesh. Methods: HBV infection was detected in 25 apparently healthy subjects, who visited our hospital with different motives. These patients were given lamivudine (100 mg) orally, once daily for 12 months. Hepatitis B vaccine containing hepatitis B surface antigen (10 lg) was administered intra-muscularly along with lamivudine for five times (at 0, 1, 2, 6 and 12 months). Patients were regularly followed up to assess whether the combination was safe or not. The therapeutic efficacy of combination therapy was evaluated 13 months after commencement of the clinical trial. Result: Combination therapy was safe for all patients and local or systemic adverse effects were not recorded in any patient. At the end of the trial, the levels of HBV DNA were decreased in all patients, and become undetectable in 16 patients (64%). Flare of alanine aminotransferase was not detected in any patient. Conclusion: Combination therapy with antiviral agent and immune modulator represents one of the therapeutic options for containment of the virus in asymptomatic HBV carriers. Background and Aims: Chronic hepatitis B virus (HBV) infection is a major global health issue, and the prognosis of patients with HBV-associated acuteon-chronic hepatic failure (ACLF) is extremely poor. In this study, the efficacy of lamivudine was investigated in patients with ACLF. The effects of HBV DNA load and its related factors on the prognosis were also further explored. Methods: A matched retrospective cohort study using data on ACLF patients derived from our hospital database was conducted. 130 patients receiving lamivudine were selected into the lamivudine treatment group with another 130 without lamivudine treatment studied as control. They were matched for sex, age and imaging finding with lamivudine treatment group. All the patients were followed up for 3 months and the survival rates were compared. The influential factors on the mortality were studied by Cox proportional hazards model. Results: The cumulative survival rates of patients in lamivudine group were higher than those of the control group (v 2 = 9.50, P = 0.0021). The mortality of patients with high virus load group (71/95, 74.7%) was higher than that of those with low virus load group (15/29, 51.7%) (v 2 = 5.536, P = 0.019). For patients with MELD score 20-30, by week 4, the mortality of those with HBV DNA undetectable or declined for more than 2 log 10 (2/12, 16.7%; 18/40, 45.0%) was lower than that of those with a less than 2 log 10 decline (18/23, 78.3%) (v 2 = 10.106, P = 0.001). In Cox proportional hazards model, for patients with MELD score 20-30, treatment method (P = 0.002), pretreatment HBV DNA load (P = 0.007) and decline of HBV DNA load during therapy (P = 0.003) were independent predictors; for those with MELD score above 30, MELD score (P = 0.008) was the only independent predictor. Conclusions: Lamivudine can significantly decrease the 3-month mortality of patients with MELD score 20-30, and a low pre-treatment viral load and rapid decline of HBV DNA load are good predictors for the outcome of the treatment.

A 600 HIV-HBV co-infected mothers. Little is known about the efficacy, tolerability and safety of TDF in HBV monoinfected mothers. We report the outcomes of five pregnant women with CHB being treated with TDF at different stages of pregnancy. Methods: Data was collected retrospectively from five women with CHB being treated with tenofovir at the New York Downtown Hospital January-October 2009. Treatment was initiated in two cases for active disease, two for perinatal transmission prophylaxis, and in one case pregnancy occurred during treatment (case#3). All were registered with The Antiretroviral Pregnancy Registry. Their HBV-DNA and ALT were recorded at initiation of therapy, monthly, and at delivery, along with neonatal outcomes.

Result: See Table 1 .

Conclusions: TDF use in the third trimester of pregnancy was associated with a 3-4 log 10 reduction in HBV-during the first 4-weeks and an absolute level B10 5 copies/ml at delivery. No newborn birth defects were noted. Infants' HBsAg and anti-HBs is being monitored. TDF, appears to be safe, effective and well tolerated in pregnant CHB patients. Prospective studies on its role in preventing perinatal transmission are warranted. Background and Aims: Lamivudine is a potent antiviral agent against hepatitis B. This study was aimed to find out the significance of long-term lamivudine therapy for patients with chronic HBV infection. Methods: Two hundred seventy-six cases of various stages of chronic hepatitis B were included. Of them 174 had chronic hepatitis (group A), 56 compensated cirrhosis (group B), and 46 decompensated cirrhosis (group C). Baseline serum HBV-DNA [ 5 log(10) copies/mL and with elevated alanine aminotransferase (ALT) over the upper normal limit or complications of hepatic insufficiency, were administered 100 mg of lamivudine daily and monitored for HBV markers, biochemistry, and prognosis. Results: Lamivudine reduced HBV-DNA (group A 79%, group B 75% and group C 68%) and ALT (group A 69%, group B 65% and group C 70%) in all groups after one year of therapy (p \ 0.05). Cumulative incidence of virologic breakthrough was 12, 32, 39, and 51% at 12, 24, 36, and 48 months, respectively, and the strongest predictive factor for lamivudine resistance was persistent HBV-DNA at 6 months. Ascites, encephalopathy, and jaundice improved in the majority of patients with decompensated cirrhosis. On the other hand, hepatic failure developed or deteriorated in nine patients after virologic breakthrough, and seven of them had decompensated cirrhosis.

Conclusions: Lamivudine was effective in reducing HBV-DNA and improving hepatic reserve at all stages and was most beneficial and significant for decompensated cirrhosis. Meanwhile, close monitoring of viral load and immediate rescue treatment for lamivudine resistance is necessary to prevent hepatic failure in decompensated cirrhosis. Objective: To analyze the efficacy of adefovir dipivoxil combined with bicyclol in the treatment of chronic hepatitis B (CHB) and determine its safety. Methods: A total of 125 patients with CHB were randomized into the experimental group and the control group to be treated.The patients in the experimental group (63 samples) received adefovir dipivoxil orally 10 mg daily and bicyclol orally 75 mg daily for 48 weeks and those in control group (62 samples) received adefovir dipivoxil orally 10 mg daily alone for 48 weeks.The serum aminotransferase (ALT/AST), HBV-DNA, HBeAg/anti-HBe were observed before and after treatment. Results: Compared with pre-treatment, the serum aminotransferase were all decreased obviously in two groups, the experimental group is better (P \ 0.05, .01). HBVDNA negative conversion rate was significantly higher in the experimental group than that in the control group (58.7 vs. 40.3%, P \ 0.05).

HBeAg loss rate was significantly higher in the experimental group than in control group (31.8 vs. 16.1%, P \ 0.05). Although the experimental group is higher than the control group in the aspect of HBeAg serumconversion rate, there were no statistical difference between the two groups. There were no obvious adverse events which were probably related to the drug in the study. Conclusion: Adefovir dipivoxil combined with bicyclol is effective and safe in the treatment of chronic hepatitis B. were included in this study after meeting the following criteria: age 18-60 years, HBsAg positive for more than 6 months, serum HBV-DNA was [5 log (10) copies/ml and ALT more than two times the upper normal limit at least two subsequent occasions. They were given peginterferon alfa-2b (80 lg once weekly) for 24 weeks and followed for an additional 24 weeks. Results: After 24 weeks of follow-up, the number of patients with normalization of alanine aminotransferase levels was similar in both groups (51 vs. 54%, p = ns). However, serum HBVDNA levels below 500 copies/ml was significantly higher in HBeAg negative patients (59%) Background: In patients with chronic hepatitis B virus (HBV) infection, upregulation of the expression of the programmed death molecule (PD-1) and its ligand (PD-L1) on T cells is associated with impaired T-cell activation and function. We examined the relationship between telbivudine-induced viremia reduction and HBeAg seroconversion with regard to PD-1/PD-L1 levels and Tcell reactivity in hepatitis B e antigen (HBeAg)-positive patients with chronic hepatitis B (CHB). Methods: Ten (8 male, 2 female) patients seropositive for hepatitis B surface antigen and HBeAg, with HBV DNA levels [10 5 copies/mL and alanine aminotransferase levels [29 the upper limit of normal, received telbivudine 600 mg/day. Peripheral blood mononuclear cells were extracted from heparinized blood samples obtained from patients at baseline and weeks 12 and 24 of telbivudine treatment. PD-1 and PD-L1 expression were assessed by flow cytometry. HBV-specific CD8+ T cells were quantitated by pentamer staining. T-cell reactivity to HBV antigens was determined by interferon-c (IFN-c) assessed by enzyme-linked immunosorbent assay. Results: During treatment, two patients experienced HBeAg loss, two patients achieved HBeAg seroconversion, and six patients remained HBeAg positive. Reduction in PD-1 and PD-L1 expression correlated closely with levels of viremia at treatment weeks 12 and 24. PD-1 and PD-L1 expression decreased markedly on total CD8+ T cells and HBV-specific CD8+ T cells; this decrease was associated with an increase in IFN-c production. HBeAg seroconversion resulted in a further reduction in PD-1 and PD-L1 expression, an increased frequency of HBV-specific CD8+ T cells, and increased IFN-c production. Conclusions: The telbivudine-induced suppression of HBV replication reduces PD-1 expression, which partially restores the function of HBV-specific CD8+T cells.

The Potential of Dendritic HBV genotypic resistance testing could not be performed due to relatively low viremia. Telbivudine 600 mg/day was then added to adefovir. Serum HBV DNA was undetectable within a month and remained undetectable at 6 months. The patient was continued on combination telbivudine and adefovir without adverse effects. Serum HBV DNA was monitored every 6 months, and hepatocellular carcinoma and esophageal varices surveillance was also performed. Conclusions: Early add-on telbivudine therapy suppressed HBV DNA and normalized ALT in a compensated cirrhotic patient who failed sequential treatment with lamivudine/adefovir. HBV DNA level at the time of add-on therapy may have an important impact on the development of sequential resistance.

dipivoxil (ADV) and Telbivudine (LDT) are highly effective in the suppression of HBV replication. however, host immune responsible for sustained virological respones in HBV-infected patients treated with nucleos(t)ide analogs are not understood. This study aims to evaluate the immune response and antiviral effect after nucleos(t)ide analogs or thymosin-alpha 1 (TA1) treatment. Methods: Serial analysis of intracellular or serum cytokines including IL-2, IFN-c, TNF-a, IL-4 and regulatory T cells (Tregs), CD4 and CD8 were measured using a three-color flow cytometry before and at 12, 24, 36 and 48 weeks after treatment. Samples were also tested for HBV DNA, HBsAg, HBeAg, ALT and AST. Results: The cytokine productions were significantly increased from 24 to 48 weeks after nucleos(t)ide analogs treatments, higher levels of Th1 cytokines, IL-2, IFN-c and TNF-a were observed with ETV (n = 29) compared to ADV (n = 28), LDT (n = 41) or TA1(n = 19). By contrast, the levels of Tregs in all groups were markedly decreased after 24 weeks. This altered cytokine profile and cellular component is accompanied by a decreased HBV DNA and HBsAg levels in nucleos(t)ide analogs treatment group.

Conclusions: The enhanced immune response related to ADV, LDT and ETV treatment suggests that the anti-viral effect of the drugs may not only be attributed to its direct effect on virus suppression but also to its immune regulation capability.While the underlying mechanism remains to be understood, reduced virus load and Tregs may favor the restoration of anti-virus immunity in CHB. Furthermore, in CHB patients, the altered cytokine levels and T cell subsets, in particular the Tregs, may provide additional biomarkers for the immune response diagnosis and prediction.

Outcome Background: Lamivudine (LMV) and adefovir dipivoxil (ADV) combination therapy is a pertinent rescue in pateints with LMV-resistant chronic hepatitis B (CHB). But it dose not always guarantee an excellent result in HBeAg-positive CHB patients. We intended to investigate the association of clinical factors with virological response of LMV-ADV combination therapy in LMV-resistant CHB.

Methods: A total of 60 LMV-resistant CHB patients were treated with LMV-ADV combination for more than 6 months. Virological response (serum HBV-DNA\100 copies/mL) was evaluated during follow-up period and analyzed in the accordance with clinical characteristics. Results: All of 60 patients had YMDD mutations with virological breakthrough and were followed for a median period of 18 (ranged 6-34) months [age = 44 (ranged 20-68); Male:female = 46:14). HBeAg was positive in 47 (78%) patients and the baseline serum HBV-DNA level was 7.23 (ranged 3.9-9.7) log 10 copies/mL. The initial ALT level was 91 (ranged 16-1,665) IU/L and 38 (63%) patients were treated after biochemical breakthrough (ALT [80 IU/L). One patient was found to have ADV-resistant mutation after 22 months combination therapy. Cumulative virological response was 8, 18, and 30% at 6, 12, and 18 months, respectively. HBeAg loss occurred in two of the subjects. In multivariate analysis, female gender (P = 0.014) and HBeAg negativity (P = 0.024) were independent predictive factors for virological response. YMDD mutants test was followed at 16 months (ranged 9-27) in selected eight of the patients with HBeAg-positive among 25 patients of partial viral responder. However, all eight of the patients persistently showed YMDD mutants.

Conclusions: LMV-ADV combination therapy insufficiently suppressed viral load and the initial YMDD mutants were persistent in HBeAg-positive patients with LMV-resistant CHB. Background and Aims: ALT flare is a well-known phenomenon during interferon treatment for chronic hepatitis B. However, little is known about these flares during nucleos(t)ide treatment and the effects on long-term clinical outcomes.. Methods: A total of 170 naïve HBeAg-positive chronic hepatitis B patients were treated with one of the nucleos(t)ide analogues, lamivudine, adefovir, entecavir, or telbivudine, for at least 2 years and followed up for 1 more year. Clinical characteristics were detected and analyzed at baseline and at 3-month intervals.

Results: Two patterns of ALT flares, viral-induced and host-induced flare, were found. Viral-induced ALT flares were more common than host-induced ALT flares (15.9 vs.6.5%, P \ 0.05), characterized by rapid increased ALT and HBV DNA usually after two-years treatment. The median of ALT increased fold was 5.2 times the upper limit of normal (ULN), most of these patients had detected HBV mutations. Host-induced ALT flares had moderate elevated ALT (median 2.5-fold ULN) with a slow decreased HBV DNA procedure, occurred mainly in the first year (63.6%) with favorable long-term treatment outcomes. Besides the 3-years HBV DNA undetectable rate in the viral-induced and hostinduced ALT flare at 8 vs. 85%, HBeAg seroconversion and histological improvement was 10 vs. 85%, and 2 vs. 40%, respectively. Moreover, interferon-gamma expressed T helper cells were higher in host-induced viral ALT flares which indicated that immune system may involved specifically. Objectives: To evaluate the efficacy of combination of (TCM) and western medicine for severe hepatitis. Methods: Searching in CNKI and CBMdisc from 2002 to 2006, indexed words including severe hepatitis, Hepatic necrosis, Hepatic failure, combination of TCM and western medicine, traditional Chinese medicine, random and so on, objects investigated were limited within those whose diagnosis standard were either in accordance with the standard produced by infectious disease and Parasitosis academic conference in 1995 (Beijing) or consistent with the standard of viral hepatitis by infectious disease and Parasitosis academic conference in 2000, intervention study of both combination of TCM and western medicine and modern medicine was required, also randomized controlled trials were needed; Results: 18 articles were studied. there were two groups covering 1,525 Cases involving combination of TCM and western medicine and modern medicine, it was shown that differences were obviously (P \ 0.001): for combination of TCM and western medicine,the effective rate was 60.15% and fatality rate was 35.36%, respectively,in contrast with them,the effective rate was 41.88% and fatality rate was 53.20% for modern medicine.

Conclusion: There are more advantages for combination of TCM and western medicine than modern medicine for severe hepatitis in improving effective rate and reducing fatality rate, and selected combination therapy of TCM and western medicine is of clinical significance and wide application for severe hepatitis.

Short- Introduction: Adefovir dipivoxil (ADV) has been used widely in the treatment of chronic hepatitis B (CHB). It is known to cause osteomalacia in high doses but rarely does so in the context of CHB therapy. We report here, two cases of severe osteomalacia in patients on ADV given at the standard dose of 10 mg daily.

Both patients had compensated cirrhosis (Child-Pugh Class A) caused by CHB and were initially treated with lamivudine. ADV was added at the emergence of lamivudine resistance. The patients were Cambodian Chinese (female aged 53 yr and male 40 yr). Relevant background include partial gastrectomy in the female, hyperthyroidism and possible prior NSAID use in the male. Symptomatic osteomalacia manifested after 4 years and 2 years of ADV. Osteomalacia was diagnosed on clinical, biochemical and imaging grounds. -Serum ALP peaked at 610 U/L and 255U/L (NR 30-115). Both biochemical abnormalities predated clinical manifestation of osteomalacia by more than a year. Hypophosphatemic renal impairment was evident with phosphate 0.57 mmol/L (NR 0.8-1.5), e-GFR 46 (N[ 59 mL/min/1.73m2) and 0.49 and 56, respectively. Renal tubular dysfunction was manifested by phosphaturia, glycosuria and proteinuria. ADV was withdrawn and supplements of calcium, phosphate and vitamin D given. There was good clinical recovery in both cases. Review of 67 consecutive patients on ADV in one single clinic revealed 6 cases with mild decrease in eGFR (50-58 mL/min/1.73m2) and 1 patient with minor hypophosphataemia (0.76 mmol/L). Discussion: These cases represent a rare adverse reaction to ADV with severe osteomalacia associated with Fanconi's type renal tubular dysfunction. Patients with cirrhosis and other co-morbidity such as partial gastrectomy may be most vulnerable and should therefore be monitored closely when ADV is prescribed. Measurement of ALP and phosphate should be part of routine follow up. Background/Aims: A large proportion of patients fail lamivudine treatment and guidelines recommend switching to nucleotides or add-on therapy. The aim of this report is to evaluate the efficacy and safety of the switch to adefovir (ADV) monotherapy versus telbivudine (LDT) plus adefovir in CHB patients with confirmed YMDD mutation with prior lamivudine treatment. Methods: 150 HBeAg positive patients were planned to be included into an open label randomized 96-week trial evaluating ADV versus ADV + LDT. The study was terminated early due to difficulties enrolling patients into the ADV monotherapy arm. However, 42 patients were randomized 1:1 (91% Korean). Results: Median treatment duration was 48 weeks and 20 patients per arm completed 48 weeks of treatment. HBV DNA decline at week 24 was higher in the combination arm versus ADV monotherapy arm: -6.7 versus -5.0 log 10 copies/ml (baseline ADV + LDT:10.3; ADV:10.1 log 10 copies/ml). At week 48, HBV DNA decline by -7.4 (ADV + LDT) and -4.9 (ADV) log 10 copies/ml, with 38.5% (5/13) and 0% (0/9) of patients, respectively, achieving undetectable HBV DNA (\300 copies/ml). Virologic breakthrough occurred in 0% (0/21) of ADV + LDT and 9.6% (2/21) of ADV patients. HBeAg loss was achieved in one patient per arm at week 24 and in 23.1% (3/13) and 0% (0/9) at week 48 in the ADV+LDT versus ADV groups. The safety profile was similar in both treatment arms with 16 AEs reported per group. ALT flares (AASLD criteria) occurred in one patient per arm, increased creatine phosphokinase was reported in 3/21 patients ADV + LDT and 1/21 ADV arm, but all were grade 1/2. One case of musculoskeletal pain and parasthesia each occurred in the combination arm. Conclusion: In these 42 patients with established YMDD resistance on lamivudine treatment, the switch to the combination of telbivudine and adefovir showed better outcomes compared with adefovir alone, and both had a similar safety profile. levels (28% \300 copies/ml and 28% \4 log 10 copies/ml). Of the remaining 11 patients who did not sustain HBeAg seroconversion two maintained HBeAg loss and remained HBV DNA negative. Seroreversion (HBeAgpositive and HBeAb-negative) occurred in nine patients during the first 36 weeks follow-up. No seroreversion was observed after 36 weeks. Durability of HBeAg seroconversion was higher in Genotype B (100%, 19/19) than in Genotype C (74%, 28/38) patients. All nine patients who experienced seroreversion were Genotype C. Conclusion: HBeAg seroconversion induced by telbivudine is durable after stopping treatment for 2 years. The majority of patients have HBV DNA levels \4 log 10 copies/ml and do not require treatment as recommended by international guidelines. These data indicate, that treatment with telbivudine can be discontinued in patients with HBeAg seroconversion and low levels of HBV DNA. Results: At week 48, mean reduction in serum HBV DNA levels from baseline was -5.3 log 10 IU/mL in entecavir group and -4.9 log 10 IU/mL in clevudine group (P = 0.089). The proportions of patients with undetectable serum HBV DNA levels (\13 IU/mL by real-time PCR) at week 48 was 81.2% in entecavir group and 67.2% in clevudine group (P = 0.062). In addition, the rates of ALT normalization and HBeAg loss or seroconversion were not significantly different between two groups (75.4 vs. 77.6%, 23.3 vs. 12.8%; P = 0.757, 0.194, respectively). However, the incidence of virologic breakthrough in the entecavir group was significantly lower than in the clevudine group (0 vs. 7.5%; P = 0.021). Multivariate analysis showed that serum HBV DNA levels at week 24 C2,000 IU/mL was an only independent factor that affected virologic breakthrough (P = 0.005). Conclusions: Entecavir therapy showed a non-significantly higher virologic response and significantly lower resistance at week 48 than clevudine therapy. and improve outcomes. Treatment options are limited, with tenofovir associated with risk of further renal dysfunction and increased risk of lactic acidosis recently reported in cirrhotic patients with entecavir. The aim of this study is to evaluate efficacy and safety of telbivudine (LDT) and lamivudine (LAM) in decompensated CHB patients using a non-inferiority statistical analysis. Aims: To assess initial virological response (IVR) to lamivudine (3TC) and entecavir (ETV) treatment for naïve chronic hepatitis B patients, to identify patients with suboptimal response and to assess IVR to ETV and combination of 3TC and adefovir dipivoxil (ADV) for 3TC-resistants. Methods: One hundred seventy-four patients who received nucleos(t)ide analogues were retrospectively investigated. Initially 155 treatment-naïve patients were enrolled. Seventy-one were administered 100 mg 3TC for 51 ± 28 months; 84 were 0.5 mg ETV for 18 ± 16 months. Thereafter, efficacy of ETV and that of combination 10 mg ADV with 3TC were examined in 28 patients resistant to 3TC for 39 ± 15 and for 27 ± 18 months, respectively. Antiviral failure was evaluated based on the definition reported by A. Gallego (J Viral Hepat, 2008) . Results: Primary antiviral treatment failure for naïve cases in ETV is similar to that in 3TC, however, IVR is superior in ETV to in 3TC (91.9 vs. 78.4%, p = 0.0458, Fisher's test). Naïve patients treated with 3TC, in which 71.4% were e antigen positive, showed a higher risk of secondary antiviral treatment failure (SATF) than those treated with ETV (P = 0.007, Logrank test). SATF for these treated with 3TC was significantly higher in e antigen-positive cases (P = 0.033, Fisher's test). 3TC-resistant patients treated with ADV and 3TC showed a lower risk of SATF than those treated with ETV (P = 0.0143, Logrank test). These patients were e antigen-positive, and their nucleotide sequences had ETV-resistant mutations including 3TC-resistances. Conclusions: ETV is superior to LAM for e antigen-positive naïve Japanese patients. However, combination therapy of 3TC and ADV is recommended for those resistant to 3TC. Background: Chronic immunosuppression is a mainstay of treatment for rheumatologic diseases. However, it has been associated with hepatitis B reactivation. This review focused on the effectiveness of lamivudine as prophylaxis or treatment of hepatitis B reactivation among patients with rheumatologic diseases on chronic immunosuppression.

Methods: A search of Medline, Pubmed and Cochrane databases was performed to retrieve trials on rheumatologic patients given lamivudine for Hepatitis B. Search yielded 18 studies including 4 observational cohort studies (1 prospective, 3 retrospective). Case reports and review articles were excluded. Authors were contacted to get fulltext articles. Results: Forty-four rheumatologic patients on prednisolone alone or in combination with DMARDS or biologics, received Lamivudine 100 mg/day as treatment (n = 22) for reactivation or as prophylaxis (n = 22). One studyincluded five Lupus Nephritis patients while the other three studies included patients with Rheumatoid Arthritis (n = 14), Systemic Lupus Erythematosus (n = 6), Ankylosing Spondylitis (n = 4), Polymyalgia Rheumatica (n = 5), Psoriatic Arthritis (n = 3) and one patient each with Systemic Sclerosis, Sjogren's Syndrome, Dermatomyositis/Polymyositis, Takayasu Arteritis, Henoch-Schonlein Purpura and Behcet's syndrome. Elevated levels of alanine transferase (ALT) at baseline (n = 22) normalized shortly after lamivudine therapy. HBV-DNA levels were significantly suppressed in 17 patients after treatment. Two patients developed treatment-resistant YMDD mutation of HBV and had to be shifted to adefovir. There were no major adverse events reported and lamivudine treatment appeared safe and well-tolerated. Conclusion: Lamivudine as treatment and prophylaxis for hepatitis B reactivation seems to be a promising strategy in rheumatologic patients on chronic immunosuppression. However, the studies are limited by small sample sizes and heterogeneity, in terms of the type of rheumatologic disease, type and dosage of immunosuppressive drugs, as well as the duration of lamivudine treatment. There is a need for further prospective, preferably randomized controlled trials including a larger set of patients. Background: Adefovir mono-switch has been accepted as an effective rescue therapy in patients with lamivudine-resistant chronic hepatitis B. Recently, adefovir add-on therapy is recommended in aspect of the antiviral potency and resistance. We aimed to compare the clinical efficacy between adefovir monoswitch and adefovir add-on therapy in lamivudine-resistant patients. Methods: Sixty-one patients with chronic hepatitis B, who underwent a rescue therapy due to lamivudine resistance, were retrospectively investigated. After confirmation of genotypic resistance for lamivudine, twenty-nine patients were shown to receive adefovir monotherapy and 32 patients were treated with adefovir add-on therapy. We compared the biochemical, virological, and serological responses during 48 months of the investigation. Results: Baseline clinical, biochemical, virological characteristics presented no significant difference in two groups prior to rescue therapy. ALT normalization rate at 48 months was 89.7% in monotherapy and 87.5% in add-on therapy.

The virologic response at 48 months was 89.3% in monotherapy and 87.1% in add-on therapy. Patients with monotherapy or add-on therapy had 58.6 and 50.0% of undetectable HBV DNA titer at 48 months, respectively. The rates of HBeAg loss with seroconversion were 26.5 and 29.4% in monotherapy and add-on therapy, respectively. Time to seroconversion was shorter in combination therapy (11.8 months) than monotherapy (7.2 months), but not significance. There were no factors that showed significant difference in treatment efficacy. Following 48 months of treatment, greater HBV DNA breakthrough was observed in monotherapy than add-on therapy (24.1 vs. 9.7%, p \ 0.05). Adefovir-related genotypic resistance (rtA181 V) was confirmed in two monotherapy patients with viral breakthrough. Conclusions: The biochemical, virological and serological responses in both rescue regimens showed not much difference in patients with lamivudineresistant chronic hepatitis B, except viral breakthrough being more often in adefovir mono-switch. Background: Pokeweed antiviral protein (PAP) is a natural products that inhibits the replication of a number of virus, our recent study indicated PAP exibited potent anti-HBV activity, the major obstacle in applying PAP as an anti-HBV weapon is the challenge to deliver it to the liver specifically and efficiently without cytotoxicity. Aims: To develop a new strategy that ensue specfic and effective down regulated HBV gene expression and replication by PAP. Method: The full-length, N-terminal and C-terminal truncated PAP expression vector were co-tansfected with 1.3-fold replication competent HBV construct (pHBV1.3) to HepG2 cell respectively, HBsAg and HBeAg in supernatants were determined by using ELISA, Viral DNA intermediates were detected by Southernblot and cytotoxicity were evaluated by MTT assays. Recombinant HBV vector was constructed by modifying the wild type HBV (genotype A), a 405-nt fragment within core gene was replaced by a in frame C-terminal 25 amino acid truncated PAP (PAPc), with polymerase ORF intact.Expression of PAP was identified by way of Western blot assay, rescue of rHBV replication and its anti-HBV activity were evaluated by co-transfected with HBV core expression plasmid and wt-pHBV1.3, or transfected to HepG2.2.15 cell. Results: The full-length and PAPc could effectively down regulate extracellular production of HBsAg and HBeAg and the levels of intracellular coreassociated viral DNA. More interestingly, PAPc almost has no cytotoxicity than that of full-length by MTT assay. A recombinant HBV (rHBV) containing a modified viral core gene that specifically delivers PAPc to the liver. This recombinant virus could be self-maintained in hepatocytes through capsid complementation by wild type HBV or core expression plasmid, and exhibit anti-viral activity in pHBV1.3 co-transfected HepG2 or HepG2.2.15 cell . Conclusion: The N-terminal 69 amino acid of PAP is necessary for its anti-HBV acvity, and C-terminal 25 amino acid is related to its cytotoxicity. HBV as gene delivery vector, that specifically targets the liver and mediated liver specific anti-HBV activity of PAPc. It might represent a promising strategy capable of knocking-down and eradicating the resident HBV in chronically infected patients

The Results: The sequences of four human A3 moleculars containing double cytosine deaminase domain were cloned from PBMC. A3G suppress HBV replication in cell culture and HBV based mouse model.However, the catalytically inactive A3G derivatives and the RNA binding site mutation of A3G conserve its wild-type activity on HBV replication. In co-transfected human hepatoma cells and HBV based mouse model, A3B, A3DE and A3F also have inhibitory effect on HBV production. Conclusions: A3G, containing double cytosine deaminase domain could inhibit HBV replication in vitro and in vivo, however, A3G-mediated editing dose not seem to represent the mechanism against HBV replication.Like A3G, the editing enzymes (A3B, A3F, A3DE) which contains double cytosine deaminase domain could also inhibit HBV replication in vitro and in vivo, and contribute to the innate immunity against HBV. A total of 84 patients were divided into antiviral group and control groups (without antiviral therapy). 44 patients were treated with nucleosides (including lamivudine 100 mg/day, adeforvir dipivoxil 10 mg/day, entecavir 0.5 mg/day) and routine supportive therapy. 40 patients were only treated with routine supportive therapy. The clinic data were observed before and after treatment at weeks 2 and 4 for the biochemical data, viral load, MELD score, survival rate, actuality mortality of two groups. Result: After treatment of 2 weeks the Tbil, PTA, INR in antiviral group had been improved more than in control group (P \ 0.05). MELD score and martality of the antivival group at weeks 2 and 4 had been significantly improved comparing before and after treatment (P \ 0.05), but control group had no significant difference (P [ 0.05). After treatment 4 weeks decrease and negative rate of HBV DNA in antiviral group were markedly more than in control group (P \ 0.05), meanwhile the incidents of complication in antiviral group were less than in control group (P \ 0.05). Conclusion: Antiviral therapy can improve the prognosis of chronic hepatitis B in short term and is benefitable for chronic hepatitis B patients.

Research on Anti-HBV Activities of Four Flavonoids from Halenia Elliptica D. Backgroud: As one of the eight majior Tibetan medicines, Halenia elliptica D. Don has been used to treat icterohepatitis for hundreds of years, of which flavoniods has been confirmed as active ingredients. Methods: HepG2215 served as experimental cell line, and HepG2 served as control. Supernatants were collected 6 days after cultivation in DMEM con-taining 1-HO-2,3,4,8-MeO-Xanthone(I), 1-HO-2,3,4,5-MeO-Xanthone(III), 1-HO-2,3,5-MeO-Xanthone(IV),5,7,3 0 ,4 0 -tetrahydroxyl flavonoid(X) and lamivudine(5ug/ml), respectively. Hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) were measured by enzyme linked immunosorbent assay (ELISA). Hepatitis B virus DNA was measured by fluorescent quantitation polymerase chain reaction (PCR). Cells were harvested at 6 days, HBV replication intermediate was detected by southern blot. Cytotoxicity test was carried out via cell count kit-8(CCK8). Results: After 6 days, the rate of I, III, IV, X and lamivudine for cellular HBsAg secretion was 37.00 ± 19.75% (P \ 0.05), 8.25 ± 3.78% (P \ 0.05), 7.25 ± 3.60% (P \ 0.01), 4.50 ± 4.20% (P = 0.122) and 21.00 ± 11.40% (P \ 0.05), respectively. The inhibition rate for HBeAg secretion was 41.00 ± 16.01% (P \ 0.05), 16.33 ± 5.13% (P \ 0.05), 21.67 ± 8.15% (P \ 0.01), 10.33 ± 4.04% (P \ 0.05) and 27.00 ± 1.73% (P \ 0.01), respectively. The inhibition rate for HBV-DNA secretion was 52.67 ± 5.77% (P \ 0.05), 39.00 ± 12.29% (P \ 0.05), 42.33 ± 8.51% (P \ 0.01), 21.67 ± 2.31% (P \ 0.05) and 62.00 ± 16.27% (P \ 0.05), respectively. The inhibition rate for HBV-DNA replication intermediate (RI) was 22.33 ± 8.62% (P \ 0.05), 16.07 ± 4.33% (P \ 0.05), 16.83 ± 4.54% (P \ 0.01), 40.83 ± 4.55% (P \ 0.05) and 55.30 ± 6.69% (P \ 0.05), respectively. Conclusions: Anti-HBV effects of the four flavonoids from Halenia elliptica D. Don were confirmed, which were associated with the site of methoxyl.

Background and Aims: De novo HBV-related hepatitis has been reported in patients after hematopoietic stem cell transplantation (HSCT) and cytotoxic chemotherapy treatment. We conducted a retrospective study with aim of identifying clinical characteristics associated with de novo HBV-related hepatitis. Methods: Fifty seven patients with hepatitis B surface antigen negative who underwent allogeneic HSCT in our hospital, from January 2005 to September 2007, and were followed at least 1 year after the transplantation were enrolled in this study. Seventeen of these patients were positive for anti-hepatitis B core antigen (anti-HBc) and 40 patients were negative. Results: Five of 17 anti-HBc positive patients had de novo HBV-related hepatitis after the transplantation and none of 40 anti-HBc negative did. Median time from transplantation to the event was 14 months (range 7-18 months) and the median peak elevation of AST, ALT and bilirubin were 291 U/l (range 79-1,103 U/l), 243 U/l (range 130-1,340 U/l), and 1.4 mg/ml (range 0.3-8.6 mg/ml), respectively. All five patients were treated with nucleotide analog and resolved after treatment. There was no significant clinical difference between the patients with de novo HBV-related hepatitis (n = 5) and those without hepatitis (n = 10) in age, sex, titer of anti-HBc, with or without HBs-antibody, however the patients with de novo HBV-related hepatitis tended to have more severe chronic GVHD (extended type) than the patients without hepatits (p = 0.08). Conclusion: Among 57 patients with HBs antigen negative who underwent allogeneic HSCT, 5 patients (8.6%) had de novo HBV-related hepatitis. The patients with extended type of chronic GVHD will need more careful attention to de novo HBV-related hepatitis. Aim: To evaluate the efficence of Adefovir alone or combined with Adefovir or Entecavir in the treatment of chronic hepatitis B patients following Lamivudine initial no response. Methods: fifty nine chronic hepatitis B patients with lamivudine initial no response were enrolled,and divided into group A (n = 40) who received the therapy of Lamivudine combined with Adefovir, group B (n = 10) who switched to Adefovir alone and groups C (n = 15) who switched to Entecavir alone or Entecavir combined with Adefovir for 15 months. Liver function tests and serum virologic responses were evaluated at regular interval in all patients. Results: Normalization rate of liver function in the three groups were similar, but HBV DNA seroconversion rate and HBeAg/anti-HBe seroconversion rate in the treatment group A were significantly higher than in group B and C (p \ 0.05), HBV DNA negative rate was up to 100% and HBeAg/anti-HBe seroconversion rate was up to 17.5% at 15 month in group A and group C, which was significantly higher than group B (p \ 0.05). Conclusions: The results of different regimen for CHB suggest that the combined therapy with Adefovir as subsequent antiviral therapy following lamivudine initial no response may be beneficial for the chronic hepatitis B. It needed further clinical trail in multicenter to verify the results. Methods: Patients treated with PEGASYS according to local label recommendations are eligible for inclusion and will be followed for up to 3 years post-treatment to assess parameters of response including quantitative HBsAg and HBeAg levels. Baseline data from the first 100 patients enrolled in the study are presented. Results: A total of 380 patients from nine countries have been recruited. Analysis of the first 100 patients enrolled shows: the majority are male (73%) and of Asian/ Oriental origin (75%). Mean age is 34.8 (range 18-67) years. In patients with available data, 55.3% have HBeAg-positive disease, the most common infecting genotypes are HBV genotype C and D, and the mean assumed duration of infection is 15.9 ± 12.0 years. Baseline laboratory data are provided (Table) .

Conclusions: This international observational cohort will provide 'real life' data to identify baseline and on-treatment predictors of sustained response and eventual HBsAg clearance with PEGASYS in routine clinical practice. Analysis of the first 100 patients enrolled shows that we can anticipate a large amount of evaluable data, including data on antigen quantification. This should provide valuable information regarding the characteristics of patients most suitable for treatment as well as strategies to individualize therapy in order to improve the cost/benefit ratio of therapy.

Baseline laboratory data evaluations for the first 100 patients enrolled year post-treatment, 20 (28%) went on to achieve HBsAg clearance 5 years post-treatment. Rates of response 1 and 5 years post-treatment were significantly associated with on-treatment HBsAg decline (Table) . Rate of HBsAg clearance 5 years post-treatment was highest in patients with a C10% decline in HBsAg at week 12 or 24 and sustained immune control 1 year post-treatment, 40% (10/25) and 45% (13/29), respectively. Conclusion: A C10% on-treatment HBsAg decline with PEGASYS in HBeAgnegative patients is associated with high rates of sustained immune control 1 year post-treatment-an important step towards subsequent HBsAg clearance. Results: Overall rates of HBeAg seroconversion 6 months post-treatment (sustained immune control) and HBsAg clearance were 31% (122/399) and 4% (17/399), respectively. Patients were categorized according to HBsAg level at weeks 12 and 24; the proportion of patients achieving HBsAg B1,500 IU/mL increased from 23% at week 12 to 34% at week 24. On-treatment HBsAg level was significantly associated with sustained immune control 6 months posttreatment ( Figure) . The rates of HBsAg clearance 6 months post-treatment among those who achieved HBsAg B1,500 IU/mL at week 12 or 24 and sustained immune control 6 months post-treatment were 18% (9/51) and 20% (15/74), respectively. Conclusion: On-treatment quantification of HBsAg levels during treatment with PEGASYS can identify patients with a high chance of achieving sustained immune control and HBsAg clearance as well as those less likely to achieve sustained response. This may help guide patient management in the future. As compared with retreatment patients, initial treatment patients had no significant differences in undetectable serum HBV DNA levels, HBeAg loss and seroconversion,and normalization of ALT at week 48, in the entecavir group and the adefovir group respectively. Conclusion and discussion: The entecavir group had significantly higher rate in HBeAg scroconvcrsion and superior reduction in HBV DNA than that in the adefovir group at weeks 24 and 48. As for retreatment patients, entecavir should be given priority to treatment. Background/Aim: Current guidelines recommend antiviral therapy for chronic hepatitis B (CHB) patients with elevated ALT. Little clinical treatment data exists for CHB patients with persistently normal ALT levels (PNAL). The aim of the study is to find virologicll effect of peginerferon a-2a treatment in CHB patients with PNAL and significant histological findings (significant necroinflammation with a score of HAI 4 or greater and/or significant fibrosis Ishak's staged at 3 or greater). Methods: The study of all patients with CHB who underwent percutaneous liver biopsy and who had detectable viral load (Taqman Real Time PCR, lower limit of detection is 10 3 copies/ml) was performed. 14 patients who had PNAL (defined as normal ALT measured on at least 3 occasions in the intervals of more than two months over a period of 12 or more months apart prior to biopsy) were identified. These patients were compared with 13 patients with abnormal ALT (C2ULN) during the same period and who had no prior antiviral treatment. Background: Currently available anti-virals are either contraindicated or ineffective in children, specially of Asian origin. The two drugs recommended for children, namely lamivudine and interferon yield frustrating results. The aim of this study was to see the outcome of sequential, combination therapy with lamivudin plus interferon in immunotolerant Bangladeshi children. Methods: We recruited 12 children in this prospective study. They were between 4 and 14 years. Of them, six were boys and six girls. Written informed consent was taken from their parents. They received lamivudine (3 mg/kg body weight) for 2 months, followed by lamivudine (3 mg/kg) plus interferon (3 MIU/sq.m) for another 10 months. They were under follow up during treatment to assess response and manage complications if any. Result: 33% had positive family history of HBV infection, while in rest immediate family contacts tested negative. At baseline, HBV DNA was between 3.9 9 10 4 -4.7 9 10 11 copies/ml by PCR. HBeAg was positive in all at baseline ALT was between 31 and 123 U/L at baseline. At 6 months on treatment, significant reduction of HBV DNA (i.e.[2 log 10 ) was noted in 100% patients, with DNA becoming undetectable (i.e. \200 copies/ml) by PCR in 25%. HBeAg seroconversion was also noted in similar percentage. At 12 months on treatment, 50% had undetectable HBV DNA as well as HBeAg seroconversion. Significant reduction of HBV DNA was seen in rest 50% with 100% patients achieving ALT normalization.

Conclusion: This study shows that sequential combination treatment with lamivudine and interferon yields promising results in CHB children of Asian origin. However larger, multi-centre clinical trial is required before this treatment can be recommended. Background: Safety and therapeutic efficacy of a combination therapy of antiviral drug and immune modulator was assessed in IDHAS in Bangladesh. Methods: HBV infection was detected in 25 apparently healthy subjects, who visited our hospital with different motives. These patients were given lamivudine (100 mg) orally, once daily for 12 months. Hepatitis B vaccine containing hepatitis B surface antigen (10 lg) was administered intra-muscularly along with lamivudine for 5 times (at 0, 1, 2, 6 and 12 months). Patients were regularly followed up to assess whether the combination was safe or not. The therapeutic efficacy of combination therapy was evaluated 13 months after commencement of the clinical trial. Result: Combination therapy was safe for all patients and local or systemic adverse effects were not recorded in any patient. At the end of the trial, the levels of HBV DNA were decreased in all patients, and become undetectable in 16 patients (64%). Flare of alanine aminotransferase was not detected in any patient. Conclusion: Combination therapy with antiviral agent and immune modulator represents one of the therapeutic options for containment of the virus in asymptomatic HBV carriers.

Changing of HBVDNA HBeAg conversion \1.0 9 10 3 \1.0 9 10 3 \1.0 9 10 3 --HBeAg positive 5.36 9 10 4 2.57 9 10 3 \1.0 9 10 3 20.15 0.002 Background/Aim: At present, combination treatment and monotherapy with high genetic barrier were recommended for hepatitis B e antigen (HBeAg) negative chronic hepatitis B (CHB) patients who need long-term therapy and has high risk resistance. The aim of this study was to evaluate and compare the effect of initial lamivudine (LAM) plus adefovir dipivoxil (ADV) combination therapy and entecavir (ETV) monotherapy on naïve HBeAg-negative CHB patients. Methods: Seventy one CHB patients with HBeAg negative, ALT levels between 2 and 10 times up normal limit (UNL) and HBV DNA levels higher than 10 4 copies/mL were enrolled in this study. Among them, 31 patients were given LAM 100 mg and ADV 10 mg per day and other 40 were given ETV 0.5 mg per day for 48 weeks. In two groups 30% and 32.26% patients were diagnosed cirrhosis respectively. The bark from three Enantia chlorantha was used as therapeutic alkaloid raw material. Bark was dried, milled and dissolved in methanol overnight, and then filtered. The residue had concentrated, and the proteins were precipitated with a small amount of water. The residue was evaporated to dryness. The method was used for isolation of flavonoid, quersetin in pharmacognosy. The yield consisted of five very similar bis-benzyl-isoquinoline alkaloids, maintaining functional control in aqua solution, so that the palmatine structure is the most sensitive to compensate for the others. The proportion of alkaloids remains constant when isolated from natural sources. In this schematic picture the electrons associated with nitrogen, are orphan and the easiest to transform. A capillary zone electropherogram of this five-alkaloid mixture, (the UV-200 nm scan), was used to monitor the changes in electrons. The movement used was slight when alkaloid extract was treated with a strong excess of water by evaporating the alkaloid slurry to dryness at 135°C. However, a distinct change in electropherograms was obtained. Polarisations taking place simultaneously were recognized in pherograms, which had an opposite function: The tissue degradative and regenerative function. The anastomosis is verified with liver-injured laboratory rodents. Three different lesions were selected as proof: D-GalN., ThAA and Allyl-alcohol. The curative influence on the liver was observed in all cases under study. The special observations were concentrated on sinusoids, which regulate the blood flow and give impact to anastomoses (the capability to provide alternative blood flow to portal veins despite obstructions There is an increased need of transport for the waste of cell apoptosis and other degradation product like fibrotic tissues, collagens and procollagens. At the same time, mitosis of hepatic cells developed.

A 96-Weeks Study of Antiviral Therapy for Patients with HBeAg-Negative Cirrhosis Yu-Sheng Jie 1 , Qi-Huan Xu 1 , Xin Shu 1 , Lu-Biao Chen 1 , Yu-Tian Chong 1 , Qi-Feng Xie 1 , Gang Li 1 1 The Third Affiliated Hospital of Sun Yat-sun University, 600 TianHe Rd, Guangzhou, China

Background: Compared with HBeAg-positive cirrhosis, patients of HBeAgnegative cirrhosis with active HBV replication are at higher risk of developing liver failure and hepatocellular carcinoma (HCC). This study was to investigate the efficacy of nucleot(s)ide analogues therapy in patients with HBeAg-negative cirrhosis in China. Methods: 111 patiens with HBeAg-negative cirrhosis were divided into antiviral group (58 cases, 25 entecavir, 19 adefovir dipivoxil, 13 lamivudine, 1 telbivudine) and control group (53 cases, supportive and symptomatic treatment). These two groups were matched for demography, liver function and Child-Pugh score. But the baseline HBV DNA level of antiviral group was higher than that of control group (v 2 = 3.51, P = 0.001). Results: At the 96th week, the rate of ALT normalization and HBV DNA drop (lg copies/ml) in antiviral group were higher than that in control group (v 2 = 4.292, P = 0.038; t = -9.001, P = 0.001, respectively). The rates of HBV DNA negative (\500 copies/ml) were 88.7% (47/53) and 32.5% (13/40), respectively (v 2 = 31.427, P = 0.001). 3.4% (2/58) patients in antiviral group and 7.5% (4/53) patients in control group developed HCC (Fisher's exact test, P = 0.423). 15.5% (9/58) and 20.8% (11/53) underwent variceal bleeding, respectively (v 2 = 0.514, P = 0.473). 15.4% (2/13) patients with lamivudine treatment emerged YMDD mutations, who were received an ''add-on'' antiviral therapy with adefovir dipivoxil. 10.5% (2/19) Background: Guangdong province is a highly endemic area for hepatitis B virus (HBV) infection in South of China where HBV vertical transmission from mother was common. HBV vaccine is the main means to prevent HBV vertical transmission. It is not long for the use of recombinant yeast HBV vaccine made in China. This study aimed to find out the seroconversion and immunity duration following recombinant yeast hepatitis B vaccination in newborns from Guangdong province. Methods: Immune response and side effects of recombinant yeast HBV vaccine in neonates were investigated. 166 neonates were divided into two groups according to HBsAg positive state of mothers (31 in HBsAg positive group and 135 in HBsAg negative group) and followed up for 3 years. The response rates were compared between two groups. Result: The total response rate was 90.4% at 12th months and 92.2% at 36th months. There is no significant difference of response rate between HBsAg positive group (80.6%) and HBsAg negative group (92.6%) (P = 0.090) at 12th months. The same result is also seen in the response rate (83.9 vs. 94.1%, P = 0.124) at 36th months. 150 subjects of responders at months 12 remained anti-HBs positive at 36th months. Among 6 infants identified as non-responders in HBsAg positive group, 2 infants were found HBsAg positive and the other 4 subjects received re-immunization. No HBsAg positive infants were found in 10 non-responders in HBsAg negative group. One subject of 4 re-immunization subjects in HBsAg positive group and 2 of 10 re-immunization subjects in HBsAg negative group turn anti-HBs positive. Background: To determine whether there is an inhibitory efficacy of C-specific siRNAs on HBV expression and replication in BHK-21 cells. Methods: The PCR products of HBV C genes were cloned to generate the expression vector pEGFP-C for sequencing. Two siRNA-expressing vectors of C-specific siRNA-S1 and S2 with the length of 23 nt homologous in sequence to the AY517488 C gene were designed and synthesized, according to the HBV genome (genotype B, ayw1 subtype) of CHB patients (GenBank Accession Numbers AY517488). Also, a expressing vector of nonspecific siRNA-S3 was also designed randomly for negative control. They were cloned into vector pU6 and then cotransfected into BHK-21 cells with target gene expression vector of pEGFP-C.

Results: It was found by fluorescence microscopic detection done at 12, 24, and 48 h after cotransfection that the expression of EGFP of group S1, S2 and S1 + S2 were reduced significantly in BHK-21 cells at the 24 h, in comparison with negative control group 1 (transfected with pC-EGFP-N1 only) and 2 (transfected with S3 and pEGFP-C) (P \ 0.01). The results showed that transfection of siRNA-expressing vectors S1and S2 caused an 80-90% reduction in the expression of HBV C in BHK-21 cells.The results of sequencing and RT-PCR prove the same effects further. Conclusions: For the first time it has been found that RNAi induced by siRNA targeting HBV C gene is continuous and stable inhibition of HBV expression and replication in BHK-21 cells, and our data suggest that RNAi may provide a viable prophylaxis and approach to treat HBV infection. This work was supported by grants from the Natural Science Foundation of China 30972629 (to Zhong-Qi Bian)

The Background: Virological response on/off antiviral treatment of CHB patients is associated with the enhancement of the host immune function. Currently, no ideal immunological parameter is available to evaluate host antiviral immunity. This study was to investigate whether TCR spectratype can be used as an immunological predictor for antiviral treatment. Methods: Twenty CHB patients were involved in a telbivudine versus laminvudine, double-blind, multicenter trial. Patients were divided into either virological responders (VR) or a combination group including both partial virological responders and patients with virological breakthrough (NVR) according to the 2009 EASL Guidelines. CD8 T cells were sorted from peripheral blood mononuclear cells at week 12 and 24 by magnetic beads. Twenty-four TCR b chain variable region gene (BV) families transcripts were amplified by RT-PCR and genescaned on ABI sequencer. Results: At week 48, 13 patients were VR (65%) and 7 patients were NVR (35%). Both at week 12 and 24, all 20 patients' scanograms showed various perturbations in some TCR BV families including monoclonicity, oligoclonicity and skewed peaks. The number of perturbation families of CD8 T cells in VR at week 24 was significantly larger than that at week 12 (10.15 ± 3.53 vs. 8.62 ± 3.18, P = 0.024), while the number of perturbation families in NVR at week 24 was significantly less than that at week 12 (7.14 ± 3.67 vs. 8.71 ± 4.42, P = 0.01). Six out of 7 patients in NVR showed descendent pattern on the number of perturbation families from week 12 to 24, while no VR showed descendent pattern. Moreover, there was a significant difference on the percentage of patients with descendent pattern between VR and NVR (P \ 0.001).

Conclusion: This finding suggests that the dynamic pattern of TCR BV spectratype perturbation between week 12 and 24 would be a novel immunological parameter to predict the prognostic of antiviral treatment of CHB. Background: A clinic cohort of chronic hepatitis B cirrhotic patients that were treated with lamivudine/adefovir were monitored for clinical outcomes. Method: Patients were enrolled in a treatment program of lamivudine switching to adefovir upon resistance. Patients were analysed for clinical outcomes and predictors of these outcomes were evaluated by multivariate analysis. Clinical outcomes of ascites, encephalopathy, hepatocellular carcinoma (HCC), progression in Child-Pugh score, MELD score and mortality were assessed. Data were analysed by Kaplan Meier graphs, log rank test and Cox regression. Results: There were 143 chronic hepatitis B cirrhotics of which 19.6% had decompensated cirrhosis. At 5 years, the mean survival was 83.6%, development of ascites, HCC, encephalopathy and progression in Child-Pugh score was 7, 15.9, 10.8 and 16.9% respectively. Overall progression of liver-related complications was 32.8% at 5 years. Multivariate analysis showed that ascites, albumin B28 g/L, Child-Pugh score C7.9, MELD score C10.9 were significantly associated with liver-related complications. Low albumin and low HBV DNA were independent factors for liver associated mortality. Lamivudine resistance did not affect mortality or liver disease progression. When stratified by Child-Pugh status, the mean survival of those with Child-Pugh C was significantly worse than those with Child-Pugh A or B (p \ 0.001). Early deaths (within 12 months) were due to liver failure or sepsis while those after 12 months were due mainly to HCC. Conclusion: Patients with decompensated chronic hepatitis B cirrhosis may suffer early mortality despite antiviral therapy and should be considered for early liver transplantation.

Effect of Lamivudine and Telbivudine on Creatine Kinase in Hepatitis B Antiviral Treatment Qing-Long Jin 1 , Hong-Qing Yan 1 , Jun-Qi Niu 1 1 Hepatobiliary and Pancreatic Department, Xinmin Street 71 of Changchun, China

Objective: To compared the impact of creatine kinase (CK) betweent telbivudine and lamivudine in antiretroviral treatment. Methods: 52 patients who participated in 015 study and 007 study were randomly divided into two treatment groups. Respectively gave telbivudine 600 mg daily and lamivudine 100 mg daily for 104 weeks. To observe CK elevation and clinical symptoms in the two treatment groups at different time.

Results: CK elevation was observed in the two treatment groups, CK elevation frequency was higher in telbivudine than in lamivudine (76%: 44.4%). There was no symptom observed in the patients with CK elevation. Conclusions: CK elevation is common in nucleoside analogue antiviral therapy. CK elevation frequency in telbivudine treatment groups was higher. Majority were Grade 1-2 CK level elevation (1-7 9 ULN). The cause of CK elevation was not induced by myocardial injury. All the paitents had no symptoms. CK elevations in patients were transient and resolved without treatment. Interleukin-33 (IL-33), a new member of the IL-1 family, signals through its receptor ST2 and induces T helper 2 (Th2) cytokine synthesis and mediates inflammatory response. This study investigated the role of IL-33 to inhibit HBV replication in vitro. Methods: HBV transgenic mice, Styela plicata, were divided into two as IL-33 treatment group and control group. Serum hepatitis B surface antigen was detected by enzyme-linked immunosorbent assay (ELISA) method and serum HBV DNA was detected by real-time polymerase chain reaction (RT-PCR). Spleen cytokine IL-4, IL-5, IL-17 and IFN-c, and NK cells, CD4 and CD8 were detected by flow cytometry and a three-color flow cytometry, respectively. The histology of liver tissue was evaluated before and after treatment. Results: Two weeks after starting the therapy, serum hepatitis B surface antigen was significantly lowered in IL-33-treated mice compared to the control group (F = 88.81, P \ 0.01). Serum HBV DNA was significantly lowered in IL-33-treated mice (F = 20.71, P \ 0.01). However, the effect of IL-33 could not inhibit the replication of HBV completely. A rebound phenomenon of serum hepatitis B surface antigen and HBV DNA was found 4 weeks after withdrawal of medication. Eight weeks after the beginning of IL-33 therapy the level of IL-4, IFN-c and IL-17 was measured to be 13.56 ± 0.78, 15.62 ± 0.98 and 17.8 ± 0.93%, respectively, compared to the normal control group which was measured to be 3.41 ± 0.38, 3.05 ± 0.89 and 4.84 ± 0.48%, respectively. Histological analysis of liver of IL-33 treated HBV-transgenic mice showed significantly reduced inflammation. Conclusions: IL-33 may be an effective antiviral therapy in treating chronic hepatitis B. IL-33 also inhibits HBV replication in the liver of transgenic mice mediated by IFN-c produced by hepatic cells. Background: Natural Killer (NK) cells, controlled by multiple activating and inhibitory receptors, are an important element of the innate immune response to viral infection including hepatitis B virus (HBV). Recent data indicate that the expression of 2B4(CD244), a member of the CD2 subset of the Ig superfamily, regulate NK functions. In this study we addressed the role of the NK cells marker 2B4 and compared the frequency of peripheral blood CD3 -CD56 + NK cells in CHB patients. Methods: A total of 53 patients of chronic HBV (CHB) patients were selected, treated with Telbivudine (LDT) for 36 weeks, and examined at every 12 weeks treatment for serum HBV DNA loads, alanine transaminase (ALT) and aspartate transaminase (AST) levels. Phenotypes of peripheral blood lymphocyte subsets (T cells, CD3 -CD56 + NK cells, CD3 + CD56 + NKT cells, CD4 + CD25 + FoxP3 + Treg cells and 2B4 + CD56 + cells) were analysed using flow cytometry. Results: The results showed significantly reduced numbers of CD3 -CD56 + NK cells with CHB patients (p = 0.013), while the proportion of CD3 + CD4 + T cells, Treg cells and the ratio of CD4/CD8 were significantly higher than that of healthy controls (p = 0.025, p = 0.006 and p = 0.046). The expression of receptor 2B4 + on CD3 -CD56 + NK cells were lower in CHB patients compared to that in healthy controls (p = 0.021). The CD3 -CD56 + NK and 2B4+ expression on natural killer cells in post-treatment group were increased than that in pre-treatment group (p = 0.001). A significant inverse correlation was obtained between 2B4 + CD56 + NK cells and HBV DNA levels (R = 0.278, p = 0.035). Conclusions: These findings demonstrate that the concentration of 2B4 + CD56 + NK cells normalizes with LDT treatment in CHB patients. The finding that HBV was associated negatively with the concentration of 2B4 + CD56 + NK cells suggests that immune activation in CHB patients receiving LDT influences the target cell recognition by NK cells. Background: To observe the clinical treatment effects of the HBsAg pulsed autologous dendritic cells (anti-HBV-DC) derived from peripheral blood mononuclear cells (PBMC) combine lamivudine and Thymosin-a1 for the chronic HBV carriers. Methods: Thirteen HBeAg positive and two HBeAg negative chronic HBV carriers included in the study. Taking peripheral venous blood 50 ml and obtain PBMC, GM-CSF and IL-4 induced expansion of DC. At the sixth days, a 30 lg HBsAg was give to pulse the DCs. At the seventh days, the anti-HBV-DCs were harvested and were injected into body by subcutaneous and intravenous. Every 2 weeks one time, a total of six times. 100 mg Lamivudine every day, and 1.6 mg thymosin-a1 every one week 2 times. Results: The serum HBsAg, HBeAg and HBVDNA were significantly decreased at 4, 12 weeks. One patient's HBcAb was from negative converted to positive after treatment. One patient's HBsAb turn positive at 4 weeks. The HBeAg/HBeAb seroconversion occurred in one patient at 4 weeks, the conversion rate was 7.69% (1/13). The HBeAg/HBeAb seroconversion occurred in another patient at 12 weeks. The total HBeAg/HBeAb seroconversion rate was 15.39% (2/13). The serum HBVDNA negative conversion occurred in four patients at 4 weeks, the negative conversion rate was 26.67% (4/15). The serum HBVDNA negative conversion occurred in other four patients at 12 weeks. The total serum HBVDNA negative conversion rate was 53.33% (8/ 15). Three patient's ALT increased to 1-2 times normal value at 4 weeks, but returned to normal in two patients at 12 weeks. Another case's ALT have a mild increase at 12 weeks. Conclusions: The anti-HBV-DC combine lamivudine and Thymosin-a1 can effectively inhibit HBVDNA replication, decrease rapidly the serum HBsAg, HBeAg and HBVDNA levels, and enhance the HBeAg/HBeAb seroconversion rate, which were a safe and effective treatment method for the chronic HBV carriers.

Telbivudine Add-on Therapy in an HBeAg-Negative Cirrhotic Patient with a Suboptimal Response to Tenofovir Stephan Kaiser 1 , Bettina Lutze 1 , Cihat Sen 2 , Thomas Bock 3 1 Liver Center Stuttgart, Stuttgart, Germany, 2 Practice for Internal Medicine, Stuttgart, Germany, 3 Robert-Koch-Institute, Berlin, Germany Introduction: We report here the efficacy of telbivudine as add-on therapy in a patient with cirrhosis who demonstrated a suboptimal response to tenofovir. Case Description: A 43-year-old man with no history of alcohol abuse was diagnosed with active hepatitis B e antigen-negative chronic hepatitis B during occupational screening. He had a history of icterus during adolescence though no diagnosis had been made. Laboratory findings showed that he was hepatitis B surface antigen-positive with a serum hepatitis B virus (HBV) DNA level of 6.1 x 10 7 IU/mL. His alanine aminotransferase (ALT) level was 89 IU/mL and he had a slightly enlarged liver with normal vessels and bile ducts and no ascites. Liver histology was grade 10, stage 6 on Ishak scoring. He was started on tenofovir 300 mg/day and monitored every 3 months. His HBV DNA level declined to 5.8 9 10 2 IU/mL at 3 months and 1.4 9 10 3 IU/mL at 6 months.

Corresponding ALT values were 42 and 36 IU/mL, respectively. In light of the persistent detection of HBV DNA, the presence of cirrhosis, and the high likelihood of indefinite therapy, telbivudine 600 mg/day was initiated as addon therapy. When the patient was evaluated 6 months later, his HBV DNA had become undetectable and his ALT level had declined even further to 23 IU/ mL. HBV DNA remained undetectable 3 months after that and his ALT level was 21 IU/mL. Conclusions: The addition of telbivudine to tenofovir resulted in a sustained suppression of HBV DNA within 6 months. These finding show the benefit of telbivudine therapy in patients with liver cirrhosis. Background: Hepatitis B surface antigen (HBsAg)-based vaccines have been used as immune therapeutic approaches in patients with chronic hepatitis B (CHB). Unfortunately, the clinical outcome is inconclusive. We postulated that both HBsAg and hepatitis B core antigen (HBcAg)-specific immune responses are essential for containment of hepatitis B virus (HBV) in CHB. Methods: A nasal vaccine containing both HBsAg and HBcAg were prepared (Center for Genetic and Biotechnology, Havana, Cuba); safety and efficacy of this vaccine was confirmed in normal human volunteers. After receiving permission from institutional review board and written consents, 17 treatmentnaïve patients with CHB were immunized 5 times with a vaccine that contained human-consumable HBsAg (50 lg) and HBcAg (50 lg) through nasal route using a special device. All patients were followed up regularly, once in every 2 weeks, for 6 consecutive months. Peripheral blood mononuclear cells were isolated from these patients one month after end of the 5 vaccinations and we assessed if they harbor HBsAg and HBcAg-specific immunocytes. Results: Nasal vaccinations were safe for all patients. Flare of alanine aminotransferase was not seen in any patient due to vaccinations. Although all Hepatol Int (2010) 4:94-345 159 patients were expressing HBV DNA in the sera before vaccination, HBV DNA became undetectable (HBV DNA \200 copies/ml) in the sera in 6 patients and reduced significantly in 5 patients due to nasal vaccination. The serum levels of interleukin (IL)-8, TNF-alpha, IL-1 beta, and IL-6 were significantly increased after 5 nasal vaccinations compared to their pre-vaccinated levels (p \ 0.05). Vaccination also induced both HBsAg and HBcAg-specific lymphocytes in CHB patients. Background: Restoration of host immunity has been reported in patients with chronic hepatitis B (CHB) after treatment with lamivudine; however, the underlying mechanisms of this treatment have not been determined. This study examined the role antigen-presenting dendritic cells (DC) in restoration of host immunity in the context of lamivudine therapy. Methods: Circulating DC were isolated from peripheral blood of 23 CHB patients before and 1, 3, and 12 months after starting lamivudine therapy. The non-antigen-specific proliferation of DC was assessed in allogenic mixed leukocyte reaction. DC were cultured with hepatitis B surface antigen (HBsAg) to prepare HBsAg-pulsed DC. Proliferative capacity and production of interleukin (IL)-12 and interferon (IFN)-c of HBsAg-pulsed DC were evaluated in culture.

Results: T cell stimulatory capacity of DC were increased significantly 1, 3 and 12 months after lamivudine therapy compared to their pre-treatment levels (p \ 0.05). However, the levels of activation of DC at 3 and 12 months were not significantly higher compared to their levels 1 month after therapy commencement (p [ 0.05). HBsAg-pulsed DC also produced significantly higher levels of IL-12 and IFN-c due to lamivudine therapy compared to their levels before therapy (P \ 0.05). HBsAg-pulsed DC from lamivudine-treated patients induced proliferation of HBsAg-specific T cells of CHB patients in an antigenspecific manner (p \ 0.05). Conclusion: Immune restoration due to lamivudine therapy is regulated at least in part by activation of DC. This indicates that lamivudine is not merely an antiviral agent, but, possess direct or indirect immune modulation capacities in CHB patients. However, progressive activation of DC and immune modulation did not persisted 1 month after start of lamivudine therapy indicating that an immune therapeutic agent may increase therapeutic efficacy of lamivudine in CHB patients. Background/Aims: Clevudine (CLV) and entecavir (ETV) were recently approved in Korea as antiviral drugs against chronic hepatitis B (CHB). This study was aimed to compare treatment efficacy, resistance rate and reduction of HBsAg between CLV and ETV therapy. Methods: A total of 114 treatment-naive patients with CHB who visited Samsung Medical Center and received CLV 30 mg or ETV 0.5 mg a day for more than 12 months were analyzed retrospectively. Virological response (VR; HBV DNA \300 copies/ml), biochemical response (BR; ALT \40 IU/ml), HBeAg seroconversion, and HBsAg reduction (Abbott ARCHITECT HBsAg assay) were assessed. Results: Initial mean HBV DNA levels were 7.4 log 10 (N = 58) in CLV and 7.7 log 10 copies/ml (N = 56) in ETV groups. VR and BR were achieved in 31/38 (82%) and 31/38 (82 %) patients in CLV group and 49/56 (88%) and 44/56 (79%) patients in ETV group, respectively at month 12. For 54 patients who were followed up to month 24, VR and BR were achieved in 25/31 (81%) and 24/31 (77%) patients in CLV group and 21/23 (91%) and 18/23 (78%) patients in ETV group, respectively. HBeAg seroconversion at 24-month occurred in 5/12 (42%) patients in CLV group and 2/10 (20%) patients in ETV group. During 1-year treatment, no resistance was observed in both groups, while at year 2, viral breakthrough with YIDD mutation occurred in 4/56 (7%) patients in CLV group and no viral breakthrough in ETV group. At month 24, the patients with VR at month 12 in CLV group showed -0.2 log IU/mL of HBsAg reduction, while in ETV group, HBsAg titer was not reduced (+0.05 log IU/ml). Conclusions: Both of CLV and ETV showed potent antiviral activity against CHB. ETV was superior to CLV in resistance profile. However, serum HBsAg reduction known to correlate with hepatic cccDNA was observed only with CLV treatment. Background: Hepatitis B virus (HBV) inoculation is the best way to prevent HBV infection; however, there are 4-10% of immunized people have no specific protective antibodies. Additionally, there are lots of chronic HBV infectors including healthy HBsAg carriers, whose health are risk because of the susceptibility to hepatocyte injury. It is impending to promote the HBV vaccination effects and improve the anti-viral function in the body. Methods: Mice were vaccinated with HBV vaccine (1 mg/mouse for adult and 0.5 mg/mouse for newborn) i.m. and re-vaccinated after 2 weeks. Mouse IL-12 was studied for its adjuvant effects on HBV vaccination in several mouse models including adult and newborn wild type mice, chronic HBV carriers induced by hydrodynamic vein injection of HBV DNA plasmids, HBs-Tg mice and HBV transgenic mice. Serum anti-HBs antibody was measured to show the vaccination effect. Results: It was observed that used as adjuvant 500 ng mIL-12 could obviously increase the antibody levels in BALB/c mice and 50 ng mIL-12 could obviously increase the antibody levels in C57BL/6 mice. The antibody has been increased for 10 weeks and is still observed for its persistent time. The different injection combination of mIL-12 with HBV vaccine would cause different effects on the vaccination. In the newborn C57BL/6 mice, 25 ng mIL-12 could obviously increase the antibody levels and even a low dose of 5 ng mIL-12 could do. Chronic HBV plasmid carriers, HBs-Tg mice and HBV-Tg mice were immune-tolerance to HBV vaccine. For these mice, mIL-12 as used in wild type mice could not induce the antibody production. mIL-12 was injected to mice for more than three times and then HBV vaccine was administrated. A long time observation has been performed on these mice. Conclusions: These studies would be a great benefit to the people with failure HBV vaccination at present and chronic HBV carriers. Aims: To examine changes in the magnitude, frequency and specificity of HBV-specific T-cells following nucleoside reverse transcriptase inhibitor (NRTI) treatment of CHB in a prospective longitudinal cohort study. Methods: Whole blood from treatment naïve patients with CHB (n = 11; HBeAg-positive = 3) initiating lamivudine (n = 9) or tenofovir (n = 2) therapy was incubated with overlapping HBV peptides to the full HBV proteome. Production of TNF-a, IFN-c and IL-2 in CD4 + and CD8 + T-cells was quantified using intracellular cytokine staining (ICS) at weeks 0, 4, 12, 24, 36 and 48. Results: Prior to NRTI treatment, HBV DNA (median (IQR)) and ALT were 5.9 (4.9-6.8) log IU/ml and 59 (50-194) IU/ml respectively and both significantly declined over 48 weeks (p = 0.008 and p = 0.01 respectively). 89% of patients had undetectable HBV DNA by week 48. The magnitude of CD4 + IFN-c and dual TNF-a and IFN-c responses significantly declined (p = 0.006 and p = 0.036 respectively) following NRTI treatment. Consistent with these findings, there was also a significant decline in the frequency of CD4 + dual TNF-a and IFN-c responses (p = 0.0009). There were no significant changes in the frequency or magnitude of HBV-specific CD8 + Tcell responses following treatment. The majority of cytokine + responses were towards [1 peptide pool. At baseline the mean magnitude of CD4 + TNF-a and IFN-c responses to precore peptides was significantly greater than to X, surface or polymerase (p = 0.004), however, at subsequent time points following treatment there was no preferential response to any specific peptide pool.

Conclusions: Using multi-parameter ICS we found a significant decline in HBV-specific CD4 + T-cell responses and no change in HBV-specific CD8 + T-cell responses following effective NRTI treatment. Despite significant declines in HBV DNA with treatment there is no recovery of HBV-specific immunity. Background: Entacavir (ETV) is a nucleoside analogue that effectively inhibits hepatitis B virus (HBV) polymerase, resulting in rapid viral suppression. However, the efficacy of ETV rescue therapy in patients who had previously experienced antiviral treatment with lamivudine (LVD) and adefovir disoproxil (ADV) is unclear. We analyzed the clinical outcomes of ETV monotherapy in patients who were resistant to LVD and ADV sequential therapy. Methods: A total of 33 CHB patients who had shown viral breakthrough or suboptimal response with LVD and ADV sequential therapy were treated with 1.0 mg of ETV for at least 48 weeks. Virologic response was defined as a serum HBV DNA reduction to undetectable levels (\300 copies/mL). The mean reduction in the serum HBV DNA and the rates of virologic response, HBeAg seroconversion, and alanine transaminase (ALT) normalization were monitored until 48 weeks. Result: In the 33 CHB patients, the rates of virologic response and ALT normalization were 18.2% (6/33) and 66.7% (14/21) at 48 weeks, respectively. Viral breakthrough occurred in 21.2% (7/33) during ETV rescue therapy. In 27 HBeAg-positive CHB patients, the rate of virologic response and ALT normalization were 18.5% (5/27) and 58.8% (10/17), respectively. The mean reduction in serum HBV DNA from baseline to the end-point was 2.60 log copies/mL. Serum HBeAg seroconversion in the patient who received ETV occurred in 6.6% (2/30) at 48 weeks. In six HBeAg-negative CHB patients, the rate of virologic response and ALT normalization were 16.7% (1/6) and 100% (4/4), respectively. The mean reduction in the serum HBV DNA from baseline to the end-point was 1.31 log copies/mL. Conclusion: The 48 weeks ETV monotherapy resulted in low antiviral response and early ETV resistance in patients with LVD and ADV resistant-HBV. Background and aims: Adefovir (ADV) has been reported to be effective against both to the lamivudine (LMV) resistant HBV and wild-type HBV with similar degree of viral suppression. But, in patients carrying LMV resistant, it has been reported that viral breakthrough and the emergence of ADV resistant mutation occurred more often in ADV monotherapy group compared to ADV/ LMV combination. Most of the previous studies were confined to HBeAg negative patients. Aims of or study was to compare the antiviral efficacy and ADV resistance in lamivudine resistant HBeAg positive chronic hepatitis B patients treated with ADV mono-vs. ADV/LMV combination under the circumstances with tenofovir (TDF) was not available. Methods: 128 LMV resistant HBeAg positive chronic hepatitis B patients treated with ADV mono-(n = 92) or ADV/LMV combination (n = 36) were selected and analyzed retrospectively. Liver function test, HBV DNA, HBeAg and anti-HBe antibody were measured every 3 months. Results: In ADV or ADV/LMV patients, treatment baseline ALT (mean, IU/L, 246 vs. 196, p = ns), HBV DNA (mean, log 10 copies/mL, 7.9 vs. 7.8, p = ns) were not different. Mean duration of ADV mono-or ADV/LMV combination was 37.9 months and 28.8 months in each group. During the 36 months of treatment, ALT normalization occurred in 90% of ADV momo-and 88.8% of ADV/LMV combination groups, respectively (p = ns). DNA reduction from baseline at treatment months 12, 24 and 36 was 3.1 versus 3.3 (p = ns), 3.6 vs. 4.2 (p = ns) and 3.8 vs. 5.2 (p = 0.057) in each group, respectively. Viral breakthrough were observed in 30 of 92 (32%) patient with ADV monotherapy and 1 of 36 (2%) patient with ADV/LMV combination group and it was statistically significant (p = 0.001 Background and Aims: According to Chinese National Guidelines concerning the treatment and prevention of chronic hepatitis B (CHB) infection, treatment with interferon (IFN)-based therapy is contraindicated in patients with ALT more than 10 times upper limit of normal (10 9 ULN), although no such contraindication is given in recent European and American guidelines. There is currently a lack of published data on the efficacy and safety of IFN-based therapy in this population of patients. We therefore aimed to evaluate the efficacy and safety of treatment with peginterferon alfa-2a (PEGASYS) in HBeAg-positive CHB patients with ALT [10 9 ULN. Methods: A total of 25 patients (13 with ALT 2-10 9 ULN and 12 with ALT [10 9 ULN at baseline) received peginterferon alfa-2a 180 lg s.c. once weekly for 48 weeks, with a follow-up period of 12 weeks post-treatment. Liver function, HBV DNA levels, HBeAg and HBsAg were measured at baseline, at weeks 12, 24, 36, 48 of treatment and at 12 weeks post-treatment (week 60). Results: In patients with baseline ALT [10 9 ULN, after 48 weeks of treatment 11 (91.7%) were HBV DNA negative, 10 (83.3%) achieved HBeAg seroconversion, 9 (75.0%) had normal liver function, and 4 (33.3%) achieved HBsAg clearance. This compares with 10 (76.9%), 8 (61.5%), 7(53.8%) and 1 (7.7%) patients with ALT 2-3 9 ULN, respectively. The difference between the 2 groups was statistically significant (P \ 0.05). Treatment with peginterferon alfa-2a was well tolerated by all patients and there were no severe adverse events or other safety issues. Conclusions: Treatment with peginterferon alfa-2a was significantly more effective in terms of sustained HBV DNA suppression, HBeAg seroconversion and HBsAg clearance and normalization of liver function in patients with baseline ALT [10 9 ULN compared with patients with baseline ALT 2-10 9 ULN. Treatment was well tolerated in all patients.

Evaluation of Safety 

Background & Aim: To evaluate the long-term efficacy and safety of lamivudine treatment for patients with Anti HBe positive chronic hepatitis B Methods: 932 patients who were admitted by Ankara University faculty of medicine gastroenterology clinics with HBs Ag positive longer than 6 months were evaulated. 409 of those patients had HBV DNA (PCR) levels C10 4 copy/ ml, ALT levels higher than upper limit of normal values, proven histopathologically chronic hepatitis B infection. They were started lamuvudine therapy. They were evaluated in terms of HBV DNA (PCR) values and transaminase values after at least 1 year treatment. Statistical evaluation was made for 119 patients who are regularly controlled. The others were excluded due to unregular control.

Result: Eighty-four of those patients were male and 35 were women. The average age was 42.1 years. Patients were followed-up 12-72 months (mean follow-up time was 32 months). At basal evaluation, average ALT level was 102 IU/ml, average AST level was 77 IU/ml. Delta antibody was negative in all of the patients. Average HBV DNA level of these patients was 10 6 copy/ml. At evaluation at 12 months of these patients, ALT and AST normalization (biochemical response) rate was 50%, the negativization of HBV DNA (virologic response) rate was 45% and HBs Ag-Anti HBs seroconversion rate was 3.6%. At 18 months evaluation, 76 patients were evaluated. Average AST level was 41 IU/ml, average ALT level was 49 IU/ml. At 24 month evaluation, biochemical response rate was 43%, virologic response rate was 37%. At the end of second year evaluation, adefovir treatment had been added on 29 patients (38.15%) because of lamuvidine resistance. Forty-eight months evaluation was made among 37 followed-up patients who continued lamuvudine treatment. These patients' average ALT level was 37 IU/ml, average AST level was 32 IU/ml. Biyochemical and virologic response rates were 38 and 33% respectively. Conclusion: Our results show that lamivudine treatment is effective in half of the patients at first year and to decrease year by year. Resistance of lamivudine is also high (38.15) at the second year in Turkish population. Background: There is discrepancy between the EASL and APASL guidelines for lamivudine-resistant patients with chronic hepatitis B (CHB). The aim of this study was to evaluate the efficacy of lamivudine and adefovir (LAM + ADV) rescue therapy and compare it to entecavir (ETV) monotherapy in LAM-resistant CHB patients. Methods: From October 2007 to September 2008, 92 LAM-resistant patients with CHB were consecutively enrolled (47 LAM 100 mg + ADV 10 mg once daily and 45 ETV 1 mg once daily). All patients were followed for at least 12 months. The parameters assessed included normalization of ALT, HBeAg seroconversion, undetectable HBV DNA by PCR (lower detection limit \300 copies/ml), reduction of HBV DNA, and predictors of virologic response. Results: The median follow-up time was 15 months (range 12-24 months). In the LAM + ADV and ETV groups at month 12, virologic response occurred in 18/47 (38.3%) and 11/45 (24.4%; P = 0.182) patients, ALT normalization in 39/41 (95.1%) and 36/40 (90.0%; P = 0.432), HBeAg seroconversion in 5.1 and 2.4% (P = 0.606), and virologic breakthrough in 2.1 and 11.1% (P = 0.107), respectively. The mean reduction in the baseline HBV DNA level was greater in the LAM + ADV than in the ETV group at month 12 (3.80 ± 1.12 vs. 2.72 ± 1.32 log 10 copies/ml; P \ 0.001). Virologic response of the higher baseline HBV DNA (C8 log 10 copies/ml) was 8.0% in contrast to 53.3% in patients with lower baseline HBV DNA (5B DNA level, log 10 copies/ml \6).

Conclusions: LAM + ADV combination therapy was more effective for reducing viral load than ETV monotherapy in patients with LAM-resistant CHB. Baseline HBV DNA level was an independent predictor of virologic response. Thus, a combination of more potent drugs such as ETV + TDV may be more beneficial in the management of LAM-resistant CHB patients with a higher baseline HBV DNA level. Background/Aims: In patients with chronic hepatitis B (CHB), HBsAg titer was reported to reflect cccDNA level. Clevudine is a potent antiviral agent that was proven to be able to reduce cccDNA level and, consequently, HBsAg titer in in vivo animal studies. In this study, we aimed to evaluate the efficacy of clevudine for reducing HBsAg titer and to find an independent predictor. Methods: This retrospective cohort study included consecutive CHB patients treated with clevudine HCC between January 2007 and December 2008 at a single tertiary medical center in Korea. The reduction of HBsAg titer was defined as the decrease of HBsAg titer below 250 IU/mL. Kaplan-Meier method and Cox regression model were applied in statistical analysis. Results: A total of 131 patients were included (mean age 50.9 ± 11.1 years; male 66.4%). Sixty-two patients (47.3%) had liver cirrhosis (LC) and 29 (22.1%) had hepatocellular carcinoma (HCC). During the observation period (mean 14.9 ± 7.8 months), 8 patients (6.1%) achieved HBsAg titer \250 IU/ mL with clevudine treatment, whose initial HBsAg titer were [250 IU/mL. There was no patient who gained HBsAg-loss during the period. The cumulative probabilities of the reduction of HBsAg at 12th and 24th months were 5 and 10%, respectively. In multivariate analyses, the presence of LC was an independent predictor of the HBsAg titer reduction (hazard ratio 7.886; P = 0.034). However, initial serum levels of alanine and aspartate aminotransferases, previous antiviral therapy, initial HBeAg status, and the presence of HCC were not significantly associated with the reduction of HBsAg titer.

Conclusions: This study demonstrates that clevudine reduces HBsAg titer to \250 IU/mL with rate of about 5%/year and the efficacy is higher in patients with LC than those without LC. Further investigation is warranted to confirm the efficacy of clevudine for reducing or eliminating HBsAg.

Comparision of Entecavir (ETV) Monotherapy and Lamivudine ( Background/Aims: There are no adequate studies on the efficacy of LAM + ADV versus ETV monotherapy in chronic hepatitis B (CHB) patients who are sequentially resistant to LAM/ADV. We evaluated and compared the efficacy of LAM + ADV versus ETV in these patients. Methods: A total of 72 patients (HBeAg(+) 66/72, 93%) who had resistance (n = 30) or a partial virologic response (n = 41) to ADV with primary failure of LAM therapy and who were confirmed YMDD(+) were enrolled. They either received LAM (100 mg) in addition to ADV (10 mg) once daily (n = 31) or were switched to ETV (1 mg once daily) (n = 41). All patients were followed for at least 12 months. The parameters assessed included ALT normalization, HBeAg seroconversion, undetectable HBV DNA by PCR (lower detection limit \300 copies/ml), primary non-response (HBV DNA reduction \1 log 10 copies/ml at month 3), and reduction of HBV DNA. Results: The median follow-up time was 15 months (range 12-30 months). A primary non-response was more frequently observed in the LAM + ADV group (54.8%) than in the ETV group (29.3%; P = 0.021). At month 12, in the LAM + ADV and ETV groups, the mean reduction in the baseline HBV DNA level was 1.44 ± 1.19 and 1.92 ± 1.24 log 10 copies/ml (P = 0.101), and a virologic response was observed in 7/31 (22.6%) and 8/41 (19.5%; P = 0.777) patients, respectively. In the LAM + ADV and ETV groups, ALT normalization occurred in 11/13 (84.6%) and 16/18 (88.9%; P = 0.566), HBeAg seroconversion in 1/26(3.8%) and 4/41 (9.8%; P = 0.641) and a virologic breakthrough in 2/31 (6.5%) and 6/41 (14.6%; P = 0.452), respectively. Conclusions: Both LAM + ADV combination therapy and ETV monotherapy were not satisfactorily effective in patients with sequential LAM/ADV-resistant CHB. Therefore, sequential monotherapy with ADV should be avoided during the initial management of LAM-resistant patients with CHB.

A This study was to compare the treatment efficacy at week 96 between entecavir (ETV) and adefovir (ADV) in treatment naïve patients with chronic hepatitis B (CHB). We had prospectively randomized nucleoside-naïve CHB patients to open-label treatment with either ETV 0.5 mg or ADV 10 mg once daily. Forty-one hepatitis B e antigen (HBeAg)-positive patients randomized to ETV group (19 patients) or to ADV group (22 patients). Sixteen HBeAg-negative Hepatol Int (2010) 4:94-345 165 patients randomized to either ETV or ADV group with 8 patients, respectively). All patients were complete the treatment duration of 96 weeks. The primary objective was to compare the antiviral efficacy as achieve to undetectable serum HBV DNA (\12 IU/mL) by polymerase chain reactive (PCR) assay at week 96. In HBeAg-positive patients, the proportion of patients achieving undetectable HBV DNA was 63.2% in ETV group and 36.8% in ADV group at weeks 96 (p = 0.087). ETV group (-4.50±1.49 log IU/mL) showed more reduction of HBV DNA from baseline than ADV group (-3.05 ± 1.88 log IU/mL) at week 96 (p = 0.002). At 96 weeks, there was no difference in the rate of HBeAg seroconversion between two groups (10.5% in ETV group vs. 22.7% in ADV group). In HBeAg-negative patients, the proportion of patients achieving undetectable HBV DNA was 100% in ETV group and 62.5% ADV group at week 96 (p = 0.055). ETV group (-4.21 ± 1.82 log IU/mL) showed more reduction of HBV DNA, but there was no statistic significancy, from baseline than ADV group (-3.68 ± 1.34 log IU/mL) at week 96. In both HBeAg-positive and HBeAg-negative patients, there was no case of virologic breakthrough regardless of treatment method at entire 96 weeks. ETV treatment showed the more potent efficacy compared at 96 weeks with ADV group in both HBeAg-positive and HBeAg-negative nucleoside-naïve patients with CHB.

Long- Background and aims: The best time and hepatitis B virus (HBV) DNA level during an early adefovir dipivoxil treatment period for predicting the longterm outcome are unknown. The aims of this study were to determine the optimal time and HBV DNA load decline for the prediction of the sustained response in naïve patients treated with adefovir dipivoxil of 2 years long-term follow. Methods: 40 hepatitis B e antigen (HBeAg) positive chronic HBV patients treated with adefovir dipivoxil were studied. The HBV DNA levels at the baseline, at weeks 0, 4, 8, 12, 24, 36, 48, 76 and 104. Results: 17 patients obtained sustained response (HBV DNA level \1,000 copies/mL, and normal alanine aminotransferase levels) at year 2. When we considered the viral load reduction from baseline to each week of treatment, receiver operating characteristic curves showed good predictions as early as week 24. At the end of week 24, viral load decreased more than 4.0 log in 16 out of the 17 patients with sustained response and in 1 out of the 23 other patients. Week 24 appeared to be the optimal point time for predicting response, with a sensitivity of 88.9%, a specificity of 95.5%, a positive predictive value of 94.1% and a negative predictive value of 91.3%. Conclusion: During treatment with adefovir dipivoxil in naïve patients, viral kinetics showed that week 24 appeared to be the best time point for predicting the long-term outcome. Background/Aims: The definition of partial virological response (PVR) to nucleos(t)ide analogues(NUC) proposed by EASL guideline is based on the lower level of evidence in terms of cut-off level of HBV DNA and time-point to judge it. Therefore, we investigated the optimal PVR criteria for the prediction of virologic response during entecavir (ETV) therapy in treatmentnaive chronic hepatitis B (CHB) patients. Methods: Among 161 CHB patients who were consecutively treated with ETV as a first line therapy, 69 patients have completed 2-year ETV treatment. 64 patients were finally enrolled for analysis after excluding 5 non-responders at week 48 (\1 log reduction of HBV DNA). Four different PVR criteria [cut-off level of HBV DNA (1) undetectable (2) \3 log 10 copies/mL; time-point to define (1) at week 24, (2) at week 48] were compared for the prediction of 2year virologic response (undetectable HBV DNA by real-time PCR) Undetectable HBV DNA was defined as \12 IU/mL by real-time PCR using COBAS TaqMan 48 analyzer. Results: In 43(67.2%) HBeAg-positive patients, 9(33.3%) out of 27 patients who showed HBV DNA [3 log 10 copies/mL at week 24 and 10(30.3%) out of 33 patients who showed detectable HBV DNA at week 24, failed to achieve undetectable HBV DNA at week 96. However, 8(72.8%) out of 11 patients who showed HBV DNA [3 log 10 copies/mL at week 48 and 10(52.6%) out of 19 patients who showed detectable HBV DNA at week 48, failed to achieve undetectable HBV DNA at week 96. There was no PVR in HBeAg-negative CHB up to week 48. Conclusions: HBV DNA [3 log 10 copies/mL at week 48 was the best PVR criteria to predict the forthcoming treatment failure to remained detectable HBV DNA at week 96 in HBeAg-positive CHB patients on ETV as a first line therapy. Background: Chronic hepatitis B virus (HBV) infection is overall recognised in 10% of HIV positive persons, with large differences according to geographical region. Since the widespread use of highly active antiviral therapy (HAART), studies have documented an association between immune reconstitution and alterations in the course of hepatitis B infection. We evaluated the long term evolution of liver disease among patients receiving HAART. Methods: We followed-up 71 HIV/HBV coinfected patients treated with HAART containing, during a period of 5 years. Clinical and biological data were collected every 3 months, immunological and virological data every 6 months, ultrasound every year. Due to availability, in the last year we performed transient elastography (Fibroscan) in every patient. Determination of HBV drug resistance was performed in cases with detectable serum HBV-DNA level. Results: No patients had signs of liver disease in the begining of the study period. After 5 years of follow-up, advanced liver disease was diagnosed in 7/ 71 patients during follow-up: 4 cirrhosis, 2 hepatocellular carcinoma (HCC), and 1 fulminant hepatitis. All these patients had high level of serum HBV-DNA, lamivudine resistance and undetectable serum level of HIV. Five of these patients died. The rest of 64 patients had no signs of active liver diseases (normal or less than 2 9 ULN level of ALT, and F0-F1 according to Fibroscan). Lamivudine resistance in these patients was founded in 10 cases. Conclusions: In HIV/HBV coinfected patients treated with HAART, lamivudine resistance is less frequent (23.9%) than in immunocompetent patients (higher than 60%), but when occurred, was associated with an accelerated course of liver disease, with faster progression to cirrhosis, liver insufficiency and HCC. Appropriate monitoring of chronic viral hepatitis B in HIV positive patients include the recognition of lamivudine resistance in every case of detectable HBV-DNA level. Background: Currently, HBeAg-negative chronic hepatitis B (CHB) is increasing. But there are still controversial on the treatment of HBeAg-negative chronic hepatitis B (CHB) with ALT\2 9 ULN. We have investigated the clinical efficacy of nucleotide analogues (NAs) in the treatment of HBeAgnegative CHB with ALT \2 9 ULN. Methods: The data of patients who were treated by NAs for more than 2 years and with ALT 1-2 9 ULN (n = 102), ALT B1 9 ULN (n = 59) and ALT C2 9 ULN (n = 125) were collected. Virologic response (undetectable HBV DNA) at 12, 24, 48 and 96 W, virologic breakthrough and clinical resistance were analyzed. Results: Compared with the base line, HBV DNA level in all three groups were significantly decreased (P \ 0.01), and there was no significant difference between ALT 1-2 9 ULN group and ALT C2 9 ULN group. The change of HBV DNA level was significant decreased in ALT B1 9 ULN group at 12, 24 and 36 W (P \ 0.05). The rat of HBV DNA undetectable at 12, 24, 48 and 96 W was 63.5, 78.5, 79.5 and 86.7% respectively in ALT 1-2 9 ULN group and was 46.5, 53.7, 70.7 and 76.6% respectively in ALTB19ULN group. There was no significant difference between ALT 1-2 9 ULN group and ALT C2 9 ULN group. Compared with the previous two groups, the rat of HBV DNA undetectable 12 and 24 W were significant decreased (v 2 = 7.65, 0.05; v 2 = 20.83, P \ 0.01) in ALT B1 9 ULN group. There was no significant difference among the three groups in virologic breakthrough and clinical resistance. Conclusion: HBV replication can be satisfactory inhibited by NAs in HBeAgnegative CHB patients with ALT \2 9 ULN, which suggests that in these patients the indication of ALT is different from HBeAg-positive patients.

The Study about the Reasons of Bad Dependence of HBV Background: China has so many chronic hepatitis B (CHB) patients. When patients use these drugs like ADV, ETV, Ldt and LAM, the regularity of patients using drugs is a significant factor of therapeutic effect. But we found parts of patients did not complied to the guidance of doctors and induced some bad consequences like bounce of HBV DNA and so on. So we study the reasons of the bad dependence and drug withdrawal when is not to get to the best time to stop drug. Methods: The total 208 CHC patients were investigated. The number of male were 152 and female were 56. Results: The reasons include (1) the number of drugs which patients used were not enough (most in these patients who were came from other cities) (72.6%); (2) economy (4.6%); (3) the patients regarded the condition was improved and it was already not need to have drugs by themselves (9.6%); (4) using drugs irregular (4.5%); (5) listening to the views of other people (include specialist, general practitioner and so on) (4.6%); (6) other (4%). Conclusions: The rate of male is bigger than female in these patients who have bad dependence. It is necessary to enhance the guidance for the patients who is stopping to have drug by themselves and believing the opinions of general practitioner, other patients and so on. The work to help the patients who stop to have drug because they cannot buy enough drug in their cities or have no time to buy the drug and so on is an important thing of our hospitals.

Intrahepatic HBV Status after 10 Year of Sustained Viral Suppresion by LAM: Analysis of cccDNA Sequence and Expression Laura Belloni 1,2 , Giuseppina Brancaccio 3 , Letizia Cimino 1,2 , Gianfranca Stornaiuolo 3 , Lorenzo Andreana 3 , Rossana De Iaco 1,2 , Maria Guido 4 , Massimo Levrero 1,2,5 , Giovanbattista Gaeta 3 1 Dept of Internal Medicine and the AIRC Rome Oncogenomic Center (ROC), Sapienza University, Rome, Italy, 2 Laboratory of Gene Expression, Fondazione A. Cesalpino, Rome, Italy, 3 Dept. of Infectious Diseases, II University of Naples, Naples, Italy, 4 Department of Diagnostic Medical Sciences and Special Therapies, Pathology Unit, University of Padua, Padua, Italy, 5 EAL Inserm 785, Rome, Italy and Villejuif, France

Background and Aim: Treatment of chronic hepatitis B therapy with lamivudine is limited by the emergence of drug resistant mutants. In a minority of LAM-treated patients HBV remains suppressed. Whether a sustained suppression of HBV replication may lead over time to cccDNA elimination (eradication) is still debated. Aim of this work was to study intrahepatic HBV status (cccDNA levels, pgRNA transcription and cccDNA sequence) in chronic HBV patients with a sustained complete virological response to LAM. Methods: Liver biopsies were obtained four patients after 9-11 years of sustained viral suppression under lamivudine treatment (100 mg/day). Patients baseline characteristics were: age 24-57; 2 histological and 1 clinical decompensated cirrhosis; ALT 47-1304 IU/L; platelets 100,000-120,000/ mmc), HBV-DNA from 10 5 to [10 6 cp/m; one patient was HBeAg positive. Nuclear and cytoplasmic extracts from liver biospy samples were used to quantitate both cccDNA levels (real-time PCR with specific primers) and HBV pgRNA (RT real-time PCR). Results: At the last follow-up visit all patients were HBV-DNA negative (\15 IU/mL), had normal ALT and were anti-HBe positive (patient #3 seroconverted to anti-HBe). HBsAg serum levels ranged from 9.4 to [250 IU/ml and patient #2 was HBsAg negative. cccDNA was present at very low levels in 2/4 patients and was positive in one. pgRNA was detectable in 3 out of 4 patients. Patient #2 was negative for both pgRNA and cccDNA and had very low HBsAg titer. Sequencing, performed both on nuclear DNA and cytoplasmic cDNA, could not detect any of the known LAM resistance mutations in the HBV cccDNA Pol region. Conclusions: No LAM-selected mutations were archived in the cccDNA; cccDNA persists in the majority of patients even after several years of complete suppression of viral replication; combining cccDNA, pgRNA and HBsAg quantification may help selecting candidate patients to suspend ativiral therapy. Background: This prospective study evaluated the clinical outcomes of entecavir treatment in Chinese cirrhotic patients with chronic hepatitis B infection. Methods: Nucleos(t)ide-naive patients with HBV-related compensated (n = 31) or decompensated (n = 73) cirrhosis received entecavir 0.5 mg once daily for 96 weeks. Efficacy assessments included mean change from baseline in serum HBV DNA and the proportions of patients achieving HBV DNA \1,000 copies/mL and ALT normalization. Changes in Child-Pugh score and liver function improvement (serum albumin, total bilirubin and prothrombin time) were also evaluated. Liver histology was evaluated in 37 patients with paired biopsies. Results: At week 96, 98 and 81% of patients achieved undetectable HBV DNA and ALT normalization, respectively. Mean reduction from baseline HBV DNA was 5.1 log 10 copies/mL. No cases of virologic breakthrough or genotypic resistance to entecavir occurred through 96 weeks. The proportion of patients with Child-Pugh class A disease increased significantly at Week 96 (51%) versus baseline (30%; P \ 0.003). Liver function improved significantly in patients with decompensated cirrhosis. Mean serum albumin increased from 27.6 ± 4.4 g/L at baseline to 37.5 ± 6.7 g/L at week 96 (P \ 0.0001), mean total bilirubin decreased from 49.6 ± 38.3 lmol/L at baseline to 25.3 ± 10.4 lmol/L at week 96 (P \ 0.0001) and prothrombin time decreased from 15.1 ±2.3 s at baseline to 13.8 ± 2.5 s at week 96 (P = 0.0013). Among the 37 patients with paired biopsies, mean decreases in Knodell necroinflammatory score were 2.6, 2.0 and 0.9 points in Child-Pugh class A, B and C patients, respectively. Histologic improvement was strongly correlated with reductions in viral load at week 96 (r = 0.93). Conclusions: Entecavir is an effective treatment option for patients with HBVrelated compensated or decompensated cirrhosis, resulting in sustained virologic suppression, histologic improvement and stabilization or improvement of liver function. Background: Certain pretreatment characteristics and early response to nucleoside analogs (NAs) predict optimal outcomes in patients with chronic hepatitis B (CHB). If hepatitis B e antigen (HBeAg) seroconversion and profound and sustained viral suppression is achieved with NA therapy, drug resistance may become less of a concern. In patients with HBeAg-positive CHB, baseline hepatitis B virus (HBV) DNA levels \9 log 10 copies/mL, alanine aminotransferase (ALT) levels C2 9 upper limit of normal (ULN), and week 24 on-treatment polymerase chain reaction (PCR)-negative viral load predict better HBeAg seroconversion rates and less viral resistance. Similarly, in patients with HBeAg-negative CHB, baseline HBV DNA levels \7 log 10 copies/mL and week 24 on-treatment PCR negativity predict better outcomes. However, many patients with CHB do not possess these characteristics. This survey was conducted to determine how the pretreatment predictors of response based on the GLOBE study relate to CHB patients considered appropriate for therapy by Filipino clinicians. Methods: Results of a survey of 6 experienced physicians were compiled in a database that included baseline patient demographic characteristics, HBeAg/ anti-HBe status, ALT levels, and HBV DNA levels. Patients with pretreatment ALT and HBV DNA levels predictive of favorable treatment response to NAs were determined. Results: Fifty men (78%) and 14 women (22%) were included. Mean ages were 38 ± 13 and 28 ± 8 years, respectively. Thirty (63%) of the 48 patients with HBeAg-positive CHB had HBV DNA levels B9 log 10 copies/mL and ALT levels C2 9 ULN. Of 16 patients with HBeAg-negative/anti-HBe-positive CHB, 6 (38%) had HBV DNA levels B7 log 10 copies/mL and ALT levels C2 9 ULN. Conclusions: The proportion of Filipino CHB patients with characteristics predictive of favorable outcomes to NA therapy is similar to that of patients in global phase III trials, particularly among HBeAg-positive patients.

Yun-Fan Liaw 1 , Maria Raptopoulou-Gigi 2 , Hugo Cheinquer 3 , Shiv Kumar Sarin 4 , Tawesak Tanwandee 5 , Nancy Leung 6 , Robert P. Myers 7 , Robert S. Brown Jr 8 , Jolanta Bialkowska 9 , Yi-Cheng Chen 1 , Shijie Tang Background: There are limited data on safety and efficacy of hepatitis B virus (HBV) nucleos(t)ide therapy in patients with CHB and decompensated cirrhosis. We present Week 48 results from a randomized, open-label, comparative study of ETV versus ADV therapy in this population. Methods: 195 patients with HBeAg-positive or -negative CHB and decompensated cirrhosis (Child-Pugh score C7) were randomized (191 were treated) to ETV 1.0 mg or ADV 10 mg daily and treated for up to 96 weeks from date of last patient randomized. No prior anti-HBV therapy was allowed, with the exception of lamivudine (LVD) and interferon-alpha. The primary efficacy endpoint was the mean change from baseline in HBV DNA by PCR at week 24, analyzed by linear regression and adjusted for baseline HBV DNA and LVD-resistance status. Secondary efficacy endpoints included the proportion of patients with HBV DNA \300 copies/mL and with normalized ALT at week 48. Results: The study population was: 74% male; mean age 52 years; 54% Asian, 33% Caucasian; 54% HBeAg-positive; and 34% LVD resistant.

Week 24

Week ALT alanine aminotransferase, MELD model for end-stage Liver Disease, CTP Child-Turcotte-Pugh a The difference in HBV DNA respinses favoring ETV persisted when analyzed by subgroup (LVDr or HBeAg status), although the magnitude of the difference varied across subgroups b Analysis limited to patients with abnormal ALT at baseline c Analysis limited to HBeAg(+) patients at baseline, noncompleter = failure d Non-completer = failure

Rates of adverse events (AEs) and discontinuations related to study drug were comparable between the treatment groups. Cumulative HCC rates were: ETV 12%, ADV 20%; and cumulative death rates were: ETV 23%, ADV 33%. Conclusions: ETV demonstrated superior antiviral activity to ADV and provided clinical benefit in patients with CHB infection and decompensated cirrhosis. Both therapies were well tolerated and had a comparable safety profile; the majority of AEs in both groups were liver-related events, as expected for this population. Background: Global Phase III registration trials have shown that ETV is more efficacious than LVD in achieving histologic, virologic and biochemical improvements in nucleos(t)ide-naïve CHB patients. We present here a 48-week interim analysis comparing the efficacy of ETV versus LVD in nucleos(t)idenaïve HBeAg-negative Korean patients (ETV 105 study).

Methods: One hundred and twenty nucleos(t)ide-naive CHB patients were enrolled from 14 centers in Korea and randomized to receive either ETV 0.5 mg/day (n = 56) or LVD 100 mg/day (n = 64). The primary outcome measure was the proportion of patients with undetectable HBV DNA (\300 copies/mL by PCR) at week 24. Secondary endpoints included the proportion with undetectable HBV DNA and mean HBV DNA reduction from baseline through week 240, and the proportion with normalized serum ALT (\1 x ULN) at weeks 48 through 96. Results: Mean baseline HBV DNA levels were 6.1 ± 0.8 and 5.8 ± 0.9 log 10 copies/mL for ETV-and LVD-treated patients, respectively (P [ 0.05). At week 48, mean HBV DNA levels (estimated from linear regression model covariated with treatment group and baseline DNA level) were 2.5 ± 0.3 log 10 copies/mL in ETV-treated patients and 3.2 ± 0.3 log 10 copies/mL in LVDtreated patients (P \ 0.0001).

Week 24 Conclusions: In the ETV-treated group, significantly more subjects achieved an undetectable HBV DNA level and normalized ALT than patients treated with LVD, supporting earlier data that ETV has greater antiviral activity than LVD through 48 weeks. The safety profile of ETV is comparable to LVD, and similar to previous reports in HBeAg-negative patients. Objectives: To investigate the epidemiology characteristic of HCV infection and the related risk factors for HCV infection and possible transmission route in a prevail community of Fujian province. Methods: A community based epidemiology survey was taken and serum sample was collected to detect of HCV-Ab at the same time. Then amplified part of HCV NS5B gene of seropositive serum by RT-PCR. Phylogeny tree was constructed base on the sequenced result to genetyped the HCV gene and described the gene sequence homology between varied seropositive serum. Results: We investigated and collected 1009 serum sample from 1,351 people of the community. The seropositivity rate of HCV-Ab is 28.15%. There is no significance in seropositivity rate between sex (25.22 vs. 30.56%). It grew with age increased, it was the lowest in age of younger than 10-year-old group (2.14%) and highest in 50-60-year-group (56.70%), there is a significance difference between them. The rate was 3.35% in unmarried versus 43.64% in married (P \ 0.001). Living with a seropositive family member get a higher positive rate of HCV-Ab than no identified seropositive family member (34.36 vs. 20.70%, P \ 0.001, OR = 2.01). Farther more living with a seropositive spouse get a positive rate of 69.18% (110/159). Multiple factor logistic regression analysis revealed that age, living with seropositive family member and tooth extraction are independent risk factors of infection HCV. All of the genetype is 1b 88.89% (96/108) and 2a 11.11% (12/108), no any other subtype was found. It is high gene homology between the varied sequenced positive sample and also compared with sequence from mainland of China, Taiwan and Japan. Conclusions: The seropositivity rate of HCV-Ab in the community is 28.15%. All subgenotype of the HCV is 1b and 2a and there are highly gene homology between them. Age and living with seropositive family member are independent risk factors of infected with HCV. Invasive therapy before 1992 may play a key role in the diffusion of HCV when the absence of sterile hypodermic needles for single use. But the route of HCV transmission could not been certainty and farther research are needed. 1. During the 5-year follow-up, only 1.64% (3/183) HCV mono-infected subjects died on the cohort study, which was significantly lower than 21.21% (7/33) for HIV-1 group and 18.05% (26/144) for HCV/HIV-1 group. 2. The cause of death was mainly PCP (14, 38.89%),end-stage liver disease (8, 22.22%), cerebrum-related diseases (7, 19.44%), heart-related diseases (3, 8.33%), cryptococcal meningitis (2, 5.56%) and accident death (2, 5.56%) in this cohort survey (P = 0.0422). HCV co-infected patients are more prone to die with liver associated diseases compared with HIV mono-infected group (1 vs.7, RR = 11.23, 95%CI: 1.34-94.63). 3. AST, ALP and GGT which were indicators of liver function were highest in HCV/HIV-1 co-infected group (P \ 0.05, P \ 0.0001 and P \ 0.0001). Total protein (TP) was also highest in HCV/HIV-1 co-infected group ( P = 0.0088). ALT, ALB, TBIL, DBIL and A/G ratio were no significant differences among the three groups ( P [ 0.05). 4. Periphery CD4+ T cell count were lower in HCV/HIV-1 group (446.33 cells/ll) than in HCV group (864.78 cells/ll, P \ 0.0001) and HIV-1 group (518.89 cells/ll, P \ 0.0001).

Conclusions: HIV infection can accelerate liver damage and change the natural history of HCV infection.

Chronic Results: Among all patients 38 (21% of total) have opinion that they have been infected during surgical operation, 31 (17%) when visiting dentist and 5 (3%) during blood transfusion procedure, 7 (4%) in the cosmetic salon or during tattooing procedure, 1 (0.5%) got infected from sex partner and 9 (5%) through intravenous drug use. Majority of patients (79 or 44% of total) do not have opinion on where they got infected. Mean time from infection till this time according to patients' opinion is eight years (range 1-35 years). Conclusion: Our study draws a conclusion that majority of patients (74 or 41% of total), based on their own opinion, have been infected during medical procedures-surgical operation, visiting dentist or through blood transfusion. Background: Mongolia is known for its high endemicity for hepatitis B virus (HBV), hepatitis C virus (HCV), and hepatitis delta virus (HDV) infections among apparently healthy individuals. However, there are little or no data on the prevalence and genotype distribution of HBV, HCV, and HDV among patients with chronic liver disease in Mongolia. Therefore, serum samples obtained in 2004 from 207 patients including those with chronic hepatitis (n = 90), liver cirrhosis (n = 41), and hepatocellular carcinoma (n = 76) were tested for serological and molecular markers of HBV, HCV, and HDV infections.

Methods: The presence of HBV DNA was determined by the method described previously. The amplification product of the first-round PCR was 461 bps and that of the second round PCR was 437 bps, the nucleotide numbers are in accordance with a genotype C HBV isolate of 3,215 nt. Sera from individuals with anti-HCV were assayed for HCV RNA by reverse transcription (RT)-PCR using primers derived from the 50 untranslated region (UTR) of the HCV genome as previously described. The presence of HDV RNA was determined by RT PCR with nested primers derived from conserved areas of all reported HDV genomes of genotypes I, II, and III as described previously. Results: Of the 207 patients, 144 (69.6%), 106 (51.2%), and 117 (56.5%) tested positive for hepatitis B surface antigen (HBsAg) and/or HBV DNA, HCV RNA, and HDV RNA, respectively. Collectively, 172 patients (83.1%) were viremic for one or more of these viruses, including dual viremia of HBV/HDV (26.6%) or HBV/HCV (7.7%) and triple HBV/HCV/ HDV viremia (30.0%).

All 117 HDV isolates were classified into genotype I. The 106 HCV RNApositive samples were typed as genotype 1b (92.5%), 2a (0.9%), or 1b plus 2a (6.6%). Conclusion: The present study revealed that ongoing dual or triple infection of HBV, HCV, and HDV is highly prevalent among patients with chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma in Mongolia. Background/Aims: Genotyping of Hepatitis C virus (HCV) is important for designing therapeutic strategies and region specific diagnostic assays. The aim of this study was to determine the distribution of HCV genotypes among patients with chronic hepatitis C in China. Methods: Serum samples from 329 patients with chronic HCV infection from 12 areas in China were analyzed in this study. Viral genotypes were determined by a restriction fragment length polymorphism (RFLP) analysis of HCV 5' non-coding region (5'UTR). Results: The distribution of HCV genotypes in China was shown in Table 1 . Genotype 1b was common among the chronic hepatitis C group and was observed in 212 (64.4%) of these 329 patients. The subtypes following 1b were 2a (11.8%), 1b/1a (6.1%), 1b/2a (4.3%), 1a (4.0%), 3b (3.3%), 1c (2.1%), 2b (1.8%), 6a (1.2%) and 3a (0.9%). However, in Yanbian, 1a/1b was the dominant subtype (17/43, 40%) followed by 1b, 2a, 1a and 1b/2a. Moreover, subtypes 1b, 2a, 1b/2a, 3b and 1a were detected in more than 6 areas, while 1c was isolated only in Lanzhou and 6a was identified in merely southern areas (Guangzhou and Chongqing). Conclusions: Genotype 1b is the most frequently encountered subtype in China and genotypes 4 and 5 are not observed. However, some differences are noticed between diverse areas. The relationship between HCV genotypes and antiviral therapies or HCV vaccine remains further investigation in the near future. Introduction & Aim: Hepatitis C virus (HCV) is a major cause of chronic liver disease (CLD) in western world, where there is a steady increase in HCVrelated cirrhosis and related complications. In contrast, Hepatitis B is a major cause of CLD in Asia. In India, anti-HCV testing is available since 1993, but is made mandatory in blood banks since 2000. This study was planned to see impact of mandatory testing on clinical practice. Conclusion: After introduction of mandatory testing, awareness for HCV has gone up. This resulted in detection of more patients in asymptomatic rather than decompensated stage; thus increasing chances of treatment eligibility. In recent years, history of specific event leading to transmission of HCV was less commonly seen.

The Background: HCV RNA detection could reduce the risk of HCV transmission and aided in the early detection of HCV infections. The HCV genome is extremely heterogeneous, so the current available commercial detection kits that employ singleplex primer/probe hardly detect all the isolates of HCV and the missing detection will arise when the viral was mutated. Methods: By aligning HCV sequences recorded in the Los Alamos National Laboratory HCV sequence database, we developed a real time multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) assay using 2 sets of primer/probes and a specific armored RNA as internal controls. The 2 detection probes were labeled with the same fluorophore (FAM) in the 5' position and the same quencher in the 3' position, which could combined mutually creating greatly improved power of test. The sensitivity and specificity were compared between the singleplex primer/probe assay and multiplex primer/probe assay. Results: The sensitivity of the multiplex primer/probe assay improved significantly, with no effect on its specificity. The limit of detection of the multiplex primer/probe assay was 38.99 IU/ml. All HCV genotypes in the HCV RNA Genotype Panel (NIBSC, code 02/202, UK) could be detected. In the detection of 109 serum samples, the performance of the real time multiplex RT-PCR assay was identical to the COBAS Ampliprep/COBAS TaqMan (CAP/CTM) HCV assay and was superior to KeHua HCV RNA detecting kits (Shanghai KeHua Bio-engineering Co., Ltd.) and BIOER HCV fluorescence detection kits (Hangzhou BIOER Techonology Co., Ltd). Conclusions: The real time multiplex RT-PCR assay is a more efficient and effective viral assay as Compared to the singleplex primer/probe assay. Its assay is comparable to the CAP-CTM assay in the power of test and is appropriate for blood donor screening and laboratory diagnosis of HCV infection. Background: Hepatitis C virus (HCV) infection has been identified as the major cause of chronic liver disease among patients on chronic haemodialysis (HD). Information about the genotypes of HCV infected patients in Iran is limited. The aim of this study was to identify the HCV genotypes in group of HCV infected patients on HD in Guilan, North province of Iran. Materials and Methods: Patients on HD in all of 11 Guilan HD centers were enrolled; all sera were screened for anti HCV antibodies by enzyme-linkedimmunosorbent assay (ELISA) polymerase chain reaction (PCR) and genotyping was performed in positive anti HCV cases. SPSS 14 was used for analysis and P \ 0.05 were statistically considered significant. Results: Of 514 patients, 61 had positive anti HCV, (prevalence 11.9%) and among them 32 had positive HCV RNA (52.5%). The most frequent genotypes were 1a and 3a with 59.38 and 40.62% prevalence respectively. There were no significant relations between the type of genotypes and gender or age. Of note was the high prevalence of genotype 3a in Rasht, the capital city of province. Conclusion: HCV genotype pattern in our province, like other parts of Iran, is characteristic of the non-Arab countries of Middle East, where genotype 1 predominates. 

. Cutoff ([6.9) sensitivity for significant fibrosis (f 2 f 3 f 4 ) and extensive fibrosis (f 3 f 4 ) were (100%) specificity (0%) with accuracy (40%) and (20%) respectively.

-APRI score; AUROC (0.792) with 95% CI (0.568-1.015) comparing (f 0 f 1 ) vs.

(f 2 f 3 f 4 ) while was (0.875) with 95% CI (0.703-1.047) for (f 0 f 1 f 2 ) vs. (f 3 f 4 ). Cutoff (.5) sensitivity (0%) and specificity (100%) with accuracy (60%) for significant fibrosis and (80%) for extensive fibrosis.

-Fibrosure(fibro-acti test); showed best AUROC (1.00) in different fibrotic stages with 95% CI (1.00-1.00). Cutoff ([0.59) sensitivity (50%) for significant fibrosis and (100%) for extensive fibrosis while specificity (100%) in all fibrotic stages. The PPV (100%) for significant and extensive fibrosis. NPV and accuracy (75, 80%) respectively for significant fibroses, while NPV and accuracy (100%) for extensive fibrosis. Significant correlation between liver biopsy and Fibro-test (P 0.002) and Acti-test (P 0.000). Significant correlation between liver biopsy hepatitis activity score and APRI (P 0.047) and FORNS score (P 0.000). Conclusion: FORNS score was not considered since does not discriminate between significant and extensive fibrosis. Low sensitivity of APRI prohibtes detection of minmal fibrosis and allow undetermined results. Fibrosure classified all cases of chronic HCV sufficient to wave liver biopsy.

Evaluation of the Liver Stiffness Using Transient Elastography during Peg-Interferon Alpha-2b and Ribavirin Combination Therapy in Patients with Chronic Hepatitis C Katsuharu Hirano 1 , Takuya Genda 1 , Takafumi Ichida 1   1 Departement of Gasteroenterology, Juntendo University, Shizuoka Hospital, Japan

Background/Aims: Recently, many reports about elastography as the noninvasive evaluation of liver fibrosis were introduced. We investigated the change of liver stiffness during interferon therapy. Methods: We studied the patients who suffered from HCV-related chronic hepatitis and started PEG-IFNa-2b and ribavirin combination therapy between October 2007 and May 2009. A total number of patients registered prospectively were 100. We performed a blood test and liver stiffness measurement using elastography before the treatment and during the treatment every 4 weeks.

Results: The characteristics of the all 100 patients were median age 59.0 years old, included 55 patients had HCV genotype 1b, 45 patients had genotype 2. Pretreatment median liver stiffness was 8.1 KPa (2.8-34.8). We classified 39 patients with completed treatment. RVR was 14 patients, EVR was 16 patients, LVR was 4 patients, SVR 30 (76.9%), relapse 2, and NR was 7. The characteristics of SVR group were median age 51.5 years old, included 11 patients had HCV genotype 1b, 19 patients had genotype 2. Pre-treatment median liver stiffness was 9.2 KPa, post treatment median liver stiffness was 6.5 KPa, we showed significantly the decrease of liver stiffness (p \ 0.05) . The characteristics of non SVR group were median age 60.3 years old, included 8 patients had HCV genotype 1b, 1 patients had genotype 2. The change of liver stiffness ratio was 1/0.90/0.90/0.83 in SVR group, and 1/0.80/1.14/1.06 in non SVR group. 12 W stiffness ratio was statistically significant difference (p \ 0.05), we revealed the tendency that the decrease of liver stiffness appeared early in the treatment and persisted in SVR group, but did not persist in non SVR group. Conclusions: We concluded transient elastography was useful prediction of the effect of treatment of PEG-IFNa-2b and ribavirin combination therapy for the chronic hepatitis C.

Histological Staging and Grading of liver disease was done using modified histological activity index staging (score 0-6) and modified histological activity index grading (score 0-18) respectively. Results: A total of 594 patients were included in this study. Mean age of patients was 36 ± 8.5SD (range 18-63 years). 291 (49%) patients were male and 303 (51%) patients were female. In our study min. grade (necroinflammation) was 0 and max. Grade was 15 (mean = 5 ± 2.5SD). None of the patients has grade more than 15. 321(74.8%) patients were between grade 1 to 6. similarly 56, 23, 10, 5, 1.2, 1.7 and, &.7% patients were having stage (fibrosis) 1, 2, 3, 4, 5 and 6 respectively. Information regarding portal tracts was available in only 394 reports of liver biopsy. Average number of portal tracts in adequate samples was 6.6 ± 4.6SD. Conclusion: In our population most of the patients with chronic hepatitis c have mild disease with early histopathological changes. Background: Hepatitis C virus (HCV), a hepacivirus, is a member of the Flaviviridae family. Approximately 2% of the population worldwide is chronically infected with HCV and many of these infections will lead to recurring hepatitis, liver cirrhosis and hepatocellular carcinoma. Loop-mediated isothermal amplification (LAMP) is a novel technique for nucleic acid amplification. This simple and rapid technique relies on strand-displacing synthesis performed using the Bst DNA polymerase under isothermal conditions in the temperature range of 60-65°C. The method has also been rendered applicable to RNA genomes by combining it with reverse transcription reactions (RT-LAMP). Method: RT-LAMP is characterized by the use of 6 different primers. Amplification and detection of gene can be completed in a single step, by incubating the mixture of samples, primers, DNA polymerase with strand displacement activity and substrates at a constant temperature of 65°C. Sensitivity and specificity of primers was tested by RT-LAMP and Nested RT-PCR with RNAs extracted from HCV-infected as template. Results: We performed RT-LAMP reactions using genomic RNA s extracted from 150 HCVRNA-positive serum samples. Results showed RT-LAMP is more sensitive (2000 times) than Nested RT-PCR. Conclusion: RT-LAMP is rapid, sensitive, and specific test and also it could be a convenient tool for detecting HCV infection because it requires minimal laboratory facilities and is relatively simple and inexpensive to perform.

Study of Iron, Copper and Alpha-1 Antitrypsin in Egyptian Patients with Chronic Hepatitis C Ashraf Abugabal 1 , Naglaa Allam 1 , Mohamed Tawfeek 1 , Elhamy Abdel-moneim 1 , Mohsen Salama 1 1 National Liver Institute, Shebeen Al-koam, Menofeya

Background/Aims: Factors influencing the development of hepatic fibrosis in chronic hepatitis C are incompletely understood. Some metabolic abnormalities have been proposed as being additive factors for the pathology in chronic liver disease. The aim of the present study was to assess the association between some metabolic disorders (alpha-1 antitrypsin deficiency, disorders of iron indices, ceruloplasmin and copper metabolism) and histological characteristics in patients with HCV related chronic liver disease and determine their influence, if any, on progression of liver disease. Methods: This study was conducted on 40 patients with chronic hepatitis C (18 patients Child-pough A, 10 Child-pough B, 12 Child-pough C) and 10 healthy volunteers (control group). All were subjected to the determination of serum iron indices (iron, ferritin, TIBC, transferrin saturation), serum copper, ceruloplasmin and serum alpha-1 antitrypsin. Abdominal ultrasound was done. Liver biopsy was done for Child A. Results: Results revealed an increased prevalence rate of elevated serum iron, ferritin and transferrin saturation in HCV patients (20, 22.5, and 10% respectively) . In addition, there was a significant increase in the mean serum copper in these patients compared to the controls (p = .05). Alpha-1 antitrypsin was significantly lower in these patients (p = .05). A positive correlation between serum ferritin and transaminases was detected (p = .05, p = .001 respectively). Serum iron, copper, alpha-1 antitrypsin correlated insignificantly with liver function tests. Transferrin saturation correlated significantly with the fibrosis score (p = .001) and ceruloplasmin correlated significantly with the necroinflammatory grade (p = .05).

Conclusion: Transferrin saturation may be used as a marker of liver fibrosis. Although serum copper did not correlate with the pathologic changes, its effect may be more related to the duration of its accumulation. The possible role of copper in hepatocellular carcinoma justifies the follow up of its level in patients with HCV-related cirrhosis.

The Results: Comparing to HBV patients with liver cirrhosis, the ages of HCV patients with liver cirrhosis are bigger (the peak prevalence occurring among adults 31-40 and 51-60 years old, respectively). We found in these 25 HCV patients with liver cirrhosis: (1) the predominant genotypes are 1b and 2a, but there is no significant differences. (2) There are 9 (36%) patients have HBV and HCV coinfection; (3) there is only one patient happened hepatic failure who just infected HCV; (4) 9 patients (56.3%) of infecting HCV alone use interferon to treat, 4 patients (44.4%) of HBV-HCV coinfection patients use interferon, and one patient stop to use interferon because economy condition; (5) there are 10 patients (40%) are liver cirrhosis in broken compensation stage, and in these patients there are 5 (50%) patients are HBV and HCV coinfection.

Conclusions: Comparing to HBV with liver cirrhosis, the patients number of HCV with liver cirrhosis are few, pathogenetic condition always light, and mostly patients have chance to treat. The rate of HBV and HCV coinfection are high (36% and 50% in all patients and the patients who are in broken compensation stage, respectively). Results: Genotype 1 was found in 50 patients, with 1b in 42 (36%) and 1a in six cases (5%). Two patients were infected with mixed subtype of genotype 1: 1a/1b, 1b/3a. In 43 cases were enrolled 3a (37%), 3c in one patient. Nonclarified subtype of genotype 2, include 2a/2c, was revealed in 68% of cases. Prevalence of genotype 1b was approximately same in patients less than 30 years old (34%), from 30 to 40 (38%) and more than 40 (36%). Percentage of patients with genotype 3 was following in the same groups: 28, 44 and 41%. Conclusion: The present study showed circulation of genotypes 1, 2 and 3 in HCV-infected patients in Armenia with predominant role of subtypes 3a (37%) and 1b (36%). 

Background/Aim: Sterol regulatory element-binding protein-1 (SREBP-1) is key transcription factors which activate the transcription of lipogenic genes, including fatty acid synthase (FAS). As such, SREBP-1 plays a critical role in the pathogenesis of steatosis. Steatosis is an important clinical manifestation of hepatitis C virus (HCV) infection. The molecular mechanisms of HCV-associated steatosis are not well understood. Non-structural protein-5A (NS5A) of HCV is a multi-functional protein which is required for HCV genome replication and plays an important role in HCV-host interaction. The aim of this study is to investigate the role of HCV NS5A protein in modulating SREBP-1 activity. Methods: The NS5A coding sequence of HCV genotype-3a (HCV-3a) was cloned and expressed after transfection into human hepatoma Huh-7 cells. The effect of HCV-3a NS5A on SREBP-1 transcription and maturation was determined by measuring SREBP-1c promoter activity and the level of mature SREBP-1 protein by luciferase reporter assay and immunoblotting analysis, respectively. The impact of NS5A on the activity of FAS promoter was also studied.

Results: We showed that HCV-3a NS5A up-regulated SREBP-1c transcription as measured by promoter activities. Further analysis showed that transcriptional factor Sp1 is necessary because inhibition of Sp1 activity by mithramycin A or a dominant-negative Sp1 construct abrogated SREBP-1c promoter activation by HCV-3a NS5A. We further showed that HCV-3a NS5A could increase the level of mature SREBP-1 protein. Finally, we showed that HCV-3a NS5A could activate FAS promoter activity in an SREBP-1-dependent manner. Conclusion: Our results indicate that HCV-3a NS5A is likely a contributing factor for steatosis caused by HCV-3a infection. Recently several studies reported that hepatitis C virus (HCV) genotype 1b core amino acid (AA) substitutions at positions 70 and 91 are related with the treatment response of peg-interferon (IFN) and ribavirin. HCV core protein is also known to interact with interferon signaling pathways. We examined whether the HCV core substitutions affect IFN and cytokine-related signaling pathways in human hepatoma cell line HepG2. Materials and Methods: We made stably core-expressing cells having 4 kinds of AA substitutions at 70 and 91 and named as follows: WW (70R/91L), WM (70R/91 M), MW (70Q/91L), and MM (70Q/91M). Using these cellular RNAs, we screened IFN and cytokine signaling-related gene by real-time RT-PCR assay using the ddCt methods. We also performed reporter assay of IFN-beta promoter, interferon-stimulated response element (ISRE) and NFkB response. Results: HCV core proteins were expressed in all cell lysates. With regard to endogenous IFN-related gene, IFN-alfa and -beta mRNAs in cells with WM, MW, or MM (non-WW) were upregulated compared to those with WW (3.3 ± 1.8 and 2.1 ± 1.0 fold). As for cytokine related gene, IL1-beta and IL10 mRNAs in cells with WW were expressed higher than those with WW (3.7 ± 1.3 and 16.5 ± 10.3 fold). However, there were no different effects of IFN-beta promoter, ISRE-response and NFkB-response among these core mutants.

Conclusions: Some of IFN and cytokine-related gene expressions in HepG2 cells with non-WW were different from those with WW. However, our study did not show any different effects of HCV core AA substitutions at 70 and at 91 on IFN-beta promoter, ISRE-response or NFkB-response. We are now investigating the effects of core substitutions on these pathways in further detail.

Ren-Tian Cai 1 1 Nanjing Second Hospital, Zhongfu Street 210000, China

Aim: observe variation of expression of Programmed death-1(PD-1) on the surface of CD4 + T lymphocytes and CD8 + T lymphocytes in peripheral blood and the correlation between PD-1 and HCV RNA in chronic hepatitis C patients and to surmise the clinic significance. Methods: The PD-1 on CD4 + T lymphocytes and CD8 + T lymphocytes in peripheral blood were measured using flow cytometric assay. Fluorescent quantitation PCR measured HCV RNA. Gene Chip measured the genetypes of HCV.

Results: The expression of PD-1 on CD4 + T lymphocytes and CD8 + T lymphocytes in peripheral blood were up-regulated significantly in chronic hepatitis C patients. There were no correlation between PD-1 and HCV RNA and ALT. Conclusion: PD-1 was high expression on the surface of CD4 + T lymphocytes and CD8 + T lymphocytes and inhibited the ability of clean HCV in immunocytes result in chronicity of infection. Background: More than 170 million people worldwide are persistently infected with hepatitis C virus (HCV). It is yet unclear how this RNA virus establishes a persistent infection. However, it has been suggested that persistent infection appears to be due to virus heterogeneity and weak immune responses during acute infection. Recently, there is increasing evidence that innate immune defense is critical for the control of virus infection. The mechanism of HCV involvement in host innate immune responses, however, has not been fully understood yet. In the present study, we evaluated the expression profile of toll-like receptor (TLR) signaling pathway genes in human hepatocytes following HCV infection in vitro. Methods: The HCV replicon system (kindly provided by Dr. CM Rice) was used to generate infectious HCV virons. Viral infection experiments were performed using human hepatic cell line Huh7.5.1 (kindly provided by Dr. FV Chisari). The expression patterns of 84 genes related to TLR-mediated signal transduction were analyzed by real-time reverse transcription/polymerase chain reaction in both the naïve and the HCV-infected Huh7.5.1 cells. Gene silencing by siRNA was carried out for functional analysis. Result: All TLR genes expressed at relatively low level in Huh7.5.1 cells regardless of HCV infection. Although only one TLR gene had an altered expression after HCV infection, statistically significant differences were detected in expression of some proinflammation cytokines between the HCVinfected hepatocytes and control. Functional analysis revealed that some proinflammation cytokines had opposing proviral and antiviral effects. Aim: To develop a cell culture system capable of producing high titer hepatitis C virus (HCV) stocks with recombinant vaccinia viruses as helpers. Methods: Two plasmids were used for the generation of recombinant HCV: one containing the full-length HCV cDNA cloned between T7 promoter and T7 terminator of pOCUS-T7 vector, and the other containing the HCV polyprotein open reading frame (ORF) directly linked to a vaccinia late promoter in PSC59. These two plasmids were co-transfected into BHK21 cells, which were then infected with vTF7-3 recombinant vaccinia helper viruses. Results: After 5 days of incubation, approximately 3.6 9 107 copies of HCV RNA were present per milliliter of cell culture supernatant, as detected by fluorescence quantitative RT-PCR (FQ-PCR). The yield of recombinant HCV using this cell system increased 100-to 1,000-fold compared to in vitrotranscribed HCV genomic RNA or selective subgenomic HCV RNA molecule method.

Conclusion: This cell culture system is capable of producing high titer recombinant HCV. Acknowledgment: This work is supported by the National Natural Science Foundation of China (Nos. 30872244, 30771899). Result: 501 HCV infection patients were retrieved from our databases, of which 37 patients with PCT were retained for further analysis. Patients were middle aged (mean age 51 ± 3) and the male-female ratio was approximately 6:1 (32 males vs. 5 females). Almost all patients had detected porphyrin in the urine (35/37). 19 patients had iron overload, all patients had hyperpigmentation of face and blistering skin lesions. In 16 patients, PCT was associated with hemoglobinemia and 4 patient had chronic hepatitis B virus. The majority of PCT patients with HCV (*80%) had elevated transaminases (100-400 U/L). 17 patients were treated with phlebotomy, with success in 12. 67% of PCT patients with job related photosensitivity, i.e., farmer or herder. Conclusion: PCT prevalence in patients with HCV is approximately 7.3%. The environmental factors may also be involved in the pathogenesis of this disorder. Iron overload is present in the majority of patients and the majority of patients can be successfully treated with phlebotomy.

Top 10% Posters Objectives: Cytokines triggering is a common feature in chronic hepatitis C infection (CHC). Visfatin, as a visceral fat-specific adipokine, exerts insulinmimicking effect and activates the intracellular signaling cascade for insulin. The study aimed to assess the role of serum visfatin in virological profiles and glucose abnormalities in CHC patients. Methods: One hundred and two consecutive treatment-naïve Taiwanese CHC patients (56 males, age = 52.9 ± 10.5 years) and 97 sex-and age-matched healthy adults (42 males, age = 54.2 ± 9.7 years) were recruited. Serum visfatin levels were examined in duplicate by a commercially available ELISA (Phoenix Pharmaceuticals, Burlingame, CA, USA). The correlation between serum visfatin level and HCV infection in terms of virological profiles and insulin resistance (IR) by the homeostasis model assessment (HOMA-IR) method was analyzed. Results: The 102 CHC patients had a significantly higher mean visfatin level than that of the 97 controls (10.55 ± 7.11 vs. 9.05 ± 6.27 ng/mL, P = 0.004 System diseases were excluded additionally: RF-neg., CRP-3,5, antiDNAnneg., antiDNAd-neg., HCV RNA-positive, genotype 1b, viral load-83000 U/ ml. Liver biopsy-A10, F2. Patient got Pegasys: Pegasys 180 mkg in a week and 1,000 mg of ribavirin a day during 48 weeks. After first injection-muscular weakness, hand oedema in pogress. Patient was examined by the traumatologist on the third week, XR: microvascular hands arthrosis. Increased sclerodermitis clinical symtoms:oedema, pain, microvascular hands stiffness, edema-indurative face skin changes, dry cough, heartburn, lungs and esophagus XR-without changes.Skin biopsy (6th week)-evident sclerosis with skin appendages atrophy, vessels sclerosis with local lympohystiocitus infiltration. SSD therapy is vasoprostan, vasonit 600 mg, madecasag 10 mg a day, placvenil 0.2 g a day.

Specific markers: 10th week: anti-Scl-708 s!-70-anti RNA pol III, capillaroscopy-a Reno's syndrome. HCV RNA is not detectable on 12th, 24th and 48th weeks of treatment. No SSD symptoms progression. Titers anti-Scl-708 s!-70 and anti RNA pol III and capillaroscopy are without negative dynamics. HCV RNA is neg. in 4 months after the treatment follow-up. No clinical signs of system sclerodermitis progress. Increase of titres anti-Scl-708 s!-70-antiRNA pol III are not detected. Evidence of Reino's syndrome on capillaroscopyindurative changes of skin, telangiectasis have decreased. Background & Aims: The development of subgenomic and genomic replicons provided a major breakthrough to the understanding of HCV replication and viral-cell interactions. Huh-7.5.1 cells transfected with in vitro transcribed JFH-1 genomic RNA will successfully secrete virions. Therefore, transfection of JFH-1 RNA into the Huh-7.5.1-derived cells allows for the recovery of a viable JFH-1 virus that can then be serially passaged and used for infection-based experimentation. To determine the virus-host relationship, we also established this replication system and used this system for further study. Methods: Genotype 2a HCV full-length replication system was established using plasmid pJFH1 containing the full-length JFH1 cDNA. We used Tran-Singnal Protein/DNA Array to examine the effect of HCV replication on 345 transcription factors. We also performed luciferase reporter assays and electrophoretic mobility shift assay to confirm TranSingnal Protein/DNA Array results. We further examined NF kappa B associated factors such as I kappa Balpha and IL-8. Results: As results, 12 kinds of transcription factor were increased by HCV replication, such as c-Rel (NF kappa B p75 kDa protein), and ISRE (interferon stimulated response element). On the contrary, 11 kinds of transcription factor were decreased by HCV replication, such as Stat4 (signal transducer and activator of transcription 4), and GAS/ISRE (interferon activated factors). Activation of NF kappa B and ISRE has been confirmed using luciferase reporter assays and electrophoretic mobility shift assay. We have also confirmed c-Rel increased in the nuclear extracts from HCV replication cells. We have found phospho-I kappa B-alpha; was increased in the HCV replication cells, and IL-8 was also increased in the HCV replication cells culture. Conclusions: HCV replication affected NF kappa B system that triggers immunity system and inflammation response. Full-Length HCV replication system enables systemic analysis of intracellular response in human hepatocytes. Background & Aims: We and others have observed that levels of hepatic SOCS3 are significantly higher in chronic hepatitis C (CHC), particularly among persons who are nonresponders to subsequent IFN treatment. However, the relationship between SOCS3 and HCV replication remains unclear. We therefore sought to dissect the relationship between SOCS3 and HCV using HCV replication models. Methods: We used the genotype 1b full-length HCV replicon OR6 cell line, and the genotype 2a HCV full-length JFH1 infection system. We compared SOCS3 levels between cured and uncured OR6 cells, and between JFH1infected and uninfected Huh7.5.1 cells. In addition, we analyzed the effect on HCV replication by (1) transient overexpression of SOCS3 in OR6 and JFH1infected cells, and by (2) stably knocking down SOCS3 using shRNAs in Huh7.5.1 cells. We further analyzed the role of mTOR by treating cells with rapamycin. Results: OR6 cells and JFH1 infected Huh7.5.1 cells expressed significantly less SOCS3 than cured or uninfected cells, respectively. Inhibition of HCV replication with the HCV protease inhibitor BILN2061 restored SOCS3 protein levels. SOCS3 overexpression in OR6 cells and JFH1 infected Huh7.5.1 cells resulted in significantly lower HCV replication than in the control cells. JFH1infected, stable SOCS3 knockdown cells expressed more HCV than did control cells. The shSOCS3 also knocked down mTOR and phospho-mTOR. The mTOR inhibitor rapamycin reversed SOCS3's inhibitory effects on HCV RNA and core protein levels.

Conclusions: SOCS3 suppresses HCV replication in an mTOR-dependent manner. Our data instead suggests a model by which increased SOCS3 levels observed in chronic IFN nonresponders may reflect a compensatory host response to persistent infection and impaired defenses elsewhere in the innate antiviral pathway. Introduction: This is a seroprevalence study of HCV genotypes amongst infected haemodialysis patients in Pahang, Malaysia and the development of genotyping method using real-time PCR. Methods: Patients were screened using ELISA by detecting the anti-HCV in the sera. All negative first-round PCR products were re-tested by nested PCR. The base sequence of the PCR products was determined using the same primers as for the RT-PCR. By comparing the obtained nucleotide sequence data with sequences of known genotypes from the NCBI homepage, we deduced that our local isolates could be assigned to genotypes 1, 3, 4 and 6. Based on the data, TaqMan Ò probes were designed for the simultaneous identification and quantitation of these genotypes. A new batch of blood samples were recollected from patients and real-time PCR (Applied Biosystem) assay was conducted for genotyping. Results: Out of 472 patients, 43 (26 males) were diagnosed positive for anti-HCV by ELISA. Two seropositive patients were excluded as they refused to give consent. HCV RNA was detected by RT-PCR technique targeting 5'UTR region. Excluding those who were seroconverted to HCV negative and patients with low titer, 66.6% (26/39) were of genotype 3, 23.1% (9/39) of genotype 1, 5.1% (2/39) of genotype 6, followed by 2.6% (1/39) of genotype 4 and one patient gave a discordant result with the sequencing analysis. Conclusion: Genotype 3 was the most prevalent genotype followed by genotype 1, 6 and 4. TaqMan real-time PCR has the potential as an alternative method for rapid HCV genotyping.

Hepatol Int (2010) Background/Aim: The third generation anti-HCV EIA test (S/CO [1, AxSYM HCV 3.0; Abbot, IL, USA) has been demonstrated to be a semi-quantitative methods for prediction of viremia in HCV patients. Subject with S/CO value B10 were more likely to be a pass infection or non-specific reaction. However, the titer might be more heterogenous in patients with hemodialysis (HD) because of poor immune response. In this study, we aim to evaluate the validation of third generation anti-HCV EIA test in patients with HD. Methodology: We retrospectively reviewed 199 HD patients with anti-HCVpositive (S/CO[1) for at least 6 months. The serum HCV viral load, genotype, ALT levels were analyzed for anti-HCV titer correlation. Total 157 cases with obtained serum were enrolled for analysis. Results: The S/CO distribution was biphasic, with two S/CO peaks in the range 1-10 (24.6%) and[90 (41.7%). Among the 157 patients, detectable HCV RNA was found in 113 (56.8%). Of these with anti-HCV positive, the optimal cut-off for the prediction of HCV viremia, as derived from the ROC curve, was 36.5, with sensitivity and specificity of 75 and 25%. Of the 43 patients with S/CO values210, 22 (51.2%) was HCV RNA detectable. The sensitivity, specificity, PPV, NPV and accuracy of S/CO values210 were 31.9, 81.4, 81.8, 31.3 and 45.5%, respectively. Conclusions: These results suggest that in contrast to immunocompetent patients, anti-HCV antibodies titer was not a reliable marker to predict HCV viremia in HD patients.

Rathindra mohan Mukherjee 1 , P. Balkumar Reddy 1 , Padaki Nagaraja Rao 2 , Mitnala Sasikala 1 , Rajesh Gupta 2 , D. Nageshwar Reddy 2 1 Asian Health Care Foundation, Hyderabad-500082, India, 2 Asian Institute of gastroenterology, Hyderabad-500082, India

Background/Aims: The relationship between viral replication, genotypes, host biochemical characteristics, clinical outcomes and mode of transmission in Indian subjects with incidental detection of hepatitis C virus antibody (anti-HCV) remains unclear. to gain insight, we evaluated the above parameters in incidentally detected otherwise asymptomatic anti-HCV positive subjects. Methods: A total of 93 persons with incidental detection of anti-HCV antibody upon scrutinization for their family/clinical history were subjected to endoscopic and ultrasound evaluations. Blood samples were assessed for various biochemical and serological markers as per standard procedures.Quantification of serum HCV RNA were performed by real time PCR assay. HCV genotypes were evaluated in HCV RNA positive samples by line probe reverse hybridization assay. Results: Majority (52.7%) of the anti HCV positive subjects were identified for investigations of non specific symptoms other than HCV related disease, followed by the pre surgical check up (19%) and blood donation (14.7%). Previous history of dental procedures (47.3%), surgical procedures (34.7%) and blood transfusions (25.2%) were noted as major risk factors in these subjects.Elevated mean ALT (71.2 ± 46.7), AST (61.7 ± 39.5) and INR (1.32) values were evident among the HCV infected subjects. Genotype 3 was predominant (46%), followed by genotype 1(37%) and genotype 4 (15%) respectively while genotype 5 was identified in a single subject for the first time from southern India. A group (22%) of HCV infected subjects had diabetes. Older subjects (30%) had lower serum albumin and platelet counts while their INR and AST values were significantly higher. Cirrhotic features were observed in a number of subject (45.2%)s having both low and high viral load.

Conclusion: Among these apparently asymptomatic anti HCV positive subjects, many (45.2%) had features indicating cirrhosis of liver which is more pronounced in older subjects. Association of diabetes with HCV infection was also noted. However, the viral load and HCV genotypes had no significant association with the progression of the disease although identification of genotype 5 for the first time from a single patient with liver cirrhosis in southern India needs further study. Dental interventions appeared as most predominant risk factor followed by surgical procedures and blood transfusion.

Impact of Hepatitis C Virus Infection on Glycemic Control and Lipid Profiles of Diabetic Patients Nauman Arif Jadoon 1 , Rehan Yaqoob 1 , Muhammad Asif Shehzad 1 1 Nishtar Medical College, Multan -Pakistan, Nishtar Road Multan, 60,000, Pakistan

Aims: The aims of this study were to: 1. Determine the effect of Hepatitis C virus infection on glycemic control in patients with diabetes. 2. Study the impact of Hepatitis C virus Infection on lipid profile of individuals with diabetes. Methods: Five hundred and fifty diabetic patients attending diabetes clinic were enrolled in the study. Patients' data was collected after taking consent. Hepatitis C virus antibody presence was checked using ELISA in the study group. Patients' glycemic control was checked and lipid profile was analyzed. Blood pressure, body mass index (BMI) and waist-hip ratio (WHR) were measured. All the ethical requirements were met before starting the study. Results: The age of patients was 47.58 years and the duration of diabetes was 7.02 years. Out of 550 patients included in study, 304 were female, 428 were from urban locality and 143 had a positive family history of diabetes mellitus. Patients with HCV infection had significantly lower total serum cholesterol, serum triglycerides, LDL cholesterol, LDL cholesterol/HDL cholesterol ratio and a lower waist to hip ratio as compared to diabetic patients without HCV infection. In contrast, they had significantly higher random blood sugar value. Furthermore, diabetic patients with HCV infection had insignificantly lower HDL cholesterol, fasting blood glucose and HbA 1 c level. They also had insignificantly higher systolic blood pressure diastolic blood pressure and BMI when compared with diabetic patients who tested negative for HCV infection. Conclusion: The study shows that although HCV infection is associated with high random blood sugar values, the remaining metabolic and cardiovascular risk indicators show a favorable pattern. It is an intriguing finding as HCV infection has been shown to induce insulin resistance compounding diabetes course but in this case, it had a positive influence on the metabolic and cardiovascular risk profiles of diabetic patients.

for anti-HCV by ELISA. RNA extraction and RT-PCR from seropositive samples were performed in order to amplify the HCV core region. Positive sample was subsequently sequenced and the genotype was determined by phylogenetic analysis. Results: Seroprevalence of HCV infection in migrant workers from Cambodia, Laos and Myanmar were 33 (2.3%), 7 (0.8%) and 27 (1.69%), respectively. Based on phylogenetic analysis, HCV genotype in this group was classified as 1a, 1b, 3a, 3b and 6 (6e, 6f, 6 m, 6p and 6r). One sample from a Laotian migrant worker showed HCV-6 but unassigned subtype. Most of the isolates of HCV were common, though some of subtype 6 was not found in Thailand. Background: Hepatitis C virus (HCV) and Human immunodeficiency virus (HIV) can share the same road of transmission, especially in injection drug users (IDUs). It has long been accepted that HCV-HIV co-infection have more rapid progress of liver damage and to liver cirrhosis. However, the mechanism cannot be well understood since liver damage in HCV infection is associated primarily with series of immune reactions and patients with HIV infection may have certain degree of immune suppression. Aim: To reveal histopathologic appearance of severe immunodeficient patients with HCV-HIV co-infection. Method: This is a cross-sectional study in naïve patients with HCV-HIV coinfection. All patients had CD4+ cell count of \200 cell/lL. Mean of CD4+ cell was 44.5 cell/lL. Mean age was 30.6 ± 3.1 years old with median duration of HCV infection (defined from the time of the first time usage of IDU) was 9.8 ± 2.3 years. Liver biopsy scored with METAVIR showed necroinflammation of mild (score 0 and 1), moderate and severe were 41.6, 47.9 and 10.4%, respectively while fibrosis appeared to be mild (score F0 and F1), F2, and F3 in 50, 47.9, and 2.1%, respectively. No patients had F4 fibrosis although few of them had already more than 10 years infected with HCV. Conclusion: Severe immune deficiency state in patient with HCV-HIV coinfection prevent the immune mediated liver damage.

What Matters in a Liver Biopsy Core Specimen in Chronic Hepatitis C:

The Introduction: Liver biopsy (LB) has an integral role in management of chronic hepatitis C (CHC) particularly in assessing stage of fibrosis (SoF) and grade of inflammation (GoF). We evaluated the effect of size and number of portal tracts (NoP) on the grading and staging of biopsy. Methods: A cross sectional study was conducted at AKUH to compare size of liver biopsy core (LBC) and NoP with GoI and SoF on liver biopsy sample. The Batts and Ludwig scoring system was used to grade and stage CHC. Results: We examined 150 (98; male) consecutive patients with CHC undergoing LB. Mean length of LBC was 1.34 ± 0.57 cm. Mean number of portal tracts (NoP) was 11 ± 4.6. We found that mean length of LBC was 1.22 ± .37.cm in grade 1(n = 17 i-e 11.3%), 1.46 ± .5 in grade 2(n = 64 i-e 42.6%) and 1.5 ± .45 in grade 3 (n = 69 i-e 46%); (p = 0.102). Mean length of LBC was 1.28 ± 0.39 cm, 1.47 ± 0.56, 1.34 ± 0.30 and 1.60 ± 0.45 in SoF 1, 2, 3 and 4 respectively. Mean length of LBC was greater (1.60 ± 0.45) in stage 4 (n = 41 i-e 27.3%) and lesser (1.28 ± 0.39) in stage 1(n = 23 i-e 15%); (p value = 0.04). The mean NoP were 8.53, 10.61 and 13.16 in GoI 1, 2 and 3 respectively (p \ 0.0001). The mean NoP were 7.61, 11.12, 11.31 and 14.54 in SoF 1, 2, 3 and 4 respectively (p \ 0.000). Significant correlation was found between the length of LBC and NoP. There is a good correlation between number of portal tracts and length of biopsy core (r 2 0.56). Conclusion: We concluded that number of portal tracts is better predictor than size of liver biopsy core in accurately establishing both GoI and SoF in a liver biopsy. Background: In human Toll-like receptor 3(TLR3) and Mitochondrial antiviral signaling protein (MAVS) involved in innate immunity has been identified to respond to dsRNA a molecular signature of viruses like hepatitis C virus (HCV). Aim: To study the expression of TLR3 and MAVS gene in naïve and after antiviral therapy patients with chronic HCV infection (N = 30) who receive peg-IFNa-2a 180 mg/weekly and ribavirin 100 mg/daily. Method: Expression of TLR3 and MAVS was analyzed by immunohistochemistry and semi-quantitative RT-PCR using housekeeping b-actin gene as the internal control and then validating the copy number of TLR3 by real time PCR. The patients were then further subdivided into two group based on sustained virological response (SVR) and Nonresponder (NR). Results: The expression of TLR3 is upregulated in patients who are NR compared to baseline whereas in SVR it is upregulated by immunohistochemistry. The expression of MAVS is upregulated in patients who achieve SVR compared to baseline whereas in NR it is upregulated by immunohistochemistry. The baseline expression level is similar in both the group by semiquantitative RT-PCR. The expression of TLR3 is upregulated in patients who are NR compared to baseline whereas in SVR it is upregulated. MAVS Hepatol Int (2010) 4:94-345 181 expression by semi-quantitative RT-PCR is upregulated after therapy whereas it could not be detected at baseline Expression analysis of TLR3 and MAVS by real time PCR also validates the finding of semi-quantitative rt-PCR. The copy number of TLR3 is highly significant between SVR (DDC T = 2.18) and NR (DDC T = 3.74). In SVR (DDC T = 6.23) the copy number of MAVS is slightly upregulated compared to NR (DDC T = 6.89). Conclusion: Expression levels of TLR3 and MAVS by immunohistochemistry may be helpful in better patient's management for antiviral therapy. This need to be confirmed in more number of patients based on HCV genotypes which determine antiviral therapy. No statistically significant differences in the baseline HCV RNA and ALT activity were observed between responders and non-responders. The factor of good treatment response appeared presumed infection period: 3.5 ± 3.4 vs. 6.5 ± 3.4 (p \ 0.04) for R and NR, respectively. Adverse reactions were observed, with leucopenia predominant one, however without discontinuation of the treatment. Conclusions: Pegylated interferon with ribavirin appears to be an effective and safe therapeutic alternative in children with CHC. Shorter HCV infection period seems to be indicative of a higher treatment efficacy. Further studies are needed to assess the efficacy of treatment in children converted from recombinant interferon to pegylated interferon. Purpose: Double filtration plasmapheresis (DFPP) is newly developed in Japan that filter HCV and lipids from circulating blood. DFPP is reported to achieve ealy viral response (EVR) with interferon therapy. Twice daily administration of interferon-beta (IFN-b) is known as the most effective therapy to achieve EVR in Japan. We evaluated the boosting effect of HCV reduction by the combination of interferon-beta (IFN-b) and DFPP for the patients to achieve partial EVR but not viral response, who had been treated with pegylatedinterferon (PEG-IFN) and ribavirin (RBV). Methods: We evaluated 10 chronic hepatitis C patients with genotype1b and high viral load who had been received more than 20 weeks of combination therapy with PEG-IFNa2b and RBV, whose HCV-RNA had reduced more than 2Log but had not achieved viral response. We have compared the boosting effect between Group A (3MU of IFN-b, administered twice daily) and Group B (3MU IFN-b, administered twice daily with DFPP performed 5 sessions). Results: 7 patients were Group A. F/M were 5/2, average age was 61.7 years old. All 3 patients of Group B were male, average age was 59.7 years old. The period of IFN-b administration; Group A: 14-22 days, Group B: 9 days. The number of patients who achieved viral response after the end of IFN-b administration; Group A: 3 of 6, Group B: 3 out of 3. Conclusion: For patients who were refractory to conventional therapy with PEG-IFN and RBV, DFPP combined with IFN-b twice daily showed a rapid booster effect of HCV reduction and all patients had achieved viral eradication. Background: Hepatitis C virus (HCV) is a public health problem and a course of chronic liver disease. The goal of therapy is to prevent complications and death from in chronic hepatitis C. Treatment responses are defined by several virological parameter such as early virological response (EVR), end-of-treatment response (ETR) and sustained virological response (SVR). Virological relapse is the reappearance of HCV RNA in serum after treatment is discontinued and an ETR was documented. Virologic relapse occurs within the first 12 weeks and late relapse, beyond 24 weeks, is extremely uncommon. We studied to know the relapse rate after pegylated interferon alpha 2a and ribavirin therapy in patients with chronic hepatitis C. Conclusion: Relapse rate of chronic hepatitis C after combination therapy with pegylated interferon alpha 2a and ribavirin was 9.8%. All relapse occurred before SVR. We do not know the factor affecting relapse in this study. We need more long-term follow up after SVR and multi-center data to analyze the factor affecting the relapse.

Top 10% Posters Background: To predict HCV CTL epitopes that would cover HLA polymorphism of the majority of Chinese people, and design corresponding multiepitope antigen gene. Primary investigation on immunogenicity of this gene. This work will contribute vastly to the application of specific immunotherapy on the basis of HCV CTL epitope for Chinese. Methods: According to knowledge of molecular immunology that CTL epitopes presented antigen by MHC-I molecules, HCV CTL epitopes were predicted by genomics and bioinformatics analysis tools. Candidate epitopes would be in series in accordance with the genetic algorithm. A synthetic multiple epitope antigen gene was cloned into vector pcDNA3.1 (+), then they were used to immunize mice. Specific cellular immunity of mice was detected by MTS, ELISA and CTL. Meanwhile, T lymphocyte epitopes and frequencies of epitope specific T cells were determined by ELISPOT using PBMCs from HCV patients stimulated by mixture of peptides. Results: 12 predicted HCV CTL epitopes were combined according to their superiority. Gene sequence of HCV multi-epitope antigen was obtained, which include different HCV genes areas and a number of CTL epitopes. The specific lymphocyte proliferation level, IFN-c and CTL activity of mice immunized (Part 2), or who were naïve to treatment for HCV (Part 3). Doses of PEG-IFN-k were body weight-based and ranged from 0.5 to 3.0 lg/kg. Assessments include adverse events (AEs), laboratory values, and changes in HCV RNA. Results: 56 patients (mean age 51 years, 69% male, mean baseline HCV RNA 6.43 log 10 IU/mL) were randomized to treatment. Dose-dependent reductions in antiviral activity at 4 weeks were observed, with 83 and 86% achieving [2log decrease in HCV RNA at the PEG-IFN-k 1.5 lg/kg + RBV dose in IFN-a relapse and treatment-naïve patients, respectively. Treatment was generally well-tolerated with minimal flu-like symptoms and no clinically meaningful changes observed in neutrophils, platelet counts or hemoglobin levels (apart from RBV associated reductions). 6/56 (11%) of patients experienced reversible increases in ALT/AST with or without increases in bilirubin (majority at the 3.0 lg/kg single agent dose, Part 1). Conclusions: PEG-IFN-k was associated with robust antiviral activity at all dose levels tested in both IFN-a relapse and treatment-naïve patients. Treatment was generally safe with minimal flu-like symptoms or heamatologic effects observed. These favorable results support the initiation of Phase 2 studies in treatment-naïve patients. Background: BMS-790052 is a first in class and highly selective HCV NS5A inhibitor with picomolar in vitro potency against genotypes 1a and 1b. In a single ascending dose study with healthy subjects, BMS-790052 was shown to be safe, well-tolerated, and had a pharmacokinetic profile suggestive of once-daily dosing. Methods: The objectives of this randomized, double blind, placebo-controlled, single ascending-dose study were to evaluate the safety, tolerability, antiviral effect and pharmacokinetics of BMS-790052 in genotype 1 HCV + patients. Treatment naïve or experienced patients (18-49 years of age, HCV RNA C10 5 , with non-cirrhotic compensated liver disease) were randomized to receive 1, 10, or 100 mg of BMS-790052 or placebo (6 patients per dose; active: placebo = 5:1). Results: All BMS-790052 single doses were well tolerated and had a safety profile similar to that of placebo. BMS-790052 exposure increased in a doserelated manner similar to that observed in healthy subjects. The mean terminal half-life of BMS-790052 was approximately 12 hours. Mean decline in HCV RNA 24 hours after a single 1, 10 and 100 mg dose of BMS790052 was 1.8 log 10 (range 0.18-3.0 log 10 ), 3.2 log 10 (range 2.9-4.0 log 10 ) and 3.3 log 10 (range 2.7-3.6 log 10 ), respectively. Furthermore, the 100 mg dose resulted in a mean decline of 3.6 log 10 (range 3.0-4.1 log 10 -observed at 48 h after dosing, which was maintained at 144 h. Conclusions: BMS-790052 is a potent NS5A inhibitor that produces a robust decline in HCV RNA following a single dose in patients chronically infected with HCV genotype 1. BMS-790052 was safe and well tolerated in single doses of up to 100 mg and has a pharmacokinetic profile that potentially supports once-daily dosing. Phase 2 trials of BMS-790052 in combination with peg-IFN/ RBV, as well as a study in combination with an NS3 inhibitor are ongoing. Objective: To study the virological features of patients coinfected with HBV and HCV and the efficacy of combination therapy with peginterferon alpha-2a and ribavirin in these patients. Methods: The virological response rates of patients treated with peginterferon alpha-2a and ribavirin between the HBV and HCV coinfection group and the HCV monoinfection group were compared. Results: HCV-dominant virus strains accounted for 92.0% of the 50 coinfected individuals. The HBV DNA level of coinfected patients was 4.6 ± 0.9 log(10) copies/ml, which was significantly lower than that in HBV monoinfection group (5.9 ± 1.2 log(10) copies/ml) (t = 5.964, P \ 0.01). The HBeAg-positive rate (12.0%, 6/50) of coinfection group was significantly lower than (45.3%, 19/42) that of the HBV monoinfection group (chi (2) (2) = 0.090, P = 0.765). The incidence of side effects (30%, 15/50) of patients in the coinfection group was significantly higher than that (13%, 6/46) in the HCV monoinfection group (chi(2) = 4.031, P = 0.045). The reactivation rate of HBV DNA (33.3%, 9/27) with HCV SVR was significantly higher than that of patients without SVR (8.7%, 2/23) (chi(2) = 4.393, P = 0.036).

Conclusions: The replication of HBV was suppressed, and HCV was the dominant virus strain. Compared with HCV-monoinfected patients, pEVR, ETVR and relapse rates of patients with genotype 1 in the coinfection group were high, while they shared similar SVR rates. Background: Pharmacogenomic study of patient's single nucleotide polymorphism (SNP) and viral genotypes of hepatitis C virus can be used to understand the relationship between genetic inheritance and anti-HCV therapeutic response. The aim of the study was to establish a predictive model using artificial neural network (ANN) model based on genetic and clinical factors. Methods: We prospectively collected 150 patients with chronic hepatitis C (CHC). 50 patients were excluded due to previous other causes of hepatitis. Nine patients dropped out of treatment due to Intolerance to drugs. 91 remaining patients (male 43, female 48; mean age 56.5 ± 9.0 years/old) were enrolled into the study. We extracted DNAs from blood samples before initiating treatment. They had completed pegylated interferonaand ribavirin combination therapy for 24 weeks (genotypes 2 & 3) and for 48 weeks (genotype 1). Sustained virologic response (SVR) rate was obtained after six months' follow-up. After genotyping analysis, the probability of responsiveness to therapy based on the individual genotype could be obtained. Furthermore, an ANN approach was used to predict therapeutic response combining above host genetic factors and other clinical factors. Results: There were seven SNPs selected from six candidate genes. A genetic prediction model was built for initial testing. The above data associated with other clinical factors, such as sex, age, genotype, viral load, stage of liver fibrosis, body mass index, liver function tests (ASL, ALT) were analyzed using ANN algorithms. The data of ANN approach were validated based on fivefold crossvalidation methods with results of sensitivity = 98.7%, specificity = 41.7%, positive predictive value = 91.7%, negative predictive value = 83.3%, accuracy = 91.1%, and area under receiver operating characteristics curve = 0.702. Conclusions: This prospective study suggests that the ANN model is useful to predict therapeutic response accurately before starting anti-HCV treatment.

Comparison of Liver Function Tests in Responders and Non-Responders of the Treatment with PEG Interferon Alfa 2a plus Ribavirin in Chronic Hepatitis C Narina Sargsyants 1 , Hripsime Magdesieva 1 , Alvard Hovhannisyan 1 , Mher Davidyants 1 1 Yerevan State Medical University, Koryun 2, Yerevan, Armenia Objectives: The difference in dynamic of liver enzymes and ferritin in patients with chronic hepatitis C with following sustained virological response (responders R) and patients with virological failure (non-responders NR) on the treatment with PEG Interferon alfa-2a and ribavirin. Methods: 50 patients were treated with PEG Interferon alfa-2a plus ribavirin. Longitudinal study of ALT, AST, and GGT was performed every 4 weeks. For measurement of plasma FERR level on start-point, 12 weeks and end-point we used two-site immunoenzymatic assay. Statistic analyses were carried out with SPSS 11.0. Results: At the start-point level of GGT was significantly higher in HCVinfected with duration of disease more than 10 years (less than 5 years-77.0 ± 25.7; 5-10 years-88.7 ± 9.7; more than 10-159.2 ± 33.6) and AST was significantly higher in patients with obesity (body mass index less than 25-56.8 ± 8.0; BMI 25-30-59.3 ± 11.9; BMI more than 30-79.4 ± 20.1). In R mean of AST and GGT were normalized on the 4 weeks of treatment, ALTon the 8-12 weeks. Level of GGT in NR elevated on 8-16 weeks of combination therapy and ALT increased on 4 weeks. FERR increased on 12 weeks in both group, but in was higher in NR. Conclusions: Comparison of ALT, AST, GGT between R and NR on therapy with PEG Interferon alfa-2a and Ribavirin revealed normalization of enzymes in R from the 4-8 week of treatment versus of decreased, but non-normalized enzymes level in NR. Serum level of FERR during combination therapy increased more in NR.

Hepatol Int (2010) Background: Accumulating data propose that active injecting drug users (IDU) might not differ from the general population in terms of sustained virological response (SVR) when adherent to therapy for chronic hepatitis C (CHC). However, current guidelines contain restrictive recommendations for therapy in this group of patients. Therefore, we evaluated a cohort of CHC patients regarding the potent influence of active drug using on initial informed consent, compliance and SVR to treatment. Methods: One hundred and sixty-two consecutive patients (of which 62 active IDUs), who had been evaluated during the last 6 years in our center for CHC and proposed to receive treatment with pegylated interferon alpha and ribavirin, were enrolled. Initial informed consent, compliance, and SVR as well as data regarding age, gender, BMI, genotype, viral load, coinfection with HBV/ HDV/HIV, administered interferon alpha (2a or 2b), liver function tests, liver histology, urban residence, ethnicity, and concomitant use of alcohol were collected and analyzed in respect with injecting drug using. Result: Injecting drug using was positively correlated with male gender (P \ 0.001), young age (P \ 0.001), native origin (P = 0.043), and concomitant use of alcohol (P \ 0.001). Comparable initial informed consent (P = 0.836), compliance (P = 0.879), and SVR (P = 0.132) were observed between IDUs and non-IDUs. The results were confirmed using a multiple regression model. Conclusion: Our data further support that active IDUs do not constitute a distinct CHC patient group in terms of initial informed consent, compliance, or SVR. Therefore, injecting drug using should not be a major determinant influencing the decision for treatment of CHC in eligible patients. Background: The pathogenesis of chronic hepatitis C (CHC) is associated to severe oxidative stress and non-selective immunological disturbance that leads to necro-inflammation and fibrosis progression. Previous trials suggested that antioxidant and immunostimulant therapies may have a beneficial effect. Methods: Sixty patients with CHC and non-responder to standard antiviral treatment were randomly assigned to receive Viusid Ò (3 sachets daily, n = 30) or placebo (n = 30) for 24 weeks. End points were improvement in oxidative stress and immunological parameters. Results: Interleukin-1 levels decreased from 3.26 to 2.61 pg/ml (P = 0.04) in patients who received viusid, as compared with a significant increase from 3.09 to 6.62 pg/ml (P = 0.04) in placebo group. Similarly, TNF-a levels significantly increased from 6.9 to 16.2 pg/ml (P \ 0.01) in the patients who received placebo in comparison with almost unchanged levels (6.6-7.1 pg/ml, P = 0.26) in the patients treated with Viusid (P = 0.001). In addition, interleukin-10 levels were markedly increased in patients treated with Viusid (from 2.6 to 8.3 pg/ml, P = 0.04), in contrast with those patients assigned to placebo (from 2.8 to 4.1 pg/ml, P = 0.09) (P = 0.01). Similarly, the administration of viusid markedly increased the mean of IFN-c levels from 1.92 to 2.89 pg/ml (P \ 0.001) in comparison with reduced mean levels from 1.80 to 1.68 pg/ml (P = 0.70) in the placebo group (P \ 0.0001). Statistically significant reductions in serum HNE and MDA levels were observed in both groups of treatment in comparison with pretreatment values, however, those patients who received viusid showed marked reduction as compared with control group (P = 0.001). Viusid administration was well tolerated. Conclusions: Our results indicate that treatment with Viusid leads to a notable improvement in the oxidative stress and immunological parameters in patients with chronic hepatitis C. Introduction: Non-organ-specific autoantibodies (NOSAs) are commonly detected in HCV. The aim was to determine the prevalence of anti-nuclear (ANA), anti-smooth muscle (ASMA) and anti-liver kidney microsomes type 1 (anti-LKM1) antibodies in HCV and assess their impact on the response to antiviral therapy. Methods: One thousand and sixty-two HCV patients were reviewed retrospectively for autoantibodies and response to treatment. All patients received combined antiviral therapy in the form of Pegylated Interferon alpha 2 (a or b) plus Ribavirin (1,000-1,200 mg l day) for 48 weeks.

Results: Non-organ-specific autoantibodies (NOSAs) were observed in 308 patients (29%): ANA in 195 (18.36%), ASMA in 142 (13.37%) and anti-LKMl was the rarest occurring in 16 (1.51%). Concomitant positivity for ANA and ASMA was observed in 57 of these 308 cases (5.37%). The presence of NOSAs was associated with higher aspartate trasaminase (AST), alanine trasaminase (ALT) and c-globulin. In contrast no differences were observed regarding age, gender and viral load. End treatment response in HCV patients with NOSAs was 59.47% while it was 64.57% in patients negative to NOSAs, and this difference was non significant. Discussion/conclusion: ANA was the commonest while anti-LKM1 was the rarest among HCV patients. NOSAs were associated with more necroinflammatory grades. Autoantibodies did not predict any change in response to HCV treatment. (1) Pneumonia reported as an SAE, n (%) 3 (\1) 5 (\1)

Depression reported as an SAE, n (%) 3 (\1) 2 (\1) Neutropenia (.5 cells 9 10 9 /L) 31 (2) 51 (6) Thrombocytopenia (\50 cells 9 10 9 /L) 39 (3) 39 (5) Dose modification of peginterferon alfa-2a (40KD) for safety, n (%)

224 (14) 151 (17) Treatment discontinued for safety, n (%) 43 (3) 23 (3) PP336 Peginterferon Alfa-2a (40 KD Background: HIV-HCV co-infected patients achieve lower SVR rates than HCV mono-infected patients after treatment with peginterferon plus RBV. We evaluated the safety and efficacy of peginterferon alfa-2a (40 KD) (PEGA-SYS Ò ) plus RBV 800 mg/day or 1,000/1,200 mg/day in HIV-HCV genotype 1 (G1) co-infected patients. Methods: HIV-HCV (G1) co-infected adults with stable HIV disease (on stable antiretroviral therapy [ART] or not on ART) and C100 CD4+ cells/mm 3 were randomized (1:2) to receive RBV 800 mg/day or 1,000/1,200 mg/day in combination with peginterferon alfa-2a 180 lg/week for 48 weeks. The primary efficacy outcome was SVR (HCV RNA \20 IU/mL) at week 72. Impact of zidovudine treatment on hematological parameters was examined. Results: In the ITT population (n = 410) mean age was 45 years, 29% were African American, 27% were Hispanic, 54% had an ALT quotient B1.5, and 88% were receiving ART at baseline (15% on zidovudine). Overall SVR rates were similar in the RBV 800 and 1,000/1,200 mg/day dosing arms (19 vs. 22%; OR 1.15, 95% CI 0.68-1.93; p = 0.6119). Non-Hispanic African American patients and Hispanics achieved numerically higher rates of SVR with higher RBV dosing; however, the number of patients in these subgroups was small. High SVR rates were obtained in both arms among patients with HCV RNA \20 IU/mL at week 4 (64-75%) or 12 (63-66%). Anemia was a more common adverse event among patients receiving zidovudine (44%) than among those not receiving zidovudine (27%). Among patients receiving RBV 1,000/1,200 mg/day, RBV dose modifications were more common in those receiving zidovudine (45%) than among those not receiving zidovudine (25%).

Conclusions: Viral clearance at week 4 or 12 predicted SVR, but overall SVR rates did not differ significantly in co-infected patients treated with RBV 800 or Background: Egypt has the highest prevalence of Hepatitis C Virus (HCV) infection in the world, with the eventual development of chronic hepatitis, cirrhosis and hepatocellular carcinoma (HCC). Hepatic repair after injury requires fragmentation of scar, resolving vascular derangements and parenchymal regeneration. Hepatocyte growth factor activator (HGFA) plays a potent role in hepatic repair after injury. Objectives: To investigate tissue expression of HGFA from patients with HCV-related chronic hepatitis and to correlate its expression with the grade of activity, stage of fibrosis and tissue transforming growth factor beta1 (TGF-b1). Methods: Paraffin embedded sections prepared from 31 liver biopsies of patients with HCV infection (41 ± 8 years) and 20 normal subjects, donors for liver transplantation (29 ± 7.5 years) were evaluated for grading and staging of chronic hepatitis and applied for immunostaining with HGFA and TGF-b1 antibodies. Semiquantitative analysis of immunoreactive cells' intensity and percentage for HGFA and TGFb1 were interpreted to calculate the H-score. Results: Liver tissues from patients with HCV infection revealed HGFA around sites of injury and regeneration. HGFA H-score correlated significantly with the grade of activity, stage of fibrosis (p \ 0.01 each) and demonstrated a highly statistical significant direct correlation between TGF-b1 H-score. There was a high statistically significant difference between normal control group and HCV-related chronic hepatitis regarding the HGFA H-score and TGF-b1 H-score. Conclusions: Parallel expression of HGFA and TGF-b1 with the grade of activity and stage of fibrosis in HCV-related chronic hepatitis could make HGFA as an activity and regenerative marker, achieving its role in hepatic repair after injury.

Background: Combination therapy with peginterferon (PEG-IFN)/ribavirin (RBV) represents the current standard of care for chronic hepatitis C. There are two PEG-IFNs, PEG-IFNa-2a and PEG-IFNa-2b. Superiority of either PEG-IFN has not been established. To assess the efficacy and safety of the two PEG-IFNs in combination with RBV in Japanese patients, we have started a randomized, controlled trial. Methods: We have recruited patients from 14 hospitals in Japan. To be eligible for enrollment, patients must be HCV RNA positive for genotype 1b, and C5 logIU/ml (TaqMan PCR method). Patients who had previously received PEG-IFN/RBV combination therapy were excluded. The target sample size was 320. We randomly assigned patients to receive either PEG-IFNa-2a or PEG-IFNa-2b, each in combination with RBV. Patients were treated for 48 weeks (or 72 weeks if HCV RNA was positive at week 12 and negative at week 24) and then followed for an additional 24 weeks for the assessment of sustained virologic response, the primary endpoint of the study. The safety, including effects on QOL using SF-36, was also compared. This study was started in February 2008 and is now in progress.

Result: As of October 2009, more than 200 patients have been enrolled in the study. RVR (HCV RNA negative at week 4), cEVR (HCV RNA negative at week 12), and VR48 (HCV RNA negative at week 48) rates were not significantly different between PEG-IFNa-2a/RBV and PEG-IFNa-2b/RBV (18 and 10% in RVR, 57 and 57% in cEVR and 75 and 65% in VR48, respectively). HCV NS5A ISDR mutation was an independent predictor of RVR. HCV core 70 amino acid mutation was an independent predictor of Non-cEVR and Non-VR48. We will report the update results on March 2010.

(n = 200). Predictive factors of SVR were explored using a recursive partitioning analysis, referred to as classification and regression tree (CART). Results: The number of mutations in ISDR (C2) was the best predictor of SVR followed by younger age (\60), wild type sequence at position 70 of HCV Core, higher level of low-density-lipoprotein-cholesterol (LDL-C) (C120 mg/dl), and absent fibrosis (F0-1). The decision tree model containing these variables identified three groups with variable rates of SVR. Around 30, 10, and 60% of patients were classified into groups of high, intermediate, and low probability of SVR respectively and their rate of SVR was 75, 64, and 32% respectively. The reproducibility of the model was confirmed (r 2 = 0.92, p \ 0.001).

Conclusions: CART analysis highlighted the impact of HCV mutations on pretreatment prediction of SVR to therapy. Decision tree model that include mutations in ISDR and Core region of HCV, age, LDL-C, and fibrosis stage is useful for the prediction of response before therapy, has the potential to support clinical decisions in selecting patients with high probability to achieve SVR or saving those with low probability to achieve SVR to wait for more potent future therapy.

Splenectomy with Subsequent Antiviral Treatment is Beneficial for Hepatitis C, Compensated Cirrhotics with Thrombocytopenia I Shyan Sheen 1 , Chun Yen Lin 1 , Wen Juei Jeng 1 , Chang Wen Huang 1 1 Chang Gung Memorial Hospital, LinKou Medical Center, 5 FuHsin Street, KueiShan, TaoYuan, Taiwan

Background: Combination therapy of pegylated interferon and ribavirin is the current best treatment of chronic hepatitis C. Thrombocytopenia is one of the limiting factors for combination therapy. There are several measures can raise the platelet counts, such as thrombopoitein receptor agonist (Eltrombopag), splenic embolization and splenectomy. AIM: To assess the benefit of splenectomy with subsequent combination treatment for hepatitis C cirrhotics. Method: Splenectomy was suggested to those with hepatitis C compensated cirrhotics, platelet counts \100 9 10 3 /ml and had strong intention for combination treatment. The treatment was pegylated interferon and ribavirin for 24-48 weeks. HCV genotyping and quantification of HCV-RNA were checked. Sustained virological response (SVR) was defined as non-detectable HCV-RNA at 24 weeks after completion of treatment. The response were compared with other non-splenectomy cirrhotics who had combination therapy (186 of genotype 1 and 129 of genotype 2). Results: There were 16 patients (8 males, 8 females) with a median age of 55.2 years, received splenectomy and combination treatment subsequently. Their post-operative courses were uneventful. Eight were genotype 1b and 8 were genotype 2. The median counts of platelet and WBC were 49.1 x 10 3 /ml and 3.42 x 10 3 /ml before splenectomy, 26.3 x 10 3 /ml and 6.87 x 10 3 /ml after splenectomy. The SVR was 62.5% (5 of 8) and 87.5% (7/8) of genotype 1 and 2. The SVR rate of non-splenectomy cirrhotics was 35.5 and 75.9% in genotype 1 and 2. Conclusion: Splenectomy is safe in selected patients of hepatitis C compensated cirrhotics. Subsequent combination treatment can achieve a good SVR. Background: Treatment of chronic hepatitis C with interferon is known to be associated with thyroid dysfunction (TD) in 5-14%. We studied the incidence, types, outcome and risk factors predictive of TD. Methods: A retrospective analysis was performed on patients in centers in Emirates and Egypt who were treated with pegylated interferon a-2a or a-2b + ribavirin, and developed abnormal thyroid function tests. These were compared with treatment-matched controls to identify factors predictive of TD. All patients were followed up for 12 months after completion of antiviral therapy. Results: From a total of 233 patients, 21 cases with TD were identified (9%). Mean age was 42.76 ± 9.71 years; 18 (86%) were females. Hypothyroidism developed in 18 cases (86%): 16 (89%) were females. Hyperthyroidism was observed in 3 (14%) patients; 2 (67%) were females. Mean time for development of hypothyroidism was 24.8 ± 11.76 weeks. In hypothyroid cases, 15 (83%) patients were symptomatic and required L-thyroxin. Hormone replacement therapy was continued in 11 patients (73.3%) beyond follow up period. All hyperthyroid patients were symptomatic and required therapy; two (67%) subsequently developed hypothyroidism and required hormone replacement. Isolated elevation of serum antithyroglobulin antibodies and/or thyroid peroxidase antibodies was noticed in 12 (5%) patients and returned to normal in 10 (83%) patients during follow up. Female gender was found to be the only significant predisposing factor for TD. Family history of thyroid disease, autoimmune diseases or autoantibodies, ALT level, viral load, HCV-genotype, stage of fibrosis, type of interferon, duration of treatment, and treatment outcome were not significantly different between patients with euthyroidism and dysthyroidism.

Conclusions: It is mandatory to monitor thyroid function during and after interferon therapy. The majority of patients continued with hormone replacement therapy for up to 12 months after cessation of interferon therapy Female gender is an independent risk factor for TD. hepatitis C (CHC). There have been more than 20 studies examining the effect of treatment of CHC patients with peginterferon and ribavirin since 2005 in Korea. However, there is no consensus which data is best for representing the treatment of CHC with peginterferon and ribavirin. The aim of this study is to evaluate efficacy and safety of peginterferon plus ribavirin for treatment of the Korean CHC patients under real life condition. Methods: We retrospectively analyze the data of 758 CHC patients which gathered from 14 large hospitals in Gyeonggi-Incheon area, Korea. Results: A total of 758 patients treated with peginterferon alfa plus ribavirin were documented between 2005 and 2008. The mean age of the enrolled patient was 50.2 years (±12.0) and male was 60.8% (446/733). The genotype 1 group was 61.5% (449/733) and genotype 2 was 35.2% (257/733). The high baseline viral load (HVL) rate was 56% (378/672). The overall SVR was 59.6% (per protocol, 78%) regardless of genotype. The SVR was 53.6% in genotype 1 and 71.4% in genotype 2/3. In the multivariate analysis, male gender (P \ 0.01), age \40 (P \ 0.01), EVR (P \ 0.01), genotype 2/3 (P \ 0.01), lower viral load (\600,000 IU/mL, P \ 0.01), drug dose (C80%, P \ 0.01), but not diabetes, obesity (BMI C25) and elevated ALT were significantly associated with SVR. The rate of withdrawal was 35.7% (271/758) during the treatment. Conclusion: Our data suggest that the efficacy and tolerability of peginterferon and ribavirin therapy in Koreans is superior to those of whites in treatment of hepatitis C. It supports the previous studies showing the superiority of therapeutic efficacy in Asian.

Final Results of ACHIEVE-1: Albinterferon Alfa-2b Plus Ribavirin in Treatment-Naïve Patients with Genotype 1 Chronic Hepatitis C S Pianko 1 , Y Lurie 2 , Y Baruch 3 , B Leggett 4 , B Hughes 5 , D Shouval 6 , D Crawford 7 , S Bell 8 , J George 9 , R Safadi 10 , L Mollison 11 , S Roberts 12 , G Feutren 13 , M Subramanian 14 1 Monash Medical Centre, Clayton, Victoria, Australia, 2 Tel-Aviv Sorasky Medical Center, Tel-Aviv, Israel, 3 Rambam-Health Care Campus, Haifa, Israel, 4 Royal Brisbane and Womens Hospital, Brisbane, Queensland, Australia, 5 Background/Aims: ACHIEVE-1-a phase 3, randomized, active-controlled, multicenter study-evaluated the efficacy and safety of albinterferon alfa-2b (albIFN), a genetic fusion polypeptide of albumin and interferon alfa-2b, in patients with genotype 1 chronic hepatitis C (CHC). Methods: In all, 1,323 patients were randomized 1:1:1 to one of three treatment groups: peginterferon alfa-2a (PEG-IFNa-2a) 180 lg qwk, and albIFN 900 and 1,200 lg q2wk for 48 weeks, combined with weight-based oral ribavirin 1,000-1,200 mg/d. Randomization was stratified by baseline HCV-RNA C vs. \800,000 IU/mL, body mass index C vs. \25 kg/m 2 , and Black/ African heritage versus other. Primary endpoint: sustained virologic response (SVR; HCV-RNA \15 IU/mL at wk 72). The sample size provided 90% power to demonstrate noninferiority with a margin of 12%. Results: ACHIEVE-1 demonstrated the noninferiority of albIFN 900 lg (P \ .001) and 1,200 lg (P = .003) q2wk vs. PEG-IFNa-2a 180 lg qwk for SVR. By intention-to-treat analysis, SVR rates were 51.0, 48.2, and 47.3% with PEG-IFNa-2a, and albIFN 900 and 1,200 lg, respectively. Consistent with previous studies, multivariate analysis identified the following predictors of SVR: HCV-RNA \400,000 IU/mL, age \45 years, normal c-glutamyl transferase, high alanine aminotransaminase, fibrosis score 0-2, and non-Black race. Respective serious and severe adverse event (AE) rates, and discontinuation rates due to AEs with PEG-IFNa-2a, and albIFN 900 and 1,200 lg: 10.9, 11.1, and 13.6; 19.5, 20.6, and 24.1; and 4.1, 10.4 , and 10.0%. Severe or serious pulmonary AEs were rare. Mortality rates were 1/441, 0/442, and 2/440 with PEG-IFNa-2a, and albIFN 900 and 1,200 lg, respectively. The rates of hematologic abnormalities were numerically lower with albIFN 900 lg versus PEG-IFNa-2a.

Conclusions: Treatment with albIFN 900 lg q2wk demonstrated comparable efficacy to PEG-IFNa-2a 180 lg qwk in patients with genotype 1 CHC, with a similar safety profile.

Final Results of ACHIEVE-2/3: Albinterferon Alfa-2b Plus Ribavirin in Treatment-Naïve Patients with Genotype 2/3 Chronic Hepatitis C W.L. Chuang 1 , C.M. Lee 2 , S. Thongsawat 3 , M. Cho 4 , C.Y. Peng 5 , G. Shanmuganathan 6 , J. George 7 , I.S. Sheen 8 , R. Safadi 9 , S.R. Shah 10 , T. Tanwandee 11 , V. Mahachai 12 , S. Pianko 13 , H.T. Kuo 14 , G. Feutren 15 , G. Subramanian 16 Background/Aims: ACHIEVE-2/3-a phase 3, randomized, active-controlled, multicenter study-evaluated the efficacy and safety of albinterferon alfa-2b (albIFN), a genetic fusion polypeptide of albumin and interferon alfa-2b, in patients with genotype 2/3 chronic hepatitis C (CHC). Methods: In all, 932 patients were randomized 1:1:1 to one of three treatment groups: peginterferon alfa-2a (PEG-IFNa-2a) 180 lg qwk, and albIFN 900 and 1,200 lg q2wk for 24 weeks, combined with oral ribavirin 800 mg/d. Randomization was stratified by baseline HCV-RNA C versus \800,000 IU/mL and genotype 2 versus 3. Primary endpoint: sustained virologic response (SVR; HCV-RNA \15 IU/mL at wk 48). The sample size provided 90% power to demonstrate noninferiority (margin of 12%). Results: ACHIEVE-2/3 demonstrated the SVR noninferiority of albIFN 900 lg (P = 0.009) and 1,200 lg (P = 0.006) q2wk versus PEG-IFNa-2a 180 lg qwk. By intention-to-treat analysis, SVR rates were 84.8, 79.82, and 80.0% with PEG-IFNa-2a, and albIFN 900 and 1,200 lg, respectively. Consistent with previous studies, multivariate analysis identified HCV-RNA \400,000 IU/mL, age \45, BMI \30, genotype 2, normal c-glutamyl transferase, high alanine aminotransaminase, no steatosis, fibrosis score 0-2, and Asian region (for PEG-IFNa-2a only) as SVR predictors. In the Asian region (n = 271), SVR rates were 95. 5, 79.8, and albIFN 900 and 1, 200 lg, respectively, versus 80.5, 79.8, and 79 .3% in all other regions (n = 662). The incidence of serious (7-8%) or severe (13-16%) AEs, or death (0.3-0.6%) was similar across groups. Discontinuation rates due to AEs were 3.6, 4.8, and 5.5% with PEG-IFNa-2a, and albIFN 900 and 1,200 lg, respectively. Severe/serious pulmonary infections and interstitial lung disease were rare, with similar rates across groups. Conclusions: ACHIEVE-2/3 demonstrated that albIFN 900 lg q2wk was comparable in efficacy to PEG-IFNa-2a 180 lg qwk in patients with genotype 2/3 CHC, with a similar safety profile. Objectives: Interferon (IFN) monothreapy for 8-24 weeks lead to higher sustained virological response (SVR; defined as undetectable HCV RNA level in serum at 24 week after the end of IFN treatment) in patients with hepatitis C virus (HCV) serotype 2 and low viral load (serum HCV RNA level was less than 10 5 IU/ml by Amplicor HCV monitor assay). However, duration of treatment is still in dispute. This study evaluated the efficacy of short term treatment with IFN-beta (Feron, Daiichi-Sankyou, Japan) in patients with HCV serotype 2 and low viral load. Methods: A total of 18 patients with HCV serotype 2 and low viral load were treated with 600 million units IFN-beta by intravenously administration every day for 8 weeks. Result: Seventeen of 18 patients were negative for HCV RNA at 2 week of treatment. All patients were negative for HCV RNA at 4 week of treatment and at the end of treatment. The overall SVR rate was 83.3% (15/18). Serum ALT levels were declined by IFN-beta treatment. Univariate logistic regression analysis showed that disappearance of HCV RNA at 2 week of treatment, mild fibrosis staging and no complications such as diabetes mellitus and hypertension were important factors for SVR by IFN-beta treatment (each P \ 0.05). Treatment discontinuation for adverse events was not observed. Conclusion: This study shows that short term treatment with IFN-beta is effective and tolerable treatment in patients with HCV serotype 2 and low viral load. Patients and Methods: Study medication was administered for 72 weeks (48 wks. in association with PegIFN-ribavirin plus 24 wks in follow-up) in all patients, irrespective their lipid profile. Statin use 1 year prior study admission, significant concomitant medication, as well as any other metabolic, renal and myopathic disease were among exclusion criteria. Patients were randomly assigned either to placebo (group P, n = 105) or to fluvastatin (F) group (n = 104). Viral load (VL) was assessed at inclusion, 12 and 72 weeks and the percentage decrease of VL (PDVL) between inclusion and the 12-weeks determination was calculated in each case. We also evaluated metabolic syndrome (MS) using the NCEP-ATP III criteria whilst BMI and ALT variation was monitored every 3 months. Results: EVR was 68.9% (144/209) and SVR 59.5% (118/209) while mean VL at inclusion was 3.81 ± 0.22 9 10 6 IU/mL, decreasing at 0.27 ± 0.02 9 10 6 IU/mL at 12 weeks in overall patients (p \ 0.05). No statistically significant differences regarding ALT variation were noticed among the two groups. Background: Although cytotoxic T-lymphocyte (CTL) responses are thought to be important components of the immune system in the control of acute hepatitis C virus (HCV) infection, the key contributions of CTL cell response in chronic HCV infection and the correlation between HCV specific CTLs and viral clearance in antiviral treated patients remain elusive. To evaluate this question, we examined the CTL responses in 54 patients with chronic HCV infection on baseline and monitored their rapid virological response (RVR) after undergoing 4-week interferon therapy. Methods: PBMCs were used in ex vivo IFN-c ELISpot assays with 171 overlapping peptides spanning the proteome of HCV-NS3, NS4 and NS5 to detect antigen-specific T cell responses. Plasmas were collected before and after undergoing 4-week interferon therapy to detect HCV viral loads and evaluate RVR in these patients. Result: In 54 patients, 18 (33.33%) of patients could mount a HCV-specific immune response with recognising at least one gene product at a magnitude above 50 SFU/10^6PBMC after background subtraction. Amongst the patients with HCV-specific immune response, the most frequently recognised peptides were in the NS3 region of the proteome (72.22%, 13/18), followed by NS5 (50%, 9/18) and NS4 (27.78%, 5/18) . NS3 was also the gene product eliciting the highest mean magnitude of ELISpot responses (380 SFU/10^6PBMCs), followed by NS4 (282 SFU/10^6PBMCs), and NS5 (273 SFU/10^6PBMCs). In addition, it was apparent the HCV viral load on baseline in patients with HCVspecific immune response were lower than that in patients without immune response, although the difference did not reach statistical significance (p = 0.083). There was no correlation between the magnitude of response and RVR in patients with antiviral therapy. Conclusion: Antigen-specific T cell responses can be identified in chronic hepatitis C patients and HCV-NS3 seems contribute more than NS4 and NS5 in the control of disease progression. The effect of T cell responses to RVR is not clear and further study is necessary to clarify the correlation between the HCV specific CTLs and viral clearance. This work was supported from National S&T Major Project for Infectious Diseases Control (2008ZX10002-013).

A Randomized Trial of Interferon Alpha, Ribavirin Plus Interferon Gamma or Amantadine for HCV Genotype 3 Non-responders and Relapsers Zaigham Abbas 1 , Sajjad Raza 1 , Saeed Hamid 1 , Wasim Jafri 1 1 The Aga Khan University Hospital, Karachi, Pakistan

Background: The aim of the present study was to evaluate the efficacy and safety of triple combination regimens comprising of interferon alfa-2b and ribavirin plus either interferon gamma or amantadine in HCV genotype 3 infected patients who have not previously responded to interferon alpha (standard or pegylated) in combination of ribavirin. Methods: Patients was randomized to receive interferon alpha 2b 3MU t.i.w, ribavirin 800-1,200 mg per day with either interferon gamma 2 MU t.i.w or amantadine 100 mg twice daily. Treatment was continued for 48 weeks in patients showing complete or partial (2 log reduction) early virological response (EVR) at 12 weeks and negative PCR at 24 weeks. Results: Total enrollments were 44. Mean age 44.1 years (28-60); 25 were previously non-responders out of them 12 were in the gamma arm. Nineteen were relapsers, out of them 10 received Gamma interferon. F3 or F4 fibrosis was seen in 14 (34%) and 9 (23%) were diabetic. By intention-to-treat analysis, the EVR for interferon gamma arm was 50% (11 out of 22) and for amantadine arm 36.36% (8 out of 22) (p = 0.272). The end of treatment responses were 45% (10/22) & 27% (6/22) for interferon gamma and amantadine arms respectively (p = 0.174). Overall sustained virological response (SVR) with triple regimens was seen in 34% (15/44), SVR was 45% (10/22) in the gamma arm and 23% (5/22) in the amantadine arm. In the subgroup analysis, this figure was 60% (6/10) and 44% (4/9) for relapsers, and 33% (4/12) and 8% (1/13) for non-responders in both arms respectively. Treatment was well tolerated in both arms. Conclusions: About one third of genotype 3 patients who had not previously responded well to the interferon and ribavirin responded to the triple regimens. However addition of interferon gamma was a better option with an acceptable safety profile. Its combination with pegylated interferon and ribavirin needs further evaluation in a larger clinical trial. Background/Aims: Recently the liver-specific miR-122 was reported to stimulate HCV replication and translation in cell culture. It has still remained unclear whether IFN can change microRNA expression as a new anti-HCV mechanism. In addition, the mechanism how the addition of the broad-spectrum antiviral agent ribavirin greatly improved IFN responses to anti-HCV therapy clinically is unknown. Here, we report the relationship between IFN-b treatment in combination with ribavirin and microRNA expression in vitro. 

Female, n (%) 887 (47) 138 (49) 160 (33) 49 (40) Race, n (%); Caucasian 1699 (90) 223 (78) 427 (88) 64 (53) Black 69 (4) 14 (5) 2 (\1) 30 (25) Asian 31 (2) 4 (1) 16 (3) 3 (2) Hispanic 35 (2) 20 (7) 10 (2) 5 (4) Other 53 (3) 23 (8) 32 (7) 19 ( (35) pEVR, n (%) 401 (21) 14 (5) 34 (7) 16 (13) Ncn-nesponse/ missing, n (%)

394 (21) 14 (5) 48 (10) Background/Aims: Interferon and ribavirin are effective treatment for chronic hepatitis C (CHC) and will remain the backbone of newer specifically targeted antiviral treatment. Contraindications to interferon and ribavirin are well recognized, and previous data has indicated the consequent suboptimal treatment uptake. We aim to evaluate the treatment rate of CHC patients referred to a tertiary referral centre in Hong Kong, and to examine the reasons for nontreatment.

Methods: A retrospective review of all referred CHC patients to the outpatient clinic was conducted. Treatment uptake rate was evaluated and patients' sociodemographic, biochemical and histological data were examined to identify reasons for treatment decision.

Results: A total of 252 CHC patients were referred and assessed for antiviral therapy over a 5-year period. 57.1% were male. The mean age was 53.6 ± 12.9 years. Antiviral therapy was commenced on 133 ( Taiwan Background: To evaluate the prognostic value of Fibrotest for predicting liver fibrosis and the efficacy of peginterferon treatment on fibrosis formation in patients with CHB. Methods: This study was an open-label, multi-center, non-interventional, post-marketing observational study. All patients were followed-up for 72 weeks. Fibrotest is a non-invasive blood test that combines the quantitative results of five serum biochemical markers (alpha-2macroglobulin, apolioprotein A1, haptoglobulin, gamma-glutamyltransferase and total bilirubin) with the patient's age and gender in a patented artificial intelligence algorithm. Results: The mean age of the patients (55 male, 24 female, 18 missing data) were 35 ± 12 (17-69) years and the mean mean duration of hepatitis B disease was 4.4 ± 3.4(1-13) years. Most of the patients were under peginterferon alfa2a therapy (65 of 97 patients, 32 missing data) and peginterferon alfa2a dosage was 180 mcg. At 72nd week of the study (24 weeks after the end of therapy), the ratio of HbsAg and HbeAg positivity decreased to 34.0 and 8.2% from the baseline values of 77.1 and 34.0% respectively. Anti-Hbe positivity increased from 53.2% at baseline to 72.7% at 72nd week of the stud. Mean HBV DNA level was 254.6 x 10 6 ± 999.5 x 10 6 (median 5.5 x 10 6 ) copies at baseline and 34.5 x 10 6 ± 142.8 x 10 6 (median 2,800) copies at 72nd week and the difference was statistically significant (P = 0.008). Among patients whose HBV DNA level at 72nd week were recorded, 48% of patients were negative for HBV DNA (\10 4 copies/ml). Mean ALT level of 829 ± 6,141 U/L at baseline decreased to 63 ± 60 U/L at 72nd week (P = 0.001). The percentage of patients without fibrosis (F0), regarding Metavir scoring system, increased from 20.8% at baseline to 57.1% at 72nd week (P = 0.036). The median histological activity index (HAI) scores were 8 at baseline and 7 at 72nd week, respectively (P = 0.893). The percentage of patients without fibrosis (F0), according to Fibrotest scoring, increased from 21.8% at baseline to 25.7% at 72nd week. The median HAI scores were8at baseline and 7 at 72nd week, respectively (P = 0.893). Conclusions: Peginterferon treatment had a positive effect on serological and biochemical parameters and decreased fibrosis in patients with chronic HBV infection. Purpose: Sustained virologic response (SVR) in Hepatitis C is *50% globally in all genotypes. Alinia (nitazoxanide [NTZ] ) is an interferon inducer that may increase SVR in combination with PegIFN and RBV. Very rapid virologic response (VRVR) (undetectable HCV RNA 14 days after initiation of combination therapy) may predict higher SVR rates. We assessed NTZ with highdose RBV and PegIFN alfa-2a in achieving VRVR, RVR, early virologic response (EVR), end treatment response(ETR) and sustained viral response(SVR) in naïve patients with chronic hepatitis C genotype 1 (GT1). Methods: A prospective, open-label, pilot study of naïve chronic hepatitis C GT1 patients. All received NTZ 500 mg bid -first 2 weeks; RBV 800 mg(AM) + 600 mg(night) added to NTZ for next 2 weeks; subcutaneous PegIFN alfa-2a 180 mcg weekly added to RBV and NTZ at the same dose for next 12 weeks. Those with EVR continued PegIFN alfa-2a and RBV for the next 36 weeks at the same dose. VRVR, RVR, EVR, ETR, and SVR were assessed. Results: 23 Patients: Caucasian (n = 9, 39%), Hispanic (n = 4, 17%), African-American (n = 5, 21%) and Asian (n = 5, 21%). Mean BMI of 27.9; mean HCV RNA-600,000 IU/ml; fibrosis score-F2 (57%). Growth factors were used to minimize dose reductions. Adverse effects (AE): skin rash (39%); neutropenia (47.8%); thrombocytopenia (43.4%); anemia (52%); Alopecia Background/aim: Patients with chronic hepatitis C (CHC) are at higher risk of developing abnormal glycometabolism, and this risk may increase among hepatitis C virus (HCV) patients not responding to an antiviral therapy. Our study was designed to investigate the prevalence of insulin resistance (IR) and type 2 diabetes mellitus (T2DM) and the association between insulin resistance and rapid virologic response (RVR) in CHC patients. Methods: One hundred and twenty-five CHC and 63 chronic hepatitis B(CHB) patients were evaluated. IR (Homeostasis Model for Assessment of Insulin Resistance) was assessed by using the following equation: HOMA-IR = fasting insulin (lU/mL) x fasting glucose (mmo/L)/22.5 and was defined as HOMA-IR [ 3. RVR was defined as testing negative for hepatitis C virus-RNA after 4 weeks of antiviral therapy. HCV genotyping was performed (restriction fragment method) and serum HCV RNA quantified (PCR) for all patients before antiviral treatment. Hepatitis C virus-RNA was measured after 4 weeks of treatment. Fasting levels of insulin and glucose in the serum were measured in all patients on the first day of treatment. Results: IR was present in 17.8% of the 90 nondiabetic and noncirrhotic CHC patients type 2 diabetes mellitus was diagnosised in 12.6% of 130 CHC patients without liver cirrhosis. Forty-one CHC patients took peginterferon alpha-2a and ribavirin, and 26 of them reached RVR. In the multivariate analysis the independent predictors of RVR was: HOMA-IR more than 3.00 [P = 0.014, odds ratio 8.8 (95%CI:1.5-50.5)]. IR was less frequent in CHB than in matched CHC (3.2 vs. 15.9% respectively, P \ 0.05). Conclusions: The incidence of abnormalities in glucose metabolism in CHC was significantly higher than that in CHB patients. Insulin resistance is an independent predictor of RVR, the homeostatic assessment of insulin resistance in the baseline serum should be evaluated to predict the rapid virologic response in CHC subjects. Background & Aim: Diabetes mellitus (DM) represents a poor prognostic factor for antiviral therapy response in chronic hepatitis C patients. The aim was to study the diabetic factors which can influence treatment response in patients with chronic hepatitis C and DM. Methods: We studied 100 patients with DM who underwent antiviral therapy with pegylated interferon and ribavirin for chronic hepatitis C in Institute of Gastroenterology and Hepatology Iasi, between 1 January 2006 and 31 December 2008. In our country almost all patients (more than 98%) are infected with type 1b virus C genotype. We analyzed the following parameters: type of DM, glycated hemoglobin (Hb A1c)-determined at the beginning of antiviral therapy, lasting of DM, macro and microangiopathic complications. The ''cut off'' value of Hb A1c was considered 7%. A sustained response was defined as the absence of detectable hepatitis C virus RNA 6 months after treatment was stopped. Results: There were 58 female and 42 men, aged between 21 and 60 years (mean age 38.7 years). 40 patients had Type 1 DM and 60 Type 2 DM. The global rate of response was 42%. In Type 1 DM were 45% responders, and in Type 2-40%. Abnormal Hb A1c was found in 21.42% responders (R) and in 70.68% non-responders (NR). Macroangiopathic complications were found in 33.33% R and 44.82% NR. Microangiopathic complications were found in 28.57% R and 31.03% NR. There were 13 patients (30.95%) with more than 10 years of DM in R and 24 patients (41.37%) in NR. HbA1c was the only statistical significant factor (p \ 0.05) between the R vs. NR.

Conclusion: HbA1c is an important predictive factor to antiviral therapy response in diabetic patients with chronic hepatitis C. There are more nonresponders patients with Type 2 DM comparative with Type 1, probably due to insulin-resistance. All patients with DM and chronic hepatitis C need a good glycemic control before starting antiviral therapy. Over the recent years, hepatitis C has emerged as a dreadful health challenge at national level in Pakistan and is spreading at alarming pace. Based on a year long open study on patients with hepatitis C, the results of real-time quantitative PCR assays indicate an herbal formulation to be highly effective in containing and cure of HCV infection. The formulation, HC-786, comprises of 11 traditional herbs soaked overnight and administered orally twice a day as boiled aqueous extract. Though the response ranged from mild to marked, almost all patients observing certain dietary precautions responded positively. Interestingly, the decline in HCV RNA counts was equally good in HCV patients already suffering from diabetes who normal respond poorly to the interferon therapy. No side-effect was reported unlike interferon based therapies. The effect of medication was more pronounced in HBV infected patients with comparatively low viral loads compared with HCV infected patients. The treatment ameliorate the general health indicators such as skin color, increased appetite, better digestion and weight gain etc in some non-HCV patients with acute cirrhosis, fibrosis and/or hepatocellular carcinoma, indicating hepatoprotective nature of this medication. Currently studies on antioxidant activity, hepatoprotective nature, cytotoxicity and chemical composition of the extract are underway. Background: To investigate the effect of double filtration plasmapheresis (DFPP) as the treatment chronic genotype 1b hepatitis C with high viral load, DFPP was performed followed by combined Peg-interferon and ribavirin therapy. Methods: In 7 patients with chronic hepatitis C, DFPP was performed 5 times on days 1, 2, 8, 9 and 15 . The drug administration of Peg-interferon and Result: The HCV RNA level was undetectable in 5 patients after 24 weeks, and this decrease was continued to a sustained virological response (SVR). Conclusion: DFPP that performed with interferon and ribavirin therapy together in chronic hepatitis C patients may contribute to a sustained virological response. A long-term observation is expected whether this treatment decreases the cancer incidence in patients with chronic hepatitis C With genotype 1b.

Material and Method: Patients with CHC treated by interferon and/or ribavirin between 1995 and 2009 were retrospectively analyzed. There were 196 patients with a SVR defined as negative HCVRNA (by PCR-lower limit of sensitivity 50 iu/mL) at sixth month post-treatment. Patients with SVR were followed-up at every 6 months with liver transaminases and HCVRNA. Results: There were 89 male (45%) and 107 female (55%) patients with SVR. Pre-treatment mean age was 46.1 ± 11.1 (17-73) years. Eight of the patients were cirrhotic. Mean fibrosis stage and histological activity index (HAI) were 1.55 ± 0.94 and 7.22 ± 2.41, respectively. There were 16 patients with chronic renal failure and 17 patients with diabetes mellitus. The number of patients treated by pegylated interferon + ribavirin, standard interferon + ribavirin and standard interferon monotherapy were 139, 21 and 36, respectively. In 138 patients HCV genotype results were available. The distribution of genotypes in these patients was as follows: 77.5% genotype 1b, 5.1% genotype 1a, 9.4% genotype 2, 7.2% genotype 3 and 0.7% genotype 4a. Median follow-up duration after SVR was 33.5 months (6-112). Only 2 patients (1.02%) developed a relapse at 18th and 48th months. HCVRNA titers of the patients with relapse were as low as 6,361 iu/mL and 438 iu/mL, and repeated HCVRNA tests in both patients resulted negative. Aminotransferase levels were normal during the follow-up in 96.4% of the total patients. None of the patients develop any complication related to liver disease during the follow-up. Conclusion: Almost all patients with SVR (98.98%) did not develop virological relapse during median 3 years of long term follow-up. According to our data, SVR is durable actually and patients with relapse demonstrates only a low grade and intermittent viremia. Background: BMS-650032 is a potent and selective HCV NS3 protease inhibitor with in vitro pico-molar potency against the NS3/4A protease. BMS-650032 was safe and well tolerated at single doses up to 1,200 mg and up to 600 mg Q12 h for 14 days in healthy subjects. Methods: Two randomized, placebo-controlled studies evaluated single and multiple ascending doses of BMS-650032 in subjects chronically infected with HCV genotype 1. In study 002 (SAD), BMS-650032 was administered to treatment-naïve and experienced subjects at doses of 10, 50, 200 and 600 mg (6 subjects per dose; active placebo). Study 004 (MAD) included naïve subjects only and evaluated 3 days of BMS-650032 at doses of 200, 400, and 600 mg Q12 h (5 subjects per dose; active placebo). Results: There were no deaths or discontinuations due to AEs and all AEs were mild to moderate. SAD: BMS-650032 exposures increased in a dose-related manner and were similar or higher to that observed in healthy subjects. Tmax was *2 to 4 h, mean terminal half-life was *15 to 20 h, and mean oral clearance ranged from 302 to 491 L/h. Actively treated subjects experienced a mean decline in HCV RNA of *0. Background: PEG-IFN-lambda (PEG-IFN-k) is a unique interferon that has fewer flu-like symptoms and hematologic adverse effects than are typically observed with alpha IFNs, likely due to more focused expression of the IFN-k receptor. Here we report the 4-week antiviral activity, safety and pharmacokinetic (PK) results from the Phase 1b open-label dose-ranging study in IFN-a relapse and treatment-naïve genotype-1 HCV patients. Methods: Part 1 of the study evaluated escalating doses of single-agent PEG-IFN-k administered either Q2 W or QW for 4 weeks to HCV + IFN-a relapse patients. Parts 2 and 3 of the study evaluated different dose levels of PEG-IFNk administered weekly in combination with daily oral ribavirin (RBV) for 4 weeks to IFN-a relapse patients (Part 2), or who were naïve to treatment for HCV (Part 3). Doses of PEG-IFN-k were body weight-based and ranged from 0.5 to 3.0 lg/kg. Assessments include adverse events (AEs), laboratory values, and changes in HCV RNA. Results: 56 patients (mean age 51 years, 69% male, mean baseline HCV RNA 6.43 log 10 IU/mL) were randomized to treatment. Dose-dependent reductions in antiviral activity at 4 weeks were observed, with 83 and 86% achieving [2-log decrease in HCV RNA at the Peg-IFN-k 1.5 mg/kg + RBV dose in IFN-a relapse and treatment-naïve patients, respectively. Treatment was generally well-tolerated with minimal flu-like symptoms and no clinically meaningful changes observed in neutrophils, platelet counts or hemoglobin levels (apart from RBV associated reductions). 6/56 (11%) of patients experienced reversible increased in ALT/AST with or without increases in bilirubin (majority at the 3.0 mg/kg single agent dose, Part 1). Conclusions: PEG-IFN-k was associated with robust antiviral activity at all dose levels tested in both IFN-a relapse and treatment-naïve patients. Treatment was generally safe with minimal flu-like symptoms or heamatologic effects observed. These favorable results support the initiation of Phase 2 studies in treatment-naïve patients. Background & Aim: HAV is spread via the fecal-oral route, and is more prevalent in low socioeconomic areas in which a lack of adequate sanitation and poor hygienic practices facilitate spread of the infection. Korean epidemiology of HAV hepatitis is changing, due to the improvement of living conditions. As a result, fewer children are infected, leading to a larger population of adults who lack protective antibodies against HAV. We observe the epidemic changes of HAV and assess the need for HAV vaccination. Background: Hepatitis E is a worldwide and important public health problem, especially in areas with poor sanitation. The evidences of zoonotic transmission of HEV have been mostly reported in HEV non-endemic areas. However, the issue regarding existence of animal reservoirs in HEV-endemic regions remains unresolved. In this study, we try to learn the potential HEV animal reservoirs and the current status of HEV infection among animals and human in disease-endemic areas of Xinjiang China. Methods: 12 different animal species and 296 human sera were detected for anti-HEV with in-house enzyme immunoassay, and partial HEV RNA was amplified with reverse transcription-nest polymerase chain reaction (RT-nPCR). Results: All these distinct animal species except jerboa and hoptoad were positive for anti-HEV, and the prevalence of anti-HEV among human from Southern Xinjiang and Northern Xinjiang is 46.42 and 18.33%, respectively. HEV RNA was only amplified from pigs and a sporadic hepatitis E case in human of Han nationality, and swine HEV subtypes from Southern Xinjiang and Northern Xinjiang belonged to 4a and 4n, respectively. The human HEV strain (CHN-XJ-HE29) belonged to subtype 4a, sharing 100% nucleotide identity with swine HEV strain (CHN-XJ-SW50), both of which were collected from the same district. Conclusion: HEV genotypes among swine in Xinjiang belonged to genotype 4 and there were at least 3 prevailing HEV subtypes (4a, 4d and 4n). The potential animal reservoirs responsible for HEV outbreaks or epidemics have not been found. Evidences of swine to human transmission of sporadic hepatitis E in HEV-endemic regions were provided.

Profile of Pediatric Viral Hepatitis in Kathmandu Sudhamshu K.C. 1 Methods: complete data were collected for 1,104 cases of hepatitis E from 1998 to 2,003 years, in which 28 cases of protracted duration of more than 6 months are still unhealed and followed up for 6-10 years on a regular basis liver function, blood biochemistry, all indicators of virology, B-ultrasonography or CT, some patients had liver biopsy. Results: 15 cases after 6 months there are different degrees of symptoms; 9 cases of splenomegaly occurred, 5 patients with different degrees of hepatomegaly; 17cases in 6 months after different degrees of liver function abnormalities; after six months the anti-HEV IgM was still positive in 5 cases, the anti-HEV IgG was positive in 25 cases; after 6 months, the HEV RNA was positive in 5 cases; of liver biopsy for 12 cases; other viruses were detected in 4 cases;1 case suffering from hypertension; one case from coronary heart disease; one case from pulmonary emphysema, 1 case from asthma; 1 case of lung cancer; one case of renal transplant recipients. Discussion: male of chronic hepatitis E cases is more common, not a high degree of inflammatory activity, fibrosis was not evident. Follow-up observation of 6-10 years does not appear to cirrhosis, liver cancer cases. It is significantly different with the hepatitis B virus, hepatitis C virus. Most of 28 patients is the older, more than 18 cases of patients aged 50 years and over. With older patients the ability to remove the virus may decline, which may also contribute to more than six months duration or more important reasons. In addition, the observation also found a few cases, because of the merger of other viruses cause illness or chronic persistent unhealed. Besides, one of the reasons hepatitis E to chronic trend is associated with chronic disease or long-term using of immune agents. Background: Previous studies reported that high prevalence of antibodies to HEV have been detected in many rodents. However, only few studies reported the experimental infection of mice with HEV and the results were quite different. To confirm whether or not rats are susceptible to HEV infection, in this study SD rats were infected with swine HEV. Methods: 22 healthy SD rats, weight 180-200 g were involved in this study, including 2 rats as control. The viral suspension of subtype 4a and 4d of swine HEV were prepared from pigs with HEV infection. Nine rats were administrated with 600 ll 10% suspension of swine stool containing infectious HEV, the other rats were inoculated 1,200 ll of the same suspension, via caudal vein and gastric perfusion respectively. Two control rats were inoculated intravenously with 1,200 ll of PBS. The stool and sera specimens, which were collected pre-administration and post-administration respectively, were detected for HEV RNA by the method of RT-nPCR, and anti-HEV antibody by ELISA. The infected rat liver was examined by both light microscopy and electron microscopy for detection of pathological changes. Results: Three rats with caudal vein inoculation of subtype 4a and 1 rat with subtype 4d were observed slight elevation of serum ALT 2 weeks after administration. The liver tissues of the 4 rats were found mild inflammatory Hepatol Int (2010) 4:94-345 207 changes. The transient positive of serum anti-HEV antibody was occurred to only one rat, which was inoculated with 1200ul stool suspension via caudal vein. HEV RNA was not detected in all stool and serum samples. Conclusions: Only 3 SD rats in this study showed slight inflammatory changes after infected with swine HEV 4a and 4b but no HEV RNA and anti-HEV antibody were detected. These results suggested that the SD rats in this study were not susceptible to the genotype 4a and 4d of swine HEV.

Clinical in delayed diagnosis group. Initial laboratory findings were not different between both groups but platelet counts, ALT, and bilirubin were lower in delayed diagnosis group. Hospital stay was longer (7.4 vs. 6.0 days) in delayed diagnosis group but time to prothrombin time normalization was not different. Mean interval to IgM anti-HAV positive was 3.8 days, and titer of following positive IgM anti-HAV was not different from that of initially diagnosed patients. Conclusions: Our data suggest that not a few symptomatic AVH-A patients demonstrated IgM anti-HAV negative initially, probably due to earlier visit with more severe prodromal symptom than those of initial diagnosis group. Methods: Male C57BL6 N mice (n = 30) were randomly divided into three experimental groups and three control groups (5 mice for each group), and given MCD diet and the control methionine choline sufficient diet for 4 weeks, 8 weeks and 12 weeks, then were killed to obtain liver tissue specimens. Formalin-fixed, paraffin-embedded livers were prepared for immunohistochemistry. Then progenitors markers (CK19, CD56) and Hedgehog pathway factors (Ihh, Gli2) were studied. After excluding the major bile duct in each portal tract (PT) from consideration, cell staining positively for CK19, CD56, Ihh, or Gli2 in 10 PT/side were counted at 9400 magnification. Results were expressed as mean ± SD, statistical analysis was performed with SPSS 13.0 statistical package, using the Student's t test, P \ 0.05 were considered statistically significant. Results: In MCD diet mice and control group mice for 4 weeks, 8 weeks, 12 weeks, CK19-positive cell counts for average each PT were 21.0 ± 7.8 vs. 8.7 ± 4.0, 22.8 ± 6.9 vs. 7.8 ± 4.3, 21.8 ± 7.6 vs. 8.6 ± 5.0; CD56 for 25.0 ± 7.8 vs. 14.8 ± 6.5, 25.7 ± 8.1 vs. 13 .2 ± 6.8, 24.6 ± 8.9 vs. 13.3 ± 6.9; Ihh for 18.2 ± 5.9 vs. 10.3 ± 4.7, 17.3 ± 6.9 vs. 9. 3 ± 5.9, 19.3 ± 6.9 vs. 9.4 ± 4.9; Gli2 for 34.7 ± 11.3 vs. 16.7 ± 6.2, 37.8 ± 12.8 vs.

15.3 ± 6.1, 38.9 ± 13.4 vs. 17.2 ± 6.0. (All P \ 0.05).

Conclusions: MCD diet induced liver injury stimulated the proliferation of liver progenitor cells, which may be related to activation of Hedgehog pathway. Background: Nonalcoholic fatty liver disease (NAFLD) is a progressive disease that reflects metabolic syndrome. The Chinese herbal medicine red yeast rice is a fermented product of rice with Monascus purpureus. It contains several constituents including monacolin K, which is a inhibitor of HMG-CoA reductase. Its consumption has been found to decrease cholesterol and triglyceride concentrations. However, the efficacy of constituents other than monacolin K has not been examined. We examined the efficacy of red yeast rice using a newly developed mouse model of NAFLD associated with obesity, hyperlipidemia, and type 2 diabetes. Methods: Two types of red yeast rice grown under different culture conditions were used. 1P-DU contained only 2 mg/100 g of monacolin K, whereas 3P-D1 contained 131 mg/100 g. Twenty-seven neonatal male MSG mice were divided into three groups. Group C (control) was fed standard food; Group red yeast rice-control (RYR-C) was fed standard food with 1% 1P-DU; and Group red yeast rice-monacolin K (RYR-M) was fed standard food with 1% 3P-D1. Mice were examined when they were 12-24 weeks old. Results: Groups fed red yeast rice showed significantly low serum insulin and leptin levels and pathological liver damage. They also exhibited lower body weight, as well as lower total cholesterol and LDL-cholesterol levels compared with the control group. Group RYR-C showed significantly higher serum adiponectin level than Group RYR-M and the control group. Conclusion: Red yeast rice was found to be efficacious against obesity, insulin resistance, hyperlipidemia, and NAFLD in MSG mice irrespective of the dose of monacolin K. GABA and various peptides produced by fermentation were candidates for active constituents in this rice. We conclude that red yeast rice could be a useful supplementary food for metabolic patients including those with NAFLD. Background: Spirulina is a cyanobacterium characterized by high protein content (60-70%). It also contains carotenoids, vitamin E, phycocyanin, and chlorophyll. Many reports have recently suggested antioxidative and hepatoprotective effects of Spirulina because of carotenoids and vitamin E, but the role of phycocyanin has not been well examined. Nonalcoholic fatty liver disease (NAFLD), the most common chronic liver disease in the West, is closely associated with obesity and metabolic syndrome. Nonalcoholic steatohepatitis (NASH) is the progressive form of NAFLD and involves oxidative stress. We examined the efficacy of Spirulina and phycocyanin using a newly developed mouse model of NAFLD associated with obesity, hyperlipidemia, and type 2 diabetes. Methods: Twenty-five neonatal male MSG mice were pretreated with a subcutaneous injection of monosodium glutamate. They were divided into three groups (N = 7-9). Group C (control) was fed standard food; Group P was fed standard food with 0.35% phycocyanin; and Group S was fed standard food with 5% Spirulina. Mice were examined when they were 12-24 weeks old. Results: Groups P and S showed significantly lower visceral fat macrophage aggregation level than Group C. Group S showed significant increase in serum leptin, insulin, and ALT, as well as total cholesterol levels in addition to improvement in liver disease, and body weight compared with Group C. As expected, Group P showed intermediate improvements compared with Groups C and S. A twofold increase in the serum adiponectin level was found only in Group P compared with that in Groups C and S. Conclusion: Spirulina was found to be efficacious against obesity, insulin resistance, hyperlipidemia, and NAFLD in MSG mice. Phycocyanin increased serum adiponectin levels. We conclude that Spirulina could be a useful supplementary food for metabolic patients including those with NAFLD. Methods: Six-to eight-week-old male SD rats were randomly divided into two groups and were treated with 0.2% diethylnitrosamine (DEN 10 mg/kg) interruptedly by intragastric administration and fed either normal diet or high fat diet (HFD) for 16 weeks. The levels of serum ALT, AST, alkaline phosphatase (ALP), c-glutamyltransferase (GGT), total bilirubin (TBIL), albumin

Hepatol Int (2010) 4:94-345 209 (ALB), glucose (GLU), total cholesterol(TC), triglyeride (TG), high density lipoprotein-cholesterol (HDL-C) and low density lipoprotein-cholesterol (LDL-C) were determined by automatic biochemistry analyzer. Serum adiponectin and tumor necrosis factor-a (TNF-a) levels were measured by ELISA methods. Hepatic histology was examined by hematoxylin and eosin (HE) stain.

Results: At the end of the experiment, seven rats were enrolled in either group. All rats were confirmed to develop liver tumor, while the HFD + DEN group had frank steatohepatitis with more fat accumulation, infiltration of inflammatory cells, fibrosis and cancerous nodules. In addition, compared with the DEN group, liver/body weight ratio was significantly increased in the HFD + DEN group (9.58 ± 1.10 vs. 13.08 ± 3.19, P \ 0.05). There were no differences between the two groups in the levels of serum ALT, AST, ALP, GGT and GLU (P [ 0.05), while a significantly higher levels of TBIL (72.20 ± 36.14 vs. 33.71 ± 27.08 lmol/L, P \ 0.05), TC (6.60 ± 1.44 vs.

2.87 ± 0.59 mmol/L, P \ 0.01),TG ( Background: Preparing rat model of non-alcoholic fatty liver disease by fatrich diet to observe the expression and the role of LXR-a in rat nonalcoholic fatty liver disease. Methods: Thirty-six SD rats were randomized into basic diet-control group and high-fat diet group. Each of the two groups was subdivided into 3 subgroups (4, 8, 12 weeks) . Changes in animal weight, liver exponent and the level of TG and TC in serum and in liver were observed dynamically. Meanwhile, the expression of hepatocyte LXR-aand SREBP-1c was assayed by Reverse transcript-polymerase chain reaction at 4, 8, 12 weeks. The level of steatosis was observed under light microscope after haematoxylon-eosin (HE) staening. Result: Compared with control group, body weight, liver exponent, TG and Tc in serum and in liver were increased dynamically in fat-rich diet. Rosiglitazone of low dose and high dose was given in different time to group L 1 , group high-fat diet group (P \ 0.05); Compared with control group, the mRNA of LXR-a and SREBP-1c were obviously increased dynamically in model group (P \ 0.05). Conclusion: The increase of LXR-a and SREBP-1c in liver may be concerned with energy disorder and the increase of LXR-a and SREBP-1c in liver may be concerned with energy disorder and closely associated with the activity of inflammation and the severity of the liver damage in NAFLD rats. Result: Feeding rats with HF diet established a model of NASH, with varying degrees of hepatic steatosis and hepatic inflammation. SP600125 treatment substantially decreased the incidence of insulin resistance, reduced lipotoxicity, inhibited oxidative stress and alleviated hepatocelluar injury. Conclusion: SP600125 has the potential to remarkably attenuate steatosis and inflammation and may be the novel therapeutic drug against NASH. Aims: Serum contains a complex of proteolytically derived peptides which may provide a diagnostic clue for diseases. The aim of the study was to compare the different serum peptidome patterns between twins with and without non-alcoholic fatty liver disease (NAFLD). Methods: Peptides between 800 Da and 3,500 Da were captured and concentrated using C18 reversed-phase column, and followed by a MALDI-TOF mass spectrometric readout. The peptidomics patterns of 63 cases with NAFLD were compared with their twin healthy controls in Qingdao. The sequences of peptides associated with NAFLD were further identified by MALDI-TOF-TOF. Results: Compared with healthy controls, eleven peaks (861. 1, 877.07, 904.5, 1206.57, 1350.64, 1518.7, 1690.9, 1777.94, 2931.29, 3190.4, 3261.4) were identified to be up-regulated in NAFLD group, and 7 peaks ( Background: Obese hepatic steatosis is commonly associated with central fat accumulation and hyperleptinemia; however, the connection between nonobese hepatic steatosis and leptin is unclear.We sought to uncover the connection using an animal model. Methods: Double transgenic mice, in which hepatic over-expression of HCV core is regulated by tetracycline transactivator (tTA), were given permissive chow following 1 month of a doxycycline-rich diet since their births. Nonobese hepatic steatosis was exhibited in the 2-month-old double transgenic mice but diminished with time. The levels of leptin and adiponectin, and various metabolic features, were subsequently assessed in 2-, 3-, and 4-month-old double transgenic mice (i.e., HCV core-tTA) and single transgenic mice (i.e., tTA). The total fat mass and body fat distribution of the mice were evaluated by dualenergy X-ray absorptiometry and magnetic resonance imaging. Microarray analyses and quantitative real-time polymerase chain reaction were conducted using RNA from the visceral fat of paired double transgenic mice and single transgenic mice. Leptin was administered intraperitoneally to the 2-month-old double transgenic mice. Results: After adjusting for fat mass, the protein and RNA expression of leptin was lower in the 2-month-old double transgenic mice than in the single transgenic mice, and the DTM had hypoadiponectinemia; nevertheless, these differences diminished with time. Leptin treatment significantly ameliorated hepatic steatosis in the 2-month-old double transgenic mice. Conclusions: HCV core-induced nonobese hepatic steatosis is associated with down-regulation of the leptin gene in visceral fat and subsequent hypoadipocytokinemia; however, these effects may be ameliorated by leptin treatment.

La-Mei Xu 1 , Dan-Li Sun 1 , Yu-Shu Zhang 1 1 Department of Gastroenterology, Nanjing First Hospital Affiliated to Nanjing Medical University, 68, Changle Road, Nanjing, China

Background: Tight junctions (TJs) are the primary junctions between intestinal epithelial cells. TJs served as the maintenance of epithelial cell polarity and the rate-limiting barrier to passive movement of hydrophilic solutes across intestinal epithelia. It is known that intestinal epithelial TJ protein is decreased in hepatic cirrhosis, acute liver failure and so on. Yet, there is no research on the changes of intestinal epithelial TJ protein expression in non-alcoholic fatty liver disease (NAFLD). Methods: Thirty male SD rats were divided into two groups. The control group were fed with normal diets while the model group with high fat diets. All the animals were sacrificed after 12 weeks HE staining of hepatic tissue in model group showed that the NAFLD was established. Small intestinal epithelial tight junctions were observed by electron microscope in all groups, and the distribution and expression of tight junction protein occludin were detected by immunohistochemistry Aims: To investigate the effects of Rosiglitazone or combined with Reduced Glutathione on insulin resistance related indicators in rats with nonalcoholic fatty liver disease (NAFLD). Methods: NAFLD rats were produced. Rats were divided into four groups: rosiglitazone group, group of rosiglitazone combined with reduced glutathione, model group and normal group. Insulin levels in serum and pancreas were detected by immunohistochemisty and ELISA, and the biochemical indicators were detected at the same time. Then, all indicators above were compared. Results: The levels of albumin, ALT, total cholesterol in model group increased obviously compared with normal group (P \ 0.01), but the concentrations of triglyceride and fasting blood glucose in model group decreased markedly (P \ 0.01). Rosiglitazone could relatively reverse the levels of biochemical indicators compared with that of model group, but there were no significant differences. The levels of insulin in serum and pancreatic tissue in model group were obviously higher than normal group (P \ 0.01). Rosiglitazone could reduce the insulin expression in serum and pancreatic tissue compared with model group. The trends of insulin resistance index in above four groups were similar. Conclusion: NAFLD rats occurs significant abnormality with liver function and obvious insulin resistance. Rosiglitazone or combined with Reduced Glutathione can reverse the levels of biochemical indicators, decrease the insulin resistance index, but there is no difference between the two treatment groups. Background/Aims: Nonalcoholic fatty liver disease (NAFLD) is a progressive intractable disease that reflects metabolic syndrome. It is a growing health concern because of its rapidly increasing prevalence. Nonalcoholic steatohepatitis (NASH) is a critical type of NAFLD that progress to cirrhosis and hepatocellular carcinoma (HCC). However it's not easy to taking over the disease process of NASH. Furthermore, a mouse model of NASH is required for the same. Therefore, to examine the pathogenesis of NASH and initiate an effective treatment, we examined the detailed disease process in the TSOD mice, which has been established as an inbred strain with spontaneous development of diabetes mellitus, in the present study. Methods: The TSOD mice show spontaneous obesity, hyperglycemia, and hyperinsulinemia only in males without special treatments. We followed up the histopathological characteristics of various organs from 56 male TSOD mice aged 4-17 months. Results: TSOD mice frequently suffered from NAFLD or NASH in these 4 months, as well as from crown-like macrophage aggregation and perivascular and pericapsular lymphoid aggregation in visceral fat. At 6 months, all TSOD mice suffered from NASH, showing zone-3 steatosis, hepatocellular ballooning, Mallory bodies, and mild lobular inflammation. At this time, aggregated macrophages in visceral fat expressed various cytokines. The lipid peroxidation molecule 4-HNE was aberrantly expressed in some visceral adipocytes.

Hemosiderin-laden macrophages were observed in the spleen of all TSOD mice. Background: Hepatic oxidative stress plays a key role in development of nonalcoholic steatohepatitis (NASH). Heme oxygenase-1 (HO-1), an antioxidant defense enzyme, has been shown to protect against oxidant-induced tissue injury. This study aimed to elucidate the role of HO-1 modulation on NASH in mice.

Methods: C57BL/6 J mice were fed with a methionine-choline deficient (MCD) diet for 4 and 8 weeks to induce steatohepatitis and hepatic fibrosis. HO-1 inducer hemin, HO-1 inhibitor ZnPP-IX and/or an adenovirus vector that expressed HO-1 (Ad-HO-1) were intraperitoneal injected thrice per week, respectively. Results: MCD feeding mice showed increasing serum ALT and AST, progressive hepatic injury including hepatic steatosis, inflammatory infiltration and fibrosis, which were attenuated by administration of hemin and/or Ad-HO-1. This effect was associated with increase of HO-1 protein and activity, downregulation of pro-inflammatory cytokines tumor necrosis factor (TNFa), interleukin 6 (IL-6) and lipogenic gene sterol regulatory element binding protein-1c (SREBP-1c), suppressed expression of fibrotic genes transforming growth factor-b1 (TGF-b1), matrix metalloproteinases 2 (MMP-2) and tissue inhibitor of metalloproteinase 1 (TIMP1), up-regulated expression of peroxisome proliferator-activated receptor alpha (PPARa). A contrary effect was observed in mice injected with ZnPP-IX. Conclusions: The present study provided a morphological and molecular biological evidence for the protective role of HO-1 in ameliorating oxidative stress, inflammation and fibrosis in experimental nutritional steatohepatitis and fibrosis. Induction of HO-1 might provide a new approach for treatment of steatohepatitis and fibrosis.

The Background: It is documented that Fas and Fas ligand expressions played a key role in hapatocyte apoptosis which is associated with nutritional steatohepatitis in mice. This study aimed to explore if Fas mutation ameliorates or accelerates hepatocyte apoptosis, hepatic steatosis and inflammatory infiltration in nutritional steatohepatitis in mice. Methods: Mice homozygous for the lymphoproliferation spontaneous mutation (C57BL/6 J-Fas lpr ) and C57BL/6 J mice were fed a methionine-choline deficient (MCD) diet for three weeks to induce non-alcoholic steatohepatitis (NASH). The role of Fas gene mutation on NASH was assessed by comparing severity of hepatic steatosis and inflammation in the liver sections, the mRNA and protein expression of hepatic inflammatory and fibrogenetic related factors, adiponectin, tumor necrosis factor (TNF-a), interleukin 6 (IL-6), proliferating cell nuclear antigen (PCNA), transforming growth factor-b1 (TGFb1) and matrix metalloproteinases 2 (MMP2). Results: At weeks 3, MCD-diet induced hepatic steatosis, inflammatory infiltratory in both of wide type and Fas lpr mice. Especially, severe hepatic injury was observed in Fas lpr mice compared with wild type mice, which was associated with suppressed anti-inflammatory cytokine adiponectin, up-regulated pro-inflammation cytokines TNF-a and IL-6, enhanced cell proliferation factor PCNA and fibrogenetic growth factors TGFb1 and MMP2. Conclusions: Fas lpr mice developed more severe hepatic injury including hepatic steatosis and inflammatory infiltration induced by MCD-diet compared with wild type mice, which were related to excessive release of cell proliferation, inflammatory and fibrogenetic factors. Methods: NAFLD animal model was duplicated with high fat diet. 32 male Wister rats were randomly divided into normal control group (group N), PD98059 and common diet group (group P), high-fat model group (group M) and PD98059 and high-fat diet group (group PM) with 8 for each group. At the end of 4, 6, and 8 weeks, the rats were given a tail vein injection of either PD98059 (0.3 mg/ kg, group P and group PM) or the relative solvent (group N and group M).After ten weeks, all the rats were sacrificed. The serum ALT, AST, TG, TC and the hepatic TG, TC were measured. The pathological changes in rat liver were observed, the expression of p-ERK1 /2 and SREBP-1 in liver was detected by immunohistochemistry.

Results: Compared with group N, the group M developed over 30% steatosis, with inflammation cell infiltration and necrosis, the serum ALT, AST, TC, TG, the level of liver tissue homogenates TC, TG and the expression of p-ERK1 /2 and SREBP-1 in liver markedly increased (P \ 0.01, P \ 0.05). Compared with group M, the expression of p-ERK1 /2 and SREBP-1(P \ 0.01), the serum ALT, TC, TG and the level of liver tissue homogenates TG, TC (P \ 0.01, P \ 0.05) decreased in group PM, while the histopathological change in liver of group PM ameliorated. Conclusion: MEK/ERK signal pathway play an important role on the expression of SREBP-1in NAFLD induced by high-fat diet in rats.

Top 10% Posters Background: Fatty liver disease is frequently associated with elements of metabolic syndrome. The aim of this study was to determine whether the degree of bright liver, indicating an increase in the discrepancy in echo amplitude between the liver and kidney parenchyma on ultrasonography, was associated with elements of metabolic syndrome in a healthy Japanese population. Methods: We conducted a cross-sectional study based on abdominal ultrasound examinations of 198 men and 247 women recruited from participants in a health screening program. We measured the ratio of the average optical density of the liver to that of the right kidney (L-K index) on echographic images with commercial image analysis software. We also performed biochemical tests of liver and metabolic functioning. We then assessed the association between the L-K index and factors related to metabolic syndrome with a multivariate regression model. Results: The mean L-K index was 1.21 ± 0.33 in men and 1.26 ± 0.31 in women (P \ 0.05). The mean index was 1.15 ± 0.20 in the 138 subjects whose liver function was inferred to be normal because they had a normal body mass index (BMI), normal high-density lipoprotein cholesterol and triglyceride levels, normal blood pressure, and no hepatitis C or B virus infection, and who did not report excess alcohol consumption. An increase in the number of risk factors was strongly associated with an increase in the L-K index (P \ 0.01 for trend). The multivariate analysis demonstrated that the factors independently related to an increase in the L-K index were female gender, serum adiponectin and alanine aminotransferase levels, the homeostasis assessment model ratio, and BMI. Conclusions: These results suggest that obesity and insulin resistance are fundamentally associated with an increase in liver steatosis, as determined by a liver-kidney contrast on ultrasonography.

The is frequently used in traditional Japanese and Chinese medicine to treat several symptoms and manifests anti-inflammatory and scavenging effects. Non alcoholic fatty liver disease (NAFLD) is a common manifestation of the metabolic syndrome and has the potential to evolve to liver cirrhosis through chronic inflammation and steatohepatisis (NASH). We have recently reported the KBG significant effectiveness on liver injury in a NASH animal model which prompted us to prescribe KBG (TJ-25) to.

Methods: We performed a retrospective study and reviewed the charts of outpatients who were prescribed KBG for 8-12 weeks due to non liver-related symptoms (n = 11) over the past year to evaluate the clinical outcome. In 6 of these cases biochemical and ultrasound signs of NAFLD were observed. Result: KBG led to a significant reduction in liver injury tests and blood cholesterol but had no effects on body weight in all NAFLD cases. Further, liver tests and lipid profiles returned to baseline values when KBG treatment was stopped.

Conclusion: Based on data on a small number of subjects, we suggest that the use of KBG is a safe complementary treatment in patients with NAFLD. While it is unlikely that Kampo formulas may substitute the current nutritional approaches to the metabolic syndrome, future studies should address the possibility of an additive effect, possibly through anti-inflammatory mechanisms.

Top 10% Posters has been reported that if the ratio is 2 or more, then the risk of arteriosclerosis increases and the rate of occurrence of related ischemic cerebro-cardiovascular events also rises. In this study, we investigated the arteriosclerosis index in cases of NAFLD. Methods: The subjects comprised 46 cases diagnosed to have NAFLD at our department or at a related institution between January 2006 and November 2008 (29 cases of SS (Simple steatosis), 17 cases of NASH). We comparatively examined the LDL/HDL ratios of the 2 groups, divided the cases into 2 groups according to whether the LDL/HDL ratio was less than 2 (Group I) or 2 or more (Group II), and then comparatively examined the serological characteristics.

Results: (1) The LDL/HDL ratios were 2.49 ± 0.79 and 2.74 ± 0.99 in the SS group and the NASH group, respectively, and although no significant difference was observed, the ratio did tend to be high in the NASH group. (2) Group I included 14 cases and Group II included 32 cases.

(3) There were significant differences between the group in the serological characteristics (c-GTP, T-Cho, TG, HDL, LDL, high-molecular-weight adiponectin (Ad)) and complications (BMI, visceral fat area Objectives: To investigate the influence of polygenetic polymorphisms, which play a role in the pathogenesis of metabolic syndrome, on the susceptibility to non-alcoholic fatty liver disease (NAFLD) in Chinese people. Methods: The subjects were selected from an epidemiologic survey in Guangdong province of southern China. In each polymorphism study, 50-117 subjects who met the diagnostic criteria of NAFLD and had typical clinical and ultrasonographic findings were placed into the case group. Using a nested case-control design, the same numbers of matched people without NAFLD were included as controls. Single nucleotide polymorphisms (SNPs) at nine positions in seven candidate genes were tested. These SNPs were found to be associated with the pathogenesis of metabolic syndrome. Genetic analyses were performed using genomic DNA extracted from peripheral blood leukocytes. PCR-RFLP was applied to detect SNPs. Background/aims: Serum uric acid (SUA) is significantly elevated in nonalcoholic fatty liver disease (NAFLD) patients. However, whether this elevation is a causer, a bystander, or a consequence of NAFLD remains under debate. This study aimed to investigate the association between SUA and development of NAFLD. Methods: A total of 6,974 initially NAFLD-free subjects were followed up for 2 years, to study the association between SUA level and risk of incident NAFLD. Animal experiments were also performed to study the potential benefit of hypouricemic therapy on NAFLD. Results: Overall, 625 (8.96%) subjects developed NAFLD over 2 years of follow-up. The cumulative incidence of NAFLD increased with progressively higher baseline SUA levels. Cox proportional hazards regression analyses showed that elevated SUA levels were associated with increased risk of incident NAFLD; the age-and gender-adjusted hazard ratio (95% CI) for the subjects in quintile 2, 3, 4 and 5 versus quintile 1 was 1. Background/Aims: Serum uric acid level has been suggested to be associated with factors that contribute to the metabolic syndrome and coronary heart disease (CHD). The aim of this study was to explore the relationship between hepatic injury and hyperuricemia in patients with fatty liver disease.

Methods: A total of 119 patients with fatty liver were divided into nonhyperuricemia (n = 64) and hyperuricemia group (n = 55). Both groups were analysised by demographic characteristics and biochemical parameters. In these subjects, 21 patients with biopsy-proven non-alcoholic fatty liver disease (NAFLD) were divided into non-hyperuricemia (n = 11) and hyperuricemia group (n = 10), meanwhile, histological characteristics were also compared. Logistic tests were used to evaluate the risk factors of hyperuricemia in fatty liver patients.

Results: In all of the subjects, compare to those with non-hyperuricemia, patients with hyperuricemia had higher triglycerides, total cholesterol and apolipoprotein B level significantly (totally P \ 0.05). In the patients with biopsy-proven NAFLD, those with hyperuricemia had more significant steatosis (P \ 0.05), but inflammation and fibrosis stage were comparable (P [ 0.05). Logistic regression analysis revealed that alcohol consumption, male and hypercholesterolemia were independent risk factors of hyperuricemia while hyperglycemia was protective factor. Conclusions: Fatty liver disease is correlated positively to hyperuricemia. Alcohol consumption, male and hypercholesterolemia are related to hyperuricemia in patients with fatty liver disease. Hyperuricemia is association with hepatic steatosis, but not inflammation and fibrosis. Aim: To explore changes in cardiac diastolic function in patients with nonalcoholic fatty liver disease (NAFLD) by color Doppler Echocardiography and analyze the relationship between liver function and cardiac diastolic function in these patients. Methods: One hundred and twenty NAFLD patients treated at our hospital from July 2005 to June 2008 and equal number of matched healthy controls were included in this study. All participants were subjected to routine liver function tests. Meanwhile, B-mode ultrasound diagnosis of fatty liver and color Doppler echocardiographic evaluation of cardiac diastolic function were carried out. Diastolic cardiac dysfunction (DCD) was defined when the patient had an E/A ratio less than 1.

Results: The incidence of diastolic cardiac dysfunction in patients with NA-FLD was significantly higher than that in normal controls (26.7 vs. 5.0%, P \ 0.01). As the fatty liver disease progressed, the E/A ratio decreased gradually (P \ 0.01). In NAFLD patients with DCD, the levels of ALT, AST and GGT and patient age were significantly higher than those in patients without DCD (P \ 0.01). Significant differences were noted in TBIL and DBIL levels between normal controls and patients with moderate or severe NAFLD (both P \ 0.01). A significant difference was also noted in IBIL level between normal controls and NAFLD patients (P \ 0.01). Logistic analysis showed that serum GGT level was an independent risk factor for the E/A ratio in NAFLD patients (OR = 1.050, P \ 0.01). Conclusion: DCD is commonly seen in patients with NAFLD. The levels of ALT, AST and GGT significantly increase in NAFLD patients with DCD. Elevated GGT may be a novel risk factor for DCD or coronary artery disease.

The Background: Identifying risk factors for the development of non-alcoholic fatty liver disease (NAFLD) is essential for its early screening and prevention. Serum ferritin (SF) level has been suggested to be associated with NAFLD. SF is a sensitive indicator of body iron stores, iron is regarded as a putative element that interacts with oxygen radicals. The aim of our meta-analysis was to summarize the association between SF level and risk of NAFLD derived from previously published control studies and to examine the effect of study characteristics on this association. Methods: Searches through large literature databases, including PubMed, CNKI, among others, the quality of case studies was evaluated independently by two reviewers. RevMan4.2.8 and Stata10.0 software was used. And metaregression analyses were performed to investigate the effect of these characteristics on the association between SF level and NASH. Results: Thirteen quality control studies were identified. In the comparison between individuals of nonalcoholic fatty liver (NAFL) and those of normal control, the pooled standardized mean difference of ferrion concentrations was 0. Background/Aims: To investigate the prevalence of fatty liver disease (FL) and its relevant factors in an adult physical examination population of Qingdao, China, and to provide a data for early intervention and improvement in prognosis of fatty liver disease in Qingdao. Method: The adult physical examination people in our hospital were invited to participate in the survey. Questionnaire and ultrasonographic survey was used. The body mass index (BMI), blood pressure (BP), blood lipids (TC, TG, LDL-C and HDL-C), blood glucose (FBG) and liver function (ALT), hepatitis B virus and hepatitis C virus serum markers were detected. Statistical analysis was performed with the SPSS program 13.0. Results: The study included 5,760 subjects (3,677 men) with a mean age of 39 years. Fatty liver was found in 1,867 (32.41%) subjects. The incidence of fatty liver disease in man was 39.38% (1,448/3,677), and it was higher than that of women (16.37%, 341/2,083), P \ 0.001. The prevalence rate in the 50-60 age group peaked at 43.95%. After gender stratification, the incidence of fatty liver disease in male who were in the 50-60 age group was 36.10% and female who were in the 60-70 age group was 42.29%. The prevalences of alcoholic, suspected alcoholic and nonalcoholic FL were determined to be 10.12, 0.59, 20.38%, respectively. Multiple regression analyses showed that only eight factors (triglycerides, body mass index, fasting blood glucose, cholesterol, waist circumference, diastolic blood pressure, age, vegetarian) were closely related to FL. Conclusion: FL in Qingdao is highly prevalent and mainly related to multiple metabolic disorders. The prevalence rate of fatty liver disease and alcoholic liver disease which related to age, sex, obesity, hypertension, hyperlipidemia, high blood sugar and bad eating habits is high in Qingdao city. Obesity, diabetes mellitus, hyperlipidemia and hypertension may be independent risk factors of fatty liver. To reduce weight and maintain normal blood glucose, lipid and blood pressure are important ways to preventing the fatty liver disease. Background and Purpose: The degree of hepatic fibrosis in cases of NASH is an important factor. Therefore, clarifying the risk factors or serological characteristics of the development of fibrosis in cases of NASH is therefore considered to provide clinically useful information. In this study, we focused on fibrosis in cases of NASH in order to clarify the risk factors through a clinical and pathological investigation. Methods: The subjects comprised 87 cases diagnosed to have NASH after performing a liver biopsy at our department or at a related institution between January 1997 and November 2008. We categorized the cases by pathology. We differentiated the cases into 4 groups comprising stage 1, stage 2, stage 3, and stage 4 of fibrosis and investigated the characteristics of those groups. Results: (1) The degree of hepatic fibrosis was stage 1 in 33 cases, stage 2 in 32 cases, stage 3 in 16 cases, and stage 4 in 6 cases. (2) The AST/ALT ratio tended to be high as the fibrosis developed.

(3) As the fibrosis developed, the proportion of female cases increased and a significant difference was observed. (4) There were no significant differences between the stages in the frequency of complications (obesity, diabetes, hyperlipemia, and hypertension). Conclusion: In cases of NASH, being a female was one of the risk factors for hepatic fibrosis. All of the cases of NASH involved at least one of the following complications, and NASH should therefore be suspected in cases presenting with more than one of these complications. Background: It has been reported that patients with hypopituitarism are at risk of obesity, impaired glucose tolerance, hyperlipidemia and nonalcoholic fatty liver disease (NAFLD). The aim of this study was to investigate the clinical features and liver histology of pediatric NAFLD associated with hypopituitarism. Methods: We reviewed the clinical data and liver histology of eleven children diagnosed as having hypopituitarism and NAFLD. Results: The mean age at the time of diagnosis of hypopituitarism was 10.4 ± 3.2 years and the mean age at the time of diagnosis of NAFLD was 13.1 ± 2.7 years. Craniopharyngioma was the most common cause of pituitary dysfunction. At the time of diagnosis of NAFLD, nine patients (82%) had a body mass index greater than the 85th percentile, four patients (36%) had elevated fasting blood glucose levels and seven (64%) had hypertriglyceridemia. The mean height SD score was significantly lower at the time of diagnosis of NAFLD than at the time of diagnosis of hypopituitarism. Of the six patients biopsied, one was cirrhotic, two had nonalcoholic steatohepatitis (NASH) with bridging fibrosis, two had NASH with mild portal fibrosis and one had simple steatosis. Conclusion: Children with hypopituitarism are at risk of short stature, obesity, hyperlipidemia and NAFLD. Early diagnosis of NAFLD is important in children with hypopituitarism because advanced fibrosis is common. Background and Aim: Non-alcoholic fatty liver disease (NAFLD) is present in up to one-third of the general population and in the majority of patients with metabolic risk factors such as obesity, dyslipidemia and diabetes. We aimed to determine the prevalence of NAFLD among elderly population in Chang Chun city of China, and to characterize the clinical risk factors for NAFLD. Methods: The present cross-sectional study was performed with data obtained from 424 subjects who participated physical examination in our hospital, whose averge age was older than 60 years of age (399 men and 25 women). Each participant underwent an abdominal ultrasound, biochemical tests, an anthropometric evaluation and a standard questionnaire. NAFLD was diagnosed with the criteria from fatty liver and alcoholic liver disease study group of Chinese liver disease association. We evaluated the clinical and biochemical characteristics of NAFLD, and particularly its association with diabetes, dyslipidemia, cardiovascular disease and anthropometric parameters. Results: Of the 424 subjects, 94 were diagnosed NAFLD. Overall, the prevalences of NAFLD were 22.17%. In NAFLD group, prevalence of diseases related to the cardiometabolic risk factors were higher than control group (P \ 0.05), obesity was 80.2%, coronary heart disease 52.0%, diabetes 30.85%, hypertension 56.38%, hyperlipidemia 70.21%, physical exercise ([2 h/day) 51.2%. In normal group, obesity 36.59%, coronary heart disease 36.6%, diabetes 45.6%, hypertension 52.53%, hyperlipidemia 46.7%, physical exercise 72.12%. There was no further comparison between gender because of female cases was little. Risk factors associated with NAFLD included obesity, hypertension, diabetes, BMI and hyperlipidemia, physical exercise, but there was no significant difference in age status. Conclusions: NAFLD is prevalent in the elderly and higher than other general population, and the prevalence of some diseases related to the cardio-metabolic factors also significant higher than the general population. The high prevalence rates noted in the study may herald increased risk factors of metabolic abnormal disease in this population. Objective: To determine the incidence and risk factors of non-alcoholic fatty liver disease (NAFLD) in apparently healthy Chinese employees. Methods: Subjects free of previous liver injury, alcohol consumption of more than 140 g/week, and hepatitis B infection were identified from employees of Shanghai BaoSteel Group who underwent voluntary medical checkups at a 2-year interval. Results: The analyzed population consisted of 5,402 non-drinking subjects (4,633 men) of age 36.5 ± 9.3 years (range, 18-65 years), who had normal livers at baseline. Among them 327 subjects (6.05%) developed fatty liver 2 years later. Those who developed NAFLD showed advanced age (especially in females), elevated body mass index (BMI), higher levels of serum triglyceride and total cholesterol, higher prevalence rates of obesity, hyperlipidemia, hypertension, hyperglyceridemia at baseline, and more weight gain and increase of serum triglycerides during the 2-year period. The incidence of NAFLD increased significantly with the changes of BMI at baseline, from 1.4% in subjects with normal weight, 6.4% in overweight, 16.8% in obese patients to 24.5% in severe obesity (v 2 = 389.01,P = 0.000 in trend analysis). Logistic regression analysis demonstrated that a significant interaction occurred between age, BMI and serum triglyceride at baseline, and subtle gain of BMI and triglyceride during follow-up. Conclusions: There is a high incidence of NAFLD in a Chinese workplace. Obesity and related metabolic disorders at baseline, and more weight gain and increased serum triglyceride during follow-up are risk factors of development of NAFLD.

Course of Non-alcoholic Introduction: Nonalcoholic fatty liver disease (NAFLD) is the most common explanation for the elevated serum alanine aminotransferase (ALT) in blood donors and sonographic surveys of the general population demonstrate fatty liver in about 25% of adults in the United States. Oats Supplementation can ameliorates Alcohol-Induced Liver Damage in Rats by Preventing Gut Leakiness. Chronic oxidative stress and endotoxinemia are thought to be important in the genesis of alcohol-related liver damage. Similarities in the histologic features and natural histories of alcoholic liver disease and NAFLD suggest that common mechanisms may be involved in the pathogenesis of these two disorders. Thus it is possible that oats Supplementation can decreases gut leakiness and therefore decreases endotoxinemia and oxidative stress to liver in nonalcoholic steatohepatitis (NASH) also. Aims and Methods: The mean of aminotransferases before trial had no significant deference in two groups (65 and 67.4 for AST and 83.5 and 84.7 for ALT in Vitamin E and Vitamin E + oat groups respectively). Also groups were match for age, sex and percentage of obese, diabetic and hyperlipidemic patients. The mean of AST and ALT had significant decrease in both groups (P \ 0.001).but in Vitamin E + oat group this decrease was more significant than in Vitamin E group.(47.6 and 33.4 for AST and 48.5 and 36.9 for ALT in Vitamin E and Vitamin E + oat groups respectively) (P \ 0.01).

Results: This study is a randomized single blind clinical trial. Sixty nine patients with nonalcoholic steatohepatitis were enrolled in this study. They were divided in two groups, in first group (33 patients) vitamin E 400 i.u and in second group (36 patients) vitamin E 400 i.u plus oat 150 g was prescribed daily for 2 months. An identical nutritional and exercise program was planned for both groups. Treatment was assessed by biochemical response at the end of trial (decrease in serum AST and ALT levels). Conclusion: Oat can use in nonalcoholic steatohepatitis patients as an assisting treatment. Background: Excess trans-fatty acids (TFA) consumption is known to be a risk factor for obesity and coronary heart disease accompanied by systemic inflammation. We aimed to clarify the impact of TFA consumption on the liver pathophysiology in terms of the inductivity of hepatic inflammation and steatosis. Methods: Female C57/BL6 mice fed either low fat (LF) or high fat diet (HF), and each diet were made of either TFA-free control oil (LF-C or HF-C) or TFA-rich (28.5%/fat) partially hydrogenated vegetable oil (LF-T or HF-T) for 24 weeks. We evaluated the liver and body weight, serological features, liver lipid content, histology and mRNA expression of the whole liver for lipid metabolism, fibrosis and cytokines.

Results: Although body/liver weight and ALT do not differ significantly among LF, but increased in HF-T 1.3, 2.2 and 4.8-folds than HF-C, respectively. Hepatic triglyceride content does not differ among LF, but increased 3.1-folds in HF-T than HF-C. Despite the similar liver weight and hepatic TG content, lipogenic mRNA expression of LF-T, peroxisome proliferator-activated receptor-c2 (PPARc2) and sterol regulatory element binding protein 1 (SREBP1), increased 1.9 and 3.8folds than LF-C, respectively, And ipogenic mRNA expression of HF-T, PPARc2 and SREBP1 increased 3.9 and 2.4-folds than HF-C, respectively. Collagen 1a1 mRNA expression as a fibrotic marker increased 2.9-folds in HF-T than HF-C, but does not differ among LF. Tumor necrosis factor nd inducible nitric oxide synthetase were upregulated in HF than LF, but not differ due to oil types. Conclusion: TFA consumption in low fat diet had little impact on liver, but excessive consumption of TFA rich diet caused steatohepatitis with marked lipid depositions and degeneration of hepatocytes, accompanied by lipogenic mRNA expression in liver. The quality and quantity of lipid in diet can be a key factor of the pathogenesis of NASH. Background/Aim: Alcoholic hepatitis has been long recognized as a potentially life-threatening complication of alcohol use, whereas natural history of this disease is not clarified so far. The aim of this study was to investigate early mortality of placebo-treated alcoholic hepatitis patients. Methods: The databases PubMed, EMBASE and Cochrane Library were searched for randomized placebo-controlled trials in alcoholic hepatitis. The mortality data of placebo-treated patients were extracted and analyzed. Results: Nineteen trials with a total of 661 placebo-treated patients were included. The overall mortality rate was 34.19% with median observation time of 160 days (range, 21-720 days). Hepatic failure, gastrointestinal bleeding and infection were the top three causes of death, which accounted for 55.47, 21.17 and 7.30% of overall deaths, respectively. One-month mortality data was reported in 10 trials with a total of 324 placebo-treated patients, and the pooled mortality rate was 20.37%. With regard to one-month mortality rate, moderate to severe patients tended to have higher rate than general patients (22.69 vs. 10.93%, P \ 0.05), whereas no significant difference was observed between the patients from North America or Europe (22.43 vs. 18.45%), neither any difference was found between the studies published before or after 1990 (18.18 vs. 21.88%). Conclusions: Alcoholic hepatitis is a serious liver disease with high early mortality rate, which especially holds true among moderate to severe patients. Medications should be taken to prevent hepatic failure, gastrointestinal bleeding and infections, in order to prevent early death of alcoholic hepatitis patients.

Methods: In vitro, 48 h after treatment with or without leptin (31.2 nM), ethanol (500 mM) and leptin + ethanol to mouse HCC cell lines. Results: Ethanol exposure significantly reduced the cell viability as evidenced by 3-(4,5 dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) (P \ 0.05) and trypan blue dye exclusion method (TBE) (P \ 0.05). Moreover, ethanol treated cells significantly lowered thymidine incorporation (P \ 0.05) and increased DNA fragmentation. Ethanol incubations also significantly increased the percentage of apoptotic cells (P \ 0.05). These results were compared with that of untreated control cell lines. Leptin cotreatment with ethanol showed significantly raised (P \ 0.05) cell viability, DNA synthesis and significantly (P \ 0.05) decreased apoptotic cells and DNA ladder formation. In addition, ethanol significantly increased the reactive oxygen species (ROS) and the expressions of mRNA of caspase-3 (2-fold), procollagen type I (about 24-fold), MMP 2 (3.8-fold), MMP 9 (2.5-fold) and TIMP-1 (1.3-fold) as compared to untreated control mouse HCC cell lines. Leptin coadministration significantly down regulated the mRNA expressions of caspase-3 (3-fold), procollagen type I (3-fold), MMP 2 (2-fold), MMP 9 (4.5-fold) and TIMP-1 (10fold) as compared to that of ethanol alone treated mouse HCC cell lines. Conclusions: Thus, our experimental data provide evidence above the potential protective effect of leptin on ethanol elicited damage in the mouse HCC cell lines, warranting population based and mechanistic studies.

Nadana Saravanan 1 , Namasivayam Nalini 2 1 Rani Meyyammai college of Nursing, Annamalai University, Tamil Nadu, India, 2 Department of Biochemistry and Biotechnology, Annamalai University, Tamil Nadu, India

Aim: The aim of this study was to evaluate the effect of Hemidesmus.indicus (HI) and its active principle 2-hydroxy 4-methoxy benzoic acid (HMBA) on lipid profile and fatty acid compositions in the liver of ethanol fed rats. Methods: Male albino Wistar rats were given with 20% ethanol (5 g/kg body weight) for 8 weeks and/or naive rats fed with glucose only. Four weeks later, ethanol fed rats were either coadministered with 500 mg/kg HI extract (orally) or with 200 lg/kg HMBA for the next 4 weeks and/or HI extract/HMBA alone (naive groups). On the eighth weeks all the rats in different groups were sacrificed, and plasma and liver tissue were collected for analysis. Results: The level of hepatic marker enzymes such as Alanine transaminase (ALT) and peroxidation marker, hydroperoxide (LOOH) were significantly elevated (P \ 0.05) in ethanol administered rats when compared to naive animal fed glucose only. Following treatment with both HI and HMBA significantly lowered (P \ 0.05) ALT and LOOH as compared with lone ethanol fed rats. Moreover the observed lipid profile such as cholesterol, free fatty acids and triglycerides were significantly increased (P \ 0.05) in ethanol treated animals when compared to control. Coadministration of HI and HMBA significantly reverts (P \ 0.05) these values towards control rats. In addition, fatty acid composition which includes palmitic, palmitoleic, stearic, oleic and linoleic acids were significantly increased (P \ 0.05) whilst polyunsaturated fatty acids such as arachidonic acid was significantly decreased (P \ 0.05) in ethanol treated animals when compared to control, while coadministration of HI and HMBA significantly reverts (P \ 0.05) these values when compared to naive rats. Conclusion: These results suggest that treatment with HI and HMBA improved alcohol induced hepatic steatosis. Background/Aims: Quercetin, a common flavonoids rich in plant kingdom, has been showed to exert substantially protective effect against ethanolinduced liver damage. The protection is involved in its induction on hemeoxygenase-1 (HO-1) and then the enhancement of adaptive protective response against excessive reactive oxygen species during ethanol metabolism mainly in liver. Ethanol also induces HO-1, however, the difference of signaling pathway to induce HO-1 by ethanol and quercetin are still remained fully unknown. Methods: Human hepatocytes were incubated with ethanol (100 mM) and/or quercetin (10-100 lM) for 24 h, following the incubation with various MAPK inhibitor(s). HO-1 expression and activity, together with oxidative cellular damage were assayed by spectrophotography and Western blot. Results: Compared with untreated human hepatocytes, ethanol exposure (100 mM, 24 h) resulted in a dramatic leakage of cellular AST and LDH, a sustained GSH depletion, and a parallel increase of cellular MDA. Ethanol itself induced HO-1 expression and activity, meanwhile, HO-1 inhibition by zinc protoporphyrin 9 further exacerbated ethanol-induced oxidative damage. Quercetin dose-dependently induced HO-1 expression and activity, and protected human hepatocytes from ethanol-induced damage, which was abrogated by zinc protoporphyrin 9. Querctin itself did not show any cytotoxicity, which was different from ethanol. Both PD98059 (ERK \ extracellular signal regu- Aims: To observe the effects of puerarin on chronic alcoholic liver injury and gut-leakage in rats. Methods: Chronic alcoholic liver injury and gut-leakage was induced with Lieber-Decarli diet for 8 weeks. Forty-four SD male rats were divided into control diet (C, n = 10), ethanol diet (E, n = 12), puerarin high-dosage (Ph, n = 11) and low-dosage (Pl, n = 11) group. At the beginning of the third week, rats in puerarin high and low-dosage groups were administrated with puerarin 179.4 mg or 89.7 mg/kg.body weight per day by gastric perfusion. Serum ALT, AST and hepatic GGT activity, pathologic changes in liver and intestine (HE staining and scanning transmission electron microscopy), hepatic TG, endotoxin in plasma and hepatic TNF-alpha were observed. Results: The serious steatotic hepatocytes (Fig. 1) , increased activity of serum ALT, AST and hepatic GGT, enhanced hepatic TG were observed in ethanol diet group (Table 1) . Further more, the microvillus on the surfaces of intestinal columnar epithelial cells were observed to be disconnected and thin, as well as the tight junction between the epithelial cells was broken (Fig. 2) . Similarly, increased endotoxin in plasma and TNF-alpha in liver tissue were also detected in ethanol diet group (Table 1) . However, puerarin administration ameliorated these pathological changes, significantly, especially, in high-dosage group ( Table 1 ; Fig. 2 ). Aims: To observe the effect of Pueraria Flavonid on chronic alcoholic liver injury in rats. Methods: Lieber-Decarli liquid diet induced-chronic alcoholic liver injury model (8 weeks) in rats was introduced. Forty-four SD male rats were divided into Lieber-Decarli control diet group (C, n = 10), Lieber-Decarli ethanol diet group (E, n = 12), pueraria flavonid high-dosage group (PFh, n = 11) and pueraria flavonid low-dosage group (PFl, n = 11). The Lieber-Decarli control and ethanol diet were drank freely and respectively by the rats in control group and the other three groups for 8 weeks. At the beginning of the third week, rats in pueraria flavonid high and low-dosage groups were administrated with pueraris flavonid 240 mg or 120 mg/ kg body weight per day by gastric perfusion. At the end of the eighth week, serum and liver tissue were collected for assy. The liver function, including serum ALT, AST, AKP and GGT activity in liver tissue, pathologic changes in liver tissue and hepatic lipid aggradation, TG in liver tissue, were observed. Results: The serious fatty degeneration in liver tissue, significant increased activity of serum ALT, AST, AKP and GGT in liver, enhanced TG concentration in liver were observed in Lieber-Decarli ethanol diet group. And the pueraris flavonid administration ameliorated these pathological changes, significantly, especially, in high-dosage group. Conclusions: Pueraris flavonid inhibit Lieber-Decarli liquid diet induced chronic liver injury in rats significantly.

The Alcohol affects to many organs, especially nerve system, liver and hematology.

Most of alcohol metabolized in the liver so the abuse of alcohol harms to the liver such as fatty liver disease, alcoholic hepatitis cirrhosis and anemia. In case of severe ethylic hepatitis that affected to the liver enzymes and mean corpucule volume. The aims: To study the relation between the duration and volume of alcohol dringking and the level of liver enzymes and MCV. Results:

• There was a positive correlation between the quantity of alcoholic drinking and the level of GGT (r = 0.38). • There was a positive correlation between the quantity of alcoholic drinking and the volume of red blood cell (MCV) r = 0.42. • There was a negative correlation between the quantity of alcoholic drinking and the level of prothrombin time (r = -0.39). • There was a negative correlation between the duration of alcoholic drinking and the level of prothrombin time (r = -0.35). • There was a positive correlation between the level of GGT enzyme and the level of SGOT enzyme (r = 0.36). • There was a positive correlation between the level of GGT enzyme and the volume of red blood cell (MCV) r = 0.34.

Conclusions: Alcoholic hepatitis an anemia are common in alcoholic abuse, so in case of patient with alcoholic abuse entry to the hospital we should notice to the hematic and liver function. The increasing of liver enzymes (GGT, SGPT, SGOT, Prothrombin time…) and mean hematic corpucule volume (MCV) are the good predictors to the severe of alcoholic hepatitis. Background and Aim: Alcohol dependency is one of the world's major health and socio-economic problem and is the third biggest cause of early death and illness, behind tobacco and high blood pressure. Similarly, intake of excessive energy from an energy-condensed-diet with high percentage of fat is associated with increased incidence of cardiovascular disease, obesity, diabetes, insulin resistance and certain cancers. Alcohol together with high fat-containing-diet affects the extent of hepatotoxicity. Natural products or their active components may serve the best approaches to minimize these disease conditions. Therefore, a natural phytoalexin, Resveratrol, was selected to reduce the toxic effects induced by combined consumption of alcohol and high-fat-diet (HFD).

Methods: Male rats were randomly distributed into different groups of six each (Control, Alcohol, HFD, HFD + Alcohol and HFD + Alcohol + Resveratrol). Subchronic HFD (30%) + Alcohol (10% v/v) intoxication was induced for 4 weeks. Resveratrol was administered (5, 10 15 and 20 mg/ml, orally) daily for 4 weeks. Condition of fatty liver was measured by cholesterol, triglyceride, biliruin, HDL and LDL content in liver and serum. Activities of transaminases, alcohol-dehydrogenase and acetaldehyde-dehydrogenase were studied. DNA damage, level of TNF-a, Interleukin-6, caspase-3, IGF-1 and CYP2E1 activity were measured in liver to understand molecular mechanism. Results: All biochemical variables were significantly altered by HFD + Alcohol intoxication. DNA damage was significant reduced and level of TNF-a, Interleukin-6, caspase-3, IGF-1 and CYP2E1 activity were found towards control with resveratrol treatment. Activities of transaminases, alcohol-dehydrogenase, acetaldehyde-dehydrogenase and bilirubin concentration were significantly maintained towards control with resveratrol at 15 and 20 mg/kg doses. Transmission and scanning electron microscopic evaluation confirmed a great extant of protection by resveratrol therapy. Conclusion: Resveratrol helps in mitigating hepatic abnormalities induced by combined consumption of alcohol and high fat diet, and as such, may be considered seriously for clinical use. Background: The risk of an individual to develop Alcoholic Liver Disease (ALD) is regulated by a complex interplay of genes and environmental factors. We therefore conducted a case-control study in an ethnic population aimed to identify association of SNPs in some candidate genes in the genesis of ALD.

Methods: This study included 201 cases with clinically proven ALD and 108 controls with alcohol abuse but clinically free of any liver disease, from Ethnic Bengali population of West Bengal, India. RFLP and DNA resequencing were done for genotyping several marker loci in the following candidate genes: genes influencing alcohol metabolism (ADH1C, ADH1B), detoxification (MnSOD, GSTM1, GSTT1), immune response (CD14, CTLA-4, IL-1b) and fibrosis (TGFb-1). Results: Polymorphic markers (rs1789920, rs1693425, rs698) of ADH1C gene were significantly associated with ALD. Again, markers like rs2066701 in gene ADH1B, rs4880 in exon2 of MnSOD, rs2569190, rs3087258 and C-159T in 5'promoter region of CD14, IL-1b and TGFb-1 respectively were also significantly associated with ALD. However, no association was found between rs231775 in exon1 of gene CTLA-4 and insertion/deletion polymorphism of GSTM1, GSTT1 with ALD. The table below provides pertinent details. Conclusion: In this study we have identified SNPs in six relevant genes that may confer increased susceptibility to ALD. Polymorphic markers of ADH genes on chromosome-4 show significant association with ALD even at haplotypic level. Susceptibility to development of ALD is increased when more than one marker is considered, as evident from gene-gene interaction effect. Background: Transforming growth factor-beta (TGF-beta) is the most potent profibrogenic cytokine in the pathogenesis of liver fibrosis or cirrhosis. Several functional single nucleotide polymorphism (SNP) sites have been identified within the TGF-beta gene; however, their associations with alcoholic liver disease (ALD) were inconsistent in previously published studies. This metaanalysis was undertaken to summarize the effect size of TGF-beta SNPs associated with susceptibility of ALD hepatic disorders. Aim of the study was to evaluate the hepatoprotective activity of MD against alcohol induced liver toxicity in rats. Methods: Swiss albino mice (30 ± 5 g) were divided into various groups of six animals each and received alcohol ad libitum in different concentration (10, 20, 30, 40% in I, II, III and IV week). MD was given at the dose 1,000 mg/kg, po, for 7 days after IV week. AST, ALT, ALP, albumin, urea, uric acid and creatinine in serum; ADH, LPO, GSH, triglyceride, cholesterol, SOD, CAT, GPx and GR in liver; genotoxicity (comet assay) and histological studies were performed. Results: Alcohol induced liver damage significantly increased AST, ALT, ALP, albumin, triglyceride, cholesterol, HDL, urea, uric acid and creatinine in serum and decreased hepatic GSH whereas increased LPO. MD significantly protected these parameters. MD also reversed the altered hepatic GSH, SOD, CAT, GPx, GR and ADH and suppressed LPO. Comet assay and histopathological examination also showed marked improvement after MD treatment. Conclusion: Thus, the study proposed the hepatoprotective potential of Majoon-e-dabeed-ul-ward against alcohol-induced liver damages due to its excellent antioxidant activity. Background: Diabetes is one of the stress related disorder. Diabetes damages all the organs in body like liver, kidney, heart and brain. Present investigation was undertaken to evaluate antioxidant and antihyperglycemic properties of ginger in streptozotocin (STZ, 50 mg/kg iv single dose) induced diabetic rats. Methods: For this purpose Wistar strain male albino rats were divided in to five groups and treatment was gives as schedule in experimental protocol. The antioxidant enzymes superoxide dismutase (SOD), glutathione (GSH), glutathione s transferase (GST) activities and lipid paroxidation (MDA) levels were assayed in the liver tissue. Results: STZ produced a significant increases in blood glucose, hepatic antioxidant enzymes GST, MDA levels and decreases SOD, GSH in diabetic rats. Treatment with ginger ethanolic extract (100 and 200 mg/kg) for 30 days produced decrease in STZ induced blood glucose, GST, MDA levels and increased SOD, GSH as compared to control. Administration of ginger to diabetic rats significantly reduced the levels of lipid paroxidation and increased the activities of antioxidant enzymes. Conclusion: These results suggested that ginger possessed anti-diabetic and anti-oxidant activity and these activities may be interrelated. Furthermore, the histopathological studies also proved that ginger protected the liver tissue form diabetic oxidative stress. The results were compared with antidiabetic drug glibenclamide. Laparoscopic fenestration is a standard procedure for the treatment of nonparasitic liver cysts. After the fenestration of giant cysts, remarkable regeneration of the remnant liver is sometimes observed, while the mechanism accounting for the regeneration has been unclear so far. In the present study, pattern of liver regeneration after the laparoscopic fenestration of liver cysts was analyzed. Methods: Seven patients with giant non-parasitic liver cysts and four patients with polycystic liver disease (PCLD) underwent laparoscopic fenestration between June 2002 and February 2009. All patients possessed abdominal symptoms such as upper abdominal discomfort, abdominal pain, and appetite loss. For the fenestration of liver cysts and the resection of the cyst wall, laparoscopic coagulating shears (LCS) and the LigaSure Atlas vessel sealing system were used. Liver regeneration in each lobe or segment was analyzed after the fenestration using CT or MRI images. Results: Liver regeneration after the fenestration was dependent on the size and position of the cysts, and not uniform throughout the remnant liver. Liver regeneration in segments which had been compressed by giant cysts was significantly greater than those in other regions after the fenestration, accompanied by the increased diameter of portal branches in the segments. This phenomenon implies that normalization of liver circulation, such as portal blood flow, in the compressed hepatic area may possibly regulate the liver regeneration after the fenestration. Fenestration of liver cysts in patients with PCLD also stimulated regeneration of the remnant liver. Discussion: Laparoscopic fenestration of liver cysts stimulates regeneration of the remnant liver while removing various abdominal symptoms with minimal surgical invasion. Normalized liver circulation, such as portal blood flow, possibly regulates the regeneration implying that surgical procedure considering subsequent liver regeneration would be important. In the present report we describe moderate hyperbilirubinemia in a man with combined disorders of hereditary spherocytosis (HS) and Gilbert's syndrome (GS). It was difficult to diagnose HS in absence of signs of anemia. Therefore, we needed to evaluate both the genetic mutation in the UGT1A1 gene and erythrocyte membrane protein abnormalities; the former was heterozygous for a UGT1A1 allele with three mutations [T-3279G (UGT1A1 * 60), A(TA) 7 TAA (UGT1A1 * 28), and G211A(UGT1A1 * 6)] and the latter was partial deficiency in ankyrin expression determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). This is the first report of the concomitance of HS and 3 heterozygous mutations (T-3279G, A (TA) 7 TAA, and G211A) in the UGT1A1 gene. Background/Aims: Wilson's disease (WD) is a rare autosomal recessive disease caused by tissue copper accumulation due to mutations in copper-transporting ATPase (ATP7B) gene. We investigate a large cohort with 62 WD patients from Chinese HAN population to expand the repertoire of ATP7B mutations and find any potential new genotype-phenotype correlation in Chinese patients with WD. Methods: The entire ATP7b coding region and promoter region were analyzed by direct sequencing in 62 Chinese patients with WD from 2 to 61 years old, including 37 male and 25 female. Results: We identified 40 different mutations (14 novel) in Chinese patients. Two mutations, Arg778Leu and p.Pro992 Leu, were relatively frequent, representing 31.9 and 11.2% of patients, respectively. By Systematic document review, about 74% of mutations locate in exon8, exon13, exon12 and exon16 in Chinese patients. Patients with R778L homozygotes and nonsense mutation/frameshift mutations had more often a primary hepatic manifestation (85.7 and 100%, P = 0.0286 and P = 0.0381, respectively) and higher ALT levels at diagnosis (P = 0.0361 and P = 0.005, respectively) than other patients. Patients with nonsense mutation/ frameshift mutations showed lower serum ceruloplasmin (P = 0.0318). Conclusions: In this study, we identified 14 novel mutations and found that the spectrum of mutations of ATP7B in Chinese patients are quite distinct from Western ethnic populations. To some extent, the type of mutation plays a role in predicting the clinical manifestation. Our data provide us valuable information to understand the heterogeneity of Wilson's diseases and formulate an economical and effective strategy of molecular diagnose.

Background: The worldwide prevalence of Wilson disease is estimated to be 1 in 30,000-100,000, and it is more frequent in China. The majority of patients with Wilson disease have a mutation of the ATP7B gene; this causes a defect of copper excretion, leading to copper accumulation in the liver, brain and cornea. Since ATP7B was cloned, more than 379 variants have been reported to be causative of Wilson disease. Here we present our sequencing findings of the whole coding region of the ATP7b gene in Chinese patients with Wilson disease.

Methods: A total of 17 Chinese Han Nationality patients with Wilson disease were included in this study; 5 cc of peripheral blood was drawn from each patient, and DNA was extracted from these samples. DNA samples were amplified by PCR using primers corresponding to all 21 exons and they were subsequently sequenced. Results: Thirteen out of 17 patients had two disease-causing mutations; another three had only one mutation found, one patient had no mutation identified, and two siblings of WD patients were diagnosed with WD. Also, we have identified three new mutations and two new polymorphisms. Conclusions: Our results suggest that heterogeneity of the mutation exists in our small patient population in northeast China. At the same time, we have noticed there might be a correlation between the p.R778L mutation and liver impairment, and p.R778L homozygotes are associated with an early onset of hepatic symptoms. Another interesting finding is the fact that affected patients within the same family even twins who have the same mutation can have very different phenotypes. This suggests that there are other factors impacting the presentation of the autosomal recessive disorder. Also, the directly sequencing the exons of patients' family members which had found mutation was a strict method for family member screening. Backgrounds and Aims: According to many researches and studies, the increase of r-GT has not only been considered as risk factor of cancer, type-2 diabetes mellitus, coronary artery disease and cerebrovascular disease but also as prognostic factor of all death. r-GT significantly increases through alcohol consumption but its relationship with obesity and smoking are not well-known. Hence, the authors have determined to identify the interactions between serum r-GT and alcohol, smoking and obesity. The average r-GT value significantly increased as the alcohol, current smoking, smoking history, and BMI increased (P \ 0.01). In the case of women, smoking and r-GT value did not show significant correlation. We analyzed the odds ratio of abnormal r-GT according to three factors. The odds ratio between smoking and BMI was 11.4, that of smoking and alcohol was 20.9, that of alcohol and BMI was 23.8. Conclusions: The effects of smoking, alcohol and obesity to serum r-GT value were observed and the positive synergic effect of alcohol and obesity were shown as the greatest. Understanding and education about this synergic effect is in need.

Liver Transplant in Children with Congenital Factor VII Deficiency Souheil M. Shabib 1 , Howard Parsons 1 , Azzam Alzoebie 1 1 SKMC, Dept. of Pediatrics, P.O. Box 51900, Adu Dhabi, UAE Background: Factor VII (FVIID) a rare autosomal recessive inherited disorder that is potentially a lethal defect in blood coagulation. A level of 35% (n: 70-130%) is usually required to achieve hemostasis. Incidence is 1/500,000 populations in USA. Higher incidence in countries where consanguineous marriage is common (3X higher in Iran compared to Italy). We report three related children successfully treated with liver transplantation and remain with normal coagulation profiles and complete resolution of their symptoms. Result: We concluded the five cases' common features that all patients had hepatomegaly, significant elevation of AKP and r-GT, mass proteinuria and hyperlipoidemia, four cases had hypoprotein. The markers of hepatitis A, B, C, D, E virus and autoantibody and antinuclear antibodies were negative, AFP level was normal. Four patients had ascites. Three patients have weight loss, hepatic ultrasound and CT showed the liver was enlarged and density was low, and the hepatic echo was fine and closely, three cases CT showed the liver vessels were a little thin. kidney ultrasound showed bilateral renal parenchymal echo was intensified, the structure was not clear, renal parenchymal blood distribution was decreased. Two cases pathlogic results of liver biopsy showed sedimentation of lots of pink amyloid matters in the hepatic cells and reticular framework. Congo red dyeing was positive, four cases pathlogic results of kidney biopsy showed sedimentation of lots of pink amyloid matters. Congo red dyeing was positive, all the patients' diagnosis were made by liver or kidney biopsy and specific dyeing Conclusion: If the patient is suspicious of this disease, liver or kidney puncture and specific dyeing (congo red dyeing) is necessary.

Poster Presentation: Autoimmune Hepatitis and Cholestatic Liver Disease Background: Celiac disease has been associated with other autoimmune disorders such as autoimmune hepatitis, moreover it is known that T cell mediated immune response to dietary gluten and released cytokines are important for the entheropathy seen in celiac disease. We investigated celiac autoantibodies in patients with autoimmune hepatitis (AIH), and chronic hepatitis B (CHB). Methods: Sera from 84 patients with Autoimmune Hepatitis (AIH) type 1 and 88 patients with Chronic Hepatitis B (CHB) were tested for Immunoglobulin A and G antibodies to Gliadin, Immunoglobulin A antibodies to tissue transglutaminase using enzyme immunoassay, and Immunoglobulin A antiendomysial antibodies by both indirect immunofluorescence, and enzyme immunoassay. The patients positive for anti-endomysial antibodies and/or anti tissue transglutaminase antibodies were considered for deuodenal biopsy. Results: Immunoglobulin A anti-endomysial and Immunoglobulin A anti-gliadin antibodies were positive in two of 84 patients with AIH. Moreover, Immunoglobulin A anti-gliadin antibodies were positive in another patient who was also positive for anti tissue transglutaminase antibodies. Tissue transglutaminase antibodies were positive in eight (9.1%) of 88 patients with CHB, two of which were also positive for anti-endomysial antibodies. One of the patients with CHB was only positive for anti-endomysial antibodies. Discussion: Compared with the normal population, the prevalence of celiac autoantibodies in CHB and AIH patients is relatively high, and it is noteworthy that most positive patients are asymptomatic for celiac disease. We suggest screening for celiac disease before and during treatment in patients with viral and autoimmune hepatitis. Background: Genetical B cell deprivation exacerbated liver inflammation in dnTGF-bRII mice, a mouse model of autoimmune cholangitis, raised a concept of regulatory B cells in liver inflammation. However, effect of therapeutic B cell depletion using anti-CD20 still remains to be clarified in PBC. Methods: To examine the effect of B cell depletion, we treated dnTGF-bRII mice from 4-6 and 20-22 weeks of age by intraperitoneal injection of anti-CD20 antibody (Ab) every 2 weeks, and compared the resulting disease phenotype with control Ab-treatment. Results: The anti-CD20 treatment initiated from 4-6 weeks of age demonstrated a significantly lower incidence of liver inflammation and a fewer number of activated hepatic CD8 + T cells in dnTGF-bRII mice. Also, B cell depletion increased levels of pro-inflammatory cytokines in sera, liver, and colon. Moreover, serum reactivity of anti-PDC-E2 was fully depleted after the anti-CD20 treatment. However, colon inflammation was significantly exacerbated. In aged mice, treated from 20 to 22 weeks of age, anti-CD20 was less effective either on liver or colon inflammation. Conclusion: Anti-CD20 treatment demonstrated therapeutic efficacy to regulate liver inflammation in young dnTGF-bRII mice, suggesting autodestructive B cell function in early human PBC. ultrasound and magnetic resonance cholangiopanceratography showed chronic parenchymal liver disease without bile duct changes. Liver histology demonstrated granulomatous destructive cholangitis with piecemeal necrosis and moderate lobular activity. The clinical data of high titer of ANA, elevated ALT to ALP ratio, positivity of AMA and histology were compatible with PBC dominant overlap syndrome. UDCA of 15 mg/kg/day was given and the levels of liver enzymes improved markedly after 2 months. She withdrew UDCA treatment by herself for 4 months and liver enzymes raised again (Fig. 1) . Background: Emodin, 1, 3, 8-trihydroxy-6-methyl-anthraquinone, is an anthraquinone derivative from the roots of Rheum officinale Baill that plays an important role in hepatoprotective treatment. This study is to investigate the effect and mechanism of emodin on acute intrahepatic cholestasis induced by alpha-naphthylisothiocyanate (ANIT) in rats. Methods: Normal control group, emodin group without ANIT treatment, model group and emodin group with ANIT treatment were set up. Liver function, pathological changes of hepatic tissue, the mRNA levels expression of the hepatic transport protein genes mdr1a (multidrug resistance protein 1a), mdr1b (multidrug resistance protein 1b) and mdr2 (multidrug resistance protein 2) and the expression of P-gp (P-glycoprotein) were detected. Results: Compared to the model group, Emodin treatment resulted in significant reductions in serum total bilirubin (TBiL), direct bilirubin (DBiL), alanine aminotransferase (ALT) and aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bile acid (TBA) (P \ 0.01 or P \ 0.05). By examining the liver pathology, it was found that hepatic cellular change and necrosis, inflammatory cell infiltration and bile duct proliferation were notably alleviated in emodin model with ANIT treatment. Analysis of gene expression in livers from emodin-treated cholestatic rats revealed that mdr1a, mdr1b and mdr2 could be up-regulated (P \ 0.01 or P \ 0.05), expression of P-gp was increased in accordance with its mRNA (P \ 0.05).

Conclusions: Emodin has a protective effect on hepatocytes and a restoring activity on cholestatic hepatitis. Mechanism of its action may be related to induce expression of the bile-metabolism-related transporter P-gp in the liver to prevent bile acids and other toxic compounds overaccumulation in hepatocytes and hepatic toxicity. Background: Emodin, 1, 3, 8-trihydroxy-6-methyl-anthraquinone, is an anthraquinone derivative from the roots of Rheum officinale Baill that plays an important role in hepatoprotective treatment. This study is to investigate the effect and mechanism of emodin on acute intrahepatic cholestasis induced by alpha-naphthylisothiocyanate (ANIT) in rats.

Methods: Normal control group, emodin group without ANIT treatment, model group and emodin group with ANIT treatment were set up. Liver function and pathological changes of hepatic tissue were examined. Real-time fluorescent quantitative RT-PCR was used to detect the mRNA levels of the hepatic transporter bile salt export pump (BSEP), multidrug resistance-associated protein 2 (MRP2), Na + /taurocholate cotransporting peptide (NTCP), multidrug resistance protein 2 (MDR2), multidrug resistance-associated protein 3 (MRP3) and the nuclear receptor farnesoid X receptor (FXR). Results: In the emodin group with ANIT treatment compared with the model group, the concentrations of serum total bilirubin (TB), direct bilirubin (DB), alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP) and total bile acid (TBA) were decreased (P \ 0.01 or P \ 0.05); Only mild histopathological change in liver could be seen; the genes expression of NTCP, MDR2 and MRP3 were higher than those of the model group (P \ 0.05), but the expression of BSEP, FXR and MRP2 could not be affected (P [ 0.05).

Conclusions: Emodin has a protective effect on hepatocytes and a restoring activity on cholestatic hepatitis. Mechanism of its action may be related to induce expression of the bile-metabolism-related transporters NTCP, MDR2 and MRP3 in the liver to prevent bile acids overaccumulation in hepatocytes and hepatic toxicity. Aim: T helper cells that produce IL-17 (Th17 cells) have recently been identified as the third distinct subset of effector T cells, which differentiation depends on specific transcription nuclear factor ROR-ct. We showed the Th17 cells in livers immunohistochemically and in the peripheral blood by flow cytometry. The levels of related cytokines such as Il-17, IL-6 and IL-23 were shown by RT-PCR and ELISA. Results: Our study showed that serum levels of IL-17 and IL-23 were upregulated in AIH patients than CHB patients and normal controls. In the peripheral blood, the ratio of Th17 cells in AIH patients was also significantly higher than that of CHB patients and normal controls (n = 20) (P \ 0.05). The frequencies of IL-17 + cells in AIH patients (n = 36) were significantly higher than that of chronic hepatitis B (CHB) patients (n = 15) in liver tissues (P \ 0.05). The mRNA expression of IL-17, IL-23, ROR-ct and IL-6 and IL-1b in the livers of AIH was significantly increased when compared to CHB and health. The frequency of IL-17 + cells is associated with hepatic inflammatory degree and fibrosis grade in AIH patients. IL-17 induced IL-6 expression of HepG2 cells in a dose and time-dependent manner. In a-Galactosylceramide (a-Galcer)-induced liver injury mice, which could be consider as an AIH model, anti-IL-17 antibody pretreatment attenuated significantly the inflammatory cell infiltration in the livers and hepatocyte injury. Conclusion: IL-17 induced a positive-feedback loop of IL-6 expression in hepatocyts. Anti-IL-17 treatment is worth to be explored as a new therapeutic method for immune-related liver diseases. Background: The characteristics of autoimmune hepatitis (AIH) are hypergammaglobulinemia and autoantibody positive. Some patients with AIH have anti-mitochondrial antibodies (AMA), but AMA, especially AMA-M2-type positive is found more in primary biliary cirrhosis (PBC). This study will evaluate the existence and clinical significance of AMA in autoimmune liver disease. Methods: Patients were chosen from our hospital from April 2004 to September 2008 and were diagnosed autoimmune liver disease after liver biopsy or related examinations, 39 patients (including 7 cases of male and 32 cases female, with an average age of 54.5 ± 10.8 years) were studied, AMA and other autoantibodies in their serum were detected. Results: Among 39 patients, 31 patients were diagnosed as AIH, and 8 patients were diagnosed as PBC; in the AIH patients, 28 patients with anti-nuclear antibody (ANA) positive (90.3%, 28/31); 6 patients with AMA positive (19.4%, 6/31), which 6 cases of AMA positive patients, their AMA-M2-type were all negative; there are 9 cases of anti-parietal cell antibody positive (29.0%, 9/31); there are 2 cases of AMA-M2-type positive (6.5%, 2/31); in 8 patients which were diagnosed as PBC, 6 patients with ANA positive (75.0%, 6/8); 5 cases of AMA positive (62.5%, 5/8); 5 cases of AMA-M2-type positive (62.5%, 5/8); there are 2 cases of anti-parietal cell antibody positive (25.0%, 2/ 8). The positive rate of ANA was no significant difference between AIH and PBC (P [ 0.05), AMA positive rate was significant difference between AIH and PBC (P \ 0.05), AMA-M2-type positive rates was also significant difference between AIH and PBC (P \ 0.01). Conclusion: ANA have a higher positive rate in both AIH and PBC, AMA and AMA-M2-type have a high positive rate in PBC, but there also have a certain positive rate in AIH, whether the patients with AMA-M2-type positive AIH will develop to PBC or overlap syndrome need to be further observed. Background: In the Asia Pacific region, autoimmune hepatitis (AIH) is uncommon compared to chronic viral hepatitis. AIH in the paediatric population is even less common. We describe our experience with AIH in the paediatric population that we have encountered. Methods: All patients with AIH were identified from our prospective database and reviewed. Results: There were 21 patients with AIH, two of which were in the paediatric population giving a prevalence of 9.5%.The first patient (12-year old girl) was referred with a year history of intermittent jaundice which became persistent over the last month. She had hepatosplenomegaly and other stigmata of chronic liver disease. Investigations showed deranged liver functions test (LFT), coagulopathy, positive SMA (1:40) and ultrasound scan confirmed liver cirrhosis. HLA typing was positive for DRB1*04 (DR4) and DRB1*16 (DR2). She was started on prednisolone but unfortunately died from complications of the novel influenza A H1N1 infection. The second patient (15-year old girl) was referred with three months history of jaundice and lethargy. Investigations showed deranged LFT, coagulopathy, positive SMA (1:640) and a liver biopsy showed grade III and stage III (METAVIR score) AIH. HLA typing was positive for DRB1*03 (DR3) and DRB1*10. She responded well to steroid therapy. Both patients were negative for anti-LKM, viral and metabolic markers.

Conclusions: Our AIH in the paediatric population although uncommon presented at advanced stages despite a short history of symptoms. Similar to adult patients, paediatric patients also respond to steroid therapy. Background: Adipokines which relate to metabolic circumstance become clear to have pro-inflammatory or anti-inflammatory properties and participate in the pathogenesis of immune mediated diseases such as rheumatoid arthritis and inflammatory bowel diseases. However, the serum levels of adipokins in the autoimmune liver diseases remain unknown. We measured the serum level of pro-inflammatory and anti-inflammatory adipokines in autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) to evaluate their role in the pathogenesis of autoimmune liver diseases. Methods: Forty AIH patients, 40 PBC patients and 20 healthy subjects were enrolled in this study. Serum levels of pro-inflammatory adipokines (Leptin, TNF-a, Visfatin, and Resistin) and anti-inflammatory adipokines (Adiponectin, Vaspin, and Chemerin) were measured by ELISA. The relation between the levels of adipokines and BMI, HOMA-IR index, biochemical variables (CRP, ALT, ALP, IgG, IgM) were evaluated. Results: Serum levels of Leptin, TNF-a, Visfatin, Resistin, Adiponectin and Vaspin were significantly higher in AIH and PBC compared with control (P \ 0.01). Only Leptin had a positive correlation to BMI. In AIH, Visfatin had a positive correlation to ALT (P \ 0.01) and IgG (P \ 0.001). The level of Adiponectin of predonisolon treated group was significantly higher compared with UDCA treated group (P \ 0.01). The level of Chemerin was significantly lower compared with PBC (P \ 0.05) and control (P \ 0.05). In PBC, Visfatin had a positive correlation to ALP (P \ 0.05) and IgM (P \ 0.05). Conclusion: Serum levels of adipokines except for Chemerin were significantly higher in autoimmune liver diseases compared to control. The level of proinflammatory adipokine Visfatin had a positive correlation to disease activities in autoimmune liver diseases. These results indicated that Visfatin might be involved in the course of autoimmune liver diseases. Results: Acute onset AIH showed heterogeneous hypoattenuating areas (P \ 0.001) and acute viral hepatitis diffuse ones (P \ 0.001). The diffuse hypoattenuating areas were present in none of group 1, 9 (60%) of group 2, and 5 (31%) of group 3 (P \ 0.001 between groups 1 and 2, P = 0.009 between groups 1 and 3). The heterogeneous hypoattenuating areas were present in 15 (68%) of group 1, none of group 2 and 1 (6%) of group 3 (P \ 0.001 between groups 1 and 2, P \ 0.001 between groups 1 and 3). Conclusions: Heterogeneous hypoattenuation on unenhanced CT was a characteristic CT imaging feature of acute onset AIH compared with viral acute hepatitis. This finding could be one of the tools for diagnosing acute onset AIH in combination with the international AIH scoring system. Background and Aims: In chronic liver diseases such as primary biliary cirrhosis (PBC), it is vital to improve and maintain the quality of life (QOL). The available self-report questionnaire for the assessment of QOL in patients with PBC, PBC-40 (Jacoby, 2005), consists of 40 items concerning several aspects of QOL and is currently validated only in the British population. The aims of the study were to validate the Italian and Japanese versions of PBC-40 and to assess the validity of the original six domains structure of PBC-40 by a confirmatory factor analysis. Methods: PBC-40 was translated to Italian and Japanese using the forwardbackward method and then reviewed in focus groups in the framework of a large multicentric study. A sample of 290 Italian and Japanese patients with PBC was administered two questionnaires, PBC-40 and SF-36, previously validated for assessment of general QOL both in Italian and Japanese. Results: The internal consistency of the original six domains turned out to be acceptable whereas the confirmatory model failed to fit adequately. A principal component analysis led to a seven factors structure, and suggested the exclusion of 13 items characterized by lower load; PBC-27 questionnaire was the final instrument. Pearson correlation between PBC-27 and SF-36 scores for both Italian and Japanese samples showed that the PBC-27 factors correlated significantly with the SF-36 domains. Conclusion: We here propose an alternative structure of the quality of life questionnaire for PBC, namely PBC-27, which appears to be valid and effective in detecting the impact of PBC on quality of life in Italian and Japanese patients. Background and Aims: Autoimmune hepatitis (AIH) is a disease of unknown etiology characterized by a progressive destruction of the hepatic parenchyma, the molecular mechanisms involved in the development of AIH are still unclear.This study aims to characterize functional variations of immune status in AIH and its immunologic features. Methods: Peripheral blood was collected from patients with AIH (n = 15) and healthy individuals (n = 21). Phenotypes of peripheral blood lymphocyte subsets (T cells, NK cells, NKT cells and regulatory T cells) were analysed using flow cytometry. Results: Both CD4 + CD25 + CD127 low/-Treg cells (mean 1.09%, range 0.94% to 1.96%) and CD4 + CD25 + FoxP3 + Treg cells (mean 1.27%, range 1.02-2.08%) were significantly lower in AIH patients as compared to normal individuals(mean 3.65%, range 1.52-7.14%, P \ 0.01),while the proportion of CD3 -CD56 + NK cells and CD3+CD8+ T cells were significantly higher than that of healthy controls (P \ 0.01). The expression of 2B4 + (CD244) on CD3 -CD56 + NK cells were lower in AIH patients compared to that in healthy controls (P = 0.001).

Conclusions: These findings provide evidence that there is significant difference in regulatory T cells between the patients with AIH and normal individuals. Tregs are defective numerically and functionally in AIH. The deficience of Tregs may contribute to the development of autoimmunity.

Zhi-Gang Tian 1 Background/Aims: The discovery of innate immune receptors and the emergence of liver immunology (high content of NK and NKT cells in liver) led to the second research summit in innate immunity since the finding of NK cells in the middle 1970s. Liver disease is one of the most dangerous threats to humans, and the progress in innate immunology and liver immunology made it possible to re-explain the cellular and molecular immune mechanisms of liver disease. Methods and Results: In the past ten years, we have found that innate recognition of hepatic NK and NKT subsets were involved in murine liver injury. We established a novel NK cell-dependent acute murine hepatitis model by activating Toll-like receptor-3 (TLR-3) with an injection of poly I:C, which may mimic mild viral hepatitis (such as Chronic Hepatitis B). We observed that a network of innate immune cells including NK, NKT and Kupffer cells is involved in liver immune injury in our established NK cell-dependent murine model. We noted that TLR-3 on Kupffer cells activated by pretreatment with poly I:C might protect against bacterial toxin (LPS)-induced fulminant hepatitis by down-regulating TLR-4 function, while TLR-3 pre-activation of NK cells might reduce Con A-induced NKT cell-mediated fulminant hepatitis by blocking NKT cell recruitment to the liver. We also found that the oversensitivity to injury by immune stimulation in HBV transgenic mice (full HBV gene-tg or HBs-tg) correlated to the over-expression of Rea1, an NKG2D ligand of NK cells or CD1d, a ligand of TCR-V14 of NKT cells, on HBV + hepatocytes, which leads to an innate immune response against hepatocytes and is critical in liver immune injury and regeneration.

Hepatol Int (2010) Background: 3b-hydroxy-C27-steroid dehydrogenase (C 27 3b-HSD) is the second enzyme that catalyses the synthesis of bile acids from cholesterol. Deficiency of this enzyme fails to synthesize bile acids and develop a form of progressive liver disease. This deficiency has not been previously described in China, our aim was to identify mutations in the HSD3B7 gene in a patient who was cholestatic jaundice and normal serum gamma glutamyl transpeptidase (GGT) and total bile acid (TBA). Methods: We studied a male infant with neonatal onset cholestatic liver disease and normal GGT and TBA who accepted bile acid replacement therapy resulted in clinical and biochemical improvement. Genomic DNA of peripheral blood leukocytes was extracted routinely. The entire six coding exons together with at least 100bp of adjacent intronic sequences of the HSD3B7 gene were amplified by polymerase chain reaction (PCR) and directly sequenced. Results: The patient was homozygous for a missense mutation (1031A [ G) in exon 6, which leads to amino acid transition from tyrosine to cysteine at amino acid position 344. The mutation was detected in heterozygous form in his sibling and parents. The patient with cholestatic liver disease was diagnosed as having 3b-hydroxy-C27-steroid dehydrogenase deficiency by HSD3B7 gene analysis.

Conclusions: This is the first report of an infant with 3b-hydroxy-C 27 -steroid dehydrogenase deficiency in China. Genetic studies revealed a novel mutation in the HSD3B7 gene. In China, when pediatricians encounter patients with cholestatic liver disease whom serum GGT and TBA was normal, 3b-hydroxy-C27-steroid dehydrogenase deficiency should be considered and genetic analysis should be carried out early, because this deficiency can be successfully treated by oral administration of primary bile acids.

Poster Presentation: Drug Induced Liver Disease (DILD) Background: Tamoxifen (TAM) is a synthetic non-steroidal antiestrogen. Now it is frequently used as adjuvant therapy in breast cancer patients, and proved to effectively improve their survivals. But it causes hepatic steatosis in 43% of recipients. Further more, steatohepatitis, hepatic fibrosis and cirhrrosis can develop. However, the mechanism for tamoxifen-induced steatosis is essentially unknown. Insulin resistance (IR) was considered to be the most important mechanism of nonalcoholic fatty liver, but whether TAM can induce IR was unknown. This research was to observe the effects of tamoxifen on insulin sensitivity in L02 hepatic cells, and explore the role of oxidative stress in it. Methods: Cultured L02 cell was divided into four groups according to the different dose of TAM (balnk, 1, 3 and 5 lM) in the media, and each group was incubated for 24 h in presence or absence of NAC.Then insulin was added in culture media. Residue glucose concentration was measured by the method of glucose oxidizes peroxides (GOD-POD), and the level of SOD and GSH was determined. Results: 1.Glucose absorption in L02 cell was reduced by TAM (P \ 0.05), and the effect was dose-dependent.2.The levels of GSH (P \ 0.01) and SOD (P \ 0.05) were significantly decreased by TAM compared to control group 3. Introduction: Six Chinese patients with the diagnosis of drug-induced hepatitis due to taking the medicine for hemorrhoids, ranged in age from 30 to 58, were hospitalized in Shenzhen Shekou people's Hospital in 2008. These cases were investigated further in order to diagnose and treat these special cases more effectively. Taking medicine history, clinical presentation, clinical laboratory findings, treatments and effects were summarized, and followed-up for 3-6 months. Case description: The medicine for hemorrhoids is a Chinese patent medicine containing plenty of Snphora angustifolia. It has been reported that some Hepatitis B patients had suffered from more serious liver function damage after taking Snphora angustifolia. Four of these 6 patients were admitted with jaundice 2 months following taking medicine. One patient was admitted with jaundice 1 month earlier. The other one took the medicine 3 weeks after haemorrhoidectomy then appeared hypodynamia, nausea symptoms. Laboratory and ultrasound examinations showed that they were cholestatic hepatitis. Treatment of protecting liver function and promoting bile excreting (such as UDCA) was helpful to the recovery of liver function. Two patients who appeared persistent jaundice and progressive increase in bilirubin were administered with glucocorticoid. The liver function of all these patients' returned to normal after various durations of therapy. Glucocorticoid administration was proved to be effective in the recovery of the drug-induced cholestatic hepatitis. The liver function and bilirubin returned to normal completely after 6-month therapy for three patients, 4-month therapy for two patients and more than 2-month therapy for one patient, without recurrence till now. Non-surgical therapy including oral administration and external application were commonly used in treating hemorrhoids. The compositions of some medicines are complex with involvement of hepatic function in drug metabolism. These cases remind the wariness of taking Chinese patent medicine for treating hemorrhoids or medicine containing large amount of snphora angustifolia Backgrounds: Lots of chemical drugs, herbal medicines and heath care products have potential toxicity on human health. Liver is the biggest organ for the biochemical metabolism, thus also the main target for the attack of druginduced injury. The categories, mechanisms, diagnosis and research methods of drug-induced liver injury (DILI) is still confused to a great extent, and should be paid more attention to. Methods: Review the related articles systemically. Result: Various mechanisms contribute to DILI. Though DILI is very common, clinicians and general public have not paid enough attention to its importance. In European and American countries, approximately 30-40% of patients with acute liver failure were caused by DILI, and 36% of DILI cases were caused by non-steroid anti-inflammatory agents in the US, especially acetaminophen. DILI could be divided into predictable and unpredictable types, and the former is usually caused by the direct toxicity of drugs. The unpredictable type of DILI is mediated by hypersensitive or allergic idiosyncrasy; though this is rarely seen in the clinical trials, it can lead to the abolishment of new drugs from the market. The mechanisms of DILI could be summarized as injuries from: (1) direct toxicity of the drugs;

(2) immune-mediated idiosyncrasy;

(3) metabolic idiosyncrasy; and (4) oxidative stress. A few individuals who suffered from DILI previously may be used as the most appropriate models to explore the mechanisms of idiosyncrasy-based DILI. Conclusion: There exist great puzzles in the diagnosis of DILI until now, though the diagnostic methods and criteria of DILI have got some modification, evolution and development. In China, special attention should be paid to the liver injury caused by herbal medicines. Many related questions and challenges have to be faced with, and the current urgent task is to systematically establish and improve the database of toxic and drug-induced liver diseases in China. Introduction: Hepatic drug metabolism, often with an imbalance between the generation of toxic metabolites and detoxification processes, can influence the degree of hepatotoxicity. This drug toxin causes sudden onset of portal hypertension, with very prominent hepatomegaly and ascites in a previously healthy infant or child. The majority of toxicities depend upon the bioactivation of the drug in the liver catalyzed by the cytochromes P450 (CYP). Methods: Many drugs cause mild elevations in blood levels of liver enzymes in children without symptoms or signs of hepatitis. Liver histology is the ideal tool to date for defining the pattern of hepatotoxicity. There are very few reports of hepatic injury in children caused by environmental toxins in Bosnia and Herzegovina.

Results: This indicates severe hepatotoxicity and is likely to lead to mortality in 6% of kids patients in Bosnia and Herzegovina, especially if the offending drug is not stopped. Drug-induced liver injury can mimic any pattern of primary liver disease. Girls are more susceptible than boys are to most forms of druginduced liver disease. Discussion: Measurement of blood level is important in assessing prognosis, higher levels predicting worse prognosis. Conclusions: The diagnosis of drug-induced liver injury (DILI) is a challenging problem in children. Acute drug-induced hepatitis is defined as hepatitis that lasts less than 3 months, while chronic hepatitis lasts longer than 3 months. There is no specific treatment for most cases of drug-induced hepatitis other than stopping the drug that is causing the problem.

Neetu Sharma 1 , Sangeeta Shukla 1 1 Jiwaji University, SOS in Zoology, Jiwaji University, Gwalior 474011 India

Background and Aim: Flowers part of Butea monosperma associated with various remedial properties such as, antistress, antiestrogenic and chemopreventive. Aqueous extract of Butea monosperma flowers was evaluated for its hepatoprotective efficacy against acetaminophen induced hepatotocellular damage in rats. Method: Hepatotocellular damage was induced by acetaminophen for acute (2 g/kg, po, once only) and subchronic (20 mg/kg, po, for 21 days) exposure. Butea monosperma flowers extract at different doses (200, 400 and 800 mg/ kg, po, once only) was given in acute study and selected dose was administered in subchronic study (800 mg/kg, po, for 5 days). AST, ALT, albumin and bilirubin in serum; hepatic antioxidant status (GSH, SOD, CAT, GPx and GR), CYP450 activity and genotoxicity were estimated. Histopathology and ultrastructural changes were studied along with various biological activities (free radical scavenging activity, total phenolic contents and antipyretic activity). Result: Acute and subchronic exposure to acetaminophen showed toxic effects on various blood and tissue biochemical variables. Acetaminophen significantly increased AST, ALT, albumin and bilirubin in serum. Hepatic lipid peroxidation was increased significantly, whereas reduced glutathione was decreased after acetaminophen exposure. Butea monosperma extract significantly improved hepatic antioxidant status (GSH, SOD, CAT, GPx and GR) and inhibited lipid peroxidation in liver at higher dose (800 mg/kg). CYP450 activity was increased and also protected DNA damage induced by acetaminophen with reversed histopathological alterations. Conclusion: Thus, it may be concluded that the extract of Butea monosperma flowers can be used to reduce hepatic damage and may serve as complimentary or alternative herbal drug against acetaminophen induced liver damage.

Evaluation 152 Objective: Drug induced liver injury (DILI) are the common drug adverse reaction. We find some patients diagnosed as DILI with the abnormal autoimmune antibody raising. Antinuclear antibody (ANA) is often tested as positive and we try to investigate the difference between ANA positive and negative in DILI. Methods: 152 patients clinical diagnosed as DILI and divided to two groups by ANA positive and negative are analyzed retrospectively and compared by age sexual gender level of biochemical indicators. Result: There are 23 cases with ANA tested positive and the percentage is 15.13%. Age sexual gender DILI types are not different in two groups (T test). The level of alanine transaminase is higher and the level of total bilirubin is lower in ANA positive group. The level of alkaline phosphatase seems same between two groups (Mann-Whitney test). Conclusions: We can find ANA positive cases appear usually in drug induced liver injury. Age and gender are not different between two groups. In the early clinical stage of DILI without medical treatment, we can find different change in biochemical indicators. We may suppose that DILI with ANA positive may reflect more serious hepatic cell injury. Background: Hepatic fibrosis is a major complication of various chronic liver diseases. Activated hepatic stellate cells (HSCs) play a critical role in the development of liver fibrosis, and the axis of platelet-derived growth factor (PDGF)/PDGF receptor (PDGFR), a member of receptor tyrosine kinases (RTKs), is closely associated with the activation of HSC. Insulin-like growth factor (IGF)-1 receptor (IGF-1R), which also belongs to RTKs, interacts with the PDGF/PDGFR axis, thereby cooperatively promoting hepatic fibrosis. EGCG, the major constituent of green tea, assumes RTK such as EGFR or IGF-1R a target and restrains the increase of tumor cells. Aims: We herein examined the effects of (-)-epigallocatechin gallate (EGCG), which inhibits the activation of several types of RTKs, on the development rat liver fibrosis induced by carbon tetrachloride (CCL 4 ). Methods: Jcl Male Wistar rats were given subcutaneous injections of CCL4 (0.5 mg/kg/BW) twice a week for 8 weeks. And during the same period, they received water containing 0.1% EGCG. We did a sacrifice at 14 weeks old. Results: At sacrifice, drinking water with 0.1% EGCG significantly decreased the serum levels of both aspartate aminotransferase and alanine aminotransferase raised by CCl 4 , thus indicating an improvement of liver injury. In CCl 4 -injected rats, EGCG markedly attenuated hepatic fibrosis and decreased the amount of hydroxyproline in the experimental liver. The expression of PDGFR-beta and IGF-1R mRNAs in the liver was significantly reduced by the treatment with EGCG. EGCG also decreased the expression of PDGFR-beta and alfa-smooth muscle actin proteins, thus indicating the inhibition of HSC activation. These findings suggest that EGCG can exert, at least in part, an antifibrotic effect on the liver by targeting PDGFR-beta and IGF-1R. Conclusion: EGCG might therefore be useful in the prevention of hepatic fibrosis. Activation of hepatic stellate cells (HSCs), which is regulated by multiple signal transduction pathways, is the key event in liver fibrosis. Moreover, members of these pathways are important targets for microRNAs (miRNAs).

To better understand the critical pathways of HSCs activation, we carried out comprehensive comparative bioinformatics analysis of microarrays of quiescent and activated HSCs. Changes in miRNAs are associated with HSC activation status revealed that thirteen pathways were upregulated and twenty two pathways were downregulated by microRNA. Furthermore, mitochondrial integrity based on highly up-regulated Bcl-2 and downregulated caspase 9 was confirmed in HSCs and fibrotic livers by immnofluorescence assay, quantitative RT-PCR and Western blot analysis. These findings provide in vitro and in vivo evidence that the mitochondrial pathway of apoptosis plays a significant role in the progression of liver fibrogenesis via HSCs activation.

Hu Guo-Xin 1 1 First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi Province, China

Aim: To study hydroxycamptothecin (HCPT) toxicity in hepatic stellate cells (HSC-T6) cultured in vitro and normal hepatic cells (BRL-3A) in rats, and determine its effects on proliferation and apoptosis, thereby identifying the optimal therapeutic concentration. Methods: Rat HSC-T6 and BRL-3A were cultivated as normal or in medium containing different concentrations of HCPT (0.008, 0.016, 0.031, 0.063, 0.125, 0.25, 0.5, 1, 2, 4, 8, 16 , and 32 mg/L) for 24, 48, and 72 h. Cellular proliferation was detected by MTT assay; morphology was observed by microscopy; apoptosis was detected by flow cytometry; apoptotic cell morphology was observed under transmission electron microscopy, and DNA fragmentation was determined by agarose gel electrophoresis. Results: The inhibition rate of HCPT on HSC-T6 and BRL-3A proliferation gradually increased with increasing drug concentration and prolonged medication time. When HCPT concentration was [0.5 mg/L, its toxic effects on BRL-3A cells were significantly increased. Following 24-h treatment with 0.125, 0.25, and 0.5 mg/L HCPT, the cellular apoptosis rate was 13.46 ± 2.42, 26.25 ± 5.65 and 47.05 ± 8.76%, respectively. A significant difference existed between the test and control group of 4.89 ± 1.80% (F = 34.24, P \ 0.01). Following 24-h treatment with 0.5 mg/L HCPT, apoptotic cells were observed by transmission electron microscopy. They displayed a decreased cell body, absent nucleoli and chromatin aggregation along the nuclear membrane. DNA analysis on agarose gels showed gradient bands. Conclusion: 0.5 mg/L HCPT can produce maximal inhibitory effects on HSC-T6 proliferation. HCPT can markedly inhibit HSC-T6 proliferation and induce apoptosis in a dose-and time-dependent manner. Objective: To establish bone mesenchymal stem cell (MSCs) culture and amplification system and investigate the effects of MSCs transplantation on liver of rats. The role of TGF-b1 in mechanisms of bone MSCs transplantation and hepatoma was also studied. Methods: Combined with stationary culture MSCs of male SD rat were isolated and purified. The MSCs ( Background and Aims: Sustained inflammation-induced hepatic stellate cell (HSC) activation and transformation into fibrogenic and proliferative myofibroblast-like cells is the major cause of hepatic fibrosis. Our recent studies demonstrated that endogenous connective tissue growth factor (CTGF/CCN2) is a downstream mediator of basal or TGF-b1-induced collagen I production in human HSC and regulates entry of the cells into S phage. Here, we assessed the role of CCN2 in fibrogenesis in HBV-induced chronic liver diseases (CLD). Methods: ELISA was used to measure CCN2 in sera from patients with CLD (89 with chronic hepatitis B and 66 with hepatitis B-induced liver cirrhosis) and 30 healthy individuals. Liver samples from patients (26 with chronic hepatitis B, 5 with hepatitis B-induced liver cirrhosis) and 6 healthy individuals were examined for CCN2 and TGF-b1 mRNA levels by in situ hybridization, CCN2 protein expression by immunohistochemisty, and collagen fibers by Van Gieson's method. Results: TGF-b1 mRNA was mainly detected in macrophages and activated HSCs within inflammatory and fibronous areas in the liver. However, CCN2 mRNA expression in activated HSC and fibroblasts was seen in both the fibronous septa and sinusoidal lining. The levels of CCN2 in liver samples and sera from patients with HBV-induced CLD were significantly higher than those in healthy controls, and the levels of CCN2 were positively correlated with liver collagen expression and the degree of fibrosis (both P \ 0.05). Conclusions: Our data indicate that CCN2 plays an important role in HBVinduced hepatic fibrosis, and may serve as a serological biomarker for ongoing fibrogenesis. Background and Aim: Hepatocytes (PC) are a major cellular source of connective tissue growth factor (CCN2/CTGF), a downstream amplifier of TGF-b actions that has thus been implicated in the pathogenesis of hepatic fibrosis. Earlier reports showed that the prototypical rat acute phase reactant a2-macrofetoprotein acts as an inhibitor of experimental hepatitis and fibrosis, however, the impact of the acute phase reaction initiating interleukins such as IL-6 on CCN2/CTGF synthesis in PC is yet unknown. We therefore investigated the mechanisms involved in a possible modulator role of IL-6 signalling on CCN2/CTGF expression in PC and looked for a relation between serum concentrations of these two parameters in patients with acute inflammation. Methods: Expression of CCN2/CTGF, p-STAT3, p-Smad3/1 and p-Smad2 was examined in primary fresh-isolated rat or cryo-preserved human PC exposed to various stimuli by Western blotting, reporter-gene-assays and RT-PCR. Results: IL-6 strongly down-regulated CCN2/CTGF protein and mRNA expression in PC, an effect enhanced by the extracellular presence of the soluble IL-6 receptor gp80. These data were confirmed by an inverse relation between IL-6 and CCN2/CTGF concentrations in patients' sera. The inhibition of TGF-b driven CCN2/CTGF expression by IL-6 did not involve a modulation of Smad2 (and Smad1/3) signalling. However, the STAT3 SH2 domain binding peptide, a selective inhibitor of STAT3 DNA binding activity, counteracted the inhibitory effect of IL-6 on CCN2/CTGF expression much more pronounced than pyrrolidine-dithiocarbamate, an inhibitor primarily of STAT3 phosphorylation. Conclusion: CCN2/CTGF is identified as a hepatocellular negative acute phase protein which is down-regulated by IL-6 via the STAT3 pathway through interaction on the DNA binding level.

Background: Activated hepatic stellate cell (HSC) is a major participant in hepatic fibrosis, and thus, the suppression of HSC activation and/or survival has been implicated as anti-fibrotic therapies. Exisulind, a metabolite of the nonsteroidal anti-inflammatory drug sulindac, is one of recently developed apoptosis-inducing agents, which is being evaluated in cancer treatment. The purpose of this study was to assess efficacy of exisulind in modulating activation and survival of HSCs. Methods: LX-2 cells, an immortalized human HSC line, were used in this study. Cellular apoptosis was evaluated by DAPI staining and immunoblot analyses. Collagen a1 and a-smooth muscle actin expression were examined with real time RT-PCR and immunoblot, respectively. Result: Exisulind induced HSC apoptosis in an activation-independent manner. This apoptosis was mediated by activation of JNK, Bim (BH3 domainonly protein) and later mitochondrial apoptotic signaling. Exisulind treatment also attenuated NFkB-dependent cFLIP expression in HSCs. In addition to inducing apoptosis, exisulind attenuated collagen a1 synthesis and a-smooth muscle actin expression in HSCs. Conclusion: These results demonstrate that exisulind suppresses activation and survival of HSCs through JNK and NFkB-dependent mechanism. Therefore, this agent is therapeutically implicated as an anti-fibrotic strategy in hepatic fibrosis. Background: Huangqi Tang (HQT) is a well-known Traditional Chinese Medicine. We previously reported HQT exerting significant therapeutic effects on DMN-induced cirrhosis/fibrosis in rats. In this study, Proteomics approaches were performed to identify proteins that may be involved in DMN-induced cirrhosis/fibrosis and the effects of HQT intervention were observed in rats. Methods: Differential protein spots that exhibited changes in DMN-induced cirrhosis/fibrosis were selected by Two-dimensional gel electrophoresis and identified by Matrix-assisted laser Desorption/ionization Time-of-flight and Peptide Mass Fingerprinting Analysis. After that, these protein spots were recognized by Bioinformatics Information Retrieval. Lastly, the results of this proteomic analysis were confirmed by immunoblotting and detection of enzyme activity. Results: 20 protein spots were found to be differentially expressed, including 7 spots involved in disorder of lipid metabolism and oxidative damage, 4 spots involved in protein metabolism, 3 spots involved in biotransformation and 3 spots involved in material synthesis and energy metabolism in liver. Among them, 4 spots for confirmation (L-FABP14, Prdx6, Hsp70, CAT) were consistent with the results of Two-dimensional gel electrophoresis. Compared with the model group, HQT could up regulate the expression of Prdx6, Hsp70, CAT and down regulate the expression of L-FABP14 (P \ 0.01).

Conclusions: There are significant oxidative stress, endoplasmic reticulum stress, decline of biotransformation ability and metabolic alterations occurred during hepatofibrogenesis of DMN-induced cirrhosis/fibrosis in rats, and the therapeutic effects of HQT on DMN-induced cirrhosis/fibrosis in rats might be associated with its possible antioxidant properties. Background/Aims: Hepatic fibrosis is a consequence of severe liver damage that occurs in many patients with chronic liver diseases. FuZhengHuaYu recipe (FZHY) is a Chinese Medicine formula. In this study, we analysis the differential proteome of fibrotic liver tissues induced by dimethylnitrosamine (DMN) in rats after FZHY treated and explore the action mechanism of FZHY anti-liver fibrosis. Methods: Liver fibrosis in male Sprague-Dawley rats was induced by DMN, while the rats in the treatment group received FZHY. Total liver protein was separated by two-dimensional gel electrophoresis. Protein identification was done by peptide mass fingerprinting with matrix assisted laser desorption/ ionization-time of flight mass spectrometry. The expression levels of vimentin in the liver tissues were validated by western blot and real-time PCR analyses.

Results: There were about 1,120 ± 154, 1,122 ± 113, 932 ± 160 spots on the maps of normal group (a), model group (b) and FZHY group (c), respectively. A totally of 61 protein spots exhibiting significant (P \ 0.05) changes in intensity, and 22 protein spots, corresponding to 16 different proteins or polypeptide chains were finally identified. The vimentin was further examined by Western blotting and quantitative real-time PCR, resulting in coincident data with the proteomic evidence. Conclusions: The results provide 11 therapeutic-associated proteins with the FZHY, and indicated that the mechanism of FZHY anti-liver fibrosis was associated with modulation of metabolism, stress response, structural molecules. Further studies of the functional roles of those proteins indentified by this approach could contribute to more thorough understanding of the mechanism of action FZHY against liver fibrosis. Methods: The HSC cell line LX-2 was incubated with four concentration (10 -4 , 10 -5 , 10 -6 and 10 -7 mol/L) of salidroside for 18 h, respectively, PDGF was added at sixth hour before the completion of incubation with salidroside. A transwell chamber system was used to observe the changes of HSC migration after Platelet-derived growth factor-BB (10 ng/mL) treatment. Five fields were randomized selected to count the number of migrated cells. Results: The migration cells in control group were 34.13 ± 2.19. After stimulating by PDGF-BB, the migration cells reached to 74.20 ± 4.10 (P \ 0.05, compared to control group). Four different concentrations of Salidroside can inhibit PDGF-induced migration of the LX-2, the migration cells in each group were as follows: 7.73 ± 2.83 (10 -4 mol/L), 5.53 ± 2.13 (10 -5 mol/L), 15.20 ± 4.18(10 -6 mol/L) and 29.40 ± 0.80 (10 -7 mol/L). There is no significant difference between 10 -4 and 10 -5 mol/L group, however, there is significant difference in 10 -5 and 10 -6 mol/L group, 10 -5 and 10 -7 mol/L group. Conclusion: Salidroside can effectively inhibit the migration of LX-2. 10 -5 mol/L Salidroside has the best effect. Introduction: The mechanisms whereby overweight and obesity renders the liver more susceptible to alcohol-associated damage are unclear. In overweight persons, serum leptin levels are increased and we have shown that leptin promotes liver fibrosis. Acetaldehyde, a metabolite of alcohol, activates hepatic stellate cells (HSC). Aims: To clarify the mechanisms by which overweight might contribute to alcohol-associated liver injury, we examined whether leptin promotes the profibrogenic effects of acetaldehyde in cultured HSCs and sought to identify factors that might contribute to any observed effects. In addition, we determined the signalling pathways that mediate HSC activation during combined treatment with leptin and acetaldehyde. Methods: Rat HSCs in various stages of activation (2 days to third passage) were isolated from wt rats. Recombinant rat leptin (100 nM), acetaldehyde (175 lM), and acetaldehyde + leptin were added to cultured HSCs. Gene expression of type I collagen, TIMP1, TGFb1, CTGF and IL-6 were assessed by real time PCR. IL-6 protein was detected by ELISA. HSC a-smooth muscle actin (a-SMA) and MAPK (pERK1/2 and pp38) was analyzed by immunoblot. H 2 O 2 concentration in medium was assessed by H2DCF-DA. Results: Acetaldehyde enhanced collagen I, TIMP1 and TGFb1 mRNA expression in passaged HSCs, but not in primary cells. Leptin did not augment acetaldehydeinduced gene expression of the above proteins. In contrast, leptin plus acetaldehyde dramatically increased the gene and protein expression of IL-6 and a SMA (5 and 1.8-fold, respectively vs. controls, P \ 0.01 and 0.05, respectively). Furthermore, combination treatment activated HSC phosphorylation of MAPK (ERK1/2 and p38). Finally, IL-6 antibody neutralization attenuated the treatment-induced increase of a SMA and MAPK (ERK1/2 and p38). Conclusion: Leptin potentiates acetaldehyde-induced HSC activation, in part, due to increased IL-6 expression and IL-6-mediated downstream p38 and pERK1/2 activation. Objective: To study the protective effect of Blueberry against rat hepatic fibrosis and the effect of Blueberry on HO-1 expression pattern. Methods: SD rats were randomly divided into five groups namely control group, model group, blueberry group, Dan-shao-hua-xian (DSHX) capsule group and blueberry + Dan-shao-hua-xian group. Fibrous liver models in rats were induced by subcutaneous injection of CCl 4 and high-lipid/low-protein diet for 8 weeks except control group. Then, measuring the activity of the serum alanine aminotransferase (ALT), and superoxide dismutase (SOD) and malondialdehyde (MDA) of liver homogenate were determined. Observing the pathologic change of liver tissue. The expression of HO-1 was examined by real-time RT-PCR technique, Immunohistochemical staining and Western Blot.

Results: Serum ALT of every prevention group lower than model group (P \ 0.05), SOD of liver homogenate in prevention group were significantly higher and MDA were lower compared with the model group (P \ 0.05). And stage of hepatic fibrosis were all significantly reduced in prevention group. Expressions of HO-1 increased in model controls comparing with the control group (P \ 0.05), compared with model controls, the expression of HO-1 increased in prevention groups B, but increases were not significant. Conclusion: Blueberry had a significant prevention effect on rat liver fibrosis, its mechanism may be not through the induction of HO-1 expression. Background: To investigate the effect mechanism of CK8, CK18 and TCTP on pathogenesis of hepatic fibrosis based on proteomics research. Methods: Rats were randomly assigned to two groups: (1) normal control group (N, n = 8); (2) Hepatic fibrosis control group (M, n = 22). Hepatic histology and Hyp contain were observed dynamically. Using two-dimensional gel electrophoresis in conjunction with MALDI-TOF/TOF mass spectrometry, then the protein expression of CK8, CK18 and TCTP was measured by methods of Western blot and immunohistochemistry. Results: Fibrosis degree of M group was aggravated gradually compared with that N group. The protein expression of CK8, CK18 and TCTP increased gradually respectively, while there was a obvious positive correlation between the protein expression of CK8/CK18 and fibrosis degree (r = 0.9583 or 0.9909, P \ 0.01). In parallel with a significant positive correlation existed between the protein expression of CK8/CK18 and TCTP (r = 0.8805 or 0.9789, P \ 0.01).

Hepatol Int (2010) 4:94-345 239

Conclusions: There was a hepatocyte phenotype change in a rat model of hepatic fibrosis induced by CCl 4 , it may be a key pathological factor for hepatic fibrosis formation, and TCTP maybe an important regulation factor. Background/Aim: Bupleurum scorzonerifolium (BS), a traditional Chinese herb, has been widely used to treat liver diseases. Hepatic stellate cells (HSCs), the key cell type that regulates liver fibrosis, become activated and profibrogenic through phagocytosis of hepatocytic apoptotic bodies (ABs). Using an activity-guided fractionation, a lignan, kaerophyllin was isolated from BS. The aim of this study is to determine the anti-fibrotic potential of kaerophyllin and its possible mechanisms of action. Methods: Liver fibrosis in SD rats was induced by thioacetamide (TAA; 200 mg/kg, i.p.) injection twice weekly for 6 weeks. Two groups of rats received either high or low doses (15 and 30 mg/kg, twice daily) of kaerophyllin by gavage. LX-2, a human hepatic stellate cell line, incubated in the presence of UV-induced HepG2 ABs was used to investigate the engulfment of ABs by HSCs and the role of kaerophyllin in the inhibition of HSCs activation. Results: Phagocytosis of ABs by LX-2 induced activation, with production of collagen I and a-smooth muscle actin (a-SMA), increased motility and NF-jB activity. Kaerophyllin inhibited LX-2 phagocytosis of ABs and activation. TAA administration induced liver fibrosis, which was attenuated by kaerophyllin treatment, with reduction of GOT and GPT levels and a-SMA protein expression. Conclusions: Kaerophyllin reduced TAA-induced liver fibrosis and inhibited HSC activation and engulfment of ABs. Background: To investigate the source of TGF-b 1 , which is the key regulation factor in the process of hepatocytes transdifferentiated into bile duct epithelial cells (BECs).

Methods: Rats were randomly assigned to three groups: (1) sham control group; (2) BDL group. Rats were sacrificed at 1, 2 and 3 weeks. The protein expression of Hep Par, CK 7 and TGF-b1 were measured by Western blot. Hepatic histology were observed by HE and Sirius red stain. Double immunofluorescences of HepPar/CK 7 and HepPar/ TGF-b 1 were performed on frozen sections. Results: Hepatocytes of liver tissues were decrease, BECs were increased in parallel with fibrosis aggradation were increased gradually. Protein expression of Hep Par was decreased while that of CK7 was increased gradually. Protein expression of TGF-b 1 was detected increased after BDL and reached peak after BDL 2 weeks. Colocalized positive cells of Hep Par/CK 7 were founded increased gradually and colocalized positive cells of Hep Par/ TGF-b1 were observed (a majority of these cells were colocalized of TGF-b1 and Hep Par). Conclusions: Hepatocytes transdifferentiation maybe mediated by TGF-b 1 autocrined in hepatocytes after BDL rats. Background and Aims: Hepatitis C virus (HCV)-infected patients are at risk of progressing to liver fibrosis and cirrhosis. HCV core protein, which is secreted by infected hepatocytes, may interact with and thereby activate hepatic stellate cells (HSCs). In this study, we investigated the effects of HCV core protein on HSCs and the underlying mechanisms. Methods: LX-2, a human hepatic stellate cell line, was incubated with recombinant core protein with or without inhibitors. Phosphorylation of Janus kinase (JAK) 2, signal transducer and activator of transcription (STAT) 3, expression of a-smooth muscle actin (a-SMA), subunits of NADPH oxidase including gp91 and p47 were determined by Western blotting. The mRNA expressions of matrix metalloproteinase (MMP)-1 and tissue inhibitor of metalloproteinase (TIMP)-1 were measured by real-time polymerase chain reaction. ROS formation was assayed by fluorometer. Mitochondrial membrane potential was analyzed by flow cytometry and fluoromicroscope. Results: HCV core protein stimulated ROS formation, mitochondrial membrane potential, mRNA expression of MMP-1, and protein expressions of a-SMA, gp91 and p47 in LX-2. Phosphorylations of JAK2 and STAT3 were also significantly enhanced by HCV core protein, which were, reversed by JAK and NADPH oxidase inhibitors. NADPH oxidase inhibitor, but not JAK inhibitor, decreased the expressions of a-SMA and gp91. Conclusion: HCV core protein induced oxidative stress and JAK2-STAT3 signaling pathway, contributing to HSC activation. Background and Aim: Xiaozhang Cataplasm had adjuvant effect to reduce ascites as umbilicus plaster in cirrhotic patients. To investigate the change of ascites and gastrointestinal motility induced by Xiaozhang Cataplasm plastered on the navel of rats with dimethylnitrosamine (DMN)-induced cirrhosis and obvious ascites. Methods: Seventy wistar rats were randomly divided into six groups. Groups 4 to 6 (n = 12/group), the animals were given intraperitoneal injection of DMN with a dosage of 10 lg/kg of body weight per day for three consecutive days per week for 4 weeks. Groups 1 to 3 (n = 8/group) served as normal control groups, in which the rats were given no DMN. At the end of week 4, Xiaozhang Cataplasm were given to group 6 by umbilical sticking by dressing change every day for 1 week. Group 5 were served as model comfort group receiving blank cataplasm, Group 4 were served as model control given no Cataplasm. Groups 3 to 1 were respectively served as normal control. At the end of week 6, body weight, abdominal circumference, volume of ascites, the liver function including serum ALT, AST activity, Alb and total bilirubin (TBil) level, the residual rates of stomach contents and small intestine motional rate in all rats of groups 1 to 6 were observed. Results: During the process of DMN intoxication, four rats died in groups 4 and 5, and two rats died in groups 6. Compared with normal control groups rats, body weight, Alb content and small intestine motional rate decreased significantly (P \ 0.05). However, volume of ascites, abdominal circumference, TBil level and residual rate of stomach contents obviously rose (P \ 0.05). Compared with group 4 (model control group) and group 5 (blank cataplasm group), group 6 (Xiaozhang cataplasm group) had lower residual rates of stomach contents (P \ 0.01) and higher small intestine motional rate (P \ 0.05). Conclusion: Xiaozhang cataplasm showed good effects in promoting subsidise of the ascites due to liver Cirrhosis induced by dimethylnitrosamine.

Sang-Won Hong 1 , Guang-Yong Li 1 , Soon-Sun Hong 1 1 School of Medicine, Inha University, 7-241, 3-Ga, Shinheung-dong, Jung-gu, Incheon 400-712, Korea

Fucoidan is a sulfated polysaccharide extracted from brown seaweeds which possess a wide range of pharmacological properties including potent antioxidative effects. In this study, we investigated the protective effects of fucoidan on dimethylnitrosamine (DMN)-induced liver fibrosis in rats. Oral administration of fucoidan prevented weight loss in the body and liver by the treatment of DMN and inhibited the elevation of serum alanine transaminase, aspartate transaminase, total and direct bilirubin levels. Levels of malondialdehyde, superoxide dismutase, glutathione peroxidase and those relative gene expression show that fucoidan protects oxidative hepatic stress. From the evaluation of hepatic fibrosis related factors, we observed that fucoidan significantly reduced the expressions of collagen type I, transforming growth factor b1, and a-smooth muscle actin in mRNA and protein levels. In addition, significant decrease of collagen accumulation was observed by Masson's trichrome staining and hepatic hydroxyproline assay. And the expression of tissue inhibitor of metalloproteinase I, matrix metallopeptidase 9 and TGF-b/Smad also showed decreased hepatic fibrosis by fucoidan treatment. These suggest that fucoidan may be useful in preventing the development of hepatic fibrosis and cirrhosis.

Hepatol Int (2010) Background: Hepatic fibrosis is an important pathological change seen in chronic liver diseases. We observed the effects of ZhengganHuayu (ZGHY) decoction on the pathological changes of hepatic fibrosis in rats in order to probe its effect and mechanism on delay, blockage and inversion of hepatic fibrosis. Methods: 36 SD rats were randomly divided into three groups: Normal group, model group and ZGHY group, respectively. Except the rats in the normal group, all the other rats in the other two groups were administered 40% CCl 4 olive oil solution 5 ml/kg at the nape subcutaneously injection first-time; Then 3 ml/kg T.I.W. while rats of normal group were subcutaneously administered 0.9% saltwater of the same dose. At the end of the sixth week, the general conditions of the livers were observed after the bellies were cut open. Blood samples were taken through inferior vena cava. Liver tissues were stained with HE, Sirius and Uranium-lead respectively and the pathological and ultrastructural changes were observed through optical microscope or transmission electron microscope.

Results: Regarding the model group, hepatic fibrosis was obviously formed, the structure of hepatic lobules were damaged and the contents and deposit of the collagens in the liver tissues were obviously increased. The score of the ZGHY group was lower than that of the model group according to SSS scoring system. Mitochondria, lysosome and other organelles were seriously damaged, the collagens in the Disse gap were significantly increased; While in the ZGHY group, the structure of the hepatic lobules were nearly normal, the collagens in the Disse gap were significantly decreased compared to those of the model group.

Conclusions: ZGHY decoction can relieve both the hepatic cellular inflammation and damage to the mitochondria, inhibit the fibroplasia and deposition and alleviate hepatic fibrosis. Background/Aims: Elevated tissue inhibitor of metalloproteinase (TIMP)-1 expression contributes to excess production of extracellular matrix in liver fibrosis. The present study constructed a recombinant adeno-associated virus (AAV) carrying siRNAs of TIMP-1 and investigated the effects of administration of this recombinant AAV in carbon tetrachloride (CCl4)-induced rat liver fibrosis model. Methods: Five siRNA oligomers targeting rat TIMP-1 were designed and transfected into rat hepatic stellate cells (HSC)-T6. A U6 promoter followed by the siRNA which had the strongest suppression effect was cloned into the AAV vector and packed into 293 cells to construct the recombinant AAV/siRNA-TIMP-1. Olive oil CCl4 solution 40% (2 mL/kg), or olive oil control, was administered hypodermically to Wistar rats twice each week for 6 weeks. Rats were randomly divided into CCl 4 control group and recombinant AAV/siRNA-TIMP-1 administration group, which were administered the recombinant AAV at the end of first week and second week. At the sixth weekend, the rats were sacrificed for detection of histological changes of liver tissue, liver tissue hydroxyproline content and expression of TIMP-1. Moreover, the expression level of the TIMP-1 was detected at 12 weeks after the infection of the recombinant AAV in HSC-T6. Results: Three of the five designed siRNA oligomers had a suppressing effect on TIMP-1 expression in rat HSCs within 72 h. AAV/siRNA-TIMP-1 administration attenuated fibrosis severity, as determined by fibrosis scores (3.5 ± 0.9) compared with the CCl 4 -treated group (2.6 ± 0.5) (P = 0.02). Compared to controls, the hydroxyproline content and TIMP-1 expression of liver tissue in AAV/siRNA-TIMP-1 administration group was decreased by 22 and 45%, respectively. Furthermore, in vitro inhibition of TIMP-1 could last 12 weeks and decreased 90%. Conclusions: RNA interference exerts suppressive effect on the TIMP-1 gene in cultured HSCs for a longer time when a recombinant AAV is utilized as the gene delivery vector. Administration of AAV/siRNA-TIMP-1 attenuated CCl4induced rat liver fibrosis and decreased TIMP-1 expression. Background: No specific therapy is currently available to treat liver fibrosis, a common complication associated with chronic liver disease. Although rapamycin has been shown to inhibit liver fibrosis in rats, the exact mechanisms of this treatment remain unclear. Methods: Hepatic liver fibrosis was induced in rats by CCL 4 administration for 8 weeks. 39 Rats were divided into three groups: CCL 4 -induced fibrotic rats then treated for 2 weeks with 2 mg/kg/day rapamycin (Rapa) or 2 ml/kg/day 0.9% NaCl (Control), and left untreated (Normal). Primary hepatocytes and liver slices were isolated and their protein expression of p70S6K and TGF-b1pathway proteins was examined using immunohistochemistry and ELISA. Statistical significance of the expression levels between groups was defined using analysis of variance as P \ 0.05. Results: Rapamycin successfully inhibited rat liver fibrosis, as evidenced by a reduced volume of connective tissue and decreased AST (ASpartate amino-Transferase) levels in Rapa rats compared to Control or Normal animals. Fibrosis increased the expression of the EMT marker vimentin, and this effect was reversed in Rapa animals. The expression levels of several Smads (p-smad2, p-smad3, smad 2/3, and smad 4) and p-p70S6K were higher in Control than in Normal rats. Rapamycin treatment eliminated these upregulations.

Conclusions: This study is the first to our knowledge to show that rapamycin inhibits liver fibrosis by blocking the EMT, potentially through a pathway involving Smads and p70S6K. Background/aims: Hepatic stellate cells (HSCs) are pericytes of liver sinusoidal endothelial cells. Activation of HSCs into a myofibroblast-like phenotype is involved in several hepatic disease processes. The alpha-smooth muscle actin is a well known marker of hepatic stellate cell activation. It has been known that activated HSCs are the main matrix-producing cells and play an important role in the progress of liver cirrhosis. The aim of the present study was to immunolocalize muscarinic receptors in the activated HSCs of thioacetamide-induced cirrhotic rat liver. Methods: Antibodies against M1-M5 muscarinic receptors was used for immunohistochemical study in the cirrhotic liver sections. Results: In the present study, we could demonstrate immunochemically that the activated HSCs in thioacetamide-induced cirrhotic liver, which are immunopositive for alpha smooth muscle actin, express M3 muscarinic receptor but not M1, M2, M4 and M5. Conclusion: These finding suggest that M3 muscarinic receptor might be involved in triggering the intracellular signaling pathways in activated HSCs leading to the production of collagen fibers. If this assumption appears to be true this mechanism may be of future interest in therapy of hepatic fibrosis. Background: The relationship between HBV carrier state and liver fibrosis are incompletely understood, and lack a mouse model. We now develop HBVassociated liver fibrosis in HBV transgenic (HBV-tg) mice which resembles human inactive HBV carriers, and investigate the immune response contributing to liver fibrosis. Methods: We performed comparative investigations between C57BL/6 and HBV-tg mice. For acute liver injury, C57BL/6 and HBV-tg mice were injected with a single hepatotoxin carbon tetrachloride (CCl 4 ). For chronic liver injury and liver fibrosis, mice were injected with biweekly CCl 4 as long as 10 and 14 weeks. The histological changes of liver injury and fibrosis were visualized in sections stained by hematoxylin-eosin and sirius red. Liver samples from both mice were evaluated for mRNA expression of fibrosis related markers. Results: Liver fibrosis spontaneously occurred in old HBV-tg mice. After CCl 4 injections, there were much more liver injury and liver fibrosis in HBV-tg mice compared with C57BL/6 mice. We found the collagen-producing hepatic stellate cells (HSCs) remarkably over-activated in HBV-tg mice after chronic CCl 4 administration. The immune dysfunction correlated with progressive liver fibrosis in HBV-tg mice. Over-activation of liver HSCs in CCl 4 treated HBV-tg mice was largely ameliorated by natural killer T (NKT) cells depletion, but not by natural killer (NK) cells depletion. More importantly, adoptive transfer of purified liver NKT cells from HBV-tg mice into Rag1 -/mice led to accelerated HSCs activation, emphasizing a prominent role for NKT cell-mediated liver fibrosis.

Conclusions: HBV-tg mice may be used as a good model to study HBV-related liver fibrosis, and NKT cells play a critical role in HSCs activation in HBV-tg mice. Background: Resveratrol, a representative antioxidant derived from grape, has been reported to show widespread pharmacological properties. In this study, we investigated the protective effect of resveratrol on dimethylnitrosamine (DMN)-induced liver fibrosis in rats. Methods: Rats were given a single intraperitoneal injection of DMN (40 mg/ kg) and treated with resveratrol daily by oral gavage for 7 days. Results: Resveratrol remarkably recovered the body weight loss and liver weight in DMN-induced liver fibrosis. Liver histology showed that resveratrol alleviated the infiltration of inflammatory cells and fibrosis of liver tissue. Also, resveratrol decreased the level of malondialdehyde (MDA) and increased the levels of glutathione peroxidase (GPx) and superoxide dismutase (SOD). Resveratrol significantly inhibited the mRNA expression of transforming growth factor b 1 (TGF-b 1 ), collagen type I and a-smooth muscle actin (a-SMA). In addition, there was significant decrease of hydroxyproline by resveratrol in DMN-induced liver fibrosis. Conclusions: Our results suggest that resveratrol could improve DMN-induced liver injury and fibrosis and may be useful in preventing the development of liver fibrosis and cirrhosis from liver damage. Background and Objective: The herbal ingredients formula (CGA) composed of Cordyceps sinensis polysaccharide (C), Gypenosides (G) and Amygdaloside (A) was found to inhibit liver fibrosis significantly at the proportion of 6:8:5 by uniform design in our previous research. To illuminate the mechanism and Compatibility significance of CGA, the present study compared CGA to each ingredient on the anti-liver fibrosis effects Methods: DMN solution (0.5%) was injected to rats for three consecutive days per week for 3 weeks. Rats were randomly divided into vehicle groups, DMN control group, C, G, A, and CGA groups. Each group was orally administered with specific decoctions daily for 3 weeks. Rats in the vehicle group were orally administered with only water for 3 weeks. Results: The Hyp content in vehicle groups, DMN control group, C, G, A, and CGA groups were 165 ± 35, 435 ± 91, 338 ± 70, 318 ± 54, 323 ± 79,254 ± 39 lg/g of liver tissue, suggesting that each ingredient had a anti-liver fibrosis effect,however, CGA was more effective (compared with each Ingredients P \ 0.05). Compared with control rats, the protein expressions of a-SMA, TIMP-1,TIMP-2, MMP13 in DMN rats were all significantly increased. The protein expressions of a-SMA and MMP13 were reduced by C and A treatment. The protein expressions of TIMP-1 were reduced by C treatment. Compared with DMN rats, The protein expressions of a-SMA, TIMP-1, TIMP-2, MMP13 were all significantly reduced in CGA rats. Conclusion: Our results show that CGA treatment exerted antifibrotic effects,which was related to inhibition of HSC activation, regulation of collagen metabolism. There was synergetic effect among each ingredient. Background and aim: Therapeutic options for the treatment of hepatic fibrosis are limited. Picrorhiza kurrooa (Pk), an herb found in the north western parts of Himalaya, has proven hepatoprotective efficacy, however its anti fibrotic potential is not yet evaluated. The study is designed to evaluate the therapeutic potentials of Pk extract against liver fibrosis in vitro using Human hepatic stellate cell (HSC) LX2, which is responsible for increased synthesis and deposition of collagen in the liver. Methods: Crude root extracts of Pk was prepared. LX2 cell line was maintained in DMEM medium with 2% FBS in vitro. The cells were activated in-vitro by environmental toxin arsenic (2.5 lmol) for in vitro. After activation, the LX2 cells were treated with Pk extracts (200 lg) for 1-5 days. Cell proliferation was evaluated by MTT assay and cell apoptosis was detected by TUNEL staining, annexin-V/propidium labelled flow cytometry. Western blot analysis was done using BCl2, Bax, AKT and beta actin. Results: Arsenic enhanced the proliferation of LX2 cells in vitro. Addition of root extracts of Pk hindered the arsenic induced proliferation of LX2 cell line. The absolute enhancement of growth inhibition rates was significant compared to the control group and arsenic treated group. The apoptosis rates in Pk and Pk plus arsenic groups were significantly increased compared with control and arsenic groups. These effects were associated with complete down regulation of the antiapoptotic protein Bcl-2 and upregulation of the apoptotic protein Bax. Further Pk significantly inhibited Akt activation in LX2 cells. Conclusion: Root extracts of Pk can inhibit proliferation of activated LX2 cells in vitro and shows a proapoptototic effect on HSCs Bcl2/ Bax pathway as well as Akt inactivation. This study suggests root extracts of Pk as a novel anti fibrotic agent. Background and Aims: N-acetylcysteine (NAC) is potential antioxidant and provides significant protection against a broad array of modern toxins. It well recognized that NAC would benefit liver damage and fibrosis. Hypoxia is very closely associated with hepatic stellate cell activation and fibrogenesis. We found that NAC had a good action against liver fibrosis in our previous study, here we investigate NAC effect on hypoxia HSC cell line-T6 induced by CoCl 2 . Methods: HSC-T6 was cultured and incubated with 10-200lM of CoCl 2 for 4-24 h, and cell growth was assayed with MTT. Then CoCl 2 intoxicated HSC-T6 with or without serum supplementation, was incubated with 1 mM NAC, and cell growth was observed as the above too. Results: Compared to the normally cultured HSC-T6 cells, CoCl 2 inhibits HSC-T6 growth in the dosage and time dependent manner, the maximum effect CoCl 2 on HSC-T6 is almost at 150 lM for 16 h incubation. NAC could counteract CoCl 2 induced HSC-T6 suppressing growth. Conclusion: CoCl 2 suppressed HSC-T6 growth in vitro, and NAC counteracted this suppression. Background and Aim: Hepatic Stellate Cells (HSC) transdifferentiate into myofibroblastic cells, which are the main producers of extracellular matrix components in hepatic fibrogenesis. Myofibroblastic transition is associated with an altered profile of growth factors and mediators of inter-and intracellular cell communication. Therefore, HSC express a wide spectrum of neuronal signal transducers after myofibroblastic transition. Since microRNAs, which are endogenous small single-stranded non-coding RNAs, inhibit gene expression by interaction with the untranslated region (UTR) of many transcripts, we studied the role of miRNAs known to be involved in neuronal development, during HSC transdifferentiation. Methods: Primary HSCs were isolated from male Wistar rats and cultured for up to 10 days. The RNA extraction was done by the Trizol method and a polyadenylation combined reverse transcription was performed. The neuronal miRNAs miR-9, miR-125b and miR-128 were quantified by Real-Time PCR using the miScript system of Qiagen. Putative miRNA targets were validated in reporter assays by cloning the binding site of the 3 0 -UTR in fusion to the coding sequence of a luciferase reporter gene. Results: All three neuronal miRNAs miR-9, miR-125b and miR-128 were upregulated during myofibroblastic HSC differentiation. Screening of databases like Pictar, miRanda and TargetScan, revealed putative binding sites of the miRNAs in transcripts encoding for several pluripotency factors such as Lin-28 and Klf4. For validation of the target, reporter assays were performed of the cloned wildtype in comparison to corresponding mutated sequences carrying two point mutations in the binding consensus. We identified LIN28, which triggers pluripotency in embryonic stem cells, as target for miR-9, miR-125b and miR-128. Conclusion: In conclusion, the neuronal miRNAs miR-9, miR-125b and miR-128 are indicated to take part in myofibroblastic transition as well as expression and neuronal features of HSC by targeting Lin28 transcripts. 

Background and Aim of the Work: The accurate diagnosis of hepatitis C virus (HCV)-related fibrosis is crucial for prognosis and treatment decisions. Due to the limitations of liver biopsy, noninvasive alternatives including Fibroscan have been developed. This study was assigned to compare Liver Stiffness Measurement (Fibroscan) with hepatic fibrosis assessed by histologic examination in patients with chronic hepatitis C genotype 4. Patients and Methods: The study included 120 chronic HCV patients with genotype 4. They were subjected to liver biopsy and FibroScan examination. Liver fibrosis was staged on a 0-4 scale according to the METAVIR system. The diagnostic performance of FibroScan was assessed by analysis of the receiver operator characteristics (ROC) curve. Results: The area under ROC curve of FibroScan for significant fibrosis was 0.87 (95% confidence interval, 0.801-0.940). The diagnostic accuracy of fi-broScan was found to be 75.8 with 84.6% sensitivity and 76.5% specificity. The positive predictive value was 63.5% and the negative predictive value was 91.2%. The identical matching, overestimation and underestimation of Fibroscan to liver biopsy were 35, 36.7 and 28.3% in overall patients; 32.1, 35.8 and 32.1% in patients with early fibrosis (F0-F2); and 41, 38.5 and 20.5% in patients with advanced fibrosis or cirrhosis (F3, F4). Conclusion: This study suggests that, Fibroscan is a promising method for the diagnosis of hepatic fibrosis in patients with chronic hepatitis C genotype 4.

Top 10% Posters factors other than the fibrosis stage, including hepatic necroinflammation. A recent study suggested that LSMs elevated in hepatic congestion. Because respiration affects intrathoracic pressure and hepatic venous return, LSMs could be affected by respiration. This study was performed to evaluate the influence of respiration on the LSMs. Methods: Patients were requested to breathe freely during examination. LSMs were performed at the end of inspiration and then, at the end of expiration. The median values obtained during inspiration and those obtained during expiration were defined as iLSM and eLSM, respectively. Significant difference in LSMs was defined as higher LSM -IQR [ lower LSM + IQR. Results: A total of 93 patients with chronic liver disease were enrolled. Mean age was 38 years and 66 patients (71%) were men. Liver cirrhosis was combined in 14 patients (15%). eLSM (11.0 ± 8.8 kPa) was significantly higher than iLSM (10.6 ± 9.2 kPa) (P \ 0.001), while there was no significant difference between iLSM/iIQR (0.1 ± 0.1) and eLSM/eIQR (0.1 ± 0.1) (P = 0.441). eLSM was higher than iLSM in 58 patients (62.4%), lower in 31 (33.3%), and same in 4 (4.3%). eLSM was significantly higher than iLSM in 22 patients (23.7%), while iLSM was not significantly higher in any patient. When baseline characteristics and laboratory data were compared between patients with significant differences in LSMs and patients without significant differences in LSMs, only the proportion of patients with liver cirrhosis showed significant difference between two groups (P = 0.024). Conclusions: LSMs were significantly elevated during expiration in some patients. These results might suggest that fibrosis stage could be overestimated if LSMs were performed during expiration.

Gamal Shiha 1 , Ayman Eldesoky 1 , Khaled Zalata 1 , Waleed Samir 1 , Amira Elbeeh 2 1 Mansoura University, Egypt, 2 Biotechnology Research Center, New Damitta

Background and Aim of the Work: The accurate diagnosis of hepatitis C virus (HCV)-related fibrosis is crucial for prognosis and treatment decisions. Due to the limitations of liver biopsy, noninvasive alternatives including Fibroscan have been developed. This study was assigned to compare Liver Stiffness Measurement (Fibroscan) with hepatic fibrosis assessed by histologic examination in patients with chronic hepatitis C genotype 4. Patients and Methods: The study included 120 chronic HCV patients with genotype 4. They were subjected to liver biopsy and FibroScan examination. Liver fibrosis was staged on a 0-4 scale according to the METAVIR system. The diagnostic performance of FibroScan was assessed by analysis of the receiver operator characteristics (ROC) curve. Results: The area under ROC curve of FibroScan for significant fibrosis was 0.87 (95% confidence interval, 0.801-0.940). The diagnostic accuracy of Fi-broScan was found to be 75.8 with 84.6% sensitivity and 76.5% specificity. The positive predictive value was 63.5% and the negative predictive value was 91.2%. The identical matching, overestimation and underestimation of Fibro-Scan to liver biopsy were 35, 36.7 and 28.3% in overall patients; 32. 1, 35.8 and 32 .1% in patients with early fibrosis (F0-F2); and 41, 38.5 and 20.5% in patients with advanced fibrosis or cirrhosis (F3,F4). Conclusion: This study suggests that, FibroScan is a promising method for the diagnosis of hepatic fibrosis in patients with chronic hepatitis C genotype 4. Seoul, Korea, Backgrounds/Aims: While liver stiffness measurement (LSM) predicts histological cirrhosis accurately, complementary methods are needed for the better performance. Furthermore, alanine aminotransferase (ALT) influences LSM, making it necessary to modify its use in patients with high ALT levels. We developed a new LSM-based prediction model for cirrhosis and estimated cutoffs for different ALT levels. Methods: From 2008 to 2009, we prospectively enrolled 330 consecutive patients who were diagnosed with chronic hepatitis B and underwent liver biopsy and LSM on the same day. For detection of cirrhosis, we performed univariate and multivariate analysis, using v 2 test/t test and logistic regression, respectively. Thereafter, a prediction model was derived from multivariate predictors. Results: In multivariate analyses of patients with and without cirrhosis, we found significant differences in LSM, spleen diameter and platelet count. Then, we developed a LSM-Spleen diameter to Platelet ratio Index (LSPI): (LSM 9 Spleen diameter/Platelet count) 9 100. The area under the receiver operating curve was 0.956, significantly higher than LSM alone (0.919, P = 0.032). We suggested different cut-offs in patients with ALTBupper limit of normal (ULN) (NALT group, 164 patients) and ALT [ ULN (HALT group, 166 patients). In NALT group, LSPI cut-offs of 38/62 provided 95.7% negative predictive value (NPV)/95.5% PPV (positive predictive value), while in HALT group, cut-offs of 42/94 yielded 95.1% NPV/96.4% PPV, respectively. Therefore, liver biopsy could be avoided in 76.7% of subjects. Conclusions: LSPI is a reasonable, noninvasive tool that can replace liver biopsy in assessment of liver fibrosis among the lowest and highest-risk patients. Background: Assessment of liver fibrosis is important for deciding the treatment and predicting the prognosis of patients. Liver biopsy is a gold standard for assessing hepatic fibrosis. However, this method frequently has a risk of severe complications. We evaluated the clinical accuracy of fibroscan, serum hyaluronic acid, serum retinol-binding protein 4, and serum transferrin as a simple, non-invasive marker for predicting hepatic fibrosis in chronic liver disease.

Methods: We enrolled 132 patients with chronic liver disease. Fibroscan and liver biopsy were performed on all patients. Fibrosis stage was assessed according to the Batts-Ludwig scoring system. Results: The mean value of liver stiffness measured by fibroscan in each stage group (F0, F1, F2, F3, and F4) was 7.2 ± 3.7, 8.2 ± 5.7, 10.4 ± 6.9, 13.3 ± 10.3 and 22.6 ± 12.0 kPa. Liver stiffness was significantly correlated to the fibrosis stage(r = 0.530, P = 0.000). The AUROC (95% CI) of C F2, C F3 and F4 was 0.760(0.70-0.84), 0.785 (0.71-0.87) and 0.865 (0.80-0.94). Serum hyaluronic acid, serum retinol-binding protein 4, and serum transferrin also showed a significant correlation(r = 0.428; P = 0.000, r = -0.280; P = 0.030, r = -0.361; p = 0.001) with hepatic fibrosis. Conclusions: Fibroscan is a reliable, useful method for assessing liver fibrosis in patients with chronic liver disease in addition to using serum hyaluronic acid, serum retinol-binding protein 4, and serum transferrin.

Liver Stiffness Measurement Using Acoustic Radiation Force Impulse (ARFI) in Chronic Liver Disease Ki Tae Yoon 1 , Jeong Heo 2 , Hyun Young Woo 2 , Mong Cho 1 1 Pusan National University Yangsan Hospital, Yangsan, South Korea, 2 Pusan National University Hospital, Busan, South Korea Background/aims: Acoustic Radiation Force Impulse (ARFI Ò , Siemens) is a new technology to measure liver stiffness and this allow the assessment of liver fibrosis. The aim of this study was to evaluate the diagnostic accuracy of a new devise using ARFI technology and to compare with other markers such as the aspartate aminotransferase to platelet ratio index (APRI) and aspartate to alanine aminotransferase (AST/ALT) ratio. Methods: We enrolled consecutive 38 patients (M/F = 28/10) who performed the liver biopsy for evaluating cause and severity of chronic liver diseases. The fibrosis stage was determined by two independent pathologists according to the METAVIR classification. Liver stiffness measurement (LSM) was performed five times at the same site on the day of liver biopsy procedure. Results: The LSM could be obtained in all patients without measurement failure. The etiology of liver diseases included HBV infection (n = 8), HCV infection (n = 4), autoimmune hepatitis (n = 12), PBC (n = 2), NASH (n = 2), Wilson's disease (n = 1), and unknown origin (n = 9). According to the fibrosis stage, the median values of LSM were 1.63 ±0.47 for F0 (n = 4), 1.83 ± 0.30 for F1 (n = 8), 1.65 ± 0.42 for F2 (n = 15), 2.11 ± 0.60 for F3 (n = 10), and 2.93 for F4 (n = 1), respectively. Spearman correlation coefficient was 0.402 (P = 0.012). According to the Area under the curve of receiver operation characteristic (AUROC) analysis for different fibrosis thresholds, ARFI, APRI and AST/ALT ratio showed 0.697, 0.768, and 0.522 for CF2, 0.714, 0.524, and 0.616 for CF3, and 0.919, 0.054, and 0.811 for CF4, respectively. Conclusions: ARFI is a reliable, rapid non-invasive method to diagnosis the severity of fibrosis in patients with chronic liver disease. Background: To evaluate the clinical significance of serum Vascular Endothelial Growth Factor (VEGF) and Model for End-stage Liver Disease (MELD) in predicting prognosis of patients with decompensated liver cirrhosis (DLC). Methods: Serum VEGF, TBI, PT and CR were detected in 84 patients with DLC. The correlations between serum VEGF and MELD and the prognosis of them were analyzed. Results: The concentrations of Serum VEGF were (255.52 ± 92.27) pg/ml, (130.33 ± 100.28) pg/ml in died patients or the survivals with DLC respectively. The score of MELD were (25.84 ± 7.32), (15.00 ± 4.91) in died patients or the survivals with DLC respectively. Serum VEGF and MELD were both higher in the died DLC patients than those in the survivals (P \ 0.005).There was a positive correlation between serum VEGF and MELD (r = 0.444, P \ 0.0001 Introduction: Non-invasive, i.e. serum-based assessment of liver fibrosis is still an important challenge although multiple single and multiparametric panels of biomarkers have been proposed. Aim and Methods: (a) Haptoglobin, ALT, gammaGT, alpha 2-macroglobulin, apolipoprotein A1 and bilirubin in sera of 4 patients with histological proven fibrosis (F1-F4, A1-A3) were determined in 6 different quality-controlled laboratories. Inter-laboratory variations of the calculated Fibrotest Score for staging and Actitest Score for grading (both BioPredictive TM ), and their error ratios compared to the results obtained by biopsy were calculated. (b) The variability of obtained Fibrotest/Actitest Scores depending on 64 differential combinations of the allowed analyt-specific maximum/minimum permissible values as determined by the external quality control of the German Association of Clinical Chemistry and Laboratory Medicine (DGKL) was determined and the frequency distribution of the results calculated. Results: (a) Fibrotest and Actitest Scores were largely reproducible among the different laboratories. However, the error ratio was 77% for all results calculated by both, Fibrotest and Actitest when compared to the histological findings. (b) Calculated scores varied among F2 (9%), F3 (31%), F3-F4 (6%), and F4 (54%) (Fibrotest), as well as A1/A2 (48%), A2 (9%), A2-A3 (5%), and A3 (38%) (Actitest). Conclusion: Despite reproducibility of Fibro-and Actitest results among the six laboratories, large scale investigation (n = 64) displayed increasing variability of the results depending on interlaboratory differences that were still in a quality controlled, analytically acceptable range. Furthermore, calculated scores coincided with histological findings only in less than 25% of all cases. Thus, the diagnostic accuracy of these tests must be considered as low, if histology is accepted as gold standard. Background/aims: It has not been explicitly addressed whether new biomarkers, in addition to readily available clinical data, make a contribution to the prediction of liver fibrosis. Methods: A total of 209 patients with chronic hepatitis B who underwent liver biopsy were recruited. The Clinical Score (CS) model using only routine clinical data and the Biomarker Score (BS) model using seven putative biomarkers (a2-macroglobulin, haptoglobin, apolipoprotein A1, hyaluronic acid, Hepatol Int (2010) 4:94-345 249 matrix metalloproteinase-2, tissue inhibitor of metalloproteinase-1, procollagen III N-terminal peptide) were derived from the derivation set (n = 105), and these models were applied to a separate patients group (n = 104) to investigate whether the addition of BS improved the diagnostic accuracy in predicting significant fibrosis beyond an assessment based solely on CS. Results: The most informative biomarkers for predicting significant fibrosis were hyaluronic acid and matrix metalloproteinase-2. The BS was an independent predictive factor for significant fibrosis even after accounting for CS in both sets. Among the derivation set, the incorporation of the BS into the CS model did not significantly increase the ROC area, with only a small improvement of about 2% (P = 0.11 Objective: Currently, the application of metabolomics to liver disease was focused on the analysis of urine or serum. The feces offer a unique opportunity to reflect the cooperation between microbiome-mammalian metabolisms which is an essential element in the study of the human metabolon. Here, a global metabolomics approach was firstly used to find and identify metabolites differentially regulated in the feces of liver cirrhosis patients caused by hepatitis B virus (HBV). Methods: Seventeen HBV-induced liver cirrhosis patients and 24 healthy individuals were recruited. Fecal metabolite was detected through a nontargeted reversed-phase ultra performance liquid chromatography coupled to electrospray ionization quadrupole time of flight mass spectrometric metabolomics approach.

Results: A total of 9,215 peaks had been detected. Using unequal variance t tests, 2,393 peaks with P B 0.05, approximately 71.5% of which were the results of decreased fecal metabolite concentrations in liver cirrhosis versus healthy control, were observed. Integrating multivariate data analysis, we identified six major groups of metabolites: a dramatically decrease of bile acids and bile pigments were found in the patients, a strong increase of lysophosphatidylcholines, aromatic amino acids, fatty acids and acylcarnitines were found in the patients. Conclusions: The results suggested that a severe hepatic injury and malabsorption of nutrient substance and disorders of fatty acid occurred in the patients with liver cirrhosis. And the imbalance of gut microflora may play a role in metabolomic alterations in liver cirrhosis. Our findings contribute to a better understanding of clinical problems in liver cirrhosis. Background: As chronic liver disease progresses, an imbalance occurs between synthesis and breakdown of extracellular matrix (ECM). Matrix metalloproteinases (MMPs) are involved in degrading ECM while tissue inhibitors of metalloproteinases (TIMPs) prevent their fibrolytic action. We determined if serum levels of TIMP-1 and TIMP-2 are related to liver histology in patients with chronic hepatitis B. Methods: Commercially available enzyme-linked immunosorbent assay (ELISA) assays were used to study circulating values of TIMP-1 and TIMP-2 in patients with chronic hepatitis B (CHB; n = 159) and 20 healthy controls. Hepatic histology was evaluated using the Hepatitis-Activity-Index (HAI) according to Ishak et al. (J. Hepatol, 22:696-699, 1995) , quantifying separately inflammatory activity and fibrosis. Results: Serum levels of TIMP-1 and TIMP-2 were higher in the patients than those in the healthy controls, and had significant difference (P \ 0.05). Serum levels of TIMP-1 and TIMP-2 increased significantly with the development of fibrosis and had a positive correlations with HAI(r = 0.51, P \ 0.01; r = 0.42, P \ 0.01), whereas serum ALT was not related to fibrosis or to HAI. (10) 5.9-6.6 (F1) 8.3% (5) 15.4% (2) 6.7-9.7 (F2) 33.3% (20) 7.7% (1) 9. 8-17.4 (F3) 15.0% (9) 0 [17. 5 (F4) 3.3% (2) Background: To directly compare Fibroscan Ò , caffeine breath test (CBT)-a marker of hepatic cytochrome metabolism, and the serum derived ratios of aspartate aminotransferase platelet ratio index (APRI) and aspartate alanine aminotransferase ratio (AAR) in the detection of liver fibrosis in chronic hepatitis B patients. Methods: Five patients with chronic hepatitis B in the last 6 months who had a liver biopsy were enrolled into the study. Demographics, clinical history, relevant pathology and imaging results were collected. Valid Liver Stiffness Measurement (LSM) obtained from Fibroscan Ò required ten successful measurements in the mid axillary line. The plasma caffeine clearance using orally administered 13C-caffeine was obtained as per protocol. The reference ranges for LSM cut-offs correlating to stage of fibrosis were determined from local data. Conclusion: Fibroscan Ò was able to detect cirrhosis in 2/2 patients, whereas CBT and AAR detected 1/2 patients and APRI 0/2. CBT most frequently correlated to biopsy in 3/5 patients. AAR was able to correctly rule out cirrhosis in 3/5 patients but is not able to distinguish lesser degrees of fibrosis. APRI was correct in 2/5 patients. The results of this pilot study suggests that further research extended to a larger population is worthwhile, with analysis of whether non-invasive markers used in combination may improve assessment of liver fibrosis. Results:

Hepatol Int (2010) Aims: To investigate the relationship between anti-beta2-GPI antibodies and some autoantibodies and extracellular matrix in patients with post-hepatitis B liver diseases. Methods: A cohort of patients with chronic hepatitis B (CHB) and post-hepatitis B cirrhosis (PHBC) was studied for the serum anti-beta2-GPI antibodies levels by ELISA with purified beta2-GPI as antigen. The serum dsDNA, smooth muscle antibody (SMA), ribonucleoprotein antibody (RNPA), collagen IV (IV-C), hyaluronic acid (HA) and laminin (LN) were also detected by corresponding commercial kits. Results: High positive rate of anti-beta2-GPI antibodies was observed in patients with CHB or PHBC (20.9%, 9/43; 49.3%, 37/75) comparing with that in control group (3.1%, 1/32) (P \ 0.01) and anti-beta2-GPI antibodies level of PHBC group was higher than that of CHB group (P \ 0.05). High titer of autoantibodies was found in CHB and PHBC groups. The positive rate of anti-beta2-GPI antibodies was higher than those of dsDNA, SMA and RNPA (CHB: 4.7, 11.6, 9.3%; PHBC: 8.0, 22.7, and 14.7%). High serum levels of IV-C, HA and LN in patients with PHBC were found to be related to the elevated anti-beta2-GPI level.

Conclusions: HBV infection may contribute to the generation of anti-beta2-GPI antibodies. Beta2-GPI may play a role in the development of post-hepatitis B liver diseases. Background: Transient elastography (Fibroscan) is now a well recognized noninvasive method to evaluate fibrosis in patients with chronic hepatitis. Real time elastography has not yet an established role in this field. Aim: to evaluate the usefulness of real time elastography (RTE) in the assessment of the staging of liver fibrosis in chronic B and C hepatitis. Methods: Our study included 90 patients (52 females and 38 males, mean age 43.6 ± 12, 7 years), 60 with chronic hepatitis C and 30 with chronic hepatitis B. All the patients underwent liver biopsy (fibrosis stage was quantified according Metavir scoring system) and RTE Hitachi. In evaluation of the liver stiffness we calculated elastic ratio of the liver for the subcutaneous tissue; 10 RTE measurements were made in each patient and a mean value was calculated. Results: Stiffness of the liver was significantly lower as histological stages advanced. Median elastic ratio in each stage were: in chronic hepatitis C-F0: 1.40 ± 0.09 SD (N = 7); F1: 1.12 ± 0.07 SD (N = 13); F2:1.07 ± 0.09 SD (N = 21); F3: 0.88 ± 0.05 SD (N = 11); F4: 0.31±0.04 SD (N = 8); in chronic hepatitis B-F0: 1.27 ± 0.09 SD (N = 6); F1:1.18 ± 0.08 SD (N = 13); F2:1.02 ± 0.09 SD (N = 7); F3: 0.98 ± 0.08 SD (N = 2); F4: 0.51 ± 0.06 SD (N = 2). Conclusions: The elastic ratio correlated well to the advanced histological fibrosis staging (F4) in chronic hepatitis C but not so well in chronic hepatitis B. In both B and C chronic viral hepatitis the performance of method was low for discriminating mild fibrosis from significant liver fibrosis, which might limit the applicability of this technique in clinical practice. Background/Aims: Early detection of liver fibrosis in patients with chronic liver diseases is crucial to prevent cirrhotic complications. The aspartate aminotransferase (AST) to platelet ratio index (APRI) has been proposed as an inexpensive noninvasive marker to assess liver fibrosis. The aim of the present observational, cross-sectional study was to evaluate the diagnostic performance of APRI for fibrosis prediction in patients with chronic hepatitis B (CHB), chronic hepatitis C (CHC) and nonalcoholic fatty liver disease (NAFLD). Methods: This study included 207 patients with CHB, 108 patients with CHC, and 140 patients with NAFLD. The APRI index was calculated according to the following formula: APRI = [(AST level/upper limit of normal)/platelet counts (109/L)] 9 100. Stage of liver fibrosis in patients with chronic viral hepatitis was graded with the METAVIR scale. The Kleiner system for grading fibrosis was used in patients with NAFLD. Results: Simple correlational analysis revealed that the APRI correlated significantly with fibrosis scores in patients with CHC (r = 0.2634, P = 0.0059) and NAFLD (r = 0.2273, P = 0.0069) but not in those with CHB (r = 0.1005, P = 0.1495). To assess the usefulness of APRI as a predictor of the absence or presence of liver fibrosis (fibrosis score of 0 versus fibrosis scores of 1-4), receiver-operating-characteristic (ROC) curves were determined in each patient group. In patients with CHC, APRI showed a 72.7% sensitivity and 62.4% specificity for a fibrosis score of 1-4. In the NAFLD group, APRI showed a 60.0% sensitivity and 73.3% specificity for a fibrosis score of 1-4. In patients with CHB, APRI showed a 55.0% sensitivity and 75.4% specificity for a fibrosis score of 1-4. Conclusions: APRI index shows an acceptable accuracy for the assessment of liver fibrosis in patients with CHC and NAFLD but not in those of CHB.

Eplerenone Reverses Spironolactone-induced Painful Gynaecomastia in Patients with Cirrhosis: A Clinical Study Georgios Dimitriadis 1 , Vasileios Papadopoulos 1 , Konstantinos Mimidis 1 1 First Department of Internal Medicine, Democritus University of Thrace, 68100 Alexandroupolis, Greece Background: We have previously reported the use of eplerenone instaed of spironolactone in cases of painful gynecomastia in patients with cirrhosis from chronic hepatitis B. Here, we conducted a prospective study on the efficacy and safety of eplerenone in these patients. Methods: Fifteen consecutive patients with cirrhosis (8 alcohol abusers and 7 suffering from chronic hepatitis B of mean age 64.4 ± 13.3 years), who had been administered spironolactone and suffered from painful gynaecomastia, have been included in the study. Substitution of spironolactone with eplerenone followed under close observation during the next 3 months. Age, gender, Child-Pugh score, pain (in a 1-5 scale), FSH, LH, 17 (OH) progesterone, DHEA-S, testosterone and prolactine, along with standard haematological and biochemical parameters were measured at the beginning and the end of the study. Result: All 15 patients exhibited withdrawal of both pain and gynaecomastia (P \ 0.0001). Furthermore, five patients ameliorated as far as Child-Pugh score is concerned. With the exception of LH and 17 (OH) progesterone, which were found lower at the end of the study (P = 0.011 and 0.019, respectively), all other biochemical and hormone parameters remained unchanged. In a multiple regression model including the amelioration of pain as independent variable and age, cause of cirrhosis and Child-Pugh score as dependent ones, age (P = 0.012) and cause of cirrhosis (P = 0.013) were found to be independently correlated with the amelioration of pain; this was not true for Child-Pugh score (P = 0.318). Conclusion: The use of eplerenone instead of spironolactone in cirrhotic patients with painful gynaecomastia might reverse pain and presents a safe alternative treatment.

Endoscopic Treatment of Fundal Gastric Varices by Sclerotherapy with N-butyl-2-Cyanoacrylate Hong Joo Kim 1 , Jung Ho Park 1 , Dong Il Park 1 , Yong Kyun Cho 1 , Chong Il Sohn 1 , Woo Kyu Jeon 1 , Byung Ik Kim 1 1 Sungkyunkwan University Kangbuk Samsung Hospital, 108, Seoul, Background/Aims: Our study aimed to evaluate the therapeutic results of endoscopic N-butyl-2-cyanoacrylate injection (EBC) in patients with fundal gastric variceal hemorrhage (GVH). Methods: Twenty-two patients (19 male, 3 female) with large fundal varices (spurting or active oozing, 12; red spot or blood clot adherence, 10) who were treated with EBC from April 2005 to April 2009 were included in our study. Median (range) volume of 2.75 (1) (2) (3) (4) (5) cc of N-butyl-2-cyanoacrylate mixed with lipiodol (0.5:0.8 ratio) were injected into the fundal variceal lesion until all the lesions became hard. Results: The number of Child-Pugh A, B, and C cases was 5 (22.7%), 12 (54.5%), and 3 (22.7%), respectively. The classification of GV included in our study was as follows: GOV2, 17 (77.3%); GOV1 + GOV2, 2 (9.1%); IGV1, 3 (13.6%). Initial hemostasis was achieved in all 22 patients (100%) and significant rebleeding occurred in 10 patients (45.5%) after median (range) follow-up of 10.5 (1-29) months. Among these 10 patients with significant rebleeding, 2 patients and 6 patients were successfully treated by 2nd session of EBC and rescue balloon occluded retrograde transvenous obliteration (BRTO), respectively. However, remaining two patients had no chance of rescue treatment and died due to massive rebleeding. During the median (range) follow-up of 14.5 (1-56) months, complete eradication of GV and the aggravation of preexisting esophageal varices were observed in 13 (59.1%) and 5 (22.7%) patients, respectively. The significant predictors of rebleeding after initial EBC treatment were active bleeding (spurting or active oozing) on initial endoscopy and multiple sessions of EBC. Conclusions: This study showed that EBC might be effective treatment modality for patients with fundal GVH in terms of initial hemostasis. However, a significant proportion of patients with fundal GVH treated by EBC might experience rebleeding and were in need of rescue treatment. Egypt has one of the highest incidences of liver diseases in the world with prevalence of schistosomiasis. NO diffuses into cytosol of adjacent vascular smooth muscle cells play a role in the pathogenesis of vasodilation. Endothelial cells also produce the most potent vasoconstrictor agent endothelin (ET-1). Aim: Evaluation of nitric oxide and endothelin-1 and procollagen III peptide in patients with chronic liver disease and portal hypertension in both systemic and portal blood samples together with the histopathological scoring of liver biopsies. Subjects: The control group 15 subjects free from any liver disease. The patient group 30 patients with chronic liver disease and schistosomal portal hypertension. Methods: Clinical examination, abdominal ultrasonography, measurement of portal venous pressure and histopathological examination of liver biopsy. Laboratory investigations included evaluation of total nitric oxide (NO), endothelin-1 (Et-1) and type III procollagen (PIIINP) in both portal and systemic blood. In addition prothrombin, serum alanine, aspartate aminotransferase (AST, ALT), c glutamyl aminotransferase (cGT) activities, serum bilirubin, albumin, serodetection of hepatitis B surface antigen (HBsAg) and hepatitis B core antibody and (anti-HCVAb). Conclusion:

• NO and ET-1 levels in both systemic and portal blood of SHF patients were significantly higher than in the control group. • NO is a potent vasodilator.

• ET-1 increase may be a compensatory mechanism to antagonize the vasodilatory effect of NO. • Child class B subgroup had higher NO and ET-1 than class A.

• NO and ET-1 levels did not differ between anti HCV positive and negative SHF patients. Background/Aims: Bacterial infections have been documented into 35-66% of cirrhotic patients who have variceal bleeding, and were identified to be associated with early rebleeding. Patients with cirrhosis revealed significant increase in serum nitrites/nitrates, an index of nitric oxide (NO) generation. Endotoxemia secondary to bacterial infection may increase plasma NO and trigger variceal hemorrhage in liver cirrhosis. The aims of this study were to assess the impact of antibiotic prophylaxis on reduction of serum NO and prevention of early rebleeding after endoscopic therapy in esophageal variceal bleeding (EVB), and investigate the risk factors for early rebleeding within 6 weeks after endoscopic treatment in EVB. Methods: From May 2007 to April 2009, cirrhotic patients who had received endoscopic therapy in acute EVB were enrolled in the study. Patients with EVB were randomized into the two groups (antibiotic prophylaxis group and control group) after endoscopic treatment. Patients in the prophylactic group received antibiotic treatment with intravenous ciprofloxacin for 7 days. Results: A total of 44 patients (18 patients in prophylactic group and 26 patients in control group) were included for analysis. There were no differences between the two groups in parameters. Antibiotic prophylaxis did not decrease plasma renin, aldosterone, and serum NO concentrations. The early rebleeding rate within 6 weeks in the prophylactic group was not significantly lower than in control group. The presence of hepatocellular carcinoma and hepatitis B virus-related liver cirrhosis were significantly associated with early rebleeding (P = 0.022, 0.021, respectively). Conclusions: Antibiotic prophylaxis did not reduce serum NO and did not prevent early rebleeding within 6 weeks after endoscopic therapy in acute EVB. In the future, larger clinical studies about effect of antibiotic prophylaxis on reduction of serum NO and prevention of early rebleeding after endoscopic therapy in EVB will be needed. Results: Causes of cirrhosis were 55.7% (98 patients) in HBV, 5.7% (10 patients) in HCV, 35.2% (62 subjects) in alcohol, 0.6% (one subject) in autoimmune, and 2.8% (5 subjects) in cryptogenic. On univariate analysis, SBP was associated with Child-Pugh class C (P \ 0.001), high MELD score (P \ 0.001), PPI use (P = 0.018) and previous history of variceal bleeding (P = 0.038). On multivariate analysis, SBP was associated with Child-Pugh class (Class C, OR = 2.965, 95% CI 1.489-5.904, P = 0.002), MELD score (C20, OR = 3.272, CI 1.078-9.938, P = 0.036), and PPI use (yes, OR = 2.948, 95% CI 1.015-8.559, P = 0.047).

Conclusions: Besides Child-Pugh class and MELD score, PPI use was also a risk factor for SBP in cirrhotic patients with ascites.

Top 10% Posters Backgrounds: To reduce portal pressure, the only recommended medication is nonselective beta blocker (NSBB). However, NSBB has some limitation to apply clinically because of poor response rate and compliance. Angiotensin receptor blocker (ARB) inhibits activated hepatic stellate cell contraction and it is supposed to reduce dynamic portion of intrahepatic resistance. This study aimed at comparing the effects of a combined treatment using candesartan plus propranolol with propranolol monotherapy on portal pressure in patients with cirrhosis, prospectively. Methods: The hepatic venous pressure gradient (HVPG), mean blood pressure (BP), Child-Pugh score, and model for end-stage liver disease (MELD) score were assessed at baseline and were followed at 3 month later in both combination (28 patients) and monotherapy group (31 patients) from January 2008 to July 2009. The initial dose of propranolol (20 mg) was subsequently adjusted until the target HR was reached. Patients in whom HVPG reduced by[20% or to less than 12 mmHg were defined as responders. The mean dose of candesartan and propranolol were 7.3 ± 1.6 and 155 ± 62.1 mg respectively. In propranolol monotherapy group, the mean dose was 157 ± 60.3 Results: Portal pressure declined significantly in both groups (combination group 16.3 ± 3.6-14.0 ± 3.8 mmHg, P \ 0.05, propranolol monotherapy group 16 ± 4.9-12.9 ± 4.5 mmHg, P \ 0.05). However, no difference was observed in the effect of pressure reduction between both group (2.3 ± 3.9 in combination group vs. 4.1 ± 5.1 in propranolol monotherapy group, P [ 0.05).

No difference in reduction of mean BP and heart rate between both groups. Conclusions: The combination of ARB with NSBB to reduce intrahepatic pressure do not showed beneficial effect compared to NSBB monotherapy in cirrhotic patients. Background/Aims: Poor blood oxygenation despite breathing 100% oxygen is a very severe clinical condition in patients with hepatopulmonary syndrome (HPS). Blood oxygenation while breathing 100% oxygen was studied in CBDL rats, an animal model of portal hypertension and HPS with progressive liver damage, as well as in rats with partial portal vein ligation (PVL), a portal hypertension model without liver damage.

Materials/Methods: We studied 47 Sprague-Dawley rats, including 16 CBDL rats, 16 PVL rats, and 15 sham rats. Arterial blood and mixed venous blood were collected for analysis from unrestrained conscious animals breathing room air or breathing 100% oxygen for 30 min each. Qs/Qt was calculated by Fick's equation, and intrapulmonary microsphere shunting (IPMS) was determined by using radiolabeled microspheres (15 mm in diameter) in animals' breathing room air or 100% oxygen. Results: CBDL rats' breathing room air showed significant dysoxygenation and their arterial oxygen pressure (PaO 2 ) was inversely correlated with Qs/Qt, which was significantly greater than in the other two groups. The mean PaO 2 of Sham rats, CBDL rats, and PVL rats breathing oxygen was respectively 382 ± 64, 249 ± 75 and 366 ± 52 mmHg (ANOVA P = 0.001). In CBDL rats and PVL rats' breathing room air, IPMS was significantly greater than in Sham rats. After breathing oxygen, however, in CBDL rats and PVL rats showed similar IPMS to Sham rats. Macroscopic atelectasis was observed in both CBDL rats and PVL rats, and it was particularly marked in the former group.

Conclusion: Increased venous admixture may contribute to hypoxemia in HPS rats. The arterial oxygenation of CBDL rats breathing 100% oxygen was poor. Absorption atelectasis due to breathing pure oxygen may have contributed to the blunted blood oxygenation response of CBDL rats.

Epidemiological and Clinico-Biochemical Comparison between Hepatitis B and Alcoholic Liver Cirrhosis Guan Wee Wong 1 , James Li 1 , Jason Chang 1 1 Departmentof Gastroenterology and Hepatology, Singapore General Hospital, Outram Road, Singapore 169608

Background: Chronic hepatitis B and alcohol are the two most common etiologies of liver cirrhosis in Singapore. There is limited data on the different clinical and biochemical features of chronic hepatitis B liver cirrhosis (CHBLC) and alcoholic liver cirrhosis (ALC). Aim: To compare the epidemiology and clinical characteristics between CHBLC and ALC. Methods: We performed a retrospective study of patients enrolled into the liver cirrhosis registry of Singapore General Hospital from September 2006 to January 2009. Clinical and biochemical data were obtained from computerized hospital records. Cirrhotics due to CHB and alcohol were selected. Epidemiology and clinical characteristics of these two groups were analyzed. Results: The study included 210 patients, 169 (80.5%) with CHBLC and 41 (19.5%) with ALC for a median follow-up period of 51 months. There were more males (85.4 vs. 64.5%), more Indians ( More ALC patients presented with hepatic decompensation at diagnosis compared to CHBLC (39 vs. 21.3%, P \ 0.05). More CHBLC patients were diagnosed on screening compared to ALC patients (59.2 vs. 31.7%, P \ 0.05). Conclusions: ALC patients had more advanced liver disease, greater degree of portal hypertension and tend to present with decompensation at diagnosis. In contrast, CHBLC patients were diagnosed at an earlier stage of cirrhosis due to earlier detection on screening. ascites, splenomegaly. The relation between gastric varices and the portal hemodynamic alterations has been studied using doppler ultrasonography. Aims and Methods: To assess portal hemodynamics in cirrhotic patients with portal hypertension and examine the various hemodynamic values to affect gastric varices, we performed doppler US in total 624 patients with cirrhosis. 438 patients underwent esophagogastroscopy. The longitudinal, transverse diameter and thickness at the hilum of spleen were measured to get the splenic volume. Hepatic vein diameter at intercostal scan was also measured. Main portal vein (MPV) and umbilical portion of left portal vein (UPV) were scanned and maximal portal flow velocity was calculated. We defined delta portal flow velocity (delta PFV) as subtracting UPV velocity from MPV velocity. Resistive index (RI) and pulsatile index (PI) of renal arcuate artery were also measured. Hepatic vein doppler waveform was classified as triphasic, biphasic or monophasic. Using it's waveform damping index was obtained. 19) or without (n = 61) PVT were analyzed. Seventeen common ultrasonographic and serum markers were analyzed. SPSS software was used for statistic analysis.

Results: Logistic regression model showed that the d-dimer, main portal vein width and platelet were the independent risk factors of portal vein thrombosis (PVT) in liver cirrhosis patients. Conclusion: The d-dimer, main portal vein width and platelet should be monitored in patients with cirrhosis and we can take suitable therapy to prevent and cure portal vein thrombosis.

The Results: SIBO was diagnosed on 23 (57.7%) of 40 cirrhotic patients, and on 11 (27.5%) of the 40 controls (P \ 0.012). Among the cirrhotic patients, SIBO positive patients showed higher constipation score (1.30 vs. 0.47, P \ 0.05) and hard stool score (0.87 vs. 0.18, P \ 0.02) than SIBO negative patients. OCTT was slower in cirrhotic patients compared to healthy controls (140 ± 29 vs. 116 ± 20 min, P \ 0.001) and slower in SIBO positive group compared to SIBO negative group (136 ± 25 vs. 120 ± 28 min, P = 0.035). Delayed OCTT was associated with hard stool score (r = 0.44, P = 0.02). Conclusions: Delayed OCTT and higher prevalence of SIBO were noted in cirrhotic patients. SIBO positive cirrhotic patients showed slower OCTT, higher constipation score and hard stool score than SIBO negative patients.

Application There were no significant differences between the observed and expected survival curves in the stratified risk groups (low risk, P = 0.61; high risk, P = 0.77). Conclusion: Our data suggest that the proposed model is able to accurately predict survival in cirrhotic patients.

Serum-ascites Albumin Concentration Gradient (SAAG) Greater than 20 g/L can be Used as Non-invasive Indication for Starting Primary Prophylaxis of Esophageal Variceal Hemorrhage (EVH) Pises Pisespongsa 1 , Sirirattana Sukhiranwat 1 , Apinya Leerapun 1 , Taned Chitapanarux 1 , Satawat Thongsawat 1 , Ong-Ard Praisontarangkul 1 1 Chiang Mai University, 110 Inthawaroros, Chiangmai, Thailand

Purpose: Screening for esophageal varices (EV) with esophagogastroduodenoscopy (EGD) is recommended in cirrhotic patients but EGD is invasive, and not available in many hospitals. SAAG is correlated with portal hypertension and presence of EV so we aimed to determine value of SAAG that predict presence of high risk EV. Method: Prospective study included 35 newly diagnosed cirrhotic patients with ascites, who never had bleeding or hepatocellular carcinoma and never received beta-blocker or esophageal variceal ligation. Child-Pugh score (CPS) and SAAG were calculated and EGD were performed. Results: Twenty-one patients (60%) were male. One was CPS class B and the remaining were class C. AUROC for SAAG to predict EV was 0.855. In 19 patients with SAAG[20 g/L, large EV ([5 mm) were found in 7 (36.8%), and small in 11 (57.9%). Whilst in 16 patients with SAAG \20 g/L, large EV were found in only 1 (6.3%), and small EV in 9 (56.2%). Since presence of either large or small EV in patients with CPS class C was indication for EVH prophylaxis, hence presence of EV that needs prophylaxis was significantly higher in patients with SAAG [20 g/L (94.7 vs. 62.5%, P = 0.032). The sensitivity, specificity, positive predictive value and negative predictive value of SAAG [20 g/L in predicting high risk EV were 64.3, 85.7, 94.7 and 37.5% respectively; and positive likelihood ratio and negative likelihood ratio were 4.5 and 0.41. Conclusions: SAAG [ 20 g/L can predict presence of high risk EV that needs prophylaxis therapy with PPV of 94.7%. Therefore it could be used as cheap and convenient non-invasive indication for starting primary EVH prophylaxis in cirrhotic patients. The aim of our study is to analyse predictive factors of mortality at admission using the current ICU and specific liver scores in cirrhotic patients admitted to the liver ICU secondary to an acute decompensation. Patients and Methods: we studied prospectively 377 consecutive patients admitted to our liver ICU for decompensated cirrhosis from May 2005 to May 2009. Mean age was 55.5 ± 11.4 years (73.5% of male). The etiology of liver cirrhosis was mainly alcoholic (68.4%). At admission, mean prognostic values were 45 ± 21 for SAPS II score, 24 ± 10 for the MELD, 29 ± 13 for the MELD-Na, 48 ± 13 for iMELD, 1.9 ± 0.8 for MESO index, 10 ± 2 for Child-Pugh and 9 ± 5 for SOFA score. Results: ICU and hospital mortality were respectively 34.7 and 42.9%. The statistical analysis comparing cirrhotic patients who died to those who survived showed that all above mentioned prognostic scores at admission were very strong predictors of mortality (P \ 0.0001). The area under the curve (ROC) predictive mortality was 0.82 for the MELD, 0.79 for MELD-Na, 0.81 for iMELD, 0.82 for MESO index, 0.79 for Child Pugh classification, 0.89 for SAPS II score and 0.92 for SOFA score. A cutoff predicting mortality of 47.5, 10.5 and 28.5 for SAPS II, SOFA and MELD score was found, respectively. The need for respiratory support (44.8%), inotropic support (40.3%) and continuous hemodialysis for renal failure (18.6%) predictive mortality was analysed. The need for one, two or three of these organ support yielded respectively to 5.8, 77.1 and 96% mortality.

Hepatol Int (2010) 4:94-345 257

Conclusion: SAPS II and SOFA scores at admission better predict ICU mortality compared to specific liver scores. The need of two or more organ support during hospital stay would highly influence the mortality. Aim: To describe the admission and mortality pattern pertaining to liver disease (CLD) in our hospital to highlight burden of this disease and future planning for prevention, control and management. Methodology: It was a descriptive type of study. Data was collected from yearly audit reports of department for last 10 year (1998) (1999) (2000) (2001) (2002) (2003) (2004) (2005) (2006) (2007) (2008) . Data of year 2003 and first 6 months of 2004 was not available due to some technical reasons. Share of liver related diseases to total number of admissions per year was calculated. Similarly all cases pertaining to medical mortality were reviewed. Out of these cases, patients of liver disease were identified. Statistical analysis was done using SPSS. Admission and mortality rate due to liver diseases was calculated for each year. Results: According to collected data, number of admissions for liver diseases gradually increased from 1998. In 1998 total number of admission was 1,267, out of which 279 (22%) patients were with liver related diseases. Maximum admissions due to liver diseases were in 2006 i.e 1,263 (35%) out of 3,595, total number of admissions. The share of Liver diseases to total admissions in last ten years continued to progress almost at a constant ratio till 2006. In last two years (2007 and 2008) this share has decreased to 27 and 28%, respectively. In second part of study total mortalities and percentage of liver related mortalities for last 10 year was calculated. Data shows that in 1998 share of liver related mortalities to total mortalities was 37% (115 cases). Then form 1999-2007 it remained between 21 to 31%. Finally in 2008 it dropped to 15% (88 cases) which is the lowest in last 10 years. Conclusion: Liver diseases due to chronic viral hepatitis are still a burden on our health resources. Current trend of decreased number of admissions and falling mortality due to this disease may be due to better understanding of disease, programs for prevention like vaccination for HBV and effective treatment for HCV with IFN. Background: Hepatic encephalopathy is associated with increased morbidity and mortality in patients with end-stage liver disease. L-Ornithine-L-aspartate (LOLA) activates the periportal urea cycle in the liver and lowers plasma ammonia. Previous placebo-controlled, double blind trials has shown the efficacy of LOLA infusions in cirrhotic patients with hepatic encephalopathy. To date, limited data is available to confirm the efficacy of the oral formulation. Aim: Retrospective audit of the use of LOLA to assess patient outcome and change in encephalopathy related admissions. Methods: Fourteen patients with end-stage liver disease and intractable hepatic encephalopathy, unresponsive to lactulose and enemas, received LOLA. Clinical and laboratory data were collated to assess response to treatment. Results: Twelve out of 14 patient records were completely examined. The median duration of LOLA was 7 months (range 1-17). Median age was 54 years (range 48-67). 69.2% were males. Ten patients had end-stage chronic hepatitis C, three had alcoholic liver disease and one had autoimmune hepatitis. Median baseline MELD score was 12 (range 6-24) which did not change during treatment. To date, six patients have been transplanted, two have died and five remain on treatment, two of whom are awaiting transplant. The median number of admissions per month for hepatic encephalopathy fell from 0.5 episodes pre treatment (IQR 0.37-0.94) to nil (IQR 0-0.33) whilst on LOLA; P = 0.004. The median hours per admission per month pre-and posttreatment were 23.9 (IQR 11.1-97.5) and 0 (IQR 0 -20.8), respectively (P = 0.004).

No patient reported any significant side effect from the treatment requiring treatment discontinuation. Conclusions: Oral LOLA is a safe and effective treatment for intractable hepatic encephalopathy, reducing hospital admissions. Further randomised controlled trials are required to confirm that LOLA improves hepatic encephalopathy and quality of life. Background: A deterioration of renal function in cirrhotic patients with spontaneous bacterial peritonitis (SBP) is a predictor for in-hospital mortality; however, the clinical significance of renal failure during a bacterial infection other than SBP is unknown. The aim of this study was to investigate the prevalence and clinical significance of renal failure due to bacterial infections other than SBP in patients with liver cirrhosis. Methods: Retrospective data from inpatients with bacterial infections other than SBP were analyzed. Results: Eighty patients were enrolled. The types of infections were urinary tract (37.5%), pneumonia (23.8%), biliary tract (20%), cellulitis (12.5%), and bacteremia of unknown origin (6.3%). Renal failure developed in 29 patients (36.3%), and 11 patients had irreversible renal failure. The initial MELD score, neutrophil count, albumin, and blood pressure were significant risk factors in the univariate analysis, but only the MELD score was an independent risk factor for the development of renal failure (P \ 0.001).

Conclusion: The prevalence of renal failure during a bacterial infection other than SBP in patients with liver cirrhosis was 36.3%, and its development was related to the severity of the liver disease. Whether irreversible renal failure developed or not determined the prognosis of these patients.

Prognostic Factors of Mortality in Patients with Liver Cirrhosis (A Tertiary Care Hospital Experience) Masroor Qazi 1 , Hashim Raza 1 , Ali Imran 1 , Saba Anjum 1 , Ghulam Muhy-ud-din 2 1 Quaid-e-Azam Medical College, Bahawalpur, Pakistan, 2 Nishter Medcial Colleges, Multan, Pakistan

Background: Liver cirrhosis causes portal hypertension, which ultimately leads to severe complications. We conducted a study to find out the factors associated with high mortality in patients with liver cirrhosis (portal hypertension) during hospital stay. Methods: A retrospective, experimental study was conducted in Bahawal Victoria Hosptial/Quaid-e-Azam Medical College, Bahawalpur, Pakistan (1st July 2007 to 15th August 2009). In 540 cirrhotic patients (either HBS positive, anti-HCV positive or both), frequency of mortality was noted and patients were divided into two groups. Group-A, patients who died during the hospital stay and Group-B, patients who survived. Variables studied were: sex, age, body mass index (BMI), diabetes mellitus (DM), hepatic encephalopathy (HE), ascites, spontaneous bacterial peritonitis (SBP), upper gastrointestinal bleed (UGIB), total leukocyte count (TLC), prothrombin time (PT) difference, serum bilirubin, albumin and creatinine. Chi square test was applied for statistical significance. Results: Mortality was 24.4%. When two groups were compared (Group-A and Group-B), BMI [ 25 (P \ 0.0001), HE (P \ 0.0001), SBP (P \ 0.0001), TLC [ 10,000 (P \ 0.0001), serum bilirubin [2 (P \ 0.0001), serum albumin \3.5 (P = 0.044) and serum creatinine [1.5 (P \ 0.0001) were significantly associated with mortality. All other variables like sex, age, DM, UGIB and PT difference [4 min were statistically insignificant. Conclusion: In our study mortality was high among the patients who were having HE, SBP, raised BMI, raised TLC, raised serum bilirubin, raised serum creatinine and low serum albumin.

Atoko Uegaki 1 , Mina Sasaki 1 , Kenichiro Nakajima 1 , Maki Tobari 1 , Yasuko Ushio 1 , Kenji Kino 1 1 Department of Gastroenterology, Tokyo Metropolitan Geriatric Hospital, 35-2, Sakae-machi, Itabashi-ku, Tokyo, Japan

Background and Aims: It is critical to precisely evaluate renal function in patients with liver cirrhosis. However, serum/uninary creatinine (Cr) and Cr-based estimation of glomerular filtration rate (GFR) may not be a reliable marker of renal function in cirrhotic patients with lower body muscle. In this study, we investigated which parameters or formulae are appropriate for estimation of GFR in patients with liver cirrhosis. Patients and Methods: Seven patients with liver cirrhosis were enrolled in this study. All Japanese, M/F ratio 4/3, age 76.9 ± 9.9, and the etiology included 6 HCV and 1 alcoholic. The body mass index in 7 patients was 21.7 ± 1.9. All patients had neither detectable ascites nor encephalopathy at baseline, and were graded as Child-Pugh A. After an overnight fasting period, inulin clearance was determined by standard technique, and used as 'true' GFR. At the same day, serum Cr and cystatin C (Cys C) were measured from the same blood samples, and urinary Cr was measured by collecting urine for a whole day. Then we calculated GFR using Cr-based GFR estimating formulae (MDRD, Cockcroft-Gault) as well as Cys C-based formulae (Hoek, Larsson), which were statistically compared with GFR. Results: GFR determined by inulin clearance were 60.3 ± 21.1 (35.6-97.1) ml/min, suggesting the presence of renal impairment in some cirrhotic patients, even in Child-Pugh A. Cr clearance (Ccr) calculated using serum Cr was 70.7 ± 25.6, statistically unassociated with GFR by inulin (r = 0.148, P = 0.752). In addition, Cr-based GFR estimating formulae, MDRD and Cockcroft-Gault, demonstrated 95.7 ± 17.3 (r = 0.689, P = 0.087) and 59.7 ± 21.7 (r = 0.673, P = 0.098) as calculated GFR, respectively. By contrast, serum Cys C were 1.1 ± 0.1 (r = -0.742, P = 0.056), well correlated with GFR as a single parameter. As a result, Cys C-based GFR estimating formulae, Hoek and Larsson, demonstrated 68.0 ± 6.2 (r = 0.743, P = 0.055) and 67.6 ± 7.4 (r = 0.744, P = 0.055) as calculated GFR, and thus shown a better correlation with GFR. Conclusion: For the assessment of renal function in patients with liver cirrhosis, Cys C itself or Cys C-based formulae are more appropriate parameters than those using serum Cr.

Sudhir Gupta 1 1 Govt. Medical College & Super Speciality Hospital, Rige Road, Nagpur India

Background: Portal hypertension (PHT) is an important cause of morbidity and mortality in children. The spectrum of causes of PHT in our community is different from what is described in west. Methods: Children below 12 years who presented with features of PHT in Gastroenterology OPD were evaluated. All patients underwent detailed clinical examination, Doppler study, viral markers and liver biopsy (wherever feasible). Results: 62 patients were enrolloed in the study. Mean age of the patients were 8.2 years with male to female ratio of 3.5:1. Forty five patients presented with GI Bleed, 22 patients with distention of abdomen, 10 with jaundice and 8 with lump in Lt hypochondrium. Forty two (67%) were diagnosed as Extra hepatic portal vein obstruction (EHPVO), 6 (9%) patients were diagnosed as Non cirrhotic portal fibrosis (NCPF) and 14 (22%) patients had chronic liver disease. (5 HBV related 3-Wilson's disease, 2-autoimmune liver disease and 3idiopathic). History of umbilical sepsis was present in only 10% of the cases of EHPVO. Doppler study revealed portal cavernoma in all patients of EHPVO and only 10% patients had demonstrable thrombosis in portal vein. 56 patients had oesophageal varices at the initial endoscopic examination. 45 patients with GI bleed were treated with endoscopic variceal band ligation (EVL) and Betablockers and were prospectively followed. In majority of patients Grade four varices were changed to grade I or completely eradicated after single session of EVL. 19% of the patients required more than one session of EVL. Shunt surgery was done in 6 patients (failed endoscopic therapy-3, living in far Hepatol Int (2010) 4:94-345 259 places-3). Growth retardation was seen in 12% of the patients with EHP-VO. Total mortality in this study was 5% (2 HBV related CLD and 1 Wilson's Disease). Conclusion: EHPVO is most comon etiological factor of Portal hypertension among children in the region. EVL is a safe and effective modality of the treatment in patients with EHPVO presenting as GI Bleed.

FibroScan as a Predictive Tool for Esophageal Varices and Variceal Bleeding Adrian Goldis 1 , Iulia Ratiu 1 , Daniela Lazar 1 , Dimitrios Koukoulas 1 , Mihai Ionita 1 1 Emergency Clinical County Hospital Timisoara, Romania, Bulbuca 10, Timisoara Romania

Background:

Upper digestive bleeding remains one of the problems commonly encountered in medical practice and, especially in patients with liver cirrhosis it has a major influence in morbidity and mortality. The key point in approaching these patients is to determine patients with major risk for bleeding. In this study we evaluated liver stiffness measured by Fibroscan as a possible predictive factor for variceal hemorrhage. Background: Portal hypertension and its complications are the primary cause of mortality in cirrhotic patients. Hepatic venous pressure gradient (HVPG) is an indirect measure of portal venous pressure. The aim of the study was to measure HVPG in cirrhotic patients and correlate it with Child-Turcotte-Pugh (CTP) score, Model for end-stage liver disease (MELD) score, size of varices, variceal bleeding, ascites and etiology of cirrhosis in order to identify the predictors of HVPG. Methods: 30 consecutive patients with cirrhosis were studied. HVPG was measured using a Swan-Ganz catheter placed in the hepatic vein through percutaneous transjugular route. The mean HVPG in patients with different CTP classes, different grades of varices (small and large), bleeders and non bleeders, with and without ascites and cirrhosis of various etiologies was calculated. Correlation of various variables with HVPG was assessed using Spearman correlation test. Univariate analysis and multivariate analysis of variables predicting HVPG was done using Cox regression analysis. Receiver operating characteristic (ROC) curve was used to identify the HVPG measurement predicting variceal bleeding. Results: The mean HVPG was significantly higher in CTP C as compared to CTP B and CTP A (P B 0.001). The mean HVPG was significantly higher in bleeders, patients with varices and ascites (P \ 0.05). The mean HVPG was higher in alcoholic as compared to non-alcoholic cirrhosis but was not statistically significant. Variceal bleeding, CTP and MELD scores were found to be independent predictors of HVPG in multivariate models. HVPG above 14.5 mmHg had a sensitivity of 83% and specificity of 100% in predicting variceal bleeding. Conclusion: Variceal Bleeding, CTP and MELD scores were found to be independent predictors of HVPG. A higher HVPG signifies more severe liver disease. So HVPG can predict the major complications of cirrhosis thereby helping in prevention and early treatment of complications.

The Background: Various modalities are suggested for secondary prophylaxis of gastric variceal bleeding. Endoscopic variceal obliteration (EVO), endoscopic variceal ligation (EVL), and balloon-occluded retrograde obliteration (BRTO) are commonly utilized. But simultaneously comparing three modalities are rarely available. We investigated the comparative effects between three modalities. Methods: Sixty four patients, stopped variceal bleeding spontaneously after admission, were enrolled between July 2001 and May 2009. 19 patients were received EVO, 20 were EVL, and 15 were BRTO for secondary prophylaxis of gastric variceal bleeding. We checked laboratory tests and vital sign. Then we calculated Child score, Child classification, and MELD score. Rebleeding was defined as new onset hematemesis, hematochezia, melena or endoscopically proven bleeding. Time-to-rebleeding and survival time were analyzed with Kaplan-Meyer survival analysis, and p value, lower than 0.05 was considered as statistically significant.

Result: There were no statistically significant differences between three groups in baseline characteristics. Overall follow-up period was 65.13 months. During follow-up period rebleeding was occurred in a total of 7 patients, 2 in EVO, 4 in EVL, and 1 in BRTO. Time-to-rebleeding was 47.44 ± 5.51 months in EVO, 49.27 ± 18.39 months in EVL, and 51.33 ± 3.54 months in BRTO group, and there was no statistical significance (P = 0.597). Median survival time was 56.01 ± 13.12, 56.10 ± 15.15, and 42.79 ± 6.23 months, respectively, and there was no statistical significance (P = 0.531).

Conclusion: There were no statistical significant differences of time-to-rebleeding and survival time between EVO, EVL, and BRTO. This may be attributed to selection bias, caused by retrospective study, and small number of patient size. Further prospective and large scaled study is inevitable

The Application of ''U'' Retroflexion Ligation Therapy on Esophagogastric Varices (EGV1) Hong Xu 1 , Geng Chen 1 , Yan Sun 1 , Tong-Yu Tang 1 , Ying-Kai Wwang 1 1 Department of Gastroenterology, The First Hospital of Jilin University, Changchun 130021, China

Objective: To evaluate the effects of ''U'' retroflexion ligation therapy on esophagogastric varices. Methods: One hundred and ninety one patients of esophagogastric varices were randomly allotted into two groups, one hundred and one in the A group with endoscopic variceal ligation (EVL) in routine method and the rest in the B group with ''U'' retroflexion EVL (EVLR), to assess efficacies of EVL and EVLR for esophagogastric varices. Results: During the period of following-up (18 months), excellence rate, inefficacy rate and averaging therapy time had significant difference between two groups (P \ 0.05), B was better to A, and hemostasis rate of emergency in B was 100%. However, relapsed bleeding rate, varices recurred rate, complications rate and mortality were not significant difference in both groups (P [ 0.05). Conclusion: ''U'' retroflexion ligation is more effective and safer than endoscopic variceal ligation in routine method for esophagogastric varices, it can raise the rate of emergency hemostasis rate, and decrease the averaging therapy time.

Effect of Navel Application with Chinese Herbal Cataplasm on Patients with Ascites Due to Liver Cirrhosis Feng Xing 1 , Cheng-Hai Liu 1 1 Shuguang Hospital Affiliated to Shanghai University of Traditional Chinese Medicine, Zhangheng Road 528#, Shanghai 201203, China

Aim: To observe the effects of navel applications with Xiaozhang Cataplasm on patients with ascites due to liver cirrhosis. Methods: One hundred and twenty four in-patients with ascites due to cirrhosis were enrolled and randomly divided into placebo (61) and trial (63) groups. Xiaozhang Cataplasm was composed of Radix Rhizoma Rhei, Folium Raphani, and Moschus etc., and was made into transdermal plaster-cataplasma. Besides routine treatment with limited usage of diuretics, the trail group received Xiaozhang Cataplasm on navel, while the placebo group received cataplasm without medicine. The patients' symptoms, body weight, abdomen circumference, urinary amount were observed, and ultrasound examinations were taken before and after treatment, and efficacy was evaluated according to comprehensive grading criteria. The side effects were also observed. Results: 107 cases finished the full course, while 12 withdraw and 5 were removed. The age and gender ratio etc between placebo and trial groups had no differences. Compared to placebo group, patients in trial group had decreased body weight, abdomen circumference and ascites amount, but increase urinary amount and flatus. The comprehensive efficacy at the trial groups were obvious better than placebo, efficient rates of I and II grades at placebo and trial were 39.3, 68.2%. Three cases had local and slight skin rashes, which disappeared after 2 days of temporal stopping plaster and using iodine oil. Conclusions: Umbilical sticking with Xiaozhang Cataplasm may be therapeutically useful in treating patients with ascites due to liver cirrhosis. The action mechanisms may be associated with improving urination and flatus. The skin reactions happened rarely and slightly, and the cataplasma was safe for clinical use.

Tong-Yu Tang 1 , Yuqin Li 1 , Dan Wang 1 , Yang Shi 1 1 First hospital of Jilin University, NO.71 Xinmin Street, China

Background and aim: Upper gastrointestinal endoscopy (UGIE) is a useful tool that identify those who have esophageal varices. But it is not adopted by many patients. We aimed to investgate a parameter based on non-invasive index for the prediction of esophageal varices (EV) in patients with cirrhosis.

Methods: One hundred and sixty eight patients with liver cirrhosis had physical examination, laboratory tests, color-Doppler ultrasound, upper gastrointestinal endoscopy, and portal vein diameter and splenic vein diameter were calculated.Platelet count/spleen diameter ratio was also calculated for all patients.The accuracy of all the non-invasive parameters in predicting the presence of varices was evaluated by the area under the receiver operating characteristic (ROC) curves (AUC). Results: The prevalence rate of EV was 67%. The diameter of portal vein, spleen diameter, the diameter of splenic vein were different in different degrees of esophageal varices. Platelet count, spleen diameter, platelet count/spleen diameter radio, and Child-Pugh class were significantly different among patients with or without EV. By a multivariate analysis, the platelet count/ spleen diameter radio was one independently parameter associated with the presence of EV. The AUC value generated by the ROC curves for platelet count/ spleen diameter ratio was 0.89, higher than the others. Conclusions: Noninvasive prediction of EV maybe not accurate. But combining with simple laboratory variables has a high sensitivity to predict the varices. It suggested that platelet count/spleen diameter ratio is an ideal noninvasive predictive indexes of the presence of the varices. Background: Esophageal varices bleeding (EVB) is a common and usually lethal complication of liver cirrhosis. Here we made a survey for factors related to EVB onset and try to establish a simple model to predict EVB onset. Methods: Hepatic cirrhosis patients with EVB were as case group, and the cirrhosis patients without EVB were as control group. About 200 factors were surveyed by special trained physicians, including the features of present and past history, personal hobby, food habit, occupational condition, exercise habit, sleeping and rest, habitus features, psychological features, medical service condition, socio-economic condition, physical examination findings, laboratory results, medical image results, endoscopical results. Univariate Logistic regression was used to calculate odds ratio (OR) of each factor, and multivariate Logistic regression was used to establish the predictive model. Results: A total of 206 patients with hepatic cirrhosis were enrolled into this study, in which there were 125 patients with EVB and 81 patients without EVB. Univariate analyses showed that ORs of 20 factors were significant, including 7 laboratory items, 5 endoscopical items and 8 other items. Three independent factors [bleeding history (BH), serum direct bilirubin (DBIL) and alkaline phosphatase (AKP)] were screened out by stepwise multivariate Logistic analysis, and the Logistic function was Logit (P) = -3.458 + 4.799BH + 0.018DBIL -0.31AKP. This model was valuable for predicting EVB onset in cirrhosis patients, with sensitivity 88.9%, specificity 87.5%, accuracy 88.4%, positive predictive value 93.3%, negative predictive value 80.0%, positive likelihood ratio 7.11 and negative likelihood ratio 0.13. Conclusions: Many factors are related with EVB onset in cirrhosis patients, especially bleeding history, serum direct bilirubin and alkaline phosphatase. A valuable model could be established based on these factors.

Epidemiology of Patients with Liver Cirrhosis in a Single Tertiary Centre in Singapore James Li 1 , Guan-Wee Wong 1 , Jason Chang 1 1 Singapore General Hospital, Outram Road, 169608 Singapore

Background: Liver cirrhosis is a common cause of morbidity and mortality and is an important burden on the healthcare system. To date, there are no published studies on the epidemiology of cirrhotic patients in Singapore. Aim: To describe the epidemiology and clinical characteristics of cirrhotic patients from a single tertiary referral centre in Singapore. Methods: We performed a retrospective review of patients enrolled into the cirrhosis registry of Singapore's largest tertiary medical centre from September 2006 to January 2009. Diagnosis of cirrhosis was based on radiology and/or Background/Aims: Liver stiffness is estimated to be consisted not only by static factor like fibrosis, but also dynamic factors like inflammation, cholestasis, and portal blood flow. Although the relation of the liver stiffness and liver fibrosis is well investigated, the relation of the portal vein pressure is not investigated sufficiently. In this study, we measured the liver stiffness and portal vein pressure simultaneously before and after the procedure that fluctuates portal hemodynamics by BRTO (Balloon occluded retrograde obliteration) and PSE (partial splenic embolization), and estimated the degree of the importance of the portal vein pressure as the participation factor of liver stiffness. Methods: BRTO was performed in three patients with liver cirrhosis for the treatment of gastric varices for the purpose of occlusion of gastro-renal shunt. PSE was performed in seven cirrhotic patients for the treatment of hypersplenism. The value of liver stiffness and WHVP (wedged hepatic vein pressure) were measured before and after the procedure. Liver stiffness was measured by FibroScan.

Results: Mean value of liver stiffness is increased from 26.1 ± 20.7 to 42 ± 23 kPa (mean ± SD) and WHVP is increased from 266.6 ± 40.4 to 386.6 ± 49.3 mmH 2 O in BRTO. Mean value of liver stiffness is decreased from 30.7 ± 15.9 to 23.9 ± 11.9 kPa and WHVP is decreased from 305.7 ± 117.1 to 259.2 ± 125.9 mmH 2 O in PSE. The change ratio of liver stiffness and WHVP is correlated significantly (r = 0.70, P \ 0.05) and 1 kPa of liver stiffness is estimated to be equivalent of 4.6 mmH 2 O of WHVP.4) Conclusions: Portal vein pressure is estimated to be one of the constructing factors of the liver stiffness. Liver stiffness measurement will be useful to estimate the portal hypertension. We will present our data using statistical analysis.

Effects Objective: To investigate the expression of marrow hemopoiesis factor TPO, GM-CSF in the marrow fluid and its effects on the reduction in circumference blood corpuscle of liver cirrhosis patients. Methods: TPO and GM-CSF concentration of liver cirrhosis group and control group was detected by enzyme linked immunosorbent assay (ELISA). Results: The TPO concentration in the control group and liver cirrhosis group are respectively 118.414 ± 49.232 and 90.756 ± 30.92 pg/ml, and there is no statistic significance to their difference(t = 0.386, P [ 0.05).TPO concentration of marrow fluid and the number of platelet of circumference blood corpuscle in liver cirrhosis patients is negative correlative. The CM-CSF concentration in the control group and liver cirrhosis group are, respectively 24.82 ± 3.66 and 22.01 ± 6.71 pg/ml, with their difference of no statistics significance (t = 0.9, P [ 0.05). Correlation analysis of the number of white blood cells in marrow fluid CM-CSF patients and liver cirrhosis patients was made, which showed no evident relation between the two. Liver cirrhosis patients were classified into three groups according to liver Child-Pugh graduation, and their differences in marrow blood TPO and TPO CM-CSF expression among different groups are of no statistic significance (P [ 0.05). Conclusion: The decline TPO concentration of the marrow might play an important role for the number of platelet of circumference blood corpuscle reduced. ''Platelet count, spleen diameter, Platelet count/spleen diameter ratio: noninvasive parameters to predict the presence of esophageal varices in cirrhsis'' Introduction and Objective: Esophageal varix (EV) is one of the most dreaded complications in cirrhotic patients. All cirrhotics should undergo screening for EV. There are several studies about noninvasive markers to predict the presence of E.V. The study was conducted to evaluate platelet count (PC), spleen diameter (SD) and platelet count/ spleen diameter ratio (PC/SD ratio) as the noninvasive parameters to predict the presence or not of EV in cirrhotics. Design: An observational, cross-sectional study.

The study was realized in Hue Central Hospital, from April 2007 to May 2008. Patients and Methods: Ninety three patients with liver cirrhosis and 30 healthy persons were studied. Biochemical study, upper GI endoscopic findings, maximum spleen bipolar diameter measurement by abdominal ultrasound; Platelet count, spleen diameter and platelet count/spleen diameter ratio were collected in all patients. Results: The prevalence of EV was 79.6%. The platelet count/spleen diameter ratios are much lower in cirrhosis than those of the temoins (840 vs. 2,469). Especially the cirrhosis with EV the ratio is lower than the cirrhosis none EV (687 vs. 1,434).

Receiver Operating Characteristic Curve (ROC) was used for analysis. Background: Liver Cirrhosis is a chronic liver disease characterized by replacement of liver tissue by fibrous scar (regenerative nodules). Cirrhosis is commonly caused by hepatitis B, C and others. Liver biopsy is gold standard to diagnose is however it is not necessary if there is complete clinical, laboratory, and radiology data. Furthermore, there are risk and complication to do liver biopsy. The severity of cirrhosis is classified with the Child-Pugh score (bilirubin, albumin, INR, ascites, encephalopathy) to classify patients in class A, B, C; while class A has a good prognosis and C has poor prognosis. Transient elastography is a non-invasive, rapid, and reproducible method to evaluate of liver fibrosis by measuring liver stiffness. In cirrhotic patients, liver stiffness measurements range from 12.5 to 75.5 kPa. This study was to know correlation between degree of cirrhosis classified by Child-Pugh criteria and degree of liver stiffness Methods: This was a cross-sectional study. Liver cirrhosis patients classified by Child-Pugh criteria undergone the liver transient-elastography using Fibroscan Ò technique to know the value of liver stiffness.

Result: There were 13 subjects, 69.2% was male with mean age of 50.6 years.The mean value of liver stiffness was 35.53 kPa. There was correlation between degree of cirrhosis and degree liver stiffness (RR 0.774, P = 0.002). Background and Aim: Ascites becomes refractory to medical treatment in 5-10% of patients with cirrhosis showing ascitic decompensation. Refractory ascites (RA) is closely related to a high morbidity and mortality. This study investigated the predictors of the development of RA in patients with hepatitis B virus (HBV)-related cirrhosis experiencing the first episode of ascitic decompensation and determined the prognostic factors of patients with RA. Methods: This retrospective study enrolled 199 consecutive patients with HBV-related cirrhosis experiencing the first episode of ascitic decompensation who was admitted to our institute between January 1996 and December 2008. The end of follow-up was March 30, 2009. The median follow-up period was 55.0 (range 0.2-331.9) months. Results: The mean age of the patients experiencing the first episode of ascitic decompensation was 52.9 ± 10.3 years and males predominated (n = 130, 68.1%). Patients with Child-Pugh C liver function were the most common (n = 119, 62.3%). The overall median survival was 43.1 (range 0.8-331.9) months. Univariate and subsequent multivariate analyses showed that platelet count (P = 0.016), sodium (P = 0.045), and serum potassium (P = 0.037) at admission independently predicted the development of RA. Sixteen patients (8.0%) experienced RA in 8.9 (range 4.6-62.6) months after the first episode of ascitic decompensation. In a subgroup analysis of 16 with RA, the median survival, and 1-, 2-, and 3-year survival rates were 2.7 (range 0.2-16.0) months, 87.5, 75, and 37.5%, respectively. However, we could find no prognostic factor affecting the survival of patients with RA. Conclusions: In patients with HBV-related cirrhosis experiencing the first episode of ascetic decompensation, platelet count, serum sodium, and serum potassium were independent predictors of the development of RA. The survival of patients with RA was poor with no identified independent predictor of survival.

Endoscopic Band Ligation Compared to Cyanoacrylate Injection for Management of Gastric Varices: A Randomized Controlled Study Gamal Shiha 1 , Elsayed Khalil 1 , Ashraf Elfakhry 1 1 Mansoura University Hospital, Mansoura, Egypt

Background and Aim: The number of patients presenting with gastric variceal hemorrhage (GVH) have been increasing in recent years. Gastric variceal obturation (GVO) with cyanoacrylate has been proved effective for controlling bleeding from gastric varices (GV). Also, endoscopic gastric variceal ligation (GVL) for GVH appeared promising and safe. The present work aimed to compare the efficacy, safety and outcome of both modalities.

Patients and Methods: Ninety patients, who presented with haematemesis and/ or melena due to GVH, were included in the study. They were randomly classified into two treatment groups, GVO group; included 45 patients (31 males and 14 females) using iso Amyl-2-cyanoacrylate. GVL group; included 45 patients (33 males and 12 females) using Saeed six shooter. Treatment was repeated regularly until eradication of GV and the patients were followed-up until death or 12 months after the last patient was included.

Results: There were no significant difference in the control of active bleeding in GVO compared to GVL groups {88.9 vs. 93.8%, P [ 0.05}. There were non significant difference in the rebleeding rate in GVL group compared to GVO group (P [ 0.05), however, in exploratory subgroups analysis the rebleeding rate was significantly higher in IGV1 patients and large GV (F3) patients undergoing GVL than those undergoing GVO (P = 0.0363, P = 0.0293, respectively). No significant difference in the eradication of the GV between the two studied groups (P [ 0.05). The recurrence of GV was significantly higher in GVL group compared to GVO group (P [ 0.0164). Univariate analysis showed that rebleeding was significantly linked to the presence of HCC, serum creatinine and size of the GV. Conclusion: Endoscopic management of GVH using GVO or GVL is equally effective and safe. However, rebleeding and recurrence in GVL group are still a problem in patients with IGV1 and patients with large varices.

The Background: Acute variceal bleeding, uncontrolled with endoscopy may require the insertion of the Sengstaken-Blakemore (SB) tube. Inexperience and a difficult anatomy may hamper the insertion of the SB tube or result in esophageal perforation. We describe a novel technique to insert the SB tube, guided with a standard gastroscope.

Case Description: A 74-year-old Chinese male with a medical history of hepatitis B liver cirrhosis and locally advanced hepatocellular carcinoma presented with hemetemesis and melena. After resuscitation and intubation for airway protection, endoscopy was performed. Gastroscopy revealed large esophageal varices with active oozing. Endoscopic variceal ligation was performed. However hours later, he had active bleeding again. This time, endoscopy was unsuccessful. SB tube insertion was attempted, but failed despite multiple attempts, due to coiling. Endoscopic guided insertion of the SB tube was performed. The tip of the SB tube was anchored with a long suture. This was threaded through the instrument channel such that the SB tube was anchored beside the gastroscope. The gastroscope was guided down the esophagus and into the antrum. Within the stomach, the gastric balloon was inflated under direct visualization and the endoscope withdrawn. 24 h later, the SB tube was removed. Repeat variceal ligation was performed, and he was subsequently discharged.

Conclusions: This illustrates a novel method to insert the SB tube endoscopically. It is simple and feasible with a standard sized gastroscope. It allows confirmation of tube placement and the gastric balloon can be inflated under direct visualization. It is also useful in cases where blind insertion is difficult.

Hosny Salama 1 , Abdel Rahman Zekri 1 , Mortada Al Shabrawy 1 , Nahed Hassan 1 , Abeer Bahnasy 1 , Hanan Abdel Halim 1 , Eman Medhat 1 Methods: We have designed a study to assess the clinical benefit. Of amplified and differentiated adult stem cells (CD34+) infused into the hepatic artery, portal vein in patients with chronic liver insufficiency to find out their clinical benefits. We included 258 patients with chronic liver diseases: 15 children with metabolic liver diseases, 241 adults with end-stage liver diseases (218 post-viral, 23 non-viral), 2 patients with post-HCV liver cirrhosis and HCC. For all of them, granulocyte colony-stimulating factor (G-CSF) was administered (5 lm/kg SC) daily for 5 days. On day 6, the blood cells will be collected by leukapheresis. CD34+ stem cells were isolated, cultured for 1 week and injected into hepatic artery or portal vein under CT or ultrasound guidance. Follow-up using clinical and biochemical responses was done weekly for 1 month, then monthly for 18 months. Results: There was normalization of Liver enzymes and improvement in synthetic function in 50% in patients with post-viral liver failure., 94% in nonviral liver failure, 89% children with Metabolic Liver Diseases, no changes in patients with HCC (0%). Conclusion: From this study, stem cell therapy could be a promising therapeutic modality that may supplement patients with end-stage liver disease. Dose standardization and comparing different routes of injection need further assessment. Aim: Patients with advanced liver disease have a complex pattern of defects in haemostatic functions but also a profound deficit of natural anticoagulants such protein C and Antithrombin which may counterbalance the deficiency in procoagulant factors. Patients with liver cirrhosis (LC) have therefore in some cases an increase tendency to develop portal vein thrombosis (PVT). The aim of our study it is to assess the efficacy and safety of low dosage of enoxaparin in patients affected by advanced LC and with PVT. Methods: From august 2008 to august 2009 we studied by ultrasound doppler 155 cirrhotic patients as screening for hepatocellular carcinona. We found fifteen patients with PVT and without hepatocellular carcinoma (prevalence of 9.6%). Six patients with portal cavernoma were excluded from the study. The other nine patients with recent PVT were included in the study. An endoscopic examination and Computed Tomography (CT) was performed in all the patients. After informed consent, nine patients received 100 U/kg/day of enoxaparin subcutaneously for at least 4 months. Follow up by ultrasound with Doppler examination was done every month. CT was done at fourth month. Results: Etiology of cirrhosis was HCV infection in five patients, alcoholic in two, HBV+HDV in one and HBV in one. Six patients were in class A Child Pugh and three patients in class B. Seven patients presented class F1/F2 esophageal varices, while two patients presented F3 class with the evidence of red cherry spot and were previous submitted to band ligation. PVT was occludent in two patients and incomplete in seven patients. Extension to portal branches was noted in three patients while extension to splenic and superior mesenteric vein was disclosed respectively in two and one patients. Complete recanalization of PVT occurred in three patients (33%) of whom one presented a previous total PVT, partial recanalization in six patients (67% Inclusion criteria's: liver cirrhosis, and non cirrhosis dyspepsia. Exclusion criteria's: emergency condition, cardiac and pulmonary diseases, renal insufficiency, malignancy, portal vein thrombosis, liver cancer, post liver resection and liver transplant, peritonitis, medicine consumption that decrease the portal pressure. The hepatic vein velocity was measured by linear measurement of Spectral Doppler Ultrasound in right liver lobe. Child-Pugh score as the severity of liver cirrhosis. The esophageal varices OMED modifying score was determined by upper gastrointestinal endoscopy. The mean differences were analyzed with student ttest or Mann-Whitney U test. Pearson or Spearman's test was used to evaluate the association between HVFV level and Child-Pugh Score, and also between HVFV and OMED esophageal varices. The significance limit was set as at P value of.05. Statistical analysis using SPSS 11.5. Results: There were 30 subjects of liver cirrhosis and 15 controlled subject of dyspepsia enrolled. There was significant difference of HVFV between liver cirrhosis and non-liver cirrhosis (24.857 + 5.135 vs. 14.867 + 1.972 cm/s). (The mean difference was 9.99 cm/s (P \ 0.001, CI 7.836-12.145) No significant positive correlation between HVFV and severity of liver cirrhosis with Child-Pugh score (r = 0.163, P = 0.389). There was significant positive correlation between HVFV and esophageal varices OMED modifying score. (r = 0.957, P = .001) The ROC curve point of HVFV was 21.700 cm/s (sensitivity 0.895, specificity 0.636) which predicted the variceal bleeding. Conclusion: Hepatic Vein Flow Velocity measurement by using Ultrasound Doppler could be used to determine the degree of esophageal varices and predict the variceal bleeding.

Ehab Abdel-Khalek 1 , Ibrahem Abdel-Aal 2 1 Internal Medicine Department, Faculty of Medicine, Mansoura University, Egypt, 2 Clinical Pathology Department, Faculty of Medicine, Mansoura University, Egypt

Background: Chronic liver disease leading to cirrhosis is the most common cause of portal hypertension which may end in serious bleeding from gastrooesophageal varices. Recent studies have demonstrated a daily pattern of acute upper gastrointestinal bleeding in patients with liver cirrhosis evidenced by one or two peaks throughout the day. We aimed at the assessment of the circadian rhythm of acute variceal bleeding with the possible participation of circadian changes of the fibrinolytic parameters. Methods: The study included 264 patients with liver cirrhosis and upper gastrointestinal bleeding in addition to 20 healthy subjects as a control group. The assessment of fibrinolytic parameters was completed in 60 of the patient group in addition to the control group. The fibrinolytic activity was assessed by estimation of Tissue plasminogen activator antigen (tPA: Ag) and Plasminogen activator inhibitor antigen (PAI-1: Ag), the latter is considered the fast acting inhibitor of plasminogen activators. Result: We observed statistically significant 2 time peaks of upper gastrointestinal bleeding at hour 0400 and hour 1700 with overlapping peak of the fibrinolytic parameter, tissue plasminogen activator antigen, with the night peak of bleeding. Conclusion: There are two time peaks of upper gastrointestinal bleeding with a temporal association between the night peak and a relative hyperfibrinolytic state. Background/Aims: Protective immune response plays an important role in eliminating HBV and HCV infection. A weak immune response to HBV and HCV can lead to chronic liver disease. This study aims to characterize functional variations of immune status in cirrhosis and determine correlations between HBV-and HCV-cirrhosis and their immunologic features. Methods: Peripheral blood was collected from patients with HBV-cirrhosis (n = 41), HCV-cirrhosis (n = 23) and healthy individuals (n = 21). Phenotypes of peripheral blood lymphocyte subsets (T cells, NK cells, B cells, regulatory T cells) were analysed using flow cytometry and Th cytokine secretion was assessed by cytometric bead array (CBA).

Results: There was no significant difference between the cirrhotic groups except in IFN-c levels. Cirrhotic patients had a lower proportion of CD3+CD8+T cells, CD27+ B cells and NK cells, while the proportion of CD3+CD4+T cells, CD27-B cells, Treg cells and the ratio of CD4/CD8 were significantly higher than that of healthy controls. The levels of Th2 cytokines (IL-6,IL-10) in cirrhotic patients were increased dramatically, while only the Th1 cytokine (IFN-c) increased in HBV-cirrhotic patients (P = 0.037). Conclusions: These findings provide evidence that there is no significant difference in immunological characteristics between the cirrhotic groups except in IFN-c levels. In post-hepatitic cirrhosis, diffuse defects in innate, adaptive, cellular and humoral immune cells are likely to be maintained regardless of which virus is involved. A cytokine imbalance oriented toward Th2-type responses may play a role in the development of post-hepatitic cirrhosis. 

Branched-chain Amino Acids Improve the Maturation of Dendritic Cells in Patients with Advanced Cirrhosis ex vivo Eiji Kakazu 1 , Yoshiyuki Ueno 2 , Yasuteru Kondo 1 , Koji Fukushima 1 , Jun Inoue 1 , Keiichi Tamai 1 , Noriyuki Obara 2 , Osamu Kimura 2 , Yuta Wakui 2 , Tooru Shimosegawa 2 1 Tohoku University Hospital, 1-1 Seiryo, Aobaku, Sendai, Japan, 2 Tohoku University Graduate School of Medicine, 1-1 Seiryo, Aobaku, Sendai, Japan Background/Aims: An imbalance of plasma amino acids, with decreased levels of branched-chain amino acids (BCAAs), is commonly seen in patients with advanced cirrhosis. Recently, we have reported that the imbalance in plasma amino acids of patients with advanced cirrhosis suppresses the maturation of dendritic cells via mTOR/S6K signaling pathway. The aim of this study is to clarify the immunological efficiency of BCAAs ex vivo. Methods: We made two media: a serum free culture medium consistent with the average concentration of the plasma amino acids from a healthy volunteer (HC: n = 25) was defined as the healthy control medium (HCM); whereas that from patients with advanced cirrhosis (LC: n = 43) was defined as the advanced cirrhotic medium (ACM). PBMCs or DCs (BDCA1 + DCs and MoDCs) were stimulated by LPS or poly I:C under HCM and ACM. For examination of the DCs function, the phenotypes were determined by flow cytometry. As in the ex vivo study, we stimulated PBMCs from LC (n = 5) by LPS or poly I:C under autologous plasma, which was collected both before and after oral administration of BCAA granules (a mixture of valine: 1.144 g, leucine: 1.904 g and isoleucine: 0.952 g). We recovered the plasma and measured cytokines by ELISA. Results: After adding the stimulants, the CD83 and CD86 expression of DCs from LC were lower than those from HC in both HCM and ACM. In both HC and LC, the CD83 and CD86 expression of DCs stimulated under HCM was higher than that under ACM. As in the ex vivo study, the plasma concentration of BCAAs in LC was maximum 60 min after oral administration. Fischer's ratio was increased from 1.37 ± 0.98 (SD) to 4.94 ± 0.99 (SD). We stimulated PBMCs from LC using autologous plasma before and after 60 min oral administration. In all cases, PBMCs stimulated by LPS in the latter had more interferon gamma production than in the former. Conclusion: Oral administration of BCAA granules enhanced the maturation and function of myeloid dendritic cells in patients with advanced cirrhosis ex vivo.

Conclusions: Octreotide decreases portal pressure in cirrhotic patients by inhibiting HSC contractility by decreasing intracellular Ca 2+ concentration via stimulation of all SSTRs on HSCs.

The Cardiac Hemodynamics Change in Liver Cirrhotic Patients Lei Li 1 , Chun-Xia Ping 1 , Hui-Guo Ding 1 1 Beijing You'an Hospital, affiliated with Capital Medical University, No. 8, youanmenwai street, Beijing 100069 Objective: The aim of this study is to investigate the cardiac hemodynamics change in liver cirrhosis patients with various degrees and to clarify the relationship between Systolic function of cardiac muscle and prognosis of portal hypertension. Methods: Forty-five liver cirrhosis patients were investigated. According to childpugh score, they were divided into child-pugh A group (n = 12), child-pugh B group (n = 17) and child-pugh C group (n = 16). Noninvasive cardiac hemodynamics detection appearance was applied to determine hemodynamic parameters. Results: With the aggravation of liver function, mean arterial pressure (MAP), stroke volume (SV), stroke index (SI), cardiac output (CO), cardiac index (CI), systemic vascular resistance (SVR), systemic vascular resistance index (SVRI), left cardiac work (LCW) and left cardiac work index (LCWI) take on degrading, which in child-pugh C group are significantly lower than that in child-pugh A group (P \ 0.05). Conclusions: Cardiac hemodyanic parameters could be effective to determine the development degree of liver cirrhosis. Systolic function of cardiac muscle degrades with the aggravation of liver function. Objective: Activated Hepatic stellate Cells (HSCs) plays very important role during the progression of the hepatic fibrosis. HSCs have biological characters of smooth muscle cells and help to adjust intrahepatic resistance and portal venous pressure. The contraction and relaxation of HSCs can be affected by vasoactive substance such as angiotonin, endothelin-1, nitric oxide (NO) and carbon monoxide (CO). Hydrogen Sulfide (H 2 S) is a new kind of endogenous gas signaling molecule except for NO and CO. We aimed to study the production of H 2 S and its significance. Methods: Activated rat hepatic stellate cells line (HSC-T6) was used. They were divided into three groups and were repeated six times in every group. After conventional culture, cystathionine-c-lyase (CSE) mRNA expression of different cell groups was detected by RT-PCR and H 2 S production rate was detected by methylene blue spectrophotometry. Results: HSCs can express CSEmRNA and can secrete H 2 S. DL-PPG (CSE inhibitor) can suppress H 2 S production rate significantly (P \ 0.05). Conclusions: HSC can express H 2 S synthetase/ CSE and substrates can form H 2 S with effect of this enzyme in the suitable reaction conditions.

Interleukin-1A but Not Interleukin-1B Gene Polymorphisms is Associated with HBV-Related Hepatic Cirrhosis Kun-He Zhang 1, 2 , Jun Ma 1 , Gen Huang 1 , Shu-Qin Wei 1 , Xin-Ying Tan 1 , Nong-Hua Lv 1 , Ji-Xiang Zhang 2 1 The First Affiliated Hospital, Nanchang University, No 17, Yongwai Zheng Street, Nanchang, Jiangxi, China, 2 The Second Affiliated Hospital, Nanchang University, No 1, Mengde Road, Nanchang, Jiangxi, China Background: Little is known about the relationship between Interleukin-1A (IL-1A) gene polymorphism and HBV infection. The aims of the present study were to analyze the correlations of IL-1A and IL-1B gene polymorphisms with the susceptibility of HBV-related hepatic cirrhosis. Methods: One hundred and forty eight patients with HBV-related hepatic cirrhosis (case group) and 127 asymptomatic HBsAg carriers with negative HBeAg and persistent normal serum level of alanine aminotransferase (ALT) (control group) were enrolled into this study. The genotypes of IL-1A-889C/T, L-1B-551C/T and IL-B1 +3953C/T loci were determined by PCR-RFLP. The correlations of genotypes, alleles and haplotypes of the three loci with the susceptibility of HBV-related hepatic cirrhosis were analyzed. Results: Univariate analyses indicated that genotype CC and allele T of IL-1A -889C/T locus were correlated with susceptibility of HBV-related hepatic cirrhosis (OR = 0.489, P = 0.047; OR = 2.046, P = 0.047), although the significances of their ORs were weakened when controlling age and gender (OR = 0.506, P = 0.065; OR = 1.796, P = 0.065).The other polymorphisms were insignificant. Multivariate analyses also showed that genotype CC and allele T of IL-1A -889C/T were independently correlated with HBV-related hepatic cirrhosis (OR = 0.430, P = 0.024; OR = 2.328, P = 0.024) and remained significant when controlling age and gender. Genotype CC and allele T of IL-1B +3953C/T were also independent factors associated with HBVrelated hepatic cirrhosis, but just nearly significant. Haplotype C-C-T only existed in HBsAg carrier group (P = 0.001), and haplotype T-C-T almost only existed in hepatic cirrhosis group (P = 0.033), but both were rare haplotypes (\4%). Conclusion: The susceptibility of HBV-related hepatic cirrhosis was closely associated with IL-1A -889C/T locus, weakly associated with IL-1B +3953C/T locus but not associated with IL-1B -551C/T.

The Association of Polymorphisms of Transforming Growth Factor-b1 with the Susceptibility of HBV-related Hepatic Cirrhosis Dong-Hai Yan 1 , Kun-He Zhang 1 , Jun Ma 1 , Gen Huang 1 , Shu-Qin Wei 1 , Xin-Ying Tan 1 , Xuan Zhu 1 , Nong-Hua Lv 1 1 First Affiliated Hospital, Nanchang University, No 17, Yongwai Zheng Street, Nanchang, Jiangxi, China

Background: Transforming growth factor-b1 (TGF-b1) is closely correlated with liver fibrosis. The polymorphisms of TGF-b1 gene is related with TGF-b1 expression. Here, we analyzed the association of TGF-b1 gene polymorphisms with he susceptibility of HBV-related hepatic cirrhosis. Methods: One hundred and nineteen patients with hepatic cirrhosis due to hepatitis B virus infection (case group) and 129 asymptomatic HBsAg carriers with negative HBeAg and persistent normal serum level of alanine aminotransferase (ALT) (control group) were enrolled into this study. The genotypes of TGF-b1 -509C/T, +869T/C and +915G/C loci were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Correlations of genotypes, alleles and haplotypes of the three loci with the susceptibility of HBV-related hepatic cirrhosis were analyzed. Results: Univariate analyses showed that genotype CC of TGF-b1 +869T/C was more frequent in case group than control group (45.7 vs. 30.6%, P = 0.015, OR = 1.926), and allele T of this locus was more frequent in control group than case group (33.2 vs. 43.5%, P = 0.021, OR = 0.539). There were no significant differences of frequencies of genotypes and alleles of TGF-b1 -509C/T between two groups. No polymorphism was detected at TGF-b1 +915 G/C loci. Multivariate analyses showed that genotype CT of -509C/T and genotype CC of +869T/C were independent risk genotypes (OR = 1.732, P = 0.054; OR = 2.194, P = 0.005), and allele T of +869T/C was a independent protective allele (OR = 0.532, P = 0.019). The overall frequency difference of four haplotypes constructed on -509C/T and +869T/C loci was significant (P = 0.008), and haplotype C-C is a risk factor (P = 0.001, OR = 2.987) and haplotype C-T is a protective factor (P = 0.012, OR = 0.586). Conclusion: The polymorphisms of TGF-b1 -509C/T and +869T/C loci are associated with susceptibility of HBV-related hepatic cirrhosis.

The Gender Difference in Susceptibility of HBV-related Cirrhosis is Associated with Polymorphisms of Several Genes Kun-He Zhnag 1 , Gen Huang 1 , Jun Ma 1 , Shu-Qin Wei 1 , Dong-Hai Yan 1 , Xin-Ying Tan 1 , Xuan Zhu 1 , Nong-Hua Lv 1 1 The Department of Gastroenterology, First Affiliated Hospital, Nanchang University, No 17, Yongwai Zheng Street, Nanchang, Jiangxi, China Background: Gender difference is clear in the susceptibility of HBV-related hepatic cirrhosis (more frequent in male than female), which indicates that genetic difference could be one of mechanisms. Here we analyzed a group of gene polymorphisms in patients with chronic infection of hepatitis B virus and observed the differences between the male and the female. Methods: A total of 323 patients were recruited into our study, consisting of 168 patients with HBV-related cirrhosis (case group) and 155 asymptomatic HBsAg carriers with negative HBeAg and persistent normal serum level of alanine aminotransferase (ALT) (control group). The genotypes of 16 loci from 11 genes were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The odds ratio of each genotype and allele in male and female group was analyzed, respectively, by univariate Logistic regression. Results: The polymorphism loci with difference between male and female group were showed in Table 1 . Conclusion: The gene polymorphisms of estrogen receptor-a +29T/C, interleukin 1B +3953C/T, interleukin 10 -592A/C, interferon-c +874T/A and microsomal epoxide hydrolase -613C/T loci are associated with gender difference in susceptibility of HBV-related hepatic cirrhosis. Background: Only a small part of patients with chronic infection of hepatitis B virus will finally develop hepatic cirrhosis. The genetic background might be an important factor for the risk of cirrhosis development. Here we analyzed a group of polymorphisms of liver enzymes for toxin metabolism and explored their relationship with susceptibility of HBV-related hepatic cirrhosis. Methods: One hundred and sixty eight patients with HBV-related cirrhosis (case group) and 155 asymptomatic HBsAg carriers with negative HBeAg and persistent normal serum level of alanine aminotransferase (ALT) (control group) were enrolled into our study. The genotypes of matrix metalloproteinase-9 (MMP-9) -1562C/T, microsomal epoxide hydrolase (MEH) Tyr113His and -613C/T, CD14 -159C/T loci were determined by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Glutathione S-transferase (GST) M1 gene was detected by polymerase chain reaction. The frequency of each genotype or allele was calculated and its difference between two groups was analyzed.

Results Background: Bacterial infections are frequently documented in cirrhotic patients presenting with upper gastrointestinal bleeding. Bacterial infection is independently associated with failure to control bleeding in cirrhotic patients with gastrointestinal bleeding. Bacterial infection may trigger the production of endotoxin, which in turn stimulates production proinflamatory citokin (IL-6) and in the next step stimulates endothelin and expression of inducible nitric oxide synthase. Subsequently, endothelin may stimulate stellate cells to contract and results in elevation of variceal pressure. Production of nitric oxide reduces platelet aggregation and primary hemostasis. As a result, the chain reaction induces variceal bleeding. This study aim to know the association between IL-6 and upper gastrointestinal bleeding in liver cirrhosis patients. Methods: We performed A cross-sectional analytic study in liver cirrhosis patients, in sanglah general hospital Bali. We used a student t test to know the mean different of IL-6 between patient with or without upper gastrointestinal bleeding. Result: We collected 55 liver cirrhosis patients. Fourty one (74.5%) out of this was male and the rest was female. With range of age between 30 to 78 years old. Base on Child-pugh classification 56.4% with class C, only 5.5% with class A and the rest was class B. We found 12.7 and 65.4% out of patients with hepatic encephalopathy and ascites, consecutively. Thirty (54.5%) out of 55 patients with acute bleeding. We found significant higher mean level of IL-6 in liver cirrhosis patient with upper gastrointestinal bleeding compared with those without upper gastrointestinal bleeding (39.3 vs. 17.8 pg/mL; P = 0.02). Conclusions: Higher IL-6 level was associated with upper gastrointestinal bleeding. Background: Gall bladder stones are considered a public health problem allover the world. Many factors have been proposed to explain the increased incidence of gallstones in liver cirrhosis as alcoholism, changes in total bile acid pool, decreased cholesterol secretion, hemolysis secondary to hypersplenism, increased oestrogen levels and changes in gall bladder and sphincter of odd motility, but the exact mechanism has not been yet elicited. Our objective was to study gall bladder contractility and prevalence of gallstones in patients with liver cirrhosis in addition to analysis of the effect of portal hypertension on these two parameters.

Methods: Eighty patients with chronic liver disease were enrolled (60 patients with liver cirrhosis and 20 patients with pure hepatosplenic schistosomiasis). Ultrasonographic diagnosis of gallstones with evaluation of gall bladder contractility and portal hypertension were done.

Result: An increase in the number of subjects with gallstones with an increase of gall bladder fasting volume and residual volume with a decrease in gall bladder ejection fraction in the patients than the controls. Also there was an increase in the percentage of patients with gallstones among those with higher grades of portal hypertension. Conclusion: Patients with liver cirrhosis have higher frequency of gallstones with diminished gall bladder contractility and higher measures of portal hypertension.

Endoscopic Ultrasonography or Retrograde Cholangiopancreatography for Suspected Choledocholithiasis? Max Petrov 1 , Tom Savides 1 1 The University of Auckland, Private Bag 92019 Auckland 1142, Australia

Introduction: Endoscopic ultrasonography (EUS) has recently emerged as an accurate diagnostic alternative to endoscopic retrograde cholangiopancreatography (ERCP). The aim of this study was to systematically review all randomized controlled trials on EUS-guided ERCP versus ERCP-only strategy in patients with suspected choledocholithiasis. Methods: The search for eligible studies was done in three electronic databases. The results of individual trials were statistically aggregated to obtain a pooled risk ratio (RR) and corresponding 95% confidence interval (95% CI) for each outcome.

Results: There were a total of 14 complications in 213 patients (7%) in the EUS-guided ERCP group as compared with 40 complications in 210 patients (19%) in the ERCP-only group. The use of EUS-guided ERCP strategy, in comparison with ERCP-only strategy, was associated with a lower risk of overall complications (RR 0.35; 95% CI 0.20-0.62; P \ 0.001). The risk of post-ERCP acute pancreatitis was significantly reduced in the EUS-guided ERCP group in comparison with the ERCP-only group (RR 0.21; 95% CI 0.06-0.83; P = 0.03). At the same time, the risk of bleeding did not differ significantly between the groups (RR 0.49; 95% CI 0.10-2.44; P = 0.38). Also, there was no difference between the groups with regard to in-hospital mortality (RR 2.00; 95% CI 0. 19-21.56 ; P = 0.57). In the EUS-guided ERCP group, EUS failed to detect common bile duct stones in 2 of 213 (0.9%) patients and a total of 143 ERCPs were avoided when EUS did not detect choledocholithiasis (67%). Conclusion: ERCP may be safely avoided in two-thirds of patients with common bile duct stones if EUS is performed first. Application of EUS in selection of patients for therapeutic ERCP significantly reduces a number of complications. Background/Aims: The size of the diverticula and location of the papilla in relation to periampullary duodenal diverticula (PADD) are variable. We aimed to determine if PADD subtypes are correlated with clinical severity in choledocholithiasis patients undergoing ERCP. Methods: Data of 120 patients who underwent CBD stone(s) extraction by ERCP and also presented PADD from January 2005 to December 2007 were retrospectively analyzed. Analyses of variables including PADD types by location and size, CBD stone size, and incidence of SIRS or sepsis were performed. Results: Type 1 PADD patient group was associated with larger PADD (P \ 0.001) and CBD stone size (P = 0.001), with greater WBC level (P = 0.015) and more frequent Systemic Inflammatory Response Syndrome (SIRS) (P = 0.009) incidence compared with type 2 or 3 PADD group. Large PADD group presented larger CBD stone size (P \ 0.01); and showed tendency for increased SIRS incidence (P = 0.091). On multivariate analysis type 1 (OR: 18.70, CI: 1.43-243.43, P = 0.025) and larger PADD (OR: 7.15, CI: 2.179-23.46, P = 0.001) were significant contributors for CBD stones larger than 10 mm of diameter. Conclusion: Type 1 and large PADD patients are associated with larger CBD stones and showed a tendency for increased SIRS incidence. Therefore clinician must be aware about the possibility of a more complicated evolution in patients with these risk factors.

Ghous Soomro 1 , Zaigham Abbas 1 , Nasir Luck 1 , Mujahid Hassan 1 , Yousuf Memon 1 , Anwar Naqvi 1 , Adib Rizvi 1 1 Sindh Institute of Urology and Transplantation, Karachi, Pakistan

Background: The etiology of biliary atresia is poorly understood. This extrahepatic obstructive cholangiopathy is progressive even after birth resulting in worsening neonatal cholestatic jaundice eventually leading to secondary biliary cirrhosis and liver failure. The process seems to be an acquired one with a possible viral etiology. We examined this hypothesis by testing our patients for exposure to TORCH infections (toxoplasma, rubella, CMV, herpes) and Ebstein Bar Virus (EBV), Methods: Test performed on these patients included PCR for CMV and Ebstein Bar Virus (EBV), IgM antibodies for CMV, toxoplasma, rubella, and herpes infections. Results: Thirty-three infants were included in the study, male 22, median age of diagnosis 2.5 months (range 1.0-5.0 months). Median birth weight 2.9 kg (2.5-3.2), median gestational age 39 (38-40). All of these patients presented with jaundice, diarrhea, clay colored stools, poor feeding and failure to thrive. 32 (97%) were febrile and 31 (93.9%) were having pruritis,. On examination pallor, edema and hepatosplenomegaly in 100%, and ascites and microcephaly 

Oyuntsetseg Batkhuu 1 , Dagvadorj Byamba 2 , Nyamsuren Dagvadorj 2 , Bira Namdag 1 1 Health Sciences University of Mongolia, Sukhbaatar district, 2 Eleg Hospital, Bayanzurkh district

Aim: Increased gallstone prevalence in liver disease have not been studied yet in Mongolia. To explore the prevalence of cholelithiasis in chronic viral hepatitis.

Methods: A total 14,561 subjects were enrolled between January 2001 to April 2006, who was admitted in ''Eleg'' hospital of Ulaanbaatar in inpatients and outpatients clinic. Evalution for each subject included a complete history and physical examination and abdominal ultrasound. Gallstones were identified by the presence of strong intraluminal, gravity dependent echoes producing posterior acoustic shadowing. The statistical analysis was performed using SPSS version 15.0 for Windows and E views 3.1 programme. A two tailed P value of .05 was considered statistically significant between groups. Results: 5,895 patients with chronic hepatitis consisting of 3,033 (51.4%) males and 2,862 (48.6%). The prevalence of GS in the chronic hepatitis was 4.3% (5.9% for females, 2.7% for males). Patients were divided into four groups: group I: chronic hepatitis B (n = 1,122), group II: chronic hepatitis C (n = 1,336), group III: chronic hepatitis D (n = 24), and group IV: chronic hepatitis B + C (n = 174). A total of 257 patients with gallstone revealed in this study and 180 (70%) were women and 77 (30%) were men. Their mean age was 47. 1 ± 12.4 (range 22-83) . The male to female ratio was 1:2.3. Female predominant was statistically significant (P \ 0.05). The study revealed a significant increase in the incidence of gallstone in hepatitis C and B. In group I and II patients, gallstone appeared more frequently (8.7, 11% respectively) than in group III and IV (4.1, 6.3%) (v 2 = 6.742, P \ 0.002). Conclusion: Overall prevalence of GSD in patients with chronic viral hepatitis was 4.3% (5.9% for females and 2.7% for males). Cholelithiasis occurs frequently in patients with chronic viral hepatitis C and B contributes to the burden of the disease.

Endoscopic Management of Biliary Disorders during Pregnancy Anand Jalihal 1 , Vui Heng Chong 1 1 Gastroenterology Unit, Department of Medicine, RIPAS Hospital, Bandar Seri Begawan BA1710, Brunei

Background: Biliary interventions during pregnancy are associated with risks to both the pregnancy and developing fetus. Methods: Endoscopic retrograde cholangiopancreatography (ERCP) carried out during pregnancy (May 2003 to Sep 2009) were identified from our database. All procedures were carried out using conscious sedation and lead shielding. Results: During this period, 7 patients had 8 ERCP (8/586, 1.4%). The median gestation was 22.5 week (\2-36); 2, 4 and 2 patients were in their first, second and third trimester, respectively. Indications included cholangitis (n = 2), obstructive jaundice (n = 5) and acute pancreatitis/obstructive jaundice (n = 1). Two patients had endoscopic ultrasound (EUS) before ERCP, which provided additional information. Fluoroscopy was used in four procedures (median 7 s, 2-20) and overall procedure time ranged from 5-15 min. Treatment consisted of endoscopic sphincterotomy (n = 4), stenting (n = 5) and balloon clearance (n = 3). One procedure related complication occurredinduction of labor. During the course of pregnancies, there were four nonprocedures related complications (1-acute cholecystitis, 1-HELLP syndrome resulting in spontaneous abortion and 2-stent migrations secondary to followup default). Five pregnancies had uncomplicated term deliveries while two required urgent caesarian sections (1-fetal distress and 1-cholangitis secondary to stent migration). All seven babies were well at birth with median APGAR scores of 9 and 10 at 5 and 10 min, respectively. One baby died of sudden infant death syndrome at 40 days old. Conclusion: ERCP is safe during pregnancies. Different strategies can be applied depending on situations and expertise of endoscopist. Most can be managed with biliary stenting and subsequent clearance can be made after deliveries. EUS has a complementary role. However caution should be taken to minimize radiation exposure and procedure time. Objective: To investigate the prevalence of and risk factors for cholelithiasis among adult residents in Shanghai. Methods: A cross-sectional survey with multiple-stage stratified cluster and random sampling was performed. All residents aged 15 and above were invited to participate in the survey, they came from four communities of Yangpu District and Pudong New District. Questionnaire, physical examination,serum lipid-profile,and 75 gram oral glucose tolerance test and ultrasonographic examination of liver and biliary system were undertaken. Analysis of data was performed through SPSS 15.0 for Windows statistical package.

Results: A total of 3,173 residents took part in the survey, which was 2.26/ 10,000 of Shanghai municipal residents. Of the 3,173, 1,217 were males and 1,956 were females. The mean age of the participants was 52.4 ± 15.1 years and ranged from 15-88 years. Cholelithiasis was detected with ultrasound examination in 339 participants (10.7%), the age-and sex-adjusted prevalence of cholelithiasis in Shanghai adult residents was 7.5%. The prevalence of cholelithiasis was increased with aging both in males and females. The age of years, body mass index, waist circumference, blood pressure, fasting and 2 h serum glucose level, triglyceride, low-density lipoprotein cholesterol (all P \ 0.01), total cholesterol(P \ 0.05)in the cholelithiasis group was significantly higher than those in the group without cholelithiasis, and the highdensity lipoprotein cholesterol level was lower in the cholelithiasis group. Obesity, hypertension, impaired glucose tolerance, diabetes mellitus,and ultrasonographic fatty liver were positively associated with the risk of gallstone disease in both men and women. However, central obesity, hypertriglyceridemia and hypercholesterolemia were risk factors for gallstone disease only in women. Logistic regression analysis demonstrated that the prevalence of cholelithiasis was only positively correlated to three risk factors,including advanced age, systolic blood pressure and fatty liver. Conclusions: There is a high prevalence of cholelithiasis among adult residents in Shanghai. Advanced age, hypertension and fatty liver are closely associated with cholelithiasis.

than those of healthy controls (P \ 0.01). The serum VEGF levels were higher in the cases in medium-term or later stage than those in early stage cases (P \ 0.01).The level of serum VEGF was higher in the died SACLF patients than that in the survivals. Conclusion: The serum levels of VEGF in patients with SACLF were closely associated with severity and prognosis of the disease. VEGF may play an important role in the pathogenesis of SACLF.

Characteristics of Patients with Acute-on-Chronic Liver Failure (AoCLF) in Jakarta, Indonesia Cosmas Rinaldi Lesmana 1,2 , Laurentius A. Lesmana 1,2 , Lidwina Cahyadinata 2 1 Div. Hepatology, Dept. of Internal Medicine, Univ. of Indonesia, Salemba Raya no.6, Jakarta Pusat, Indonesia, 2 Digestive Disease Center, Medistra Hospital, Gatot Soebroto 59, Jakarta Selatan, Indonesia

Background: Acute-on-chronic liver failure (AoCLF) is associated with high mortality rate. Study on ACLF in Indonesia is still lacking. While mortality rate is certainly high, clinical characteristics, etiology, and precipitating factors may differ. Patients and Method: This was a cross-sectional study of patients with AoCLF (based on APASL Consensus Definition 2008) in one centre in Jakarta between 2004 and 2007. Clinical data were collected from the patients' medical record. Clinical endpoint was either the patient's death or survival. Data were analyzed descriptively using SPSS version 12. Results: There were 20 patients included in the study with a mean age of 50.3 ± 15.45 years (27-83 years). Thirteen (65%) patients were men. The mean serum bilirubin level was 25.6 ± 10.86 mg/dL and the mean MELD Score was 32.8 ± 6.5. The etiology was hepatitis B virus infection in 14 (70%) patients, followed by hepatitis C virus in 5 (25%) patients and fatty liver in 1 (5%) patient. The most common precipitating factor was Chinese herbal or traditional medicines (25%), followed by hepatotoxic drugs (15%), infection (15%), and bleeding (10%). Other clinical manifestations were hepatorenal syndrome (70%). 18 patients (90%) were died. Conclusion: Cases of acute on chronic liver failure have recently been identified in Jakarta, Indonesia. Hepatitis B was the most common cause of chronic liver disease, while herbal or traditional medicine was the most common precipitating factor. Background: To report the clinical experience, biochemical findings, complications and maternal outcome in patients with acute liver failure in acute fatty liver of pregnancy. Patients and Methods: Retrospective study over a period of 8 years (1998) (1999) (2000) (2001) (2002) (2003) (2004) (2005) . The diagnosis of acute liver failure (ALF) and acute fatty liver of pregnancy (AFLP) was confirmed by liver biopsy and/or by clinical symptoms, and laboratory findings. Results: Twelve women were included in this study with a mean age of 28 ± 3.2 years. 33.3% of cases were primigravid and all cases are singleton pregnancy. The mean gestational age was 36 ± 1.3 weeks (range 34-38 weeks). The fetus was a male infant in 66% of cases. Nausea and vomiting were the most common symptoms (75%). The complications observed were mainly: acute liver failure in 100% cases, coagulation disorders in 100%, acute renal function damage in 100%, hypoglycemia in 83.3%, postpartum hemorrhage in 50%, coma in 33.3%, anuria in 25% and acute pancreatitis in 8.3%. During the hospital, three patients (25%) presented psychiatric symptom, manifested as visual hallucination, auditory hallucination, and delusion of persecution. There was one maternal death (8.3%) and 2 fetal deaths (16.7%). Discussion and Conclusion: Intensive care facilities and an early diagnosis are essential for the management of mothers with acute liver failure of AFLP. Reversible peripartum liver failure may be diagnosed and managed on the basis of clinical and laboratory criteria. With adequate support, the most patients may have full recovery of hepatic function.

The Background: To observe the protective effect of ethyl pyruvate (EP) on acute hepatic injury rat models induced by D-galactosamine hydrochlorate. Methods: 48 male Wistar rats were used as the acute hepatic injury animal model induced by D-galactosamine hydrochlorate and divided into six groups randomly, including normal group, model group, early interference of low dose EP group (20 mg/kg), early interference of high dose EP group (40 mg/kg), low dose EP treatment group (20 mg/kg) and high dose EP treatment group (40 mg/kg). At 24 h after modeling, the serum levels of TNF-a, IFN-c, IL-10 and IL-18 were assayed by ELISA, the changes of HMGB1 mRNA in liver tissue were determined by RT-PCR, and the pathological changes of liver tissue were detected by HE staining. Results: The levels of HMGB1 mRNA of liver tissue in model group rats were significantly higher than normal group rats (P \ 0.01), while serum levels of TNF-a, IFN-c, IL-10 and IL-18 in model group had no obvious changes (P [ 0.05). Compared with model group, the levels of TNF-a, IFN-c, IL-10 and IL-18 had no obvious changes in EP early interference group or EP treatment group, but the levels of HMGB1 mRNA in EP early interference group were obviously decreased (P \ 0.01), especially in high-dose group, which was more obvious than EP treatment group (P \ 0.01). It was shown by histopathological assay that the liver histology was most significantly improved in EP early interference group, especially in high dose group. Conclusion: Ethyl pyruvate could protect effectively the acute liver injury induced by D-galactosamine hydrochlorate in rats, the protective effect was shown as dose-dependent and time-dependent manner. However, the optimal interference time point is need for further study.

(MELD) and Child-Pugh score (CPS), are mainly being accepted to determine the prognosis of patients with end-stage liver disease, However, these models are still controversial in predicting mortality of patients with acute-on-chronic hepatitis B liver failure (ACHBLF). In this study, we aim to evaluate the possibility to better predict mortality of the hospitalized patients with ACHBLF using a novel logistic regression model (LRM Background and Aim: Severe acute hepatitis B is a rapid deterioration of liver function and carries a high mortality. The aim of this study was to evaluate the efficacy of lamivudine in patients with severe acute hepatitis B. Methods: Eighty patients with severe acute hepatitis B were randomly divided into the lamivudine and control group. HBsAg, HBeAg seroconversion rates, serum HBV DNA negative rate, biochemical indicators, the incidence of liver failure and mortality of patients in two groups were compared. The influential factors on the mortality were studied by Cox proportional hazards model. Results: The improvement in serum TBiL, INR and HBV DNA levels of lamivudine group was significantly greater than that of control group. The mortality (7.5%, 3/40) of lamivudine group was significantly lower than that (25.0%, 10/40) of control group (P = 0.034). The incidence of liver failure (8.7%, 2/23) of patients receiving lamivudine within a week was significantly lower than that (35.3%, 6/17) of patients receiving lamivudine after a week (P = 0.038). In multivariate Cox proportional hazards analyses, age (P = 0.043), ratio of total to direct bilirubin (P = 0.009), treatment method (P = 0.006) and the decline of HBV DNA load during therapy (P = 0.017) were independent predictors of mortality. The HBsAg seroconversion rates (62.5%, 25/40) and HBeAg seroconversion rates (63.6%, 21/33) of lamivudine group were significantly lower than those (85.0%, 34/40), (87.5%, 28/32) of control group (P = 0.022, 0.026).

Conclusions: Early treatment with lamivudine causes a greater decrease in HBV DNA level and better clinical improvement and mortality improvement in patients with severe acute hepatitis B, but with a lower seroconversion rate. A rapid decline of HBV DNA load is a good predictor for the treatment outcome. Background: Growing body of evidences have shown that impaired liver regeneration is one of factors related to high mortality in patients with acute liver failure (ALF), however, the mechanism involved in impaired liver regeneration in patients with ALF are not fully understood. In this study, we explore the effect of plasma from ALF on growth and proliferation of HepG 2 cells as well as on the expression of intracellular cyclin D1 and cyclin dependent kinase 4 (CDK4). Methods: The plasma from 3 patients with ALF were collected during the treatment of plasma exchange and were stored at -40°C. HepG 2 cells were cultured with 50% plasma from patient with ALF for 6, 12, 24, 48 and 72 h with or without 10 ng/ml Epidermal Growth Factor (EGF) stimulation for 12 h. Plasma from normal subject was used as normal control (NC). Cell proliferation was studied by MTT assay and intracellular cyclin D1, CDK4 expression in HepG 2 cells was analyzed by western blotting.

Results: (1) The growth and proliferation of HepG 2 cells was significantly inhibited by treatment with 50% plasma from patients with ALF from 12 to 72 h when compared with 50% NC plasma. (2) 10 ng/ml EGF significantly induced HepG2 proliferation in presence of either 50% ALF plasma or NC plasma, however, the proliferation of HepG2 cells was still significantly inhibited in presence of 50% ALF plasma when compared with that of NC plasma.

(3) Western blot analysis showed that the expression of cyclinD1 in HepG 2 cells was obviously decreased and the expression of CDK4 was also down-regulated after treatment with ALF plasma from 12 to 72 h. Conclusion: Our results suggested that plasma from patients with ALF inhibit the growth and proliferation of HepG 2 cells in vitro. One of possible mechanisms by which plasma from patients with ALF inhibited the growth and proliferation of HepG 2 cells is by down-regulating the expressing of cyclinD1 and CDK4. Background: Hepatocyte growth factor (HGF) has been shown to have multiple biological properties on liver, including mitogenic, antifibrotic, and cytoprotective activities. However, the absence of HGF receptor (c-met) in the rats with D-galactosamine/lipopolysaccharide (D-GalN/LPS) treatment leads to the fact that the high expression of HGF cannot activate signal transduction from cmet on survivor hepatocytes. Whether exogenous administration of c-met by gene transfer can promote liver regeneration and protects the rats from liver failure is not clear. Methods: We established the rat model with liver failure induced by D-GalN/ LPS, and constructed c-met expression plasmid with albumin promoter (p-albc-met), and then transfer it into the rat liver by hydrodynamic injection. c-met and PCNA was detected by immunohistochemical staining, and then apoptosis in hepatocytes were measured by flow cytometry. Results: Hydrodynamic delivery of p-alb-c-met induced the targeted expression of c-met in liver cells; markedly reduced hepatocytes apoptosis; promoted the proliferation of survivor hepatocytes; and elevated the survival rate among the rats with liver failure. Conclusions: These results qualify c-met as part of a physiologic, protective response of hepatocytes to injury and a promising gene therapy candidate for clinical applications aimed at preventing and treating viral and toxic liver failure. Acknowledgements: This work was supported by NSFC (No.30800973). Background: Our studies and those of many others have implicated fibrinogenlike protein 2 (fgl2) prothrombinase and tumor necrosis factor receptor (TNFR) mediated hepatocyte necrosis and apoptosis in the development of fulminant viral hepatitis, a disease with a mortality rate of greater than 80% in cases lacking immediate organ transplantation. Methods: This study was designed to explore the efficacy of dual short hairpin RNA (shRNA) interferenceof fgl2 and TNFR1 in the treatment of murine hepatitis virus strain 3 (MHV-3) induced fulminant hepatitis in mice. Plasmids p-mfgl2shRNA and p-mTNFR1shRNA complimentary to the sequences for mfgl2 and mTNFR1 were constructed. Plasmids pEGFP-mfgl2 and pEGFP-mTNFR1 expressing mfgl2-EGFP (Enhanced Green Fluorescent Protein) and mTNFR1-EGFP fusion proteins were also constructed for screening of the inhibitory effect of p-mfgl2shRNA and p-mTNFR1shRNA on mfgl2 and mTNFR1 expression. Results: Cotransfection of individual shRNA plasmids and pcDNA3.0-mfgl2 and pcDNA3.0-mTNFR1 expression constructs into Chinese hamster ovary (CHO) cells significantly inhibited mfgl2 and mTNFR1 gene expression, as evidenced by fluorescence microscopy, reverse-transcription polymerase chain reaction (RT-PCR), and Western blot. In vivo hydrodynamic delivery of dual interference shRNA plasmids for mfgl2 and mTNFR1 significantly decreased mfgl2 and mTNFR1 expression and markedly ameliorated fibrin deposition, hepatocyte necrosis, and apoptosis and prolonged survival against fulminant viral hepatitis induced by MHV-3 in Balb/cJ mice compared to mfgl2 or TNFR1 single-gene interference. Conclusion: These results indicate that in vivo gene interference of more than one key target provides superior treatment efficacy compared to that with single-gene interference. Objective: miRNA are small noncoding RNA that negatively regulate gene expression at post-transcriptional level. Malfunction of miRNA regulation is associated with a wide variety of human liver diseases, but the role in acute liver failure is unknown. In this study, the expression of miR-122 was studied in acute liver failure mice. Methods: D-galactosamine (D-GalN, 900 mg/kg) plus lipopolysaccharide (LPS, 10 lg/kg) were used to construct the acute liver failure model,total RNA was isolated from liver tissue and the expression of miR-122 was analyzed using quantitative Real-time RT-PCR and locked nucleic acid (LNA) -based DIGlabeled Northern -blot. Results: Twenty-four hours after induced, the mortality of mice reached over 80%. At the early stage liver tissue showed punctate bleeding, at the mediumterm showed hepatocyte apoptosis and late (7 h later) a large number of liver cell necrosis. miR-122 is highly expressed in normal mice liver (ct & 14), after administration it was up-regulated significantly at first hour (P = 0.013) and down-regulation after 3 h, the changes were sigificant at 7 h (ct & 15,P = 0.002) and it was verified by Northern-blot; ALT/AST levels increased obviously at 3 h and reached peak at 7 h, after 7 h due to a large number of liver cells injury showed a sharp decline; pro-inflammatory cytokines including TNF-a and IL-6 all were increased in serum and liver tissue (P \ 0.05), the correlation analysis between miR-122 and TNF-a, ALT and AST found that there has good correlation (correlation coefficients were 0.674, 0.493, -0.505 and -0.399, respectively). Conclusion: Liver specific miR-122 can reflect liver injury and there were good correlation between miR-122 and TNF-a, IL-6, ALT and AST respectively. Our results suggest that miR-122 might be a novel biomarker for early evaluation of hepatocyte injury in liver failure in the future. Methods: Seven indicators: hepatic encephalopathy, creatinine, prothrombin activity, serum total bilirubin, infection, the dimension of liver, the maximum depth of ascites, were scored from 0 to 4 points according to their severity. We divided the 409 patients with ACLF in hepatitis B into two groups randomly. The first group was to establish the severity scoring system and define the cut-off-point, the second group was to test the severity scoring system. Results: For the first group (n = 309), there were significant differences (P \ 0.000) of the total score between the living (n = 144) and the dead patients (n = 165). The area under ROC curve was 0.953. The cut-off-point was 9.5. The sensitivity was 0.97, the specificity was 0.82. The second group (n = 100) was divided: the one was C10 score, the other was B9 score. The mortality rate of the first group was 87.5%; the second was 2.3%. There were significant differences between the two groups (P \ 0.000 Background: Glycyrrhizin, a biological active compound isolated from the licorice root, has been used for the treatment for patients with chronic hepatitis.

In Japan and other Asian countries, treatment with glycyrrhizin is administered as Stronger Neo-Minophagen C. We examined the effect of glycyrrhizin on acute hepatitis using an animal model of lipopolysaccharide (LPS) and Dgalactosamine (GalN)-induced mouse liver injury. Methods: Liver injury was induced by intravenously injecting a mixture of 25 ng LPS and 20 mg GalN to 6 week old male Balb/c mice. Hepatocelluar damage was evaluated by measuring serum alanine aminotransferase (ALT) and histological observation. Serum cytokines, matrix metalloproteinase-9 (MMP-9) and high mobility group box 1 (HMGB1) levels were measured by ELISA kits. Intraperitoneal administration of glycyrrhizin was performed 30 min before LPS/GalN treatment. Effects of glycyrrhizin on liver damage were examined 8 h after LPS /GalN treatment. Results: Glycyrrhizin dose-dependently inhibited serum ALT levels in response to LPS/GalN treatment. In addition, this compound significantly suppressed serum interleukin (IL)-18, HMGB1 levels and hepatic MMP-9 levels. Immunoreactivities for IL-18 and MMP-9 were predominantly detected in the pericentral inflammatory area containing many macrophages and neutrophils at 8 h after LPS/GalN treatment. Glycyrrhizin remarkably reduced the IL-18 and MMP-9-immunoreaction in liver tissues. Conclusion: The present results suggests that IL-18, HMGB1 and MMP-9 are involved in the development of LPS/GalN-induced mouse liver injury, and that a mechanism by which glycyrrhizin reduced ALT levels and hepatocelluar damage in response to LPS/GalN may be due to a down-regulation of these factors.

The Aims: To observe the change of CD4 + CD25 + regulatory T cells in peripheral blood and NK cells in liver tissue of patients with chronic hepatitis B(CHB) and liver failure, and to investigate the immunology mechanism of the patients with liver failure infected with hepatitis B virus. Methods: Thirty-four patients with CHB and 54 patients with liver failure were enrolled.The latter composed of 31 chronic liver failure (CLF) and 23 acute liver failure (ACLF). Calculate the percent of the CD4 + CD25 + FOXP3 + T cell in the CD4 + T cells. Detect the number and position of NK cell in the liver tissue using immunohistochemistry. Results: The percentage of Treg cell in the CLF group was significantly lower than that in the ACLF group. Dynamic observation results showed that the amount of Treg cells inceased significantly at first, then decreased slowly during the development of the disease in the ACLF group. The amount of Treg cells kept on the lower lever in the CLF group. In the liver failure group, the percentage of Tregs declined with the development of the disease. The number of NK cells in the necrotic liver tissue of patients with liver failure was found more than that in the peri-necrotic tissue. Conclusion: The changed amount of Treg cells in peripheral blood was in connection with the development and prognosis of HBV infected disease. The position that NK cells infiltrated and number of cells related to the degree of liver tissue inflammation. Background/Aims: It is frequently important to identify the prognosis of fulminant hepatic failure (FHF) patients as this will influence patient management and candidacy for liver transplantation. Therefore, a novel scoring system based on metabonomics combining with multivariate logistic regression was developed to predict the prognosis of FHF mouse model. Methods: BALB/c mice were used to construct FHF model. Parts of plasma were collected at 4, 5, and 6-h time points after treatment, respectively, and detected using gas chromatography/time-of-flight mass spectrometry (GC/ TOFMS). The acquired data were processed using partial least square discriminant analysis (PLS-DA). The metabolic markers identified were used to construct a scoring system by multivariate regression analysis. Results: 28 mice of survival group and 28 of dead group were randomly selected and analyzed. PLS regression analysis showed that both the PLS models of 5 and 6 h after d-galactosamine/lipopolysaccharide treatment demonstrated good performances. Loadings plot suggested that phosphate, beta-hydroxybutyrate (HB), urea, glucose and lactate concentrations in plasma had the highest weightings on the clustering differences at the three time points. By the multivariate logistic regression analysis, the death/survival index (DSI) was constructed based on relative concentrations of HB, urea and phosphate. It provided general accurate rate of prediction of 93.3% in the independent samples. Conclusions: The novel scoring system based on metabonomics combining with multivariate logistic regression is accurate in predicting the prognosis of FHF mouse model and may be referred in clinical practice as a more useful prognostic tool with other available information. The imaging studies included ultrasound, CECT, MRI were done whenever required. UGI endoscopy, ascitic fluid examination, blood and urine culture, arterial blood gas analysis and liver biopsy were done whenever indicated. ACLF was diagnosed by the Asia pacific Study of Liver Diseases (APASL) criteria. Result(s): One hundred and fifty patients were studied (median age 9 (2-16) years). Male to female ratio was 2.1:1.The distribution of diseases were ACLF 31 (20.7%), ALF 41 (27.3%), CLD 45(30%), acute viral hepatitis 30 (20%) AND Budd Chiari syndrome 3 (2%).The underlying CLD were AIH 13 (41.9%), WD 13 (41.9), HBV 2 (6.5%), ICC 1 (3.2%), and secondary biliary cirrhosis 2 (6.5%).The etiology of acute events were superadded HAV 13 (41.9%), HEV infection 3 (9.7%), HBV reactivation 1 (3.2%), AIH flare 3 (9.7%), cholangitis 1 (3.2%), UGI bleed 1 (3.2%) and undetermined 7 (22.6%). The manifestation of acute liver failure were ascites 31 (100%), encephalopathy 18 (58.1%), UGIB 8 (25.8%) and spontaneous bacterial peritonitis 9 (29%). The mortality was in 6 (19.4%) patients. The predictors of mortality were INR and SOFA score. The 6.5 was the best cut-off in predicting mortality with a sensitivity of 100%, specificity 80%. Underlying CLD and acute events made no difference in mortality. Results: In non-severe hepatitis, liver histology showed severe acute hepatitis in 7 and chronic hepatitis in 9. In severe/fulminant patients, histology showed massive necrosis in 6, submassive necrosis in 1, severe acute hepatitis in 4, and chronic hepatitis in 3. Centrizonal necrosis with or without plasma cell accumulation was characteristic for acute onset AIH. On immunohistochemical staining for CK7, periportal PC and intralobular IM was less commonly present in severe/fulminant patients than in non-severe ones (P = 0.02). In severe/fulminant patients, periportal IM, intralobular PC and intralobular IM were more commonly present in survivors than in non-survivors (P = 0.01).

Conclusions: The histologically acute form of hepatitis was more common in severe/fulminant patients than in non-severe ones. PC and IM may play an important role in liver cell regeneration from massive/submassive hepatic necrosis in acute onset AIH.

High Frequency of Th17 Cells is associated with Worse Prognosis of Acute-on Chronic-liver-failure Caused by HBV Background: IL-17-producing CD4+ T cells (Th17) is a newly identified pro-inflammatory T-cell subset, which plays critical role in inflammation and tissue damage, including liver damage in chronic HBV infection. However, its role in HBV related acute-on-chronic liver failure (AoCLF), a condition with severe liver damage and [70% fatality rate, remains unknown. We want to define whether Th17 cells contribute to worse prognosis of AoCLF and high incidence of bacteria and fungus infection. Subjects and Method: Fifty-nine patients with HBV related AoCLF (median age 45 years, 49 male) were recruited. Frequency of Th17 cells amongst peripheral blood was defined by CD4 and IL-17 (intracellular, eBioscience) staining (BD FACs Calibur) and Flow Jo software (Tristar, USA). The dynamics of Th17 cells was defined through measurement at admission, every two weeks in-hospital and 3-month after discharge amongst those without bacteria or fungus infection, and at time points of before, peak and infection controlled amongst those with infection. Results: Patients were divided into two groups: survival and non-survival (died, required OLT and gave up treatment). At admission, the frequency of Th17 in survivals (4.82 ± 1.19%) was significantly lower than in non-survivals (6.21 ± 2.15%, P = 0.003) and continuously decreased (2.61 ± 1.16% 3-month after discharge). In contrast, it increased or remained unchanged in non-survivals. The differences in Th17 frequency between admission and week-2 correlated with the prognosis of AoCLF (R = 0.537, P = 0.002). The Th17 frequency at admission was correlated with the mortality ratio (R = 0.36, P = 0.005). Thirteen patients developed bacteria or fungus infection during the admission period, whose Th17 frequency at admission tended to be higher than those without infection (5.81 ± 2.04 vs. 4.96 ± 1.27%, P = 0.163). Conclusion: Overall, the frequency of Th17 cells is associated with the prognosis of AoCLF: the higher of the Th17 frequency, the worse of the outcome, indicating that depletion of Th17 cells may be of therapeutic value in management of AoCLF patients. 

Age & PT as Prognostic Factors of Acute Liver Failure (16 Years of National Liver Transplant Program in Singapore) Vincent Lai 1 , Aung Myat Oo 1 , Belinda Mak 1 , Teoh Kum Faut 1 , Seng Gee Lim 1 1 National University Health System, 5, Lower Kent Ridge Road, Singapore 119074 Introduction: Acute liver failure (ALF) is rare condition with devastating consequences. We assessed the prognostic indicators at predicting adverse outcome in our cohort of ALF patients over 16 years period. Methods: All cases of adult patients with ALF referred to national liver transplant program in Singapore from January 1992 to December 2008 were included in the study. Their baseline and outcome data were collected and analyzed. Adverse outcome was defined as mortality without transplant or undergone liver transplantation. Results: There were 101 patients with ALF between study periods, 71 (70.3%) Chinese, 51 (50.5%) male, age 43. 5 (13.4) years. Major aetiologies were hepatitis B, 45 (44.5%) and drug induced liver injury (DILI), 37 (36.6%). Traditional Chinese Medicine accounts for larger portion of DILI, (42%). 13 patients underwent liver transplantation and only 3 (23%) of them died post liver transplant. Among 88 patients without liver transplant, 74 (84%) died. Age and PT were the independent factors significantly related to adverse outcome (P = 0.009 and 0.012). AUROC (sensitivity, specificity) for age C37 years, PT C30 s, age and PT combined, MELD C33 and KCH criteria are 0.641 (78, 50%), 0.759 (77, 64%), 0.727 (60, 86%), 0.722 (59, 86%), 0.616 (52, 71%) respectively. Conclusion: Age and PT were significantly related to adverse outcome and higher AUROC. Aetiology of ALF in Singapore is different from the west and some part of Asia because of endemic of chronic hepatitis B and widely use of TCM in community. New prognostic model is needed for this group of patients. Background: Acute on chronic hepatitis B liver failure (ACHBLF) is main reason of liver failure in China, and has a poor prognosis. We speculate Entecavir would play more great role in improving prognosis of ACHBLF, because of its potent inhabiting HBV replication and more lower drug resistant. The study's aim is to evaluate the therapeutic efficacy of entecavir treating patients with ACHBLF. Methods: 112 patients with ACHBLF were entered into a treatment group (ETV group) and a control group, according to whether combined treating with entecavir based on the comprehensive medical treatment, 55 cases and 57 cases, respectively. HBV DNA level, liver function and survival condition of the patients in different time-points were observed; factors possibly related to entecavir treatment efforts were also analyzed. Results: From week one to week eight, HBV DNA levels of patients in ETV group are lower than that in control group, and P \ 0.01, at week 12, P = 0.03. Whether in ETV group nor in control group, HBV DNA levels of patients at week 12 are lower than that at baseline. Significant differences of cumulative survival rate of patients in two groups were observed after the third week of treatment. The cumulative mortality rate in the ETV group at week 12 was 43.9% while the rate in the control group was 64.9% (v 2 = 4.558, P = 0.033); Cox regression analysis showed the patients who were treated at an early stage and whose lower serum bilirubin level and young age, had a high survival rate and survival time (P \ 0.05). Conclusions: Entecavir drops rapidly HBV DNA level of patients in ETV group and may improve the survival rate of patients with ACHBLF. Patient 0 s stage of the disease, serum bilirubin levels, and age at enrollment were major factors that may have affected entecavir therapeutic efficacy.

Altered Monocyte-derived Dendritic Cells Responses to poly (I:C) in Patients with Acute-on-Chronic Liver Failure Ning Li 1 , Ming-Quan Chen 1 , Meng-Qi Zhu 1 , Qian Li 1 , Zhi-Ping Qian 2 , Guang-Feng Shi 1 1 Huashan Hospital, Fudan University, 12 Middle Urumqi Road, Shanghai 200040, China, 2 Shanghai Public Health Clinical Center, Fudan University, Jinshan, Shanghai 200040, China Background: The role of the adaptive immune response, with regard to the development of hepatitis B virus infection, has been extensively studied in acute-on-chronic liver failure (ACLF). However, the importance of innate immunity has been noted only recently. The aim of this study was to investigate the effect of TLRs on pathogenesis in ACLF patients. Methods: We isolated peripheral blood monocyte-derived dendritic cells (MoDCs) from patients with chronic hepatitis B (CHB, n = 20) or HBVassociated acute-on-chronic liver failure (ACLF, n = 40), compared with agematched normal controls (n = 20) and stimulated such cells in vitro with poly (I:C). TLR3 pathway gene of extracted RNA were detected using real-time PCR. Supernatant fluids from the cultures were analyzed for levels of six different pro-inflammatory cytokines, interleukin (IL)-1b, IL-6, IL-8, IL-10, IL-12p70, and TNF-a. Results: After in vitro challenge with poly (I:C), ACLF MoDC produced imbalanced relative levels of pro-inflammatory cytokines, particularly IL-6 and TNF-a, were higer than CHB patients and controls. While, significantly lower amounts were found of IL-12p70, IL-1b, and IL-8 on MoDCs in ACLF. Conclusions: The results suggest impaired TLR3 pathway signaling in MoDC from patients with ACLF, resulting in secretion of selective pro-inflammatory cytokines integral to the inflammatory response that may be critical in the pathogenesis of ACLF. Background: Patients with subacute fulminant hepatic failure (FHF) and lateonset hepatic failure (LOHF) show relatively slow disease progression, although usually suffer serious liver damage. It sometimes seems that liver can lose its regenerative capability at the appearance of hepatic coma. No effective treatments have been established for such cases. We advocated that predicting the development of FHF and early intervention with intensive medical treatment for the underlying cause could improve the survival rate. The present study examined the efficacy of our treatment system. Method: Inclusion criteria of the present study were as follows: acute hepatitis patients admitted to our hospital from 2002 to 2008, prothrombin time below 60% of normal, and absence of hepatic coma on admission. Prediction of FHF was determined based on a previously reported formula (Yoshiba M, J Gastro 2002) . Treatment for underlying cause consisted mainly of immunosuppressive therapy and/or antiviral therapy. Patients were also given artificial liver support immediately after the appearance of hepatic coma; it comprised plasma exchange in combination with hemodiafiltration. Results: The inclusion criteria were met by 74 patients with causes of hepatitis listed as HAV (3), HBV (11), HBV carrier (23) Aims: Increasing evidence suggests the contribution of natural killer (NK) cells to pathogenesis of human hepatitis, but the detailed mechanisms have yet to be clearly elucidated. Methods: In this study, injection of polyinosinic: polycytidylic acid (poly I:C) and D-galactosamine (D-GalN) was used to establish a novel murine fulminant hepatitis model. We then examined the liver injury by determining serum transaminase ALT levels and liver pathologic changes. The FACS analysis provided us with the precise information about lymphocyte phenotype, activation and cytokine production. Hepatic NK cells and Kupffer cells were isolated by positive magnetic cell sorting (MACS) and were co-cultured with poly I:C stimulation. We used ELISA and real time PCR to examine cytokine levels.

Results: Our results showed that treatment with poly I:C induced severe liver injury in D-GalN-sensitized mice and predepletion of either NK cells or Kupffer cells could completely abolish the liver injury. Injection of poly I:C/D-GalN into mice could promote tumor necrosis factor-c production and surface retinoic acid early inducible-1 (Rae1) protein expression by Kupffer cells, which then activated NK cells to produce interferon-c via NKG2D-Rae1 recognition. NK cell-derived interferon-c and Kupffer cell-derived tumor necrosis factor-a synergistically mediated the severe liver injury. Moreover, Kupffer cell-derived interleukin-12 and interleukin-18 were also found to improve cross talk between NK cells and Kupffer cells. Conclusion: These results provide the first in vivo evidence that NKG2D/ ligand interaction is involved in the synergic effects of NK cells and Kupffer cells on acute liver injury.

Hepatoprotective Background and aim: Rheum emodi is used as herbal drug and its aqueous extract was evaluated against acute liver damage induced by carbon tetrachloride. Free radical scavenging activity, total phenolic contents and choleretic activity was also investigated. Method: Acute toxicity was induced in rats by CCl 4 (1.5 ml/kg, i.p.; once only) administration. Extract of Rheum emodi were given at different doses (150, 300, 450 mg/kg; once only) after 48 h of toxicant administration. All the animals were sacrificed after 48 h of the last administration and SGOT, SGPT, ALP, albumin, bilirubin and urea were determined in serum. Oxidative stress marker (LPO and GSH), metabolic enzymes (ATPase and G-6-pase), hexobarbitone induced sleep time and BSP retention along with histopathological alterations also studied.

Result: The levels of SGOT, SGPT, ALP, bilirubin, albumin and urea in serum; hepatic lipid peroxidation were significantly increased while reduced glutathione level, metabolic enzymes and total protein were significantly decreased after CCl 4 administration. All the doses of Rheum emodi extract lowered SGOT, SGPT, ALP, bilirubin, albumin, urea and lipid peroxide level. Lower most dose of extract significantly restored glutathione and metabolic enzymes. Inhibition in lipid peroxidation and restoration of glutathione levels indicated strong hepatoprotective activity of the plant extract. Rheum emodi extract decreased the hexobarbitone induced narcosis and BSP retention, which indicated improvement in excretory capacity of liver. Histopathological observations of the liver also supported the biochemical findings. Background: Acute-on-chronic liver failure (ACLF) is a well recognized and common clinical entity. Reactivation of chronic hepatitis B (CHB) is an important cause of ACLF. Potent antiviral drugs may help reduce the high morbidity and mortality in such patients.

Aim: To evaluate the efficacy of tenofovir in reducing HBVDNA, improvement in outcome and to determine the predictors of mortality in patients with spontaneous reactivation of CHB with ACLF. Methods: Consecutive patients of spontaneous reactivation of CHB, who presented with ACLF were randomized to receive tenofovir or placebo. The primary endpoint were survival at 3 months, decrease in HBVDNA levels and secondary endpoint was improvement in CTP and MELD scores. Results: Of the 90 patients of ACLF presenting in 2 years, 27 (26%) were due to reactivation of CHB and were enrolled. 15 (56%) patients were HBeAg and 20 (74%) were IgM anti-HBc positive. The median baseline HBV DNA level was 9 9 10 5 IU/mL. 14 patients were randomized to receive tenofovir and 13 to placebo. Baseline characteristics were comparable. At the end of 3 months, the survival was significantly better in tenofovir than placebo group (8/14

[57%] vs. 2/13 [15%] respectively, P = 0.03). In the surviving patients, there was a significant improvement in the median CTP and MELD scores in the tenofovir group (P \ 0.05) while there was no improvement in these scores in the placebo group ( P = NS). There was a significant decline in the HBV DNA levels in surviving patients in the tenofovir group at 2 weeks ( P \ 0.001) and at 3 months ( P \ 0.01) of treatment; however, there was no change in HBVDNA levels in the placebo group. In this study, we investigate protective effects of CGX on Con A-induced liver injury and its underlying mechanisms. CGX was administered once daily during 6 days and 3 h before Con A injection (15 mg/kg, iv). After 6 and 20 h of Con A injection, determination of AST, AST, and organ weight, mRNA expression analysis and histological examination were performed. Spleen weight was increased, whereas thymus weight was decreased by Con A injection. CGX ameliorated splenomegaly. CGX prevented the increase of AST and ALT in serum, In histopathological examination, necrosis area of liver tissue was significantly reduced and DNA fragment was also decreased by CGX pretreatment. MDA, final product of lipid peroxidation in liver tissues increased by fivefold compared to normal by Con A injection. But CGX treatment decreased MDA content in liver significantly in a dose dependent manner. Among antioxidant components including GSH, SOD, catalase, Only GSH is significantly increase at 200 mg/kg of CGX. In the conclusion, CGX, an herbal medicine, CGX protects liver from T cell-mediated liver injury. Background: ASPP2 is a proapoptotic member of a family of p53-binding proteins ASPPs (the ankyrin-repeat-, SH3-domain-and proline-rich-regioncontaining protein). We have previously reported that ASPP2 was frequently down-regulated by DNA methylation in HCC cells. Here, we investigate the role of ASPP2 in epithelial-mesenchymal transition (EMT) and metastasis of HCC.

Methods: The expression of ASPP2 was analysed by immunohistochemistry assay in 51 HCC patients and compared with the clinicopathological features of the patients. Expression of ASPP2 was down-regulated by short hairpin RNA, then morphology and matasasis potential of tumor cells were further analysed. Results: The expression of ASPP2 was correlated with metastasis and recurrence in HCC patients (63.0 vs. 28.6%, P = 0.037). Down-regulation of ASPP2 made cells undergoing morphological changes from a classic epithelial morphology to a spindle-like shape. Down-regulation of ASPP2 also enhanced cell adhesion, migration and invasion through Matrigel in vitro, promoted lung metastasis in HCC xenografts in nude mice. Further studies revealed that down-regulation of ASPP2 activated the non-receptor tyrosine kinase c-Src and then promoted b-catenin translocating from cytomembrane to cytoplasm and nucleus. Conclusion: Our results suggest down-regulation of ASPP2 promotes EMT and metastatic progression of HCC .These findings provide new insight into the molecular mechanisms leading to matastasis of HCC and may have potential therapeutic applications. Background: The ankyrin-repeat-, SH3-domain-and proline-rich-region-containing protein (ASPP) family of proteins regulates apoptosis through interaction with p53 and its family members. This study evaluates the epigenetic regulation of ASPP1 and ASPP2 in HBV-positive hepatocellular carcinoma (HCC) and explores the effects of down-regulation of ASPP1 and ASPP2 on the development of HCC. Methods: HCC cell lines and tissues from HCC patients were used to examine the expression and methylation of ASPP1 and ASPP2. Results: The expression of ASPP1 and ASPP2 was diminished in HCC cells by epigenetic silence owing to hypermethylation of ASPP1 and ASPP2 promoters. Analyses of 51 paired HCC and surrounding non-tumor tissues revealed that methylation of ASPP1 and ASPP2 was associated with the decreased expression of ASPP1 and ASPP2 in tumor tissues and the early development of HCC. Moreover, ASPP2 became methylated upon HBV x protein (HBx) expression. Down-regulation of ASPP1 and ASPP2 promoted the growth of HCC cells in soft agar and in nude mice, and decreased the sensitivity of HCC cells to apoptotic stimuli. Objective: The abnormal expression of many genes involved in the formation and development of hepatocellular carcinoma (HCC). Genetic and epigenetic changes regulate the expression of cancer-related genes, which is the core biological processes in HCC. We investigated the dynamic expression and alteration of hypoxia inducible factor-1a (HIF-1a) and its pathological features in hepotacellular carcinoma (HCC). Methods: Hepatomas model was induced with 2-FAA on male SD rats for investigating dynamic changes of HIF-1a. Liver specimen from HCC patients were collected by self-control method. The expression, cellular distribution, and pathological features of HIF-1awere analyzed by immohistochemistry. Results: Rat hepatocytes from granule-like degeneration to atypical hyperplasia and HCC development, and the progressing increasing of the levels of hepatic HIF-1a and HIF-1amRNA expression during the course. The levels of HIF-1a in hepatoma tissues and sera were significantly higher than those in normal and degeneration ones. There was positive relationship of HIF-1a levels between them in hepatoma tissues and sera (P \ 0.05). The positive HIF-1a was brown and granule-like, mainly presented in cytoplasm and few in nucleus. Background: Deregulation of EGFR and ERBB2 signaling is seen in many human cancers and a wealth of clinical and experimental data indicate the etiological role of these events in cancer pathogenesis. Target therapies against the deregulation of EGFR and ERBB2 signaling exert effects on many human cancers, while there are modest effect on treatment of hepatocellular carcinoma (HCC). Recent evidence that ERBB3 is responsible for tumor resistance to therapeutic agents targeting EGFR or ERBB2 in lung cancer and breast cancer highlighted its critical role in the oncogenic signaling of EGFR family. However, roles of ERBB3-mediated signaling in HCC carcinogenesis remain totally unclear. Methods: Expression and phosphorylation of ERBB3 was assayed by quantitative polymerase chain reaction and immunoblot assays. Gene number was determined using fluorescence in-situ-hybridization. The roles of EGEF, ERBB2, and neuregulin in phosphorylation of ERBB3 were assayed via transfection with small-interference RNAs to silence their expression. The effects of ERBB3 on cell proliferation, invasion and tumorigenesis were assayed in vitro and in vivo. Results: We developed a high efficiency method to isolate HCC deregulated genes and identified ERBB3 as an upregulated gene in HCC. Quantitative RT-PCR confirmed up-regulation of ERBB3 in about 70% of HCC tissues.

Overexpression of ERBB3 was strongly associated with higher tumor recurrence (P = 0.000) and lower long-term survival (P = 0.004) (via Kaplan-Meier analyses and log rank tests). Fluorescence in-situ-hybridization assays showed amplification of ERBB3 genes in most of the HCC cells and tissues. Neuregulins, the ligand of ERBB3, and phosphorylated ERBB3 were detected in most of human hepatoma tissues, indicating a constitutive activation of ERBB3 signaling in HCC. Activation of ERBB3 signaling enhanced cell proliferation and colony formation, while silencing ERBB3 suppressed xenograft tumor formation and growth.

Conclusion: Overexpression of ERBB3, which was primarily due to gene amplification, was associated with a higher tumor recurrence and a worse prognosis. Aberrant activation of ERBB3 signaling was directed involved in hepatocelluar carcinogenesis. This oncogenic signaling of ERBB3 can be the target for anti-HCC therapy.

Histone Deacetylases (HDACs) and Hepatocellular Carcinoma ( Background: Transcriptional regulation in eukaryotes occurs within a chromatin setting, and is strongly influenced by the post-translational modification of histones, the building blocks of chromatin, such as methylation, phosphorylation and acetylation. Within these modifications, acetylation plays an important role. Steady-state levels of acetylation of the core histones result from the balance between the opposing activities of histone acetyltransferases (HATs) and HDACs. In general, increased levels of histone acetylation is associated with increased transcriptional activity, whereas decreased levels of acetylation is associated with repression of gene expression. HDACs associate with a number of tumor-suppressor genes, the transcription of which was suppressed through the recruitment of HDACs, thus the cancer cells are unable to undergo differentiation, leading to excessive proliferation. The link between HDACs and tumor has been reported in acute promyelocytic leukaemia originally. HCC is one of the most frequent solid tumors occurring worldwide. At present, there are some articles revealed that HDACs is also related with HCC. Methods: miRNA expression in normal and HCC cell lines and tissues was evaluated using qPCR array and hybridization based microRNA microarray. Angiogenesis and invasion across a matrigel layer were quantitated using commercial assays. Promoter methylation, matrix metalloproteinase (MMP) 1, 2, 3, 9 mRNA and mature miRNA expression was quantitated by real-time PCR analysis. Results: Genome-wide qPCR array and microarray expression profiling identified a significant silencing of miR-125b in HCC tissues and cells, including tumor endothelial cells (T-ECs). The 5'-promoter region of miR-125b was noted to be embedded within a CpG island. The expression of miR-125b was considerably increased after 5-aza-dCyd treatment in T-ECs and PLC/PRF-5 HCC cells, suggesting that the level of miR-125b could be epigenetically modulated. Methylation-specific PCR detected that miR-125b promoter was hypermethylated in T-ECs as well as PLC/PRF-5 cells. Angiogenesis and cell invasion were significantly decreased by miR-125b precursor introduction in T-ECs and PLC/PRF-5 HCC cells. Bioinformatics and dual-luciferase reporter assays identified placenta growth factor (PlGF), another angiogenic factor of VEGF family, as a direct target of miR-125b. Whereas PIGF expression in T-ECs and PLC/PRF-5 cells was high relative to N-ECs and N-Heps, overexpression of miR-125b resulted in a significant reduction of PIGF level. Furthermore, modulation of miR-125b also altered expression of MMP-2 and 9, mediators involved in enzymatic degradation of extracellular matrix. Conclusions: The discovery that epigenetic inhibition of miR-125b plays a significant role in the initiation of angiogenesis and invasive phenotype through PIGF provides the basis for an exciting field in which the epigenomic miRNAs of HCC endothelial cells may be manipulated with potential therapeutic benefits.

Background: Foxp3 as a key molecular mediating immune-related inhibitory functions can be expressed in Regulatory T cells (Tregs). The expression of Foxp3 in Tregs is due to Foxp3 promoter demethylation. In the previous works, we validated that the Foxp3 also can be expressed by hepatoma cell lines and involved in the Foxp3 mediated immune-suppression in tumor immune microenvironment. Whether the mechanism of Foxp3 expression in hepatoma cell lines is related to the Foxp3 promoter demethylation is very important for clarifying the roles of Foxp3 in hepatoma cell lines. Methods: 10 hepatoma cell lines Huh7, Hep3B, HepG2, et. al) Emerging evidence implied that miRNAs play critical roles in diverse biological processes such as cell proliferation, differentiation and apoptosis, and development of diseases including cancer. To explore the effects of miRNAs on the pathophysiology of human liver cancer, we examined the expression patterns of some miRNAs previously reported as aberrantly expressed in clinical samples in different liver cancer cell lines. We further analyzed one miRNA with expression depletion for its potential role in cancer development and metastasis. Methods: Expression of five miRNAs was examined by real-time reverse transcription/polymerase chain reaction in seven human liver cancer cell lines. Functional analysis on miRNA was performed using gain-of-function approach in which the selected miRNA was restored for cell proliferation, viability, and migration/invasion assays. Result: Five miRNAs were differentially expressed among seven cell lines with one exception, miR-224, which expressed at extremely low level in 4 out of 7 cell lines. Functional assays based on both Huh-7 (hepatocellular origin) and Sk-Hep-1 (endothelial origin) cells that were transfected with pre-miR-224 indicated that cell migration and invasion were significantly intensified whereas cell proliferation and viability remained unaffected. Conclusion: The expression patterns of microRNA differ in each liver cancer cell line studied. Functional assays indicated that miR-224 may contribute to the metastasis of liver cancer by enhancing the cell migration/invasion. Background: Hepatitis B virus X protein (HBx) or its C-terminally truncated mutant (ctHBx) has been implicated in the development of hepatocellular carcinoma (HCC). Furthermore, TGF-beta signaling is considered to be crucial in hepatocarcinogenesis. However, the molecular mechanisms whereby HBx/ ctHBx mediates TGF-beta signaling pathway and contributes to hepatocarcinogenesis are yet to be clarified. Methods: Using the subtractive suppression hybridization (SSH) method, we identified HCC-related genes that were differentially expressed in HCC containing a mutant form of beta-catenin and functional ctHBx. Among them, BAMBI was identified and functionally analyzed to elucidate its role in HBxrelated hepatocarcinogenesis.

Result: BAMBI was found to be up-regulated, which led to the down-regulation of TGF-beta signaling in HepG2 cells with a mutant beta-catenin. In contrast HBx/ctHBx trancriptionally downregulated BAMBI, and resulted in the subsequent enhancement of TGF-beta signaling. TGF-beta transcriptionally upregulated BAMBI through SMAD3. Exogenous treatment with TGF-beta induced migratory and invasive morphologies in HepG2 cells. The forced expression of HBx/ctHBx increased colony formation and anchorage-independent tumor cell growth. Conclusion: These results suggest that funtional HBx/ctHBx contribute to hepatocarcinogenesis by modulating BAMBI-mediated TGF-beta regulation. The University of Hong Kong, 21 Sassoon Road, Pokfulam, Hong Kong, 2 National University Health Systems, National University of Singapore, Singapore YAP, a downstream effector of Hippo signaling pathway, has recently been implicated in cancer development. The overexpression of YAP is frequently observed in several types of cancer, such as prostate, ovarian, and liver. Ectopic expression of YAP promotes cell proliferation, epithelical-mesenchymal transition (EMT), and anchorage-independent growth, signs of oncogenic potential. Given the important role of YAP in cancer development, we would like to investigate the regulatory mechanism of YAP expression in HCC. In this study, the miRNA regulation of YAP was investigated. From the miRNA profiling data, we found that miR-375 was significantly down-regulated in HCC patients, and bioinformatic algorithm predicted that YAP is a potential target of miR-375. To confirm the functional interaction between miR-375 and YAP, we performed a luciferase reporter assay, and the results showed that miR-375 significantly repressed the expression of luciferase carrying the 3'-untranslated region of YAP and reduced the endogenous protein level of YAP in HCC cell lines. Further investigation showed that the downstream target of YAP, such as CTGF, was also suppressed upon the etopic expression of miR-375, suggesting miR-375 is involved in regulation of Hippo-YAP signaling pathway. Moreover, in vitro functional assay showed that ectopic expression of miR-375 slowed down the invasiveness and proliferation of HCC cells. Taken together, our data suggested that aberrant expression of miR-375 can contribute to HCC growth, and expression of YAP is regulated in part by a specific miRNA, leading to decreased tumor cell motility and proliferation.

Association of CXCL10 G-201A Polymorphism and the Risk of HCC Among Chinese Cirrhotic CHB Patients Jie Chen 1 , Xiang-Mei Chen 1 , Ting Zhang 1 , Qiang Xu 1 , Feng-Min Lu 1 , Hui Zhuang 1 1 Peking University Health Science Center, Xueyuan Road 38#, Haidian, Beijing, China

Background: As a pro-inflammatory factor, the role of CXCL10 in chronic HBV infection disease progression and in persistent inflammation related tumorigenesis has been emphasized recently. A polymorphism in the promoter of CXCL10 gene, G-201A, is reported to alter the binding affinity of nuclear protein and regulate CXCL10 expression. Therefore, an increased CXCL10 activity is thought to be associated with liver tissue damage and cirrhosis. But also as an angiostatic factor, its role in hepatocellular carcinogenesis is inconsistent and unclear. We studied the association of G-201A polymorphism with disease progression related to HBV infection and the impact of CXCL10 on cell proliferation. Results: Five types of pre-S region deletions were found: pre-S1 start codon inframe deletion; pre-S1 deletion involving S promoter; pre-S2 start codon mutation; pre-S2 in-frame deletion and pre-S2 in-frame deletion with pre-S2 start codon mutation. The overall prevalence of S promoter deletion mutant was 14.6% (6/41) but none in non-tumor tissues. Immunocytofluorescence showed that unlike wild type LHBS's whole cell diffusion, the S promoter deletion LHBS mutant showed an aggregated cytoplasmic pattern, and was found mainly accumulated in endoplasmic reticulum. Accordantly, higher level of GRP 78 mRNA was detected in exogenous mutant LHBS expressing hepG2 cells in vitro. Conclusion: HBV S promoter deletion was found solely existed in HCC tumor tissue. Mutations which affect of L/M/S envelope protein ratio were found more frequently in tumor tissues. The altered ratio and changed secretion pattern of surface proteins, the aggregation of mutant large surface proteins in ER and consequent UPR activation might possibly involve in HBV-related hepatocellular carcinoma. Methods: Percutaneous liver Bx was histologically evaluated for inflammation and fibrosis. The extent of COX-2 and CD34 expression was measured by immunohistochemistry and graded according to the number of hepatocytes expressing high intensity of COX-2. Results: HBV and HCV patients with higher fibrosis stage (0-4) presented with raised ALT and AST, however patients with fibrosis stage of 5 and 6 demonstrated fall in ALT and AST. The liver Bx from HCV patients showed higher HAI and fibrosis scores as compared to HBV group. The percentage of steatosis was higher in HCV (P \ 0.05) as compared to HBV patients. COX-2 expression was confined to hepatocytes and bile duct epithelium and was not detected in vascular endothelium or inflammatory cells. The extent of COX-2 expression was greater in HCV infected liver Bx (5.13 ± 1.11) than in hepatitis B (3.68 ± 0.92). HCC infected liver Bx showed greatest COX-2 expression (11.68 ± 6.32) and the expression of COX-2 was even greater in adjacent non-tumorous portion (25.61 ± 6.32). The expression of COX-2 was significantly correlated with the fibrotic stage both in HBV [Low (0-2) vs. High (3-6) fibrosis, 2.45 ± 0.41vs. 6.92 ± 3.12], and HCV [Low (0-2) vs. High (3-6) fibrosis, 5.13 ± 1.11vs. 10.17 ± 5.76 ] and also the HAI. There was positive correlation between the CD34 and COX-2 expression in Bx obtained from HBV, HCV and HCC patients. Conclusion: Overexpressed COX-2 expression is associated with high levels of necroinflammation, regeneration activity and disease progression of liver disease. Background: The poor prognosis of hepatocellular carcinoma (HCC) after surgical resection is due to recurrence and intrahepatic metastasis (IM). For study, HCC intrahepatic metastasis model using orthotropic implantation (OTI) of human HCC cell lines into mice liver has been established but it is time consuming. Aims: To compare the efficacies between OTI and of a direct injection method (DI) in SCID mice to induce IM. Methods: Ten mice were used in either the OTI or DI group. In the OTI method, approximately 1 9 10 7 cells in 0.2-mL culture medium were injected subcutaneously into the right flank of the SCID mice, which were then observed daily for signs of tumor enlargement. Once the subcutaneous tumor reached 1-1.5 cm in diameter, it was removed, cut into 1-2 mm cubes, and then implanted into the left liver lobe. As for the DI method, HepG2 cells (1, 5, 10, 20 and 30 9 10 4 , respectively) were directly injected into the middle lobe of mouse liver. Two mice were scarified in each month and their livers taken out for macroscopic and microscopic observations (100 sequential sections, 4 lm, were cut and stained with H&E) to confirm IM in either the left and right lobe. Results: In the DI method using 30 9 10 4 cells, IM occurred in both the left and right lobes in two months. However, the OTI model required 4 months to derive the same severity of IM as in the DI method. Conclusions: The DI method was straightforward and saved much time when compared with the OTI model that was rather time consuming. Also, OTI was not suitable for studying specific HCC cell line with over-expression of some target genes because a lot of uncontrolled molecular changes might had occurred during this long-term and complicated process. This DI model will be especially useful in the experimental design for gene therapy to prevent IM using various genetic modified HCC cell lines. Methods: Small interfering RNA targeting HBV X (x-siRNA) were synthesized. The level of HBV X mRNA was detected by semi-quantitative RT-PCR. The nude mice model bearing human hepatoma xenograft was established by subcutaneous inoculating HepG2-GFP and HepG2-GFP /HBx cells. These mices were treated with x-siRNA (25 mg/g), 5-Aza-dC (1.5 mg/g) or combination of both. The growth of the hepatoma xenograft was observed. The methylation of p16 gene promoter was determined by methylation specific polymerase chain reaction (MSP), level of p16 protein was detected by Western blot.

Results: Semi-quantitative RT-PCR analysis showed that the level of HBV X mRNA in x-siRNA-treated HepG2/GFP-HBx cells decreased markedly. The experiment in hepatoma xenograft nude model showed that the growth of tumors in HepG2/GFP-HBx group was faster than HepG2/GFP group while slow growth was observed in X-siRNA treatment group or 5-Aza-dC treatment group (P \ 0.05). MSP analysis showed that p16 gene methylation was observed in the tumor originating from HepG2/GFP-HBx, while no methylation was observed in thses tumor originating from HepG2/GFP. However,p16 methylation reduced in the tumors treated with x-siRNA or 5-Aza-dC. Western blot analysis showed that the level of p16 protein in HepG2/GFP-HBx group decreased, whie increased p16 protein was observed in these tumor treated with X-siRNA and 5-Aza-dC. Conclusions: HBV X promotes the growth of hepatoma cells in vivo by inducing p16 gene methylation and suppressing p16 protein expression. Specific siRNA targeting HBV X and methylation inhibitor can inhibit the growth of hepatoma cells via reversing p16 gene methylation and up-regulating p16 protein. Background: The present study explored the influence of AFP in alleviating the apoptosis was induced and the expression of CXC chemokine receptor 4 (CXCR4), it is the stromal cell derived factor-1 receptor was regulated by ATRA in hepatoma cells. Methods: Human hepatoma cells line, Bel 7402 and HepG2 (AFP-producing), or HLE(non-AFP-producing) were selected for performing this investigation; Confocal laser microscopy was used to observed the co-locazition of AFP with RAR-b, and the expression of CXCR4 in membrane of the cancer cells; siRNA was applied to knockdown the expression of AFP, and full length cDNA of AFP gene was inserted into pcDNA3.1(named pcDNA3.1-afp) for constructing AFP expressed vector; Interaction of AFP with RAR-b was analyzed by coimmunoprecipitation (Co-IP); Chromatin immunoprecipitation (ChIP)-PCR was applied to study the interaction of RAR-b with CXCR4 promoter. Results: The co-location of AFP with RAR-b in cytochylema; Co-IP indicated that AFP has a property to interact with RAR-b in Bel 7402 or in HepG2 cells, and the same effect also generated after HLE cells were transfected with pcDNA3.1-afp for 36 hours; The expression of CXCR4 was repressed concomitant with blockdown the expression of AFP by siRNA in Bel 7402 or in HepG2 cells, but the expression of CXCR4 was enhanced while HLE cells was transfected with pcDNA3.1-afp, and co-treatment with Ly294002, the expression of CXCR4 was partial inhibited. Analyzed by ChIP-PCR indicated that AFP was able to impeded RAR-b binding to the promoter of CXCR4, treated with ATRA (160 lmol/L) for 12 h, the coupling of RAR-b with CXCR4 promoter was increased, and Western blotting analyzed showed that the expression of AFP was suppressed.

Conclusions: This was first evidences to show that AFP up-regulated the expression of CXCR4 through blocking the transcriptional activity of RAR-b; AFP has a property to stimulate the expression of CXCR4 was the crucial event that promote the metastasis of hepatoma cells. Recently, another mechanism of intercellular communication has been proposed that involves Tunneling Nanotubules (TNTs), which contain Factin as a prominent cytoskeleton component. In contrast to gap junctions that permit direct passage of small molecules between cells, TNTs allow for exchange of larger materials, including organells, nutrients and cytoplasmic constituents. Association of such bridges with malignant transformation in vitro and their potential for supporting cancerous proliferation in vivo has been proposed. An in vitro study was conducted on HepG-2 cells to elucidate TNTs as novel conduits for intercellular communication and promising target for anti-cancer therapy using gallic acid (GA) as anticancer agent. Methods: Cell proliferation after graded concentrations of GA treatment was measured by MTT assay. Cell membranes were fluorescently labeled with lipophilic stain Vybrant Ò Dii (red) and Dio (green), whereas F-actin was labeled with Oregon green and observed under laser confocal scanning microscope and fluorescent microscope. Study was also conducted to observe intercellular communication between normal liver cells (Chang) and malignant HepG-2. Number of cytonemic structures, length and breadth of TNTs in HepG-2 were measured with computer assisted program. Results: GA significantly reduced the cellular proliferation and also inhibited polymerization of F-actin that eventually decreased the cellular protrusions, cytonemic and TNT like structures in concentration dependant manner. Cell shape was also distorted with significant decrease in its size.

Conclusions: Modulatory effect of anticancer agent on F-actin would be helpful in perturbation of cellular cytoskeleton, which will help in controlling cellular protrusions, cytonems and TNTs that would eventually affect the cellular communication, migration and/or metastasis. Thus, TNTs can be considered as novel conduit for intercellular communication and as such, promising targets for anti-cancer therapies. The results showed that the siRNA duplexes specifically reduced AR genes expression at a dose-independent manner at protein level 24 h later. And inhibitions of Caspase-12 expression were also found in a dose-dependent manner in these HepG2 cells. Being transfected with the siRNA duplexes 24 h later, the HepG2 cells were treated with 5-Fu. 12 h later, a significant increase in apoptotic cell death was found in HepG2 cells at a concentration-dependent manner.

Conclusions: As a transcriptional factor, AR is involved in the regulation of Caspase-12 genes expression in HepG2 cells. Our data showed that knocking down AR protein level leads to increase in apoptosis induced by 5-Fu in HepG2 cells. Presumably, this increased apoptosis might be partially associated with inhibition of Caspase-12 expression by silencing human androgen receptor expression. Aims: Beta2-glycoprotein I (beta2-GPI) is an autoantigen in the antiphospholipid antibody syndrome. We have found that beta2-GPI could specifically binded to HBsAg and hepatocellular carcinoma cell membrane. This study determined the effect of beta2-GPI on the HCC in vitro. Methods: Hepatocellular carcinoma cells, SMMC-7721, exposed to the following different treatments. Group one: cells transfected with beta2-GPI; group two: cells transfected with HBsAg; group three: cells cotransfected with beta2-GPI and HBsAg; group four: cells transfected with control vector; group five: normal control cells. NF-jB activation was measured by non-radioactive EMSA and immunofluorescence. Whether IjB kinase inhibitor (BAY11-7082) or tyrosine kinase inhibitor (Piceatannol) played the role by analyzing the IjB activation by non-radioactive EMSA. Phosphorylated amino acids and residues on the IjBa were detected by co-immunoprecipitation assay followed by western blot analysis and immunofluorescence. Results: Group one and group two induced the activation of NF-jB with the relative NF-jB DNA binding activity of 55.84 and 50.25, respectively. However, higher relative NF-jB DNA binding activity was observed in the group three (75.5). BAY11-7082 decreased NF-jB activation more obviously than Piceatannol in a time dependent manner. Introduction: Injuries to the liver in the form of viral assault, ethanol or drugs can turn on the production of certain growth factors and mitogens for liver regeneration, which in turn will cause activation of the receptor tyrosine kinases, leading to the activation of various signaling events. Ethanol is the most common culpable chemical agent thought to play the role of a carcinogen. Ethanol induced liver injury and alteration in growth factor profiles and the consequent deregulation in signaling can act as a driving force for uncontrolled proliferation of the hepatocytes, leading to tumor growth. In present study we evaluated the hypothesis that chronic EtOH exposure activates Wnt b-catenin signaling in liver Aims: To study the differential mRNA expression profile of b-catenin in alcoholic liver cirrhosis and ethanol induced HCC. Methods: Quantitative Real time PCR analysis was done for b-catenin using bactin as endogenious control. Total RNA extracted from liver tissue of ethanol related cirrhosis (n = 10), HCC (n = 5) and healthy liver (n = 5). Immunohistochemistry was done for b-catenin. that its down-regulation is associated with cell migration, invasion and intrahepatic metastasis. Very few is known on the pathways and transcription factors that regulate miR-122 expression in the liver. The aim of this work was to isolate and study the promoter region of the HCC related miR122. Methods: The putative intergenic pre-miR-122 promoter region (-5000/ +1000 bp) has been studied using the UCSC, motif.genome.jp, dragon and genomatix resources. Expression of miR-122 mature miRNA was evaluated by a two step stem-loop real-time RT-PCR using the U38b small RNA as endogenous control. Recruitment of transcription factors on putative pre-miRNA promoters vivo has been assessed by ChIP. DNA methylation has been studied by bisulfite treatment followed by methylation sensitive PCR and direct sequencing.

Results: miR122 is down-regulated in HCC and NT tissue as compared to normal livers in a validation cohort of 47 italian HCV-related HCC. miR122 was downregulated in 55% and upregulated in 28% of our HCCs. Low miR-122 tumors were less differentiated and more often multinodular. In silico analysis of the putative miR122 promoter revealed the presence of two conserved CpG islands. CpG island 1 (proximal to the TSS) is methylated in vivo in HepG2 cells. Accordingly, the demethylating agent azacytidine increases miRNA-122 expression but has no effect on the expression of other HCC deregulated miRNAs (miR-199, -21 and -221 There were different significances between the five proapoptotic proteins with Bcl-2 respectively (all P \ 0.01). All the expression intensities of proapoptotic proteins in our study were significantly higher than that of Bcl-2. Moreover, the former indexs were mainly found at the grade (+) or (++), among which Bax expression was the highest and its positive rate was 88.57% (31/35 cases). While the latter was mainly found at the grade (+), and its positive rate was only 47.36% (9/19 cases). The expression rates of Cyto-c in low, medium and high PHC differentiation were 25.00% (1/4 cases), 91.18% (31/34 cases) and 100% (7/7 cases), respectively. The expression intensity was increased with the improvement of tumor cells differentiation. Conclusions: It is suggested that both extrinsic and intrinsic apoptosis pathways are involved in the regulation of apoptosis in PHC. In addition, the sensitivity of HCC diagnosis according to AASLD guideline was 89.6% in pathologically cirrhotic patients and 92% in pathologically non-cirrhotic patients. No significant difference of sensitivity exists between cirrhotic and non-cirrhotic groups (either imaging or pathologic cirrhosis). Further analysis also indicated that large tumors ([2 cm) exhibited a significantly higher sensitivity of HCC diagnosis than small tumors C2 cm) (P \ 0.001) Conclusions: According to AASLD guideline, we can get a high sensitivity of HCC diagnosis for tumors larger than 2 cm. Diagnostic sensitivity for HCC less than 2 cm was still low, which may need further biopsy. The AASLD guideline can also be applied on non-cirrhotic patients.

Expression of CAS in Hepatocellular Carcinoma Tissues and Its Relationship with HBV Infection Hong Zang 1 1 302 Hospital, Xisihuan Zhonglu 100, Beijing, China

Background: The cellular apoptosis susceptibility (CAS) gene has important roles in process of tumor necrosis factor (TNF)-induced apoptosis and cell proliferation. Up-regulated expression of CAS is found in Hepatocellular carcinoma (HCC), suggesting that it may play an important role in the development of human HCC.In this study, we observed CAS protein expression and aim to investigate whether CAS protein could serve as a pathologic marker for HCC diagnosis and to explore the roles of CAS expression in HBV infection associated HCC. Methods: The expression of CAS protein in HCC and its paracarcinoma tissues, non-tumor liver cirrhosis and hepatitis tissues were detected by immunohistochemistry. Meanwhile, HBsAg, HBcAg and HBV DNA in HCC tissues with HBV infection were examined by immunohistochemistry and in situ hybridization respectively. Results: The expression of CAS protein was significantly higher in HCC than in its paracarcinomas tissues (P \ 0.01), and higher in paracarcinomas tissues than in non-tumor liver cirrhosis and hepatitis tissues (P \ 0.01).Poorly differentiated tumors immunochemically stained stronger than moderately or well differentiated (P \ 0.01). CAS protein expression was significantly higher in HBV-infected HCC tissues than that of in non-HBV infection (P \ 0.01). Meanwhile, in HBV-infected HCC tissues, the staining intensity score of CAS protein in HBV DNA positive HCC tissues was significantly higher than HBV DNA negative tissues (P \ 0.05).

Conclusions: Higher expression of CAS protein is found in HCC tissues, and the intensity of CAS protein expression is related closely to tumor differentiation.We suggested that CAS protein might serve as a marker for HCC diagnosis and differentiation estimation, and deduced that CAS might play an important role in the initiation of HBV infection associated HCC through upregulating expression of CAS.

Top 10% Posters Background: Thrombin plays an important role in the invasion and metastasis of hepatocellular carcinoma (HCC). In this study, we investigated the expression of protease-activated receptors (PARs) in HCC to define their prognostic significance after curative surgery. Methods: Real-time fluorescence quantitative reverse transcriptase chain reaction (qRT-PCR) technology was used to investigate thrombin and PAR1, 3, 4 expression on HCC cell lines. The technology was also used to detect their expressions in primary HCC tumors and their secondary metastatic lesions compared with surrounding non-cancerous tissues. We also investigated PAR1, 3, 4 expression as well as thrombin on paraffin tissue sections containing 230 HCC patients by Immunohistochemistry. Results: The qRT-PCR results show that PAR1 expression has significant difference between normal group and low-metastasis one in cell lines. On the Contrary, PAR3 expression has no significant differences among all three groups and PAR-4 expression was hardly detected in the MHCC97H cell line. We also found that PARs expressions of the cancer recurrence group had no significant difference with non-relapse group in tissues. The immunohistochemical results show that the 1, 3, 5-year overall survivals (OS) in the high thrombin expression group are significantly lower than the low expression group, and the time to recurrence (TTR) in the patients with high expression of thrombin is significantly higher than the low expression group (P \ 0.05). Conclusions: Although thrombin is associated with OS, TTR of HCC patients, the thrombin receptors (PARs) may not be involved in this process.

The Results: GOLM-1 was significantly up regulated in cancerous tissue when compared with noncancerous tissue both in mRNA (P \ 0.001) and protein levels (P \ 0.001), meanwhile GOLM-1 was also found highly expressed in patients with metastasis in mRNA level (P = 0.032) and protein level (P = 0.005). GOLM-1 was also differently expressed between high metastasis and non-metastasis HCC cell lines (P \ 0.05). Cell proliferation measured by CyQUANT Cell Proliferation Assay Kit was significantly inhibited in GOLM-1 konck-down cell lines (P \ 0.001) Conclusions: We validated the gene array results both in mRNA and protein level in patients with and without metastasis and firstly proposed that GOLM-1 was a target gene favoring HCC metastasis. Knocking down of GOLM-1 could inhibit the proliferation of MHCC97 h cells in vitro. Background: Over a decade, there was nearly a two fold increase of the proportion of HCC among patients with chronic liver diseases in Egypt with a significant decline of HBV and slight increase of HCV as risk factors.

Objective: This study was done to evaluate the role of HGF and IL-1 b in HCV infection and HCC in a trial to find a predictive value of developing HCC. Methods: Sixty individuals divided into three groups. 20 HCC patients, 20 HCV infected and 20 healthy individuals. Alpha feto protein (AFP) and HGF were assayed by ELISA. HCV-RNA was measured by RT-PCR and IL1-b was measured in culture supernatant of stimulated lymphocytes by phytohaemaglutinin (PHA) by ELISA Results: AFP was significantly increased in both HCV and HCC groups than control group (P \ 0.0001, 0.001, respectively) and was higher in HCC than HCV group (P \ 0.0014). HGF was significantly higher in HCC than both HCV and control groups (P \ 0.0001, 0.0013, respectively). IL1-b was increased in both HCV and HCC than control group (P \ 0.009, 0.04 respectively) but it was higher in HCV than HCC (P \ 0.044). HGF was directly correlated with age and AFP in all groups (P \ 0.004, 0.0001, respectively). Also HGF was directly correlated with HCV-RNA and AFP in HCC group (P \ 0.027, 0.004, respectively) Conclusion: IL-1 b was increased in HCV and HCC. Thus, an alteration of cytokine level can favor the establishment of persistent HCV and enhance liver cirrhosis and HCC. The serum level of HGF represents the degree of the carcinogenic state in HCC irrespective of AFP level .Therefore, the determination of serum HGF concentrations may be useful as a third tumor marker of HCC in detection as well as follow-up therapy.

Role of Promoter 19) . Sexual and aging factor was not meaningful (P value 0.471, 0.572). PIVKA-II level of metastatic group was marked increased rather than non-metastatic group (2,500 ± 0 vs. 256 ± 212, P value = 0.022). AFP level of metastatic group was increased rather than non-metastatic group, but it was not statistically meaningful (853 ± 208 vs. 106 ± 5, P value = 0.587). Tumor sizes of metastatic group are arranged from 1.6 to 16 cm. Five metastatic patients of larger mass (more than 7 cm) showed high PIVKA-II level ([2,000) with variable level of AFP (0.9-706). Two metastatic patients of smaller mass (less than 7 cm) showed lower level of PIVKA-II (\300) with high level of AFP ([600). All non-metastatic group showed smaller size (less than 7 cm) with low level of PIVKA-II (\2,000, 27-1,705). Eggel class 2 (mass form) and class 3 (diffuse infiltrative form) showed high PIVKA-II rather than class 1 (nodular form). Conclusion: Metastatic HCC is frequent in patients with larger mass (rather than 7 cm) and high level of PIVKA-II ([2,000). But PIVKA-II level was closed correlated with mass size and morphology. Smaller mass with morphology class 1 with metastasis was correlated with high level of AFP. Aims: To assess logp9/4 based on serum N-glycan profiling in diagnosing HBV-related hepatocelluar carcinoma (HCC) follow-up study. Methods: Patients with pathologically confirmed HBV-related HCC were enrolled in this study. Sera were collected before and 4-8 weeks after hepatoma radical resection only those whose AFP were positive but turned to be negative after operation were recruited. N-glycan profiling of serum proteins was determined by DNA-sequencer-assisted fluorophore-assisted carbohydrate electrophoresis (DSA-FACE) with a capillary electrophoresis-based sequencer.

Results: A single agalacto core-1, 6-fucosylated biantennary glycan (NG1A2F, peak 4) was increased and a branching triantennary glycan (NA3Fb, peak 9) was decreased in samples from post operation cases. In addition, the Logp9/4 of post operation samples was lower than that of pre-operation. The decreased level of Logp9/4 was correlated well with the improved clinic data and liver function. Background: Lens culinaris agglutinin-reactive fraction of alpha-fetoprotein (AFP-L3) as well as alfa-fetoprotein (AFP) and des-gamma-carboxy prothrombin are used worldwide for the diagnosis of hepatocellular carcinoma (HCC). Because a high-sensitive AFP-L3 immunoassay system was developed by Wako chemical industry (Japan), we became to evaluate accurate AFP-L3 level in serum of HCC patients with low total AFP. In this study, we assessed the clinical usefulness of the high-sensitive AFP-L3 immunoassay system with sera obtained from HCC patients and anti-HCV-positive patients without HCC. Also we measured AFP-L3 values using patients sera at 6 months before imaging diagnosis of HCC Background: Cancer stem cells are associated with metastatic potential, treatment resistance, and poor patient prognosis. Although hepatectomy is the best method to provide long-term survival for patients with hepatocellular carcinoma (HCC), high postoperative recurrence rate is a major problem. We screened expressions of 3 stem cell markers (Oct4, Sox2 and Nanog) in HCC patients by immunohistochemical staining and investigated the function and molecular mechanism of Oct4 in HCC progression. Methods: Expression of Oct4, Sox2 and Nanog were assessed by immunohistochemistry from 200 patients who had undergone hepatectomy for histologically proven HCC. Expression of Oct4 among HCC cell lines with different metastatic potential was determined by real time and western blot. Gain-and loss-of-function studies in vitro examined the function of oct4 on proliferation, apoptosis, invasion, and transformation ability of HCC cell lines.

In vivo assays for tumor growth and metastasis was conducted on nude mice. Results: Immunohistochemically, we did not detect the positive staining of nanog, but Oct4 and Sox2 were highly expressed in HCC patients. The amount of Oct4 in HCC cell line with high metastatic potential was more than that of low metastatic potential. In vitro, we confirmed that Oct4 promoted cell proliferation and invasion and also increased the ability of cell transformation. Conclusions: Oct4 and Sox2, 2 stem cell markers, were expressed in HCC patients. Both gain-and loss-of-function studies in vitro and in vivo validated Oct4 as a potent oncogene and it might play a crucial role in HCC progression. Objective: To investigate differences in protein expression of hepatocellular carcinoma and para-carcinoma tissues. Methods: Laser capture microdissection (LCM) was applied to isolate hepatic parenchymal cells of HCC tissues and para-carcinoma tissues. Twodimensional electrophoresis was used to screen out the altered proteins. The differentially expressed proteins were identified by ESI-MS/ MS. Western blotting and immunohistochemistry were used to analyze the altered expression of dimethylarginine dimethylaminohydrolase1 (DDAH-1) in HCC. Results: 12 proteins were obtained which were significantly altered in the HCC tissues compared with the para-carcinoma tissues, including DDAH-1, a critical regulator of the nitrogen monoxidum (NO) signaling pathway. Western blotting showed that the expression of DDAH-1 was markedly increased in 16 of 20 HCC tissues in comparison with para-carcinoma tissues. Immunohistochemistry showed that DDAH-1 was localized in the cytoplasm and was highly expressed in HCC tissues. Conclusion: LCM is a useful technological method in the proteomic study of cancer tissue. DDAH-1 may be used as a potential biomarker for HCC and give some clues to study the mechanism of HCC.

Increased FoxP3+ Aims: To investigate whether single nucleotide polymorphism (SNP) of IL-10 and ALDH2 genes are associated with hepatocellular carcinoma. Methods: The distributions of the allelic, genotype and haplotypes at IL-10 rs1800896(-1082G/A)/rs1800871(-819T/C)/rs1800872(-592A/C) and ALDH2 rs4646778/rs671 were detected by PCR and oligonucleotide microarray. The sample population included 70 HCC patients, 34 patients with liver cirrhosis, 50 patients infected with hepatitis B virus and 52 healthy volunteers. Results: No significant difference was found in frequencies of genotypes and haplotypes of the three positions of IL-10 and two positions of ALDH2. The C allele frequency at ALDH2 rs4646778 (A/C) position was higher in HCC patients than in controls (79.5 vs. 65.4%, P = 0.037, OR = 2.051, 95% CI:1.037-4.058). Between HCC patients with different Child-Pugh liver function grades, IL-10-819 TT genotype frequency was higher in grade B patients than that in grade A patients (66.7 vs. 44.4%, P = 0.048). Among all the patients with liver diseases, GG genotype frequency, as well as G allele frequency at ALDH2rs671A/G position were lower in HBeAg-positive individuals than that in HBeAg-negative individuals.(GG: 65.8 vs. 85.1%, P = 0.033; G: 81.6 vs. 92.6%, P = 0.027).

Conclusions: IL-10 and ALDH2 polymorphisms significantly influence the occurrence and the progression of HCC, and ALDH2 gene polymorphisms also influence HBeAg seroconversion. , hepatic vein level in 5 (7.5%), and combined level in 6 (9.0%). Thirty-three patients underwent ballooning, stenting and/or transjugular intrahepatic portosystemic shunt (TIPS). During the mean follow-up period of 109.6 months, 17 patients developed HCC. Among them, BCS and HCC were diagnosed simultaneously in three patients. The incidence of HCC in patients with BCS was 2.9%/year. No significant difference was observed between patients who developed HCC and those who did not, except for the pressure gradient between the right atrium and IVC. Conclusion: The annual incidence of HCC in patients with BCS (2.9%/year) was similar to the incidence in patients with other etiologic cirrhosis (2.5%/ year) in Korea. The pressure gradient between the right atrium and IVC was a predictor of HCC development. Background: Tumor necrosis factor-a (TNF-a) is a proinflammatory cytokine with multiple biological effects and may play essential roles in inflammationlinked tumor development. The aim of this study was to evaluate the association between TNF-a gene polymorphisms and the risk of hepatocellular carcinoma HCC in Chinese population. Methods: The study population consisted of 101 HCC patients with evidence of chronic hepatitis B virus (HBV) infection and 166 health controls,all were Chinese Han ethnic. TNF-a polymorphisms-1031(T/C), -863(C/A), -857(C/T), -851(C/T), and -308(G/A) were genotyped by PCR and direct DNA sequencing. Allele and genotype frequencies between cases and controls were compared using the chi-square (v 2 ) test with the statistical software SAS 8.0. Association testing for the haplotypes as well as the measure of linkage disequilibrium (D') between all pairs of SNPs was performed using the Haploview v3.1 software. Results: Of the five TNF-a polymorphisms investigated in our study, -1031(T/C), -863(C/A), -851(C/T), and -308(G/A) did not show correlation with HCC. However, -857(C/T) was found to be significantly associated with the increased risk of HCC. TNF-a-857T allele was more frequent in HCC patients than in control (P = 0.0093; odds ratios, 0.51; 95% confidence intervals, 0.30-0.85). High linkage disequilibrium was detected between -863C/A and -1031T/C (P \ 0.001, D 0 = 1.0). Five haplotypes were found to occur in more than 1% of the study population. However, no significant associations between any of the estimated haplotypes and HCC were found. In addition, a literature review revealed that the distribution of the TNF-a genetic polymorphisms among populations with different ethnic origins varied significantly. Conclusions: Our data imply that genetic variations in TNF-a gene may confer different susceptibilities to the development of HBV-related HCC. Carriage of the -857T allele is a significant predictor of HCC development, and therefore it may be used as a biomarker for one's susceptibility to HCC. Result: Without inhibiting all the tested tumor cell proliferation at the concentration between 0 and 20 lg/mL, pazopanib significantly inhibited cell migration and invasion, and induced cell apoptosis of HCCLM3 and PLC/PRF/ 5 but not HepG2 at the concentration of 20 lg/mL. In vivo tumor growth was significantly inhibited by pazopanib at the dosage of 40 mg/(kg day) in all the 3 xenograft models. Intratumoral CD31-positive microvessel density (MVD) decreased in pazopanib treated tumors, whereas the density of a-SMA-positive pericytes increased. Intratumoral blood perfusion detected by CEUS was significantly reduced by pazopanib treatment, peak intensity is significantly impaired in the treated tumors before the time when the treatment efficacy on tumor size can be observed. Mean transit time of the contrast media in the hotspot area of the tumors is reversely correlated with intratumoral microvessel density (r = -0.661, P = 0.001). Conclusion: These preclinical results indicate that the therapeutic effects of pazopanib in HCC xenografts might be at least partially attributed to its antiangiogenic effect, and this effect could be evaluated by quantitative CEUS. Method: 43 patients with HCC who were undergone RFA treatment were analyzed retrospectively. S-500 RF Generator, (Medsphere Co., Ltd) equipment was used for the treatment. Mean hospital stay was 3 days. Result: Number of lesions were 1.3 ± 3 SD, with the diameter of 2.6 cm ± 2.7 SD. No mortality was observed so far. Only two patients had a complication of skin burn. While the total HCC recurrence was observed in 11 patients (25.5%), the local recurrence was observed in 4 (9.3%). Six patients had previous treatments, which were TACE (n = 4) or liver resection (n = 2) respectively. Not much data was available about viral status of patients. 16 patients had their viral markers checked, out of which 8 (50%) patients had positive HBsAg, 7 (43.7%) were positive for HCV, while one patient (6.3%) had both HCV and HBV. Conclusion: Though RFA was introduced almost 15 months ago, the number of patients treated using this procedure is quite modest (2.8% of HCC patients). Many patients are still left with no treatment. According to our results RFA is a safe procedure with minimal complications. Besides technical aspects, the fact that RFA is not covered by insurance in our country is holding back the progress of the field. Background/Aims: Hepatocellular carcioma (HCC) is sometimes accompanied with an involvement of inferior vena cava(IVC). Aggressive surgical approaches for these patients has been reported to be useful. We herein investigate 8 operated cases of HCC with involvements of IVC (7 cases with tumor thrombus, one with the direct invasion). Patients and operations: The profiles of eight cases are listed on Table 1 . In all cases, hepatic resections were carried out after dissections of hepatic artery and the first branch of the portal vein on the tumor-side. Then thrombectomy and resection of invaded wall of IVC were done. Results: Major hepatic resections were performed in all cases. THVE was done during the tumor thrombectomy in five cases. The average time of THVE was 10.6 min, so no extracorporeal circulation system (ECS) was needed. The average of operative hemorrhage was 2,234.4 ml and 3 patients did not require an allogeneic transfusion by using auto-transfusion. There was neither operative death nor hospitalization death, and no major operative complication occurred. The mean survival time is 28.4 months, and 3-year survival rare was 65.6%. Two cases with tumor recurrence which are well controlled by hepatic arterial infusion chemotherapy are still alive (Table 2) . Conclusion: The method that hepatic resections are carried out before the treatment for the lesions of IVC is useful. Because the treatment for IVC is done safely under good visual field, no severe complications due to the liberation of tumor thrombi occurred, not to use ECS is economical and the prognosis better. Cox-regressional hazard model, Child-Pugh class were the independent prognostic factors affecting poor survival (RR = 6.8, P = 0.002). Preoperative TACE did not affect survival outcome. Thirty-four (85%) patients recurred. Of these 32 patients recurred within 2 postoperative years. Overall 1-, 3-and 5-year disease-free survival rate were 31.2, 14.2, and 8.5% respectively. On univariate analysis, Child-Pugh class was significantly related to poor disease-free survival. On Cox-regressional hazard model, Child-Pugh class were the independent prognostic factors affecting poor survival (RR = 4.0, P = 0.03). Only four patients survived without recurrences. Preoperative TACE did not affect survival or disease-free survival outcome.

Conclusion: Survival outcome of hepatic resection was better than non-surgical management. Although most patients experienced recurrences, a few patients had chance to cure after hepatic resection. [4 log(10) copies/ml at the time of resection were associated with HCC recurrence, and besides these variables, poorly differentiated tumor, intrahepatic micrometastasis, and HBeAg seropositivity at resection were also associated with poorer postoperative survival. On multivariate analysis, tumor size [5 cm (P = 0.050), Child-Pugh class B (P = 0.001), vascular invasion (P = 0.042), and sustained low viremia [HBV DNA[ 4 log(10) copies/ml] (P = 0.003) remained significant for recurrence, whereas tumor size [5 cm (P = 0.020), Child-Pugh class B (P = 0.002), and sustained low viremia (P = 0.021) remained significant for poor survival. Among patients in whom HBV DNA were serially checked, both HCC recurrence and postoperative deaths were more frequent in patients with fluctuating or lasting highlevel viremia than in those with persistently low or undetectable HBV DNA levels (all P \ 0.05). Conclusions: High HBV replication state is among the most important predictors of adverse outcome after resection in patients with HBV-related HCC. The sustained suppression of HBV to the lowest possible level is of great importance to achieve long-term recurrence-free survival. Our findings underscore the need for future work that tests the applicability of antiviral therapy in this setting.

Laparoscopic versus Open Hepatectomy-A Meta-analysis Markiyan Soloviy 1 1 Lviv National University of Medicine, Grusheva str. 14, Lviv, 79014 Ukraine

Background: There is an increasing interest in laparoscopic liver resection because of its potential benefits to the patient. Introduction: Complete mobilization of the right liver before parenchymal transection has been a standard approach during major right hepatectomy. However, in case of the patient with huge right hepatic tumor that had infiltrated the surrounding organs such as diaphragma or retroperitoneum, the mobilization of the right lobe of the liver may result in massive bleeding. Case: We experienced the patient with huge hepatocellular carcinoma occupying the right lobe that was estimated to be more than 30 cm in diameter. To avoid massive bleeding during the surgery, the liver hanging maneuver was performed in this case. This technique leads the anterior approach of liver transection much easier. The weight of the removed specimen was 6200 g, the operation time was 4 h 10 min. and the estimated blood loss during the operation was 3,100 ml. There was marked collateral veins in the retroperitoneum caused by portal vein tumor thrombus and a large blood flow into the tumor from right inferior phrenic artery. The most of the bleeding occurred during the resection of the retroperitoneum and diaphragma. In the patient like this case, the embolization of inferior phrenic artery before the surgery may reduce the bleeding during the resection of the tumor with diaphragma. Herein, we show the operation video and suggest the technique we used during the surgery.

died. The overall survival rate was significantly higher (P = 0.018) and recurrence rate was lower (P = 0.018) for patients in minimal fibrosis group. Among the serum markers, the aspartate aminotransferase-platelet ratio index (APRI) which yielded the highest the area under the receiver operating curves with a level of 0.867 exhibited the most reliable discriminative ability for predicting advanced fibrosis. Besides, the overall survival rate was significantly higher (P = 0.003) and recurrence rate was lower (P = 0.005) for patients with an APRI of 0.47 or less. Conclusions: For patients with small and solitary HBV-related HCC underwent resection, minimal fibrosis is associated with lower incidence of recurrence and better survival. Besides, APRI could serve as a surrogate for assessing hepatic fibrosis and predicting survival.

Hepatol Int (2010) Ruptured hepatocellular carcinoma (HCC) occurs in 3-15% of HCC patients. The mechanism of rupture is still not exactly known. It is so emergent mostly and high mortality (25-75%) in the acute stage, and prognosis is not good. The option of treatment is observation, transarterial embolization (TAE) or transarterial chemoembolization (TACE) as palliative treatment and one staged operation or two staged operation as curative treatment. Two staged operation is the most ideal for cure, because we can stabilize the acute emergent state by TAE or TACE. We report a HCC patient who had ruptured mass and took a two staged operation. He was 68 years old male, heavy alcoholics, chronic hepatitis C and liver cirrhosis. Initially, we controlled the bleeding by TAE and took a TACE, after 3 months, and took a operation of hepatic resection and radiofrequency ablation (RFA) successfully, after 11 months from TAE. It is 5 months of post operation state, and healthy child A. Although it is so short of follow-up period, It is so fortunately good prognosis, so far. Background: Liver cancer (HCC) is number two cause of death in Mongolia with roughly 4,000 newly detected cases each year. Aim: Our study is aimed to analyze the current outcomes of liver resection and to improve the performance in the future. Method: 285 patients who underwent surgical treatment between years 2003 and 2007 at the National Cancer Center (NCC) were enrolled in this study. Variables included patient age, gender, predisposing liver conditions, stage of cancer, type of surgery, post surgical complications and pathology results. SPSS (version 15) software was used for statistical analysis. Results: 60.9% of patients were male and 39.1% were female with average age of 56 years. Liver cirrhosis was present in 47.3% of the patients, majority of whom were in Child A stage 83%. US was the primary modality (93%) to assess the tumor size which was on average 5.31 9 5.67 cm in diameter. CT was used in 70.5% of patients with average tumor size of 5.4 9 6.3 cm. 28% of the patients were diagnosed at their early stages (I, II), while 71.1% were diagnosed at their late (III, IV) stages. 43% (n = 108) of patients had minor resection, while the rest underwent major resection (32%), exploratory laparotomy (20.7%), laparoscopy (2.4%) and Wedge resection (1.2%) respectively. HCC was the main type of cancer detected, comprising 96.1% of the total, with cholangiocellular carcinoma and other types of malignancy making the rest (3.9%). Liver failure was the major post-operative complication, making 72% (n = 34) of the total n = 47. Though it is rapidly decreasing, the post-operative mortality rate was 5.4%. Conclusion: Although much attention was paid to patient safety in the past 4 years, the post-operative mortality rate of liver resection remains significantly high, and there is much room for improvement in this field. mally invasive surgery. Herein we report our preliminary experience in robotic-assisted laparoscopic anatomic hepatectomy in 13 consecutive patients. Methods: Between April and July 2009, 13 consecutive patients underwent robotic-assisted laparoscopic anatomic hepatectomy for benign and malignant hepatic diseases. Major hepatectomy was performed in 9 patients (1 right hepatectomy, 1 right hepatectomy and simultaneous enucleation of lesion in segment IV, 6 left hepatectomy and 1 left hepatectomy and caudate segmentectomy, Roux-en-y hepaticojejunostomy for radical resection of hilar cholangiocarcinoma), left lateral sectionectomies in 4 patients. All the robotic-assisted hepatic resection procedures were performed anatomically with hilar dissection. Prior vascular control of the portal vessels was carried out before the parenchymal transection whenever possible. Eight major hepatectomies were for malignant disease and 5 hepatectomies (1 left hepatectomy and 4 left lateral sectionectomies) for benign disease. Results: No conversion to laparotomy occurred in all 13 patients. The mean operative time was 338 minutes (range150-720). The mean blood loss was 280 ml. No patients required intraoperative or perioperative blood transfusions. The Pringle maneuver was never needed. Without postoperative mortality, transient bile leakage was observed in the patient with hilar carcinoma. The mean postoperative stay was 6.7 days (range 2-13).

Conclusions: These preliminary results show that robotic-assisted laparoscopic anatomic hepatectomy is safe, feasible. It will be necessary for more long-term, evidence-based outcomes to prove their efficacy and further research on the cost-effectiveness is still required.

Prognostic Role of Preoperative Radiologic Morphology of Huge ( ‡10 cm) Hepatocellular Carcinoma Woo Young Shin 1 , Kyung-Suk Suh 1 , Teahoon Kim 1 , Young-min Jeon 1 , Nam-joon Yi 1 , Kuhn Uk Lee 1 1 Seoul National University Department of Surgery, 28 Yongon-dong, Jongno-gu, Seoul Korea

Background: Hepatic resection for huge (C10 cm) hepatocellular carcinoma (HCC) is challenging, however surgical outcome of huge HCC is poor. Preoperative evaluation for balancing the risk of operation against the benefit of operation was important in patients with huge HCC. Hence, we classified subgroup based on morphological feature using preoperatively imaging study, analyzed long-term outcome according to subgroup, and investigate prognostic factor of patients with huge HCC. Patients and methods: From January 2000 to December 2005, 49 patients who underwent hepatic resection for huge HCC were retrospectively reviewed. Median age was 51 years (33-73). There were 44 men and 5 women. Based on preoperative radiologic morphology, patients were classified nodular HCC with irregular margin \50% (Group A, n = 33), nodular HCC with irregular margin [50% (Group B, n = 9), and infiltrative HCC with totally irregular margin (Group C, n = 7). Univariate and multivariate analyses were performed on clinicopathological variables including radiologic morphology to analyze factors affecting survival and DFS. Results: On comparison using radiologic morphology, Group C showed significantly higher rate of liver cirrhosis (27.3% and 44.4 vs. 85.7%), macrovascular invasion (12. 1 and 44.4 vs. 85 .7%), and satellite nodules (12.1 and 11.1 vs. 42.9%). Five-year survival and disease-free survival rate according radiologic morphology were 33.7 and 27.5% in Group A, 25.0 and 11.1% in Group B, and 0% in Group C. Overall 1, 3, and 5-year survival rate were 70.5, 40.6, and 26.8% respectively. Overall 1, 3, and 5-year disease-free survival rate were 36.4, 20.5, and 20.5% respectively. On univariate analysis, macrovascular invasion, satellite nodules and radiologic morphology were significantly related to poor survival and disease-free survival. On multivariate analysis, macrovascular invasion was the independent prognostic factors of poor survival (RR = 19.5, P \ 0.001) and infiltrative HCC with totally irregular margin was the independent prognostic factors of poor survival (RR = 6.4, P \ 0.001). Conclusion: Although surgical resection for huge HCC was only hope of cure, the benefit of hepatic resection in patient with infiltrative HCC with totally irregular margin was limited. as compared to cells not pre-treated with bacitracin. Indeed, bacitracin pretreatment potentiated 3BP-induced apoptotic cell death, which was evidenced by more potent activation of caspase 9. Conclusions: These results demonstrate that hypoxia-mediated PDI induction protects hypoxic HCC cells against ER stress, and that PDI inhibition potentiates HK II inhibitor-induced JNK activation and apoptosis by increasing ER stress in these hypoxic cells. Thus, the simultaneous PDI inhibition may synergistically potentiate the anti-tumor efficacy of HK II inhibitor in HCCs, especially in advanced infiltrative type of tumors which are constantly exposed to hypoxic environment. .52, 75% in male) who had developed HCC after 6 months of antiviral therapy were included. As a control group, a total of 90 patients with CHB (mean age 60.29 ± 9.17, 80% in male) did not received antiviral therapy who had developed HCC were enrolled. The medical records were reviewed for clinical, laboratory and radiographic evaluations. Results: About 26 months (mean 26.08 ± 19.8 months) has passed for HCC to be discovered after antiviral therapy. At the time of diagnosing as HCC, 32 (88.89%) patients of liver cirrhosis and 7 (19.4%) of diabetes mellitus were established and the mean HBV DNA level was 4.6 9 10 7 ± 1.4 9 10 7 copies/mL. HBV DNA level of 20 (61.76%) was under 2,000 copies//mL. Among 16 (44.4%) patients with HCC who was HBeAg-positive has experienced seroconversion only in four cases after antiviral therapy. There were no significant difference of clinical characteristics including mean aFP level, multinodularity, portal vein thrombosis, cirrhotic change, treatment modality and median survival except tumor size (p = 0.012) between patients who received antiviral therapy and those who did not receive antiviral therapy. Conclusion: Antiviral therapy decreases the incidence of HCC in chronic viral hepatitis B. But chronic viral hepatitis B can develop HCC in spite of antiviral therapy. Clinical course of HCC in the CHB during antiviral therapy seems to be similar to those without antiviral therapy. Periodical follow-up with surveillance program for HCC shoud be performed for patients of the CHB undergoing antiviral therapy. Further large scaled study should be warranted.

Level of Alpha-fetpprotein in Diabetic Pateint Having Hepatocellular Carcinoma Ayesha Baig 1 , Sadik Memon 1 , Madiha Zaki 1 , Mashal Saeed 1 , Falak Naz 1 , Ishrat Saba 1 , Saima Laghari 1 1 Isra University, Hala Road, Hyderabad, Sindh, Pakistan

Background: The management of hepatocellular carcinoma is influenced not only by the tumor status and underlying liver function but also by any comorbid illness in the patients and diabetes mellitus is one of them. The present study was undertaken to evaluate the level of alpha-fetoprotein in diabetic patients having hepatocellular carcinoma. Methods: The study type is of Observational descriptive study. This study was conducted in the department of Medicine Isra University Hospital, Hyderabad from November 2005 to October 2008. Total 200 patients of hepatocellular carcinoma were analyzed. They were divided into groups with and without diabetes mellitus. Clinical Status, laboratory findings were noted, alpha-fetoprotein level measured, and patients were followed for one year to see the prognosis in both groups. Results: There were 200 patients in this study with mean age ±SD of the patients were 53.70 ± 12 years. Amongst all cases of hepatocellular carcinoma 165 (85.57 %) were males and 35 (17.0%) were females, 46 (23.0%) were diabetic and 154 (77.0%) were non-diabetic. Alpha fetoprotein was compared between two groups and we found that alpha fetoprotein level was 2737.84 ± 4149.564 ng/dl in diabetic group whereas it was 11827.31 ± 29287.392 ng/dl in non diabetic group (P = 0.03) which is statistically significant. In diabetic group 13(28.26%) patients expired and 33(71.73%) were alive or lost to follow up whereas in non-diabetics group 19 (12.3%) expired and 135 (87.6%) were alive or lost to follow up showing a significant difference in between two groups (P = 0.02). Conclusion: The overall survival rate in diabetic group was significantly lower than those the in non-diabetic group. Alpha fetoprotein level was significantly low in diabetic that in non-diabetic group. Background: Esophageal variceal hemorrhage is the leading cause of death in liver cirrhosis and hepatocellular carcinoma (HCC). But the result of esophageal variceal hemorrhage and ligation in HCC is uncertain. We investigated the frequency of esophageal variceal hemorrhage and the effect of variceal ligation in the patients with HCC Material and methods: Between June 2003 and June 2008, 301 patients were diagnosed with HCC. One hundred thirty-nine patients were reviewed retrospectively with the exception of 131 patients who were lost from follow up within 1 year and 31 patients who were not performed with esophagogastroduodenoscopy (EGD) or liver dynamic computed tomomgraphy (CT). Various factors including the size of HCC, portal vein thrombosis, variceal form(F0-F3), red color sign, stage (by modified UICC), Child score, albumin, bilirubin, platelet, and splenomegaly were evaluated. Results: The gender ratio was 101:38 (male:female), mean age was 57.9 ± 10 years (27-79 years), and the number of patients of stage 1, 2, 3, and 4 were 34, 32, 27, and 46, respectively. The mean follow up period was 18.4 months and the rate of bleeding during follow up period was 12.2%. The variceal hemorrhage occurred in 17 patients during the follow up periods and mortality associated with bleeding was 23.5% (4 patients). The variceal form and red color sign showed a significance for esophageal hemorrhage (p \ 0.001). In 82 patients who were followed up from initial diagnosis to death, the ratio of esophageal hemorrhage was 7.3% (6 patients) from all cause of death.In 23 patients who were performed with prophylatic variceal ligation, 3 patients (13%) showed recurrent bleeding with 3 weeks of ligation and all hemorrhage were from the ligation ulcer. Conclusions: The mortality of HCC by esophageal variceal bleeding was 7.3% and the important predictor to the bleeding was the form of varix and red color sign.

Hepatocellular Carcinoma in HDV: Does it Differ from HBV Monoinfection Zaigham Abbas 1 , Mustafa Qureshi 1 , Saeed Hamid 1 , Wasim Jafri 1 1 The Aga Khan University Hospital, Karachi, Pakistan

Background: Hepatitis D superinfection in patients with chronic hepatitis B leads to accelerated liver injury, early cirrhosis and decompensation. It may be speculated that hepatocellular carcinoma (HCC) may differ in these patients from HBV monoinfection. This study aimed to compare clinical aspects of hepatocellular carcinoma in patients of hepatitis D with hepatitis B alone. Methods: A total of 92 consecutive HCC cases seropositive for antibody against hepatitis D virus antigen (HDV group) were compared with 92 HBsAg positive but anti-HDV negative cases (HBV group). The clinical manifestations, Child Class, tumor characteristic, and staging were compared. Results: The mean age was not different in both groups of patients (55.4 years in HDV group and 53.7 in HBV group). Other features including sex, presence of ascites, serum biochemistry, gross tumor appearance, Child Class and Okuda stage were not significantly different between the two groups. Decreased liver size was noticed in more cases of HDV group as compared to HBV group where the liver size was normal or increased (p = 0.000). HDV group had lower platelets (p = 0.063) and larger varices on screening endoscopy (p = 0.005). Multifocal tumors were more common in HBV group (p = 0.057). TNM classification showed more stage III-IV disease in HBV group (p = 0.000) Conclusion: Hepatitis D virus superinfection does not accelerate the development of hepatocellular carcinoma. However, decreased liver size and indirect evidence of more severe portal hypertension and earlier TNM stage compared to HBV monoinfection might indicate that HDV infection causes HCC indirectly by inducing inflammation and cirrhosis. Objectives: There are notably limitations for resection and liver transplantation, at present. Percutaneous ethanol injection therapy (PEIT) and radiofrequency ablation (RF) have been presented in the treatment of small hepatocelluler carcinoma as curative treatments. The aim of the study was to review the available evidence comparing PEIT to RF for HCC. Material and methods: This study was carried out on 80 cirrhotic patients in Child-Pugh A/B/C with 1-4 nodes of HCC (diameter 15-50 mm), for a total of 117 lesions. Patients were divided into two groups to receive PEIT (n = 47) or RF (n = 33). Hepatitis B virus was the leading ethiologic factor for two groups (48/80, 60%). The primary end-point was complete response (CR) 1 year after the percutaneous ablation of all HCC nodes identified at baseline. Secondary end-points were early (30-60 days) CR, complications and survival. Results: It was found that, 1-year CR was achieved in 21/47 (44.7%) and in 21/33 (63.6%) patients treated by PEIT and RF, respectively. For lesions \2.5 mm in diameter, there was better CR rate in the RF group (69.2 vs. 48.5%). An early CR was obtained in patients treated by RF 28/33 (84.8%) compared with patients treated by PEIT 36/47 (76.6%). The number of treatment sessions was smaller (2.1 times vs. 5.6 times, respectively) in RF versus PEIT. Three years survival rate was better (57.6 and 53.2%, respectively) with RF than with PEIT. RF was also considered not to be more expensive technique, due to less hospital stay and number of the sessions, comparing with PEIT. Conclusions: The early and 1-year CR rates of RF was significantly better than PEIT. Although beter survival rate was noted in RF treatment, but it was not found significantly.

Yun-Hong Xia 1 , Rong-Xin Chen 2 , Sheng-Long Ye 2 , Rui-Xia Sun 2 , Jun Chen 2 , Yi Chen 2 , Tong-Chun Xue 2 , Yan Zhao 2 Background: In the injured liver, hepatic stellate cells (HSCs) secrete many different cytokines, recruit lymphocytes, and thus participate actively in the pathogenesis of liver disease. The stroma of hepatocellular carcinoma (HCC) is markedly infiltrated with activated HSCs and associated invasion and metastasis of HCC. But little is known of the role of HSCs in HCC in immune responses. Methods: Quiescent HSCs (qHSCs) were isolated from normal Buffalo rat liver, and Intratumoral HSCs (tHSCs) isolated from Buffalo rat HCC. To detect tHSC surface molecules and gene expression by flow cytometry and fluorescence quantitative RT-PCR. To observe T cell proliferation and cytotoxic activity by 3 H thymine ( 3 H-TdR) incorporating and releasing experiment. To test cytokines expression level of T cell by ELISA. T cell apoptosis was detected by TUNEL. To observe the migration and invasion of HCC by Transwell experiments. Results: qHSCs were found to express major key surface molecules. tHSCs express scant histocompatibility complex class I, histocompatibility complex class II, costimulatory molecules, and produce a scant or major variety of cytokines. Addition of the tHSCs suppressed thymidine uptake by T cells that were stimulated by alloantigens or by anti-CD3-mediated T cell receptor ligation in a dose-dependent manner. The tHSC-induced T cell hyporesponsiveness was associated with enhanced T cell apoptosis and contributed to the migration and invasion of hepatoma cell. tHSCs was associated with markedly enhanced expression of B7-H1. Blockade of B7-H1/PD-1 ligation significantly reduced tHSC immunomodulatory activity and hepatoma cell migration and invasion.

Conclusions: tHSCs can induce T cells apoptosis, suggesting an important role of B7-H1. The interactions between tHSCs and T cells may contribute to hepatic immune tolerance and the migration and invasion of HCC.

Hepatol Int (2010) a qHSCs and tHSCs isolated from Buffalo rat normal livers and HCC were cultured in uncoated plastic plates for 2 days. Expression of surface B7-H1 was determined via staining with anti-rat B7-H1 mAb,followed by PE-conjugated secondary anti-rabbit IgG2a and analyzed using flow cytometry. tHSCs vs. qHSCs, the expression of B7-H1 was significantly upregulated, P \ 0.05. b Blockade of B7-H1/PD-1 ligation reverses tHSC-induced T cell proliferation. Buffalo rat spleen T cells (2 9 10 5 /well) were cultured in flat-bottom 96-well plates coated with anti-CD3 mAb (2 lg/mL in 100lL) for 2 days in 5% CO 2 in air. The gamma-irradiated (50 Gy) Buffalo rat tHSCs were added at the beginning of culture at a T-cell/HSC ratio of 40:1. Anti-rat B7-H1 blocking mAb was added at 15 lg/mL to block B7-H1/PD-1 ligation. An appropriate isotype control antibody was used as the control. The thymidine uptake was significantly inhibited by tHSCs (P \ 0.05 compared with no HSC controls). This inhibition was significantly reversed by addition of anti-B7-H1 mAb (P \ 0.05 compared with isotype controls). The isotype control IgG2a had no effect (P [ 0.05 compared with no antibody controls). c Blocking of B7-H1/ PD-1 ligation reduces tHSC-induced T cell apoptosis. Upper panels microscopy of cell suspensions that were stained with TUNEL (green) and anti-CD3 (red). Original magnification 9200. Lower panel: incidence of apoptotic T cells. Cells were stained with phycoerythrin-conjugated anti-CD3 and TUNEL and analyzed as described in ''Materials and Methods''. The incidence of TUNEL-positive T cells was markedly increased in the presence of tHSCs (P \ 0.05 compared with no HSCs controls). The enhancement was significantly reversed by the addition of anti-B7-H1 mAb (P \ 0.05, compared with isotype controls). The isotype control antibody had no effect (P [ 0.05, compared with no antibody controls). The data are representative of three separate experiments. Effector T cells (4 9 10 6 /ml) and tHSCs (1 9 10 5 /ml) in 400 ll medium containing 10% FCS were added into the lower chamber. Monoclonal antibody B7-H1 (15 lg/ml) was added to the lower chamber. Serum-free medium was added to the lower chamber of control wells. The cells were allowed to migrate for 24 h or invade Matrigel for 48 h at 37°C in a 5% CO 2 atmosphere. a Tumor cell migration was evaluated at 24 h. Columns mean of three experiments. tHSC versus serum-free media control, P \ 0.05. b Tumor cell invasion was evaluated at 48 h. Columns mean of three experiments. tHSC versus serum-free media (SFM) control, P \ 0.05; tHSC+anti-B7-H1 versus tHSC, P \ 0.05. Original magnification 9100. Aims: This study determined the clinical significance and effect of beta2-GPI on the hepatocellular carcinoma (HCC). Methods: Double fluorescent immunostaining analysis were performed in paraffin wax embedded histological sections of nine HCC cases, seven adjacent noncancerious tissues and seven hepatitis B controls using a beta2-GPI polyclonal antibody and a hepatitis B surface antibody. NF-jB activation was measured by immunofluorescence in hepatocellular carcinoma cells, SMMC-7721 exposed to the following different treatments. Group one: cells transfected with beta2-GPI; group two: cells transfected with HBsAg; group three: cells cotransfected with beta2-GPI and HBsAg; group four: cells transfected with control vector and group five: normal control cells. AFP expression of all groups was detected by ELISA. Results: The strongest merged fluorescent signal of beta2-GPI and HBsAg protein staining was seen in endochylema and localised to the border of nucleus in the cancer tissues. Weak beta2-GPI protein staining was present in the endochylema, with a strong signal of HBsAg protein in hepatitis B controls samples. In contrast, adjacent nontumorous liver tissue samples showed stronger beta2-GPI and HBsAg staining also in the endochylema. In vitro, Hepatol Int (2010) 4:94-345 317

Group one and Group two cells induced the activation of NF-jB. However, the highest activation was observed in the group three. The positivity rate of the number of NF-jB translocated from cytoplasm into nucleus each group compared with total cells was 13. 5, 8.7, 24.9, 5.7 and 0.95%, respectively. The AFP levels were significantly higher in the group three than in the group five (P \ 0.001). It appeared higher in the group three than group one (P \ 0.05) and group four(P \ 0.05), without a significant difference between group three and group two, group four and group five. Conclusions: Beta2-GPI was strong expressed in the HBV-infected HCC. It played a role in the development of HCC by activated NF-jB with beta2-GPI and HBsAg combination. Results: Except micro HCC (B1 cm) which differed significantly from all other four groups in most clinicopathological variables analyzed, no differences were noted between Group 2 and Group 3, or Group 4 and Group 5. However, if B3 cm was set up as the cut-off value for small HCC (SHCC), and [3 cm for large HCC (LHCC), then the significant differences appeared between these 2 groups in the proportion of well-differentiated (grade I-II) HCC (48.0 vs. 19.4%, P = 0.000), the incidence of capsular invasion (15.4 vs. 36.3%, P = 0.000), the positive rates of tumor thrombi (6.9 vs. 23.5%, P = 0.000), the frequency of satellite nodules (12.3 vs. 35.5%, P = 0.000), the overall median survival (119.6 ± 34.7 vs. 68.5 ± 6.6 months, P = 0.000), and the median recurrence-free survival (67.0 ± 16.7 vs. 29.5 ± 3.2 months, P = 0.000). Conclusion: These tumor-size based results are currently good evidence showing that HCC smaller than 3 cm in diameter is closely related with other prognosis-related factors and can reflect the relatively benign pathobiological features of HCC at early developmental stage. Since HCC growing beyond 3 cm may exhibit highly aggressive behavior indicative of progressing towards a more malignant stage, we suggest that B3 cm in diameter can be considered a suitable size criterion for SHCC. Background: Radio Frequency Ablation (RFA) is one of the standard therapy for liver tumor. Generally, the RFA region is confirmed by image comparison between pre-and post-treatment by contrast CT. However it is often difficult to confirm coagulation legion of RFA by CT. Therefore, we study the determination of coagulation legion used by ultrasound volume data as the alternative method, and we developed new ultrasound volume acquisition system. Methods: The system was consisted of ultrasound scanner (EUB-7500: Hitachi Medical Corporation, Japan), magnetic position sensor and PC with developed software and frame grabber. To measure the spatial accuracy of this system, we defined three target points in the phantom. One of them was about 2.5 cm in depth. The remained two targets were about 10 cm in depth and separated 10 cm each other horizontally. The ultrasound volume data have been taken seven times on the same day by one examiner, and analyzed. In addition, we evaluated the spatial accuracy in liver of healthy volunteer as preliminary experience. At first, the volume data have been taken at prone position and then turn out. After that, we took the data at prone position again. These data sets were compared.

Results: Regarding spatial accuracy in phantom, max standard deviation (STD) for XYZ direction is 1.22 mm at 2.5 cm depth and max STD is 2.09 mm at 10 cm depth. With respect to the evaluation of accuracy in healthy volunteer data, we confirmed that there is good correlation under controlling body position and breath depth. Conclusions: The accuracy in phantom case and volunteer case seems acceptable to check the safety margin after RFA. Our endpoint should be to develop and evaluate the system for confirming the effect of treatment by using ultrasound imaging in practical clinical site.

Usefulness of Intraoperative Fluorescence Imaging Using Indocyanine Green to Evaluate Local Anatomy in Hepatobiliary Surgery Jota Watanabe 1 , Fumiki Kushihata 1 , Tetsuya Mizumoto 1 , Yoshikuni Yonenaga 1 , Akifumi Miyoshi 1 , Taiji Tohyama 1 , Kazuo Honda 1 , Yasutsugu Takada 1 1 Division of Hepato-Biliary-Pancreatic Surgery and Transplantation, Department of Surgery, Ehime University, School of Medicine, Shitsukawa, Toon Ehime, 7910295, Japan

Background: One of the major complications encountered in hepatobiliary surgery is the incidence of bile duct and blood vessel injuries. It is sometimes difficult during surgery to evaluate the local anatomy corresponding to hepatic arteries and bile ducts. We evaluated the usefulness of intraoperative exploration of the biliary anatomy using fluorescence imaging with indocyanine green (ICG). Method: A patient with polycystic liver and four patients with chronic cholecystitis were underwent laparoscopic surgery. ICG (5.0 ml/body of 2.5 ml/ml) was administered intravenously 1hr before operation and the bile duct was observed using the infrared-based navigation system. ICG was added intravenously to observe the location or flow condition of the cystic artery. Result: Common bile duct and intrahepatic bile duct in polycystic liver were clearly visualized and was easily and accurately identified after being fluoresced, where as the localization of bile duct was not visible under naked eye as a green spot. The location of the cystic artery was revealed after division of the connective tissues. There were no adverse events related to the intraoperative procedure or the ICG itself. Conclusion: Biliary tract identification based on ICG fluorescent imaging is useful without adverse effects at hepatobiliary surgery. were observed. 1-, 2-and 3-year survival rates of the patients who received radiofrequency ablation for liver metastases were 77, 49 and 26%, while those of the historical control were only 50, 0 and 0%. Multivariate analysis showed that prior chemotherapy, tumor size ([50 mm), and tumor number ([5 nodules) were significant prognostic factors. When the patients were divided into three groups according to these risk factors, the survival rates depended upon the risk grading. Conclusions: RFA could safely improve the prognosis of patients with unresectable liver metastases from colorectal cancer. The earlier introduction of RFA, the better may be prognosis of the patients.

Intra-arterial 5-Fluorouracil and Interferon Combination Therapy of for Advanced Cholangiocellular Carcinoma Tenyu Motoyama 1 , Shinpei Sato 1 , Takahisa Sato 1 , Miho Kanda 1 , Shuntarou Obi 1 , Masao Omata 2 1 Kyoundou Hospital, 1-8 Kandasurugadai Chiyodaku Tokyo Japan, 2 Ymansashi Prefectural Central Hospital, 1-1-1 Fujimi Koufu City, Yamanashi, Japan abdominal ultrasonography, computed tomography or magnetic resonance imaging. Histopathologic diagnosis was made by liver biopsy or operation. Results: All patients were asymptomatic. Mean age was 37.8 years old. Ratio of sex (male: female) were 3:7. The associated risk factors were two cases of oral contraceptives usage. All cases were single lesion. Mean diameter was 37.84 mm. On imaging studies, exophytic lesions (2 cases), stellate scars (3 cases) and lobulating shapes (6 cases) were seen. On USG, 9 patients, hyperechoic (5 cases), isoechoic (2 cases) and hypoechoic (2 cases) lesions were seen. On CT, all cases had lesions with enhancing on early arterial phase, washing-out on portal or delayed phase. On MRI, 6 patients, low or iso-signal on T1 weighted images, high, iso or subtle low (1 patient) signal on T2 weighted images was seen. Four patients were performed MRI with hepatocyte-specific contrast agents (Multihance Ò or Primovist Ò ). High or iso-signal on delayed images were seen in three cases. Conclusions: Differential diagnosis is important in FNH from other liver tumors. MRI with hepatocyte-specific contrast agent may helpful for differential diagnosis but further study is needed.

Infected Hepatic Cyst Treated with Percutaneous Transhepatic Drainage Hiroshi Yoshida 1 , Yasuhiro Mamada 1 , Nobuhiko Taniai 1 , Sho Mineta 1 , Youichi Kawano 1 , Tomohiro Kanda 1 , Junji Ueda 1 , Tsutomu Nomura 1 , Eiji Uchida 1 1 Department of Surgery, Nippon Medical School, 1-1-5 Sendagi, Bunkyo-ku, Tokyo, Japan

Simple hepatic cysts are common benign lesions that are usually asymptomatic and require no treatment. However, complications can occur. This report describes a patient with an infected hepatic cyst treated with percutaneous transhepatic drainage. A 64-year-old woman presented at a nearby hospital because of acute right-upper-quadrant pain, mild left-lower-quadrant pain, diarrhea, and fever. She was admitted and received intravenous antibiotics for 1 week, but symptoms persisted. She was, therefore, referred to our hospital. On admission, ultrasonography demonstrated multiple hepatic cysts. One 13cm lesion was hypoechoic, unlike the other simple cysts, which were anechoic. Computed tomography showed that the density of the hypoechoic cyst was slightly higher than that of the other cysts. The wall of the cyst was thickened and showed contrast enhancement. On initial laboratory tests the serum Creactive protein concentration was 18.49 mg/dL, and the white-cell count was 13,300/lL. An infected hepatic cyst was suspected, and percutaneous transhepatic drainage of the cyst was performed. A catheter was inserted into the cyst, and dark red fluid was obtained. The right-upper-quadrant pain gradually resolved after drainage. An infected hepatic cyst was diagnosed, and system antibiotics were administered. However, the mild left-lower-quadrant pain persisted. No pathogens were isolated from the drainage fluid. Minocycline hydrochloride (200 mg) was injected, and the catheter was clamped for 30 min, once daily for 3 days. The serum C-reactive protein concentration was 1.78 mg/dL, and the white-cell count was 5700/lL. The left-lower-quadrant pain resolved, and colonoscopic examination revealed multiple diverticula of the sigmoid colon. Infection has not recurred, and the hepatic cyst has not become larger. Angiomyolipoma is a benign neoplasm that has been reported frequently in the kidney but rarely in the liver. The radiologic appearances vary because of different compositions of fat, blood vessels, and muscle. We present an unusual case of hepatic angiomyolipoma that was indeterminate on radiological imaging. Case description: A 48 year-old lady (HBV negative, serum alpha-fetoprotein normal) with a history of treated breast cancer was incidentally found of a hypoechoic 3-cm mass in left lobe of liver on ultrasonography. CT confirmed a solid mass without fat density with three-phase contrast-enhancement (upper row). 18 F-FDG-PET was negative and 99m Tc-labeled Red Blood Cell study also negative (not hemangioma). 99m Tc-sulphur colloid SPECT/CT was photopenic in this lesion (lack of Kupffer cell function was not in favor of focal nodular hyperplasia). 99m Tc-mebrofenin scintiscan was also photopenic in this lesion (non-functioning hepatocytes excluded hepatic adenoma). In view of the above negative findings, 11 C-acetate-PET was performed. Markedly increased metabolism (lower row) was found in the left lobe lesion with SUVmax of 8.51 (lesion-to-liver ratio of 2.75). Our past data suggested that only renal angiomyolipoma and \10% of hepatocellular carcinoma (HCC) had lesion-tobackground ratio [2. 75. In view of low pretest probability of HCC, this patient was diagnosed as an atypical ''angiomyolipoma without fat'' after 11 C-acetate-PET. Subsequently, ultrasound-guided core biopsy showed histopathological findings consistent with angiomyolipoma with minimal fat elements. 11 Cacetate is highly intense in non-fat components of angiomyolipoma, thus useful in helping the diagnosis when CT and other imaging modalities were indeterminate in liver masses of low-fat content. Objective: To report a case of primary carcinoid tumor of the liver in a 22-yearold Filipino female. Background: Primary carcinoid tumors of the liver are rare and usually asymptomatic. The presence of an extrahepatic primary lesion, usually in the gastrointestinal tract or pancreas, must be ruled out prior to making a diagnosis of a primary hepatic carcinoid tumor. In confirmed cases, treatment of choice is liver resection, regardless of the presence of metastasis. For non-resectable tumors or those with widespread liver involvement, liver transplantation may be considered. Transarterial chemoembolization may offer good palliation. Summary: A 22-year-old female presented with rapid weight loss and an abdominal mass of six months duration. No other symptoms were noted. Liver function tests and alpha-feto protein were normal. Initial abdominal computed tomography (CT) scan showed a large hepatic mass occupying both lobes of the liver, which was diagnosed as carcinoid tumor by liver biopsy. Work-ups to search for a primary lesion in the gastrointestinal tract and pancreas by endoscopic gastroduodenotomy (EGD), colonoscopy, and holoabdominal triphasic CT scan with triple contrast failed to reveal the presence of a primary tumor. Subcentimeter pulmonary nodules, believed to be metastatic, were detected on chest CT scan. Bone scan showed no evident bone metastasis. Due to the non-resectability of the tumor, transarterial chemoembolization was offered as alternative to treatment, however, the patient opted for palliative care and remained on close follow-up. Background: Since the introduction of lamivudine to treatments of chronic hepatitis B, the prevalence of lamivudine resistance in liver transplantation (LT) candidates continues to increase. In this study, we aimed to evaluate the effect of pre-LT lamivudine resistance (LAM-R) on the post-LT prognosis of recipient, which has not been well elucidated. Methods: The LT recipients who were tested for pre-LT genotypic resistance to lamivudine between September 1999 and August 2009 at a single tertiary care center, Seoul, Korea were consecutively included. We compared the overall survival, and the incidence of graft failure, recurrent hepatitis and hepatocellular carcinoma (HCC) between the patients with LAM-R (the resistance group) and those without LAM-R (the non-resistance group). Results: Eighty-three consecutive patients were included. All patients were treated with both intravenous hepatitis B immunoglobulin and antiviral agents (lamivudine, entecavir, or adefovir). Thirty-seven patients were assigned to the resistance group and 46 to the non-resistance group. Their median follow-up duration was 21 months (range, 1-121). Mortality occurred in two cases of the resistance group and three cases of the non-resistance group, respectively. The occurrence of graft failure, recurrent hepatitis and HCC were one case, two cases, and two cases, respectively, in the resistance group; and two cases, one case, and two cases, respectively, in the non-resistance group. Univariate and multivariate analysis using Cox-regression model showed that there was no significant difference in survival, graft failure, HCC occurrence and recurrent hepatitis between two groups. Conclusions: Our result indicates that pre-LT LAM-R does not significantly affect on the post-LT prognosis. Thus, considering LT in patients with underlying hepatitis B disease as a therapeutic modality, LAM-R may not be a barrier if it is treated with adequate antiviral agents. Background: Biliary strictures are the most serious complications, especially in non-heart-beating donor (NHBD) liver transplantation (LT). MRP2 (Multidrug resistance-related protein 2) is a bile salt transporter located in canalicular membrane of hepatocyte, and it is a major determinant of bile flow. The purpose of this study is to identify the value of immunohistochemical detection of MRP2 as a marker predicting persistent and serious biliary strictures after liver transplantations. Methods: Thirty-three recipients of DDLT (including 14 recipients of NHBD LT) who underwent liver biopsy within 2 months and followed for more than 2 years were included in this study. Among the 14 patients with biliary strictures, 7 patients showed intrahepatic strictures. Recipients were divided into two groups: those who required biliary intervention for more than 2 years (Group A, n = 8) and who did not (Group B, n = 25). We evaluated paraffinembedded liver graft tissues with immunohistochemical staining using monoclonal antibodies for MRP2, and MRP2 score was obtained by the product of the staining intensity (1+, 2+ and 3+) and the extent (the percentage of cells staining). Results: MRP2 score was significantly lower in Group A (P = 0.022). The AUC to predict persistent biliary stricture was estimated to be 0.768 (95% confidence interval, 0.61-0.93). MRP2 score value with the best combination of sensitivity and specificity was 185 (sensitivity 87.5%, specificity 60.0%). Patients with lower MRP2 score (\185) were more frequently involved in Group A (P = 0.039), and the odds ratio for persistent biliary stricture was 10.50 (95% confidential interval, 1.12-98.91). Furthermore, NHBD LT recipients were more frequently involved in the lower MRP2 score group (P = 0.049). Conclusion: Our results suggest that there is a potential utility of MRP2 as an early postoperative marker for prediction of persistent biliary stricture after liver transplantation, especially in NHBD LT.

Long-term Efficacy of Stent Placement for Treating Inferior Vena Cava Stenosis Following Liver Transplantation Gi-Young Ko 1 , Kyu-Bo Sung 1 , Dong-Il Gwon 1 1 Asan Medical Center, 388-1 Poongnap-2dong, Songpa-gu, Seoul, Korea

The aims of this study were to evaluate both the efficacy of stent placement for treating inferior vena cava stenosis and the patency of hepatic veins following inferior vena cava stent placement. Twelve hepatic transplant recipients underwent stent placement to treat inferior vena cava stenosis. The median interval between transplantation and stent placement was 32 days. Stents varied from 20 to 36 mm in diameter and were 60-120 mm long. We retrospectively analyzed the technical and clinical success, changes of hepatic venous flow, and the patency of the inferior vena cava stents. Stent placement was successful in all patients. Clinical success was achieved in all patients. Four patients underwent hepatic vein balloon angioplasty or stent placement through inferior vena cava stent meshes either immediately (n = 1) or 8-110 days after (n = 3) inferior vena cava stent placement. Nine of the 12 patients were healthy until this manuscript was completed, and the last follow-up CT obtained at a median of 65.3 months after inferior vena cava stent placement, revealed the patency of the stent-placed inferior vena cava and hepatic veins. Inferior vena cava stent placement seems to be an effective treatment with an excellent, longterm patency for treating post-transplant stenosis, although the possibility of hepatic venous outflow abnormalities following inferior vena cava stent placement should also be considered.

Injury Background: The relationship between Injury of graft (serum biochemical indicator) and expression of redox factor-1 and ICOS (Inducible co-stimulator), in early period (3-24 h) and late period (10-50 days) liver transplantation of rats model has not been published. Which indicator is more related to the injury of graft is what we studied. Methods: 210 adult male Wister rats were randomly divided into three groups: liver transplant group, sham surgery group and control group. Animals were sacrificed in each group at different time points: 3 h, 6 h, 9 h, 12 h, 24 h, 10 day, 20 day, 30 day, 40 day, 50 day after liver transplantation. The changes in serum biochemical indicators (AST, ALT) were measured by biochemical dosimeter, and the expression of Ref-1 and ICOS were detected by immunohistochemistry analysis. Results: As compared to sham surgery group and control group, the expression of Ref-1 protein in transplant group was higher during early period after liver transplantation and decreased in the late period. Meanwhile, Anti-ICOS was low in the early period and then rise in the late period (Fig.1) .The AST and ALT rise both in early period and late period. Conclusions: In the earlier period the serum indicators showed a strong relationship with the expression of Ref-1, and in the late period with the ICOS. We analyzed the morphological particularities of the CL venous drainage on 150 liver corrosion casts. The casts were made by injecting the hepatic vasculoductal systems with plastic, followed by parenchyma corrosion with hydrochloric acid. In 84% cases, left hepatic vein (LHV) and middle hepatic vein (MHV) form a common drainage trunk, and right hepatic vein (RHV) drains separately. In 8% cases we found an inferior RHV, and in 4% cases a middle RHV. The intraparenchymal portion of the portal hepatic vein (PHV) is the main element of liver's afferent pedicle, guiding the other afferent elements into the hepatic parenchyma. PHV is one of the intrahepatic vasculo-ductal elements with a most constant distribution. According to the intraparenchymal distribution of the RHV, segmentation of hepatic parenchyma was homologated. We present a hepatic corrosion cast with multiple anatomical variants of distribution; the variations are compared with those encountered on a total lot of 150 corrosion casts. PHV trunk forks into the left branch (LBr) and the posterior branch (PBr) (on the study lot this variation is present in 4% cases). LBr of the PHV gives birth to: anterior branch (ABr), that gives birth to four segmental branches (SV, SVa, SVIII and SVIIIa) (aspect encountered on 1.33% of the study lot), a single superior lateral branch (SII) (aspect present in 98% of the study lot), an inferior lateral branch with distribution in bouquet (SIII) (aspect present in 56.67% of the casts) and three medial branches (SIVa, SIVb and SIVc) (aspect present in 5.33% cases of the total lot). The right branch of the PHV is absent as a morphologic entity. It is replaced only by PBr (aspect encountered in 4% cases of the total lot). From the initial portion of the PBr emerges the paracaval branch (SVIa) (present in 11.33% of the total lot). PBr forks into a posterior superior trunk with two segmental branches (SVII and SVIIa) (aspect present in 14% cases of the total lot) and a posterior inferior segmental branch (SVI). The caudate branches emerge both from LBr and from PBr. We analyzed a hepatic corrosion cast with 15 segments and 5 major anatomical variants. Knowing these morphological variations is important for liver resection and transplant surgery. Introduction: Split-liver transplantation (SLT) and Auxiliary partial orthotopic liver transplantation (APOLT) provide ideal means to expand the donor pool. APOLT is indicated for potentially reversible fulminant hepatic failure and non-cirrhotic metabolic liver diseases. In the former scenario, the graft can be allowed to atrophy once the native liver regenerates. In the latter scenario, the graft can be allowed to atrophy, following application of successful gene therapy in future. The larger benefit in both situations is withdrawal of life-long immunosuppression. We hereby report first successful SLT and APOLT from India. Case Report: Cadaveric organ from a healthy 17-year-old gentleman (cause of brain death: road traffic accident) was split in-situ to benefit two recipients. First recipient was a 22-year-old lady suffering from Criglar Najjar syndrome type-I (CNS-I) and kernicterus. Her pre-transplant serum bilirubin was in the range of 20-25 mg%. After left lateral hepatectomy, segment 2 and 3 from donor were successfully transplanted orthotopically. Technically, care was given to divert requisite amount of portal inflow to the allograft, while at the same time preserving inflow to the native liver. Second recipient was a 55-year-old lady with end stage liver disease (cryptogenic cirrhosis, MELD = 17), in whom right lobe from donor was transplanted successfully after recipient hepatectomy. Both the recipients fared well during postoperative period and achieved good graft function. First recipient is able to maintain serum bilirubin in normal range, with normal bilirubin fractions.

Conclusions: For patients with non-cirrhotic metabolic liver diseases with severe extra-hepatic complications, APOLT is technically feasible, provides adequate hepatocyte mass and hence should be considered as an alternate to orthotopic liver transplant with recipient hepatectomy. SLT and APOLT should be encouraged in appropriate circumstances for judicious use of donor organ pool. Background: High demand for orthotopic liver transplantation has prompted some centers to consider using donor livers previously considered marginal, although no consensus yet exists about the specific factors that define a graft as marginal. Graft survival in HCV-infected recipients is worse than those transplanted for other liver diseases, thus they are more vulnerable to the added adverse effects of a marginal donor graft. We studied whether donor diabetes type II contributes to worse outcomes for HCV positive and HCV negative liver transplant recipients. Methods: We obtained data from the United Network for Organ Sharing (UNOS) on all adult liver transplants performed in the United States between January 1, 1998 and December 31, 2007. In total, 36,499 transplant events were evaluated. Independent predictors of graft survival were determined using Cox proportional hazards regression analysis after controlling for factors previously found to be associated with differences in transplant outcomes. Results: Donor diabetes was a strong independent risk factor for graft failure [ Background/ Aims: The MELD (Model for End-Stage Liver Disease) score is used widely for assigning priority to liver organs for patients with chronic liver diseases. However, majority of Hepatocellular Carcinoma (HCC) patients will not die from liver failure, but more likely from the cancer itself. The UNOS (United Network for Organ Sharing)/ OPTN (Organ Procurement and Transplantation Network) allocation policy allows for MELD adjustment based on the estimated 15% probability of death within 3 months. This formula assumes that for the patients listed with HCC, each has an average risk of 15% of dying within 3 months. However, the actual risk for HCC patients is very much dependent on the waiting list time, graft availability as well as the tumor biology and staging of HCC listed which are factors that can be very different in each centre. Thus a standard method to adjustment score may not be appropriate for all. We aim to propose a simplified model for MELD score assignment for HCC. Methods: Thirty-eight patients with non-malignant conditions who died without a transplant from October 2003 to October 2009 in our centre were studied. Their MELD scores were plotted against their survival and a linear relationship was derived. Based on extrapolation from the linear graph, the median survival post listing for patients with HCC was used to obtain a corresponding MELD score.

Result: The linear relation between survival among the non-HCC group and their MELD score was expressed as Mean [Log e (survival) | MELD score] = 2.893 -0.103 9 MELD score. The MELD score corresponding to the median survival time of HCC group at 2.86 months was 17.89. Conclusion: Our proposed simplified method for assigning MELD score for HCC patients allows consideration of the tumor characteristics of HCC in individual centres.

Graft Versus Host Disease Following Liver Transplantation Yang Won Nah 1 , Chang Woo Nam 1 , Sang Jun Park 1 , Dong Woo Kang 1 , Hong Rae Cho 1 1 Department of Surgery, Ulsan University Hospital, Ulsan, Korea

Background: Graft versus host disease (GvHD) was reported to occur in 0.1-1% following liver transplantation (LTX). Approximately, 80 cases have been reported in literature by 2008. GvHD predominantly occurs in the first few weeks after LTX. Fever, skin rash, pancytopenia and diarrhea constitute typical symptoms. The mortality rate of acute GvHD following LTX exceeds 75%. Treatment remains challenging. Case: A 58 years old female who suffered from acute-on-chronic liver failure associated with HBV infection underwent deceased donor LTX. Donor was 33years-old male with head trauma. Ischemic time was 493 min. Quadruple immunosuppression with IL-2R antibody, Steroid, MMF and FK-506 was introduced. She showed unremarkable postoperative recovery until 20 postoperative days, when massive diarrhea occurred. It was followed by intermittent fever (up to 38.5°C) and skin rash. Colonoscopic examination showed denudation and shallow ulceration in the terminal ileum. Pathologic examination showed increased crypt apoptosis and crypt loss, consistent with acute GvHD. The authors decided to reduce her immunosuppresion level. MMF was stopped. Oral prednisolone was also stopped by 30 days after LTX. Trough level of FK was reduced from 12 to 6.2 ng/ml by 25 days after LTX. Then FK was switched to cyclosporine and the level was maintained at around 50 ng/ml. Her diarrhea was uncontrollable with any medication. Colostrum showed some beneficial effect of only short duration (2-3 days). On 32 days after LTX, WBC count dropped precipitously from 4,710 to 940/mcl. Leukopenia did not show any response to granulocyte stimulating factor or WBC Hepatol Int (2010) 4:94-345 327

Results: Our data showed that TSA selectively induced CYP2E1 in four studied human HCC cell lines (Huh7, PLC/PRF/5, Hep3B, and HepG2), but not in normal primary human hepatocytes. TSA-mediated up-regulation of CYP2E1 expression was associated with histone H3 acetylation and the recruitment of HNF-1 and HNF-3b to the CYP2E1 promoter in HepG2 cells. siRNA-mediated knockdown experiment showed that TSA-induced caspase-3 cleavage was decreased due to reduced expression of CYP2E1 in HepG2 cells. Moreover, down-regulation of CYP2E1 was accompanied by decreased production of mitochondrial reactive oxygen species. Conclusion: These results suggest that histone modification is involved in TSA-induced CYP2E1 gene expression and that CYP2E1-dependent mitochondrial oxidative stress plays a role in TSA-induced apoptosis.

The Methods: Healthy volunteers were chosen as control group. According to infection status, CHB-infected patients were sorted into four experiment groups: normal hepatic function and HBV DNA high group, normal hepatic function and HBV DNA low group, abnormal hepatic function and HBV DNA high group and abnormal hepatic function and HBV DNA low group. The expressions of immune effector molecules in CD4+ T cells were detected by flow cytometer, and the correlations between these immune effector molecules and infection status were analyzed. Results: The expressions of TNF-a in normal/abnormal hepatic function and HBV DNA high groups were higher than those in control group (P \ 0.05);the expression of IFN-c in abnormal hepatic function and HBV DNA high group was lower than those in normal hepatic function and HBV DNA high/low groups and control group (P \ 0.05); The difference between the expressions of PF,GrB and GNLY in all groups were not significantly; there were positive correlations between every two of GrB, GNLY, and IFN-c, and there was positive correlation between PF and GrB. Conclusion: There were no correlations between the expressions of PF, GrB and GNLY on peripheral CD4+ T lymphocytes and infection status. However, there was positive correlation between the expression of TNF-a in CD4+ T lymphocytes and HBV DNA levels, and there was negative correlation between the expression of IFN-c in CD4+ T lymphocytes and hepatic function.

The Methods: 11 healthy volunteers were chosen as control group. According to infection status,96 patients infected with CHB were sorted into four experiment groups: normal hepatic function and high HBV DNA level group (28 patients), normal hepatic function and low HBV DNA level group (18patients), abnormal hepatic function and high HBV DNA level group (41 patients) and abnormal hepatic function and low HBV DNA level group (9 patients). The expressions of some immune effector molecules in CD8 + T cells were detected by flow cytometer, and then their correlations with infection status in patients with chronic hepatitis B were analyzed. Results: The expressions of GrB, TNF-a and IFN-c in normal hepatic function and low HBV DNA level group were higher than those in abnormal hepatic function and high HBV DNA level group (P \ 0.05); the expression of IFN-c in normal hepatic function and high HBV DNA level group was higher than that in abnormal hepatic function and high HBV DNA level group (P \ 0.05); the expressions of PF and GNLY in all groups were not significantly different; there were positive correlations between every two of GrB, PF, GNLY, TNF-a and IFN-c. Conclusion: There were negative correlation between the expressions of GrB, TNF-a and IFN-c in peripheral CD8 + T cells and hepatic disfunction/HBV DNA level in patients infected with CHB.

The Results: As compared with control group, the expression of GNLY was significantly increased in normal/abnormal hepatic function and HBV DNA low groups and normal hepatic function and high HBV DNA high group (P \ 0.05); the expression of IFN-c in normal/abnormal hepatic function and HBV DNA high groups was higher than that in normal control group (P \ 0.05); the expressions of PF, GrB, GNLY in abnormal hepatic function and HBV DNA high group were lower than those in normal hepatic function and HBV DNA high/low groups(P \ 0.05); there were positive correlations between every two of PF, GrB, GNLY, TNF-a, IFN-c except TNF-a and GNLY. Conclusion: The level of HBVDNA replication was related with the expression of IFN-c in NK cells, and the damage of liver was related with the expressions of PF, GrB, GNLY in NK cells in CHB-infected patients.

Screening for Active Compound of Danshen Background and aims: Hepatocyte apoptosis is a fundamental component of almost all acute and chronic liver diseases. Danshen (Radix Salviae Miltiorrhizae) is a traditional Chinese herb widely used to treat liver diseases in Asian culture. The aim of our study is to screen for the active compound of Dashen on hepatocyte apoptosis. Methods: Apoptosis was induced by Act D/TNFa in a hepatocyte cell line HL-7702. Six ingredients of Danshen were screened by high content screening apoptosis assay (fluorescence labeling nuclear and mitochondrial mass). The potential effective compound was validated in two kinds of mice models of acute liver injury (LPS/GalN and CCl4). TUNEL assay was used to determine hepatocyte apoptosis in liver tissue. Results: Compared with normal control cells, hepatocytes treated with Act D/TNFa undergone apoptosis markedly, reflected by decreasing nuclear size(nuclear fragmentation or condensation) and increasing abnormal mitochondrial mass. Of all the ingredients screened, Salvianolic acid A and B (Sal A, Sal B) showed an effective inhibition of apoptosisspecifically, they protected hepatocytes from nuclear fragmentation and inhibited abnormal accumulation of mitochondrial mass. Subsequently, we tested whether Sal B attenuated hepatocyte apoptosis in mice which induced by LPS/GalN and CCl 4 , respectively. TUNEL staining showed that rare hepatocyte apoptosis occurred in normal liver, massive hepatocyte apoptosis were observed in model mice, Sal B significantly inhibited hepatocyte apoptosis in both models. Conclusion: Sal A and Sal B can effectively inhibit hepatocyte apoptosis in vitro, while Sal B has also been confirmed to have a good effect against hepatocyte apoptosis in vivo. Background: TGFb1, as a multifunctional cytokine, has important effects in the occurrence and development of liver neoplasm and hepatic fibrosis. At least 10 SNPs exist in TGFb1 gene, Leu10Pro is located in TGFb1 signal peptide whose base substitution will influence TGFb1 secretion theoretically. This study was designed to determine whether Leu10Pro influence TGFb1 secretion and function of hepatic cells. Method: Two TGFb1 plasmids CMV-Leu and CMV-Pro were constructed by cloning TGFb1 DNA fragment, which encoding either the Leu or Pro forms of TGFb1, into pcDNA3.1 vectors. L02 and HepG2 cells were chosen to be transfected with CMV-Leu, CMV-Pro or pcDNA3.1 empty vectors. The cell medium was collected and analysed by TGFb1 ELISA kit 24 h after transfection. The transfected cells were stained by Annexin V and PI, then analyzed by flow cytometry. All data was calculated by Student's t test. Background: Natural supplements and alternative medicine have attracted significant attention in recent years. Previously we reported that the standardized freeze-dried powder made from fresh grapes, promotes liver cell viability and proliferation, prevents H 2 O 2 -induced oxidative cell damage, ameliorates mitochondria-and death receptor-mediated apoptosis. Methods: To explore the mechanism underlying the cytoprotective effects of grape and identify the active polyphenolic components involved, we examined the signaling pathways and the active compounds in grape, such as epicatechin, cyanidin, quercetin, and resveratrol in Huh7 cells.

Result: The Akt/PKB signaling pathway functions as a critical regulator of cell survival and proliferation. Our findings demonstrated that treatment with grape increased the phosphorylation of both Akt and IjB. Silencing Akt expression with Akt-specific siRNA or cell pretreatment with a pharmacologic inhibitor of Akt signaling pathway (LY) almost abolished the grape-induced Akt phosphorylation. Additionally, we found that treating cells with grape induced expression of PCNA which is associated with cell cycle progression. Assessment of individual active components revealed that at concentrations corresponding to 300 lg/ml grape powder, only quercetin demonstrated slight cytoprotective effects against mitochondrial-mediated apoptosis. Increased concentrations of other individual polyphenolic compounds were required to produce measurable cytoprotective effect. Only combinations of all four polyphenolic compounds restored a degree of the anti-apoptotic effects seen with 300 lg/ml grape powder Conclusion: In addition to the anti-apoptotic and anti-oxidant effects which was reported previously, grape enhances liver cell growth and proliferation, which are associated with the Akt/ NF-jB cell survival pathway. We speculate that quercetin, epicatechin, and cyanidin are the active compounds within grape that are involved in attenuation of mitochondria-induced apoptosis. These findings may contribute to the understanding of the molecular mechanisms involved in protective effects of grape in a variety of liver conditions associated with oxidative cell injury and apoptosis, and may assist in developing clinical protocols to treat the stress-mediated diseases predominant state of liver. Bloodstream plays an important role in lymphocytes homing and recruitment. Comparing with other tissues which receive only arterial blood, liver has another special blood supply from portal vein which constitutes 75% of the total hepatic blood influx. The portal venous blood returns from the intestine and connects the liver and intestine anatomically and immunologically. Published studies have shown aberrant homing of mucosal T cells to liver under certain inflammatory conditions and inhibition of hepatic dendritic cells by gut-derived bacterial products. Our study aims at unveiling the contribution of portal venous blood flow to the high frequency of NK cells in liver under normal physiology condition. Methods: By flow cytometry, we detected percentage of NK cells in peripheral blood, portal venous blood, postcaval venous blood, and blood in sinusoids. Results: We found that NK cells percentage is much higher in liver sinusoidal blood than in other blood. However, we did not found any difference in NK cell percentage among lymphocytes from peripheral blood, portal venous blood, postcaval venous blood. In addition, the rate of NK cells in liver was not affected after tail intravenous injection of portal venous blood plasma. Nevertheless, we observed the existence of CD4 + CD3 -CD127 + lymphoid tissue inducer (LTi)like cells in liver. Moreover, an immature subset of NK cells present in liver was absent in both portal venous blood and thoracic arterial blood. Background/Aims: Natural killer (NK) cells as important populations in innate immune system are the first line of defense against infections. Liver is a particular anatomical and immunological organ, which is enriched in innate immune cells including NK and natural killer T cells. Previous studies indicated that mouse liver NK cells are a heterogeneous population with different functional subsets. However, there was no established phenotypic marker that can distinguish this NK subset. Method: We compared the levels of extensive surface molecules on liver NK cells with other lymphoid organ-derived NK cells by flow cytometry, such as bone marrow, spleen, and lymph nodes etc. Results: By means of phenotypic comparison, we found the liver contain a high frequency of phenotypically immature NK cells, which is much lower in other organs except mesenteric lymph nodes (MLNs). The expression levels of CD51, CD61, CD69 and TNF-related apoptosis-inducing ligand (TRAIL) on liver and MLNs are markedly higher than other organs. While reduced expression of CD62L, CD49d and KLRG1 was observed on liver and MLN NK cells compared to NK cells from other organs or tissues. Despite the fact that there are many similarities in phenotype between liver NK cells and MLN NK cells, differences also existed. Liver NK cells expressed higher levels of TRAIL and CD11c than MLN NK cells. Conclusion: These results suggested liver NK cells might have a similar function as MLN NK cells, and the slight differences in phenotype between liver and MLN NK cells may be attributed to the distinct microenvironment of the two organs. Functional analysis and comparison between liver NK cells and MLN NK cells should be further investigated.

The Trafficking of Hepatic NK Cells to Liver Draining Lymph Nodes Mei-Juan Zheng 1 , Rui Sun 1 , Zhi-Gang Tian 1 1 Institute of Immunology, School of Life Sciences, University of Science and Technology of China, 443 Huangshan Road, Hefei, Anhui 230027, China

Background: It is reported that the lymph node contains a small population of natural killer (NK) cells, suggesting an important function of NK cells in adaptive immunity. The liver with overwhelming innate immune cells including NK cells produces a large volume of lymph, representing about 25-50% of flowing through the thoracic duct. It is suggested that 80% or more of hepatic lymph drains into the portal lymphatic vessels. The purpose of the study is to examine the trafficking of hepatic NK cells to liver draining lymph nodes. Methods: Six-week-old male C57BL/6 mice received adenovirus containing enhanced green fluorescent protein (Ad-EGFP,8 9 10 9 pfu virus) injection in a volume of 100 lL (intrahepatic injection) or 2.0 mL(hydrodynamics-based injection through tail vein). The surface phenotype of NK cells isolated from liver, spleen, bone marrow, blood and lymph node were analyzed by Flow cytometry. Results: In this study, we found that the liver NK cells had an immature phenotype CD3 -NK1.1 + DX5 -TRAIL + CD11c + CD11b -CD51 + CD2 low in adult C56BL/6 mice, but rarely in the spleen, bone marrow and blood. By the hydrodynamics-based intravenous injection or intrahepatic injection Ad-EGFP, we found that the hepatic lymphocytes population with GFP fluorescence trafficked into the liver draining lymph nodes after various time points. In naïve C57BL/6 mice, NK cells in the liver draining lymph nodes, mesenteric lymph node, cervical lymph node, inguinal lymph node and popliteal lymph node were phenotypically analyzed,and a higher proportion of CD3 -NK1.1 + DX5cells were found in the liver draining lymph nodes and mesenteric lymph node than cervical lymph node, inguinal lymph node and popliteal lymph node, suggesting that mesenteric lymph node is possibly the secondary draining lymph node of the liver. Conclusion: The presence of immature NK cells in liver and its draining lymph nodes might suggest a possible role of hepatic NK cells in formatting adaptive immunity against hepatic pathogen or tumors.

Identification and Characterization of Immature NK Cells in Bone Marrow and Liver Xiao-Jun Jiang 1 , Rui Sun 1 , Zhi-Gang Tian 1 1 Institute of Immunology, School of Life Sciences, University of Science and Technology of China, 443 Huangshan Road, Hefei, Anhui, China

Background: Natural killer cells play important roles in immune host defenses as a member of the innate immune system. NK cells widely distribute in lymphoid organs and non-lymphoid organs in both human and mice. Whether NK cells present in different organs are different NK subsets becomes an important scientific question. Methods: We analyzed surface phenotype of NK cells isolated from blood, spleen, liver, lung, bone marrow and lymph node by Flow cytometric analysis. Results: It seems that NK cells in liver and bone marrow are more immature than those in blood, spleen, lung and lymph node, because of the lower expression of CD11b and Ly49 family receptors. We also found adhesion molecules and chemokines expressed on NK cells may change in different tissues. Moreover, heterogeneity of NK cells exists in the same tissue. On the basis of surface expression CD27 and CD11b, NK cells can be categorized into four stages which represent different growth stages of NK cell. Each stage can be found in all tissues we examined, though they have different proportions. Conclusion: The presence of NK cells at different developmental stages in various organs may provide a new hypothesis for NK cells development. However, we still need more straightforward evidence to prove that these heterogeneous NK cells are in different developing stage so that to exclude the possibility that they are different functional subgroups.

Glycochenodeoxycholate-induced Hepatocyte Apoptosis: Signaling Pathway Activated by Survivin and Nuclear Factor-kappaB Kewei Wang 1 , John Brems 2 , Ai-Xuan Holterman 1 Aims: To verify if retroviral vector (pSuper-retro-NEO/GFP, pSR) expressed hepatitis B virus X gene (HBx) specific shRNA (small hairpin RNA) could suppress viral replication in longer period than nonviral vector (pSuper-basic, pS) by hydrodynamic injected into murine tail vein directly without be packaged into viral particles. Methods: Several HBV X gene specific RNA interference target sequences were designed and the corresponding shRNA expression retroviral vectors were constructed. These vectors were cotransfected into Huh7 cell line with pAAV-HBV1.3 HBV replication competent clone and the best RNAi target expression vector was selected. The excellent shRNA sequence expression nonviral vector was also constructed and compared its function with retroviral counterparts. Both shRNA expression vectors were injected into C57 BL/6 mice with same quantity pAAV-HBV1.3 plasmid and detected the HBV serological marker and viral titer more than 16 weeks post injection. Results: There was no significant difference in suppress HBV replication between retroviral and nonviral HBx specific shRNA expression vectors. Sera HBsAg positive percentage, HBsAg quantity, serum HBV DNA concentration and HBcAg positive hepatocytes number can lowered significantly at least 4 weeks after retroviral vector injected mice versus about almost 1 week in nonviral vector mice. Conclusions: The retroviral shRNA expression vector can prolong the time of suppression HBV replication effectively by hydrodynamic tail vein injection without retroviral package process. It gives some light on the study method about specific gene's function in a longer period in a simple way. WB-F344 cells stimulated by CTGF, while TIPM-1 expression increased when stimulated by TGF-b. Conclusions: Although both transforming growth factor-b and connective tissue growth factor could inhibit the proliferation of rat hepatic progenitor cells, the two cytokines play different roles on the differentiation and TIMP-1 expression on hepatic progenitor cells.

The Phenotypes of Mouse Nature Killer Cells in Liver and Spleen of Different Ages Xian Wu 1 , Hai-Ming Wei 1 , Rui Sun 1 , Zhi-Gang Tian 1 1 Institute of Immunology, School of Life Sciences, University of Science and Technology of China, 443 Huangshan Road, Hefei, Anhui, China

Background/Aims: Nature killer cells, used to be called large granular lymphocytes, play an important role in innate immunity. They can exhibit cytolytic activity against a variety of targets cells which are harmful for health. Nature killer cells are heterogeneous populations which express many kinds of surface antigens, by which the NK cells development may be described. Methods: We test the phenotype of liver and spleen NK cells in mice of different ages by Flow cytometric analysis. Results: We have analyzed many kinds of surface antigens and found that liver and spleen nature killer cells express high level NKG2A, which is an inhibitory receptor, and TNF-related apoptosis-inducing ligand (TRAIL) in fetal and new born mice, but none ly49 family receptors were found. In adult mouse, liver nature killer cells and spleen nature killer cells express higher level Ly49 family receptors than fetal mouse and new born mouse, suggesting NKG2A hi TRAIL + may represent an immature phenotype, and Ly49+ may represent an much more matured phenotype. Liver nature killer cell express much low Ly49 family receptors than spleen nature killer cells, and only liver nature killer cells can express TRAIL without any stimulation. Conclusion: These results indicate that liver may supply a unique microenvironment for nature killer cells development and education. Background: Based on the sequence variation in the N-terminus of the UL55 gene, which encodes glycoprotein B (gB), human cytomegalovirus (CMV) can be classified into four gBn genotypes. Genotyping and quantity of HCMV gBn by real-time PCR was established in our previous study. Methods: This study examined the distribution of CMV gBn genotypes, and the correlation between CMV gBn DNA (detected by real-time PCR) and CMVpositive pp65 cells (identified by immunohistochemical staining) in a cohort of hematopoietic stem cell transplant (HSCT) patients. Results: The distribution of gB genotypes was as follows: gBn1, 60% of patients; gBn2, 13.3%; mixed gBn1 and gBn3 infection, 36.7%; and gBn4 and other mixed infections, 0%. The detected level of CMV gB DNA correlated well with the number of CMV-positive pp65 cells (r = 0.514). Conclusions: The gBn1 is the most popular genotype in CMV. In the future, the relationship between gBn genotype and clinical outcome would be studied.

Hepatocyte-like Cell Clusters Derived from Human Menstrual Blood-derived Stem Cells Azita Hekmatdoost 1 1 Shahid Beheshti University of Medical Sciences, Evin, Iran

Introduction: There are only a few reports that describe the hepatocytic differentiation potential of human menstrual blood-derived mesenchymal stem cells and no reports that describe the in vivo functions of hepatocyte-like cells differentiated from somatic stem cells including mense.

Methods: In this study, we established a new method for generation of functional hepatocyte-like cell clusters, which functioned effectively in vitro. Propagation and differentiation potential of stem cells into hepatocyte-like cells were investigated with morphological, cytochemical and molecular experiments.

Results: The generated hepatocyte-like cell clusters expressed various genes normally found on mature hepatocytes. The cell clusters exhibited functional characteristics of hepatocytes too. Conclusion: we established a new protocol for efficient induction of menstrual blood derived stem cells into functional hepatocyte-like cell clusters.

Poster Presentation: Miscellaneous Aim: To investigate the clinical significance of using C-Reactive Protein (CRP) as an end point in the antibiotics treatment for pyogenic liver abscess Methods: Retrospective study for the end points of 21 cases of pyogenic liver abscess with antibiotics treatment. 12 cases had normal CRP values as their end point of the treatment, and the other 9 cases had liver abscess size less than 4 cm and normal body temperature for 2 weeks as the end point of the treatment. The time of antibiotics treatment and recurrence rate were analyzed. Results: For the two groups of patients, the time of antibiotics treatment was 21.17 ± 6.83 days and 30.44 ± 3.71 days, respectively (P = 0.002). For the group with normal CRP value as end point, the time of antibiotics treatment was significantly shorter than the control group. The recurrence rate for both groups was 0. Two cases in the control group were lost tract during follow-up in this study. Conclusion: For pyogenic liver abscess treatment, other than percutaneous drainage, the value of CRP is an independent factor to determine the time of antibiotics treatment.

Tubercular Liver Abscesses Anil K. Sarda 1 , Anju Garg 1 , Rakesh Mittal 1 , Baljeet K. Basra 1 , Nikhil Talwar 1 , Mayank Jain 1 1 MAM College, New Delhi, 110002, India

Background: Primary tuberculosis of liver without involvement of other organs is uncommon with diagnosis usually made on postmortem examination. Our objective was to identify clinical parameters for diagnosis of tubercular liver abscess in patients initially diagnosed as amoebic liver abscess (ALA). Methods: Between November, 2004 and October, 2008, 213 cases with ALA were admitted. All patients were started on antiamoebic therapy. Patients with liver abscesses with initial size of more than 10 cm, evidence of pre-rupture, left lobe abscesses or no response to antiamoebic therapy after 48 h, underwent drainage procedures. In patients who had evidence of pulmonary TB or where drainage persisted without ultrasonic decrease in size of the abscess, ELISA test for TB, drain fluid for PCR for TB and smears for AFB were sent. Patients with positive PCR were started on antitubercular treatment (ATT). The response was documented by serial ultrasounds. Results: Out of 213 patients, 13 were included in the study on basis of persistence of drainage without radiological evidence of decrease in the size of abscess (n = 11),or recurrence after initial improvement (n = 2). Pus culture was sterile and smears for AFB were negative in all. ELISA was positive in four patients. There was associated pulmonary TB in four patients. In 12 patients, PCR was positive. In one patient with persistence of drainage, diagnosis of tubercular liver abscess was made on the basis of radiological evidence of pulmonary TB even though PCR was negative. After starting ATT, drainage catheter was removed after 2 weeks in 10 patients, whereas 3 others required the drain till 3 weeks. Subsequent follow-up at 3 months showed complete radiological resolution of the liver abscess. Conclusion: TB should be considered in patients of liver abscesses not responsive to antiamoebic therapy, with recurrence of the abscess or where the aspirate is sterile even in the absence of biochemical/bacteriological evidence of TB or associated pulmonary TB.

Histocompatibility Antigens in Relation to Hepatic Schistosomiasis Soad Zaki 1 , Aly Soliman 1 , Rafik Zaher 1 1 Faculty of Medican Alexandria Universty, Egypt

Hepatic Schistosomiasis is one of the most prevalent chronic liver disease in Egypt. Great variation in disease severity occurs among infested individuals due to the wide range of intensity and duration of tissue egg deposition. Aim: to determine the association between hepatic Schistosomiasis and antigens of the HLA system. ) in patients (48.08%) than in controls (14%) In group I HLA-B5 significantly increased in patients (60%) as compared to controls. In group II HLA -B5 significantly higher in patients (45.46%) than controls (14%) also HLA-AW19 significantly higher (40.91%) in patients than controls (12.67%). In group III HLA-AW19 significantly increased in patients (46.67%) compared to controls.No significant association between HLA antigens and cases with HBV or HCV infection. Conclusion: The significantly high association of HLA-AW19 and HLA-B5 in patients with hepatic Schistosomiasis as compared to normal controlstogether with the lack of any association with active intestinal Schisto. Antigens predispose to liver affection. Individuals possessing HLA-AW19 appear to be more prone to severe form of liver disease

A Pilot Study of the Effect of Desferrioxamine as Supplement to the Antibiotic Therapy in the Treatment of SBP Manal El-Hamamsy 1 , Nehal Abou-Seada 1 , Mohamed Reda Elwakil 1 1 Ain Shams University, Abbasia, Cairo, Egypt, Ain Shams University, Faculty of Pharmacy, Cairo, Egypt

Background: CEFOTAXIME has been used in the treatment of Spontaneous bacterial peritonitis (SBP). Desferrioxamine (Dfx) is an iron chelating agent and prevents free oxygen radical formation. Objective: to assess the efficacy of Desferrioxamine as supplement to the antibiotic therapy in the treatment of SBP. Methods: Thirty patients divided into two groups, Group I: 15 having SBP receiving Cefotaxime (1 g IV every 12 h) alone and Group II: 15 having SBP receiving Cefotaxime (1 g IV every 12 h) with desferrioxamine (500 mg IM twice daily). All patient were monitored for 5 and 7 days. Results: The examination of all Patients after 5 days of treatment revealed that in Group I only 4 Patients (26.67%) were relived from all signs and symptoms of SBP, whereas 11 Patients (73.33%) showed presence of several signs and symptoms of SBP. In Group II all 15 patients (100%) were relived from all signs and symptoms of SBP upon examination after 5 days of treatment which showed to be a significant difference at (P.05). PMN was highly elevated in both groups before treatment. Background: Hepatitis-B-associated glomerulonephritis is a distinct entity occurring frequently in hepatitis-B-prevalent areas of the world including Sudan (prevalence of Hepatitis B is 8%). The general objective was to determine the prevalence, sociodemographic, socioeconomic and risk factors of HBV infection among patients with renal problems in hospitals of Khartoum state from September to November 2007. Methods: A hospital-based, analytic, cross-sectional design study among patients of renal problems in renal departments of selected Khartoum State hospitals: Khartoum teaching hospital, Soba university hospital, Ja'afr Ibn Aouf pediatric hospital, Ibn Sena'a specialized hospital and Dr. Salma renal centre between September and November 2007 and they were 44 patients selected as comprehensive non-probability sample. Data collection by: Structured questionnaire, interview and blood sample applying third generation ELISA for HBsAg. Results: Nine patients were HBsAg carriers and the prevalence of HBV nephropathy is 20.5% which is much higher than the prevalence in Sudan (8%) and this support the theory of epidemiological, clinical and immunological evidence which suggest a causal association between HBV carriage and the development of nephropathy. More than 66% of them are below 18 years old. Sociodemographically, there is a significant relation between developing the disease and the place of origin and West Sudan having the highest percentage (44%). Developing HBV nephropathy has a relation with low socioeconomic status. There is a strong association with HBV vaccination; more than 77% of the cases were not vaccinated. There is no association with: parental medication, blood transfusion, cupping or tattooing, past dental history or past-medical history of jaundice. Conclusion: HBV nephropathy prevalence in Sudan is 20.5%. More than 66% of them are below 18 and 44% from West Sudan. It has a relation with low socioeconomic status. Not having HBV vaccination is strongly associated with developing the disease.

Results: LPS caused severe inflammation in liver including inflammatory cell accumulation, parenchymal cells edema and tissue exudation, which were all ameliorated in P+L Group. Mortality decreased significantly with PPC addition (6.67 vs. 46.67%, P = 0.035). Liver relative weight, enzymes and ICAM-1 expression also bettered. In BMDM, PPC at 0.1 mM totally abolished LPS induced TNF-a, IL-6 and IL12 production and decreased IL-10. mRNA levels (6hrs) supported the tendency except IFNc. Phosphorylation of Erk, IKKa/b, IjBa, was blocked, so did the induction of COX-2 and iNOS. Interestingly, NOx production diminished whenever PPC was added before, simultaneously with, or after LPS. Conclusion: PPC protects liver from endotoxin-induced inflammatory injury, maintain liver function and increase survival. This might be due to inhibition of proinfalmmatory signal transduction and cytokine, enzyme, and radical generation of inflammatory cells.

Early Management of Endotoxemia Using the Endotoxin Activity Assay and Polymyxin-B Based Hemoperfusion Gilnardo Novelli 1 , Vincenzo Morabito 1 , Francesco Pugliese 1 , Massimo Rossi 1 , Franco Ruberto 1 , Giancarlo Ferretti 1 , Pasquale Berloco 1 1 La Sapienza University of Rome, Viale del policlinico 155, Background: We evaluated the ability of the Endotoxin Activity (EA) assay to determine the need for early intervention of endotoxemia using polymyxin-B based hemoperfusion (PMX-DHP) on septic patients. Methods: Twenty-four patients were enrolled. Eleven patients had a high EA level (C0.6) and were treated with PMX-DHP every 24 h until the EA level was low (.4). The remaining thirteen had EA levels .60 and received standard therapy only. Results: Two PMX-DHP treatments were performed on four patients, three treatments on six patients and four on one. After the therapy, MAP increased (69.45-84.09 mmHg; P \ 0.01), HR decreased (111.73-77.91 beats/min; P \ 0.01), WBC decreased (18,380 to 9,550 cells/mm 3 ; P \ 0.01), PMN percentage decreased (88.45 to 67.82%; P \ 0.01) and PaO 2 /FiO 2 increased (275-308.09; P \ 0.01). All 24 patients survived to the 28-day follow-up. Conclusion: The EA assay can identify patients eligible for PMX-DHP treatment and aids its therapeutic dosing.

Co-axial Liver Biopsy is the Solution for Hepatologists to Obtain Adequate Tissue for Molecular Studies Chi-Tan Hu 1 1 Research Centre for Hepatology, Buddhist Tzu Chi Hospital, No 707, Section 3, Chung Yan Road, Hualien, Taiwan Background: Pathological diagnosis by liver biopsy remains the gold standard for many hepatic diseases. Tru-cut biopsy (TCB) is common while co-axial biospy (CAB) is not recognised by many hepatologits who need adequate fragments of hepatic tissue for molecular studies such as RT-PCR and immunostaining. Aims: To compare the success rate of complete biopsy, bleeding rate, and pain score in patients who underwent either TCB or CAB. Patients and methods: 276 chronic hepatitis C patients were randomised to undergo tru-cut or co-axial liver biopsies for hepatic fibrosis between 2006 and 2009 at Buddhist Tzu Chi Hospital, Hualien, Taiwan. Gelfoam dissolved in 1 mL of normal saline was used to inject into the biopsy tract for hemostasis only for patients in the CAB group. Exclusion criteria included patients with platelet count\50 9 109/L, of Child Class B and C liver function, or with any malignancies. Success rate of complete biopsy (SRCB) was defined as having three fragments of hepatic tissue, one for pathologic fibrosis score (FS Grades 0-4), one for RT-PCR, and one for NS3 protein direct immunostaining (IS) without bleeding complication. Bleeding was detected by sonography within 24 h after either biopsy procedure with or without hemodynamic change. A 10-point pain score was used for patients to answer immediately and 24 h after the procedure. Results: 140 patients received TCB and 136 patients underwent CAB. There was no difference of age and gender between both groups. Comparing the TCB and CAB groups, the SRCB were 87.1 vs. 97.1%, P \ 0.0001, the bleeding rates were 4.3 vs. 0% (P \ 0.001), pain scores were 3.4 ± 1.1 vs. 3.6 ± 1.4 (P [ 0.05) immediately and 2.1 ± 0.6 vs. 2.2 ± 0.7 (P [ 0.05), 24 h after biopsy. Conclusion: Co-axial biopsy caused no bleeding. It was more successful to obtain adequate tissues for both pathological and molecular studies and did not cause more pain than its conventional counterpart. This study implicates that the special design of CAB (i.e gelfoam injection) is the solution for molecular hepatologists to obtain adequate tissues for pathological-molecular studies.

Placement of a Percutaneous Transhepatic Biliary Stent Using a Silicon Drain with Channels along the Sides for the Treatment of a Stenotic Hepaticojejunostomy Hiroshi Yoshida 1 , Yasuhiro Mamada 1 , Nobuhiko Taniai 1 , Sho Mineta 1 , Youichi Kawano 1 , Tomohiro Kanda 1 , Junpei Sasaki 1 , Junji Ueda 1 , Tsutomu Nomura 1 , Eiji Uchida 1 1 Department of Surgery, Nippon Medical School, 1-1-5 Sendagi, Bunkyo-ku, Tokyo, Japan

Background: This report describes a method for percutaneous transhepatic biliary stenting with a BLAKE Silicon Drain and discusses the usefulness of placement of the drain connected to a J-VAC Suction Reservoir for the treatment of a stenotic hepaticojejunostomy. Methods: Percutaneous transhepatic biliary drainage was performed under ultrasonographic guidance in a patient with a stenotic hepaticojejunostomy after hepatectomy for hepatic hilum malignancy. The technique used was as follows. After dilatation of the drainage root, an 11-Fr tube (11Fr PTCS tube, Sumitomo Bakelite, Tokyo, Japan) with several side holes is passed through the stenosis of the hepaticojejunostomy. A 10-Fr BLAKE Silicon Drain is flexible, precluding one-step insertion. One week after insertion of the 11-Fr tube, a 0.035-inch guidewire is inserted into the tube. After removal of the 11-Fr tube, the guidewire is put into the channel of a 10-Fr BLAKE Silicon Drain. The drain is inserted into the jejunal limb through the intrahepatic bile duct and is connected to a J-VAC Suction Reservoir. Low-pressure continued suction is applied. Patients can be discharged after insertion of the 10-Fr BLAKE Silicon Drain connected to the J-VAC Suction Reservoir. Conclusions: Placement of a percutaneous transhepatic biliary stent using a 10-Fr BLAKE Silicon Drain connected to a J-VAC Suction Reservoir is useful for the treatment of a stenotic hepaticojejunostomy.

Evaluation the Concentration of TNF Alpha in Diagnosis and Prognosis of Acute Pancreatitis Hoang Trong Thang 1,2 1 Hue Medicine and Pharmacology University, 8 Ngo Quyen Street, Hue City, Vietnam, 2 Hue central Hopital, 16 Le Loi Street, Hue City, Vietnam Acute pancreatitis (AP) is common and severe diseases. In Vietnam the mortality of AP about 10%. In the past we based on the Ranson, Glasgow, Imrie, SAPS criteria in the prognosis for grave AP. Aims: Using the TNF a as a factor in the diagnosis and prognosis of AP, especially in the first 24 h of the diseases. Subjects and method: 51 AP. Patients enrolled in the study, 30 temoins. The diagnosis of AP: Based on the clinical features + Amylasemia [ 3 Normal + Imagery of ultrasound or CT Scanner of pancreas. Exclusive criteria: Increasing of amylase extra pancreas, the increasing of TNFa.dues to the others causes ex: infection, immunology, corticotherapy… Results: The concentration of TNFa.: The temoins: TNFa.. The AP: 24.39 ± 13,43 pg/l [10-72,1]. In mild AP: 18.06 ± 6.43 pg/l [10-35.8], in severe AP: 39.59 ± 13.91 pg/l [16. 5-72.1] The relationships and cut-off point of TNF: TNFa has middle positive relation with neutophile (r = 0,356; p \ 0.05); with amylasemia: (r = 0.417; p\ 0.05), with glycemia: (r = 0.417; P \ 0.05), and with LDH: (r = 0.417; P \ 0.05). TNFa has middle negative relationship with: PaO 2 (r = -0.558, P \ 0.01), weak negative relation with calcemia (r = -0.265, P [ 0.05), and albuminemia (r = -0.428, P \ 0.01). TNFa has close positive relation with Imrie criteria. Cut -off point of TNFa and LDH in acute pancreatitis: TNFa: 28.1 pg/ l, in AP corresponding to the sensitivity. 85.7% and the specificity 94.6%, instead of the cut off point of LDH: 309U/L and the sensitivity and the specificity only 92.9 and 70.3%.

The Catholic University of Korea, South Korea, 2 Sosa-dong

18 Korea Institute of Radiological and Medical Sciences

Daejeon PP254 Rescue Efficacy of Lamivudine and Adefovir Compared to Entecavir Monotherapy in Patients with Lamivudine-Resistant Chronic Hepatitis B Han Jak Ryu 1

PP605 Foxp3 Promoter Demethylation Controls Foxp3 Gene Expression in Human Hepatoma Cell Lines Hao Luo

Guangxi province, 541000, China PP701 Aberrant miR-9-1 Hypermethylation Plays a Role in HCC Yong-Feng Wang 1 , Xiang-Mei Chen 1 , Qing Xie 1 , Hui Zhuang 1 , Feng-Min Lu 1 1 Peking University Health Science Center

Beijing Background: The programmed death-1/programmed death-1 ligand (PD-1/ PD-L1) pathway plays a crucial role in tumor evasion from host immunity. The aim of this study was to evaluate the clinical significance of circulating PD-1 and PD-L1 expressions in HBV-related hepatocellular carcinoma (HCC)

There was a significant correlation between PD-L1 expression in PBMCs and PD-L1 expression in tumor tissue. (r = 0.706, P = 0.002). Both of them were the independent marker for the prognosis of HCC patients

The respective recurrencefree survival rates at 1, 3, and 5 years were 61.5, 31.1, and 11.7% in the TACE group and 84.0, 64.1, and 64.1% in the HR group (p \ 0.001). The respective overall survival rates at 1, 3, and 5 years were 96.8, 76.4, and 61.6% in the TACE group and 96.0, 88.5, and 88.5% in the HR group (p = 0.017). In a subgroup analysis, the HR group overall survival rates were significantly higher than those of the TACE group in patients with less than 2 cm or CLIP score 0 HCC. However, no significant difference was detected between the two groups for patients with[2 cm (and B3 cm) size (p = 0.2183) or CLIP 1 (p = 0.5214) HCC. Conclusions: HR showed better overall and recurrence-free survival rates than TACE in patients with single HCC less than 3 cm and Child-Pugh class A. TACE had overall survival rates similar to those achieved by HR in the subpopulation with [2 cm (and B3 cm) sized or CLIP 1 HCC. In such cases, TACE can be considered in patients who are not candidates for resection. PP754 Evaluation of Newly Developed Combination Therapy of Intra-arterial 5-Fluorouracil and Systemic Pegylated Interferon a-2b for Advanced Hepatocellular Carcinoma with Portal Venous Invasion Kazuhiro Kasai 1 , Kei Sawara 1 , Yukiho Kasai 1 , Akira Ushio 1 , Kanta Oikawa 1 , Yasuhiro Takikawa 1 , Kazuyuki Suzuki 1 1 Department of Gastroenterology and Hepatology

Soon Young Ko 1 , Byung Kook Kim 1 , Sang-Wo Park 2 , Chang Hong Lee 1 , So Young Kwon 1 1 Department of Internal Medicine

TSU-68, in the Phase I/II Study for Advanced Hepatocellular Carcinoma Fumihiko Kanai 1,2 , Haruhiko Yoshida 1 , Ryosuke Tateishi 1 , Shinpei Sato 3 , Takao Kawabe 1 , Shuntaro Obi 3 , Yuji Kondo 4 , Makoto Taniguchi 4 , Kazumi Tagawa 4 , Masafumi Ikeda 5 , Chigusa Morizane 5 , Takuji Okusaka 5 , Hitoshi Arioka 6 , Shuichiro Shiina 1

Japan and increased apoptosis of tumor endothelial cells by combination of avastin and sorafenib. Conclusions: VEGF may contribute to resistance of sorafenib, avastin significantly inhibited tumor angiogenesis by inducing apoptosis of tumor endothelial cells

Exerts Anti-mitogenic Effects and Decreases Expression of Pro-angiogenic HIF-1a and Tumor-elevated PPARc and COX-2 in Human Hepatocellular Carcinoma Cell Lines Christian Freise 1 , Ulrike Erben 1 , Ulf Neumann 2

Methods: The human hepatoma cell lines Huh-7 and SK-Hep1 were treated with different concentrations (50-200 lg/mL) of an aquaeus extract of L. obtusiloba. Proliferation was determined using [ 3 H]-thymidine incorporation. Cytotoxic effects of the L. obtusiloba extract were determined via uptake of fluorescent ethidium-homodimer-1 (EthD-1) in damaged cells. Effects on insulin-like growth factor (IGF-1; 100 nM) stimulated HIF-1a, PPARc and COX-2 expression were analyzed by Western-Blot. Effects on migration/ invasion were checked using the ''wound healing'' assay and a transwell system. Results: The L. obtusiloba extract induced a concentration dependent reduction of proliferation by *70% (IC 50 = 100 lg/mL) with only marginal cytotoxic effects. Furthermore, the migration of both cell lines were decreased strongly. IGF-1-increased PPARc (30%), HIF-1a (350%) and COX-2 (250%) expression were neutralized to control values by L. obtusiloba concentrations near the IC 50 . Conclusion: Suppressed de novo DNA synthesis, decreased cell motility and the repression of PPARc, HIF-1a and COX-2 after treatment with L. obtusiloba point to anti-tumor and anti-angiogenic activities of the L

Intermediate Conductance Calcium-activated Potassium Channel (IKCa) as Target for Anti-neoplastic Treatments: Specific IKCa Blocker TRAM-34 in HCC Blocks in vitro Invasion and Migration Rajan Somasundaram 1 , Christian Freise 1 , Ulrike Erben 1 , Ulf Neumann 1

Effects on migration/invasion were monitored by the ''wound healing'' assay and a transwell system using boyden chambers with 0.8 lM pore size and coated with Matrigel TM . Induction of apoptosis was assessed by activation of a dye-quenched fluorogenic substrate of caspases-3 and -7. Results: TRAM-34 effectively blocked the proliferation accompanied by reduced invasion/migration of HepG2 and SK-Hep1 cells in a concentration dependent manner (IC 50 *4 lM) by up to 70 and 80%, respectively. DMSO solvent control was not inhibitory but rather stimulated invasion/migration. At concentrations of TRAM-34 leading to a 70-80% reduction of invasion/ migration only 5-10% (compared to staurosporine treatment) of the cells become apoptotic. Conclusion: As previously shown for other epithelial cell lines functional IKCa are expressed by human HCC cell lines HepG2 and SK-Hep1: Inhibition of cellular growth and migration/invasion of HCC cells

and SNU761) were used in this study. Cell viability and growth inhibition were assessed by MTT, colony forming, and trypan blue staining assay. Combination effects were evaluated using CalcuSyn software. Results: Gefitinib and silibinin exhibited antiproliferative effects on HCC cells in a dose-dependent manner. Combined treatment with silibinin enhanced the gefitinib-induced growth inhibitory effects in some HCC cell lines such as Huh-BAT and SNU 761. In addition, it found that these results were similar to those of colony forming and cell counting assay. The combination of gefitinib and silibinin was synergistic in SNU 761 cell line however it was only additive in Huh-BAT cell line. Conclusions: The anti-tumor effects of gefitinib in HCC

Minimal Fibrosis and Lower Aspartate Aminotransferase-platelet Ratio Index is Associated with Better Prognosis for Patients with Small Solitary Hepatitis-B Virus related Hepatocellular Carcinoma Underwent Resection Surgery

Taiwan Background: Although advanced liver fibrosis is an important factor to develop hepatocellular carcinoma (HCC) for patients with chronic hepatitis B, whether it is associated with the recurrence of HCC after resection remains obscure. Methods: This study enrolled 76 patients with small (\5 cm) and solitary hepatitis B virus (HBV)-related HCC underwent resection. The outcomes between minimal and advanced fibrosis in non-tumor part were compared. Serum markers were validated to assess the stage of hepatic fibrosis

Christophe Duvoux 1 , Fabienne Pessione 2 , Hanaa Badran 3 , Philippe Compagnon 4 , Tullio Piardi 5 , Fréderic Muscari 6

15 Paul Brousse hospital, Villejuif, France 16 Grenoble hospital

PP791 Characteristics of Bone Metastasis as the Only Site of Metastasis are Different from Those of Being Part of Multi-organ Metastases in

Tracer uptake characteristics of bone metastases and primary HCC were evaluated by correlation analysis. Results: The incidence rate of HCC bone metastasis from Group I was 12%, Group II 7% (total 19%), with no significant difference between them in age, sex, hepatis viral status, serum alpha-fetoprotein level and treatment methods. Of 69 lesions in Group I and 38 lesions in Group II, ACT alone was positive in 36 and 13%, respectively, FDG alone 6 and 37%, and ''ACT+FDG'' 58 and 50%. ACT was significantly more sensitive for Group I bone metastasis (P \ 0.05) and FDG for Group

Tracer avidity of bone metastases in each group correlated well with that of the corresponding primary

Conclusions: Metastatic bone lesions are largely similar in cellular differentiation and tracer avidity with their primary HCC. Well-differentiated HCC may have a greater tendency to metastasize to bone first than poorly-differentiated HCC, which has a more random pattern of metastases

Gastroduodenal Complications after Concurrent Chemoradiation Therapy in Patients with Hepatocellular Carcinoma: Endoscopic Findings and Risk Factors Young Eun Chon 1 , Beom Kyung Kim 1 , Seung Up Kim 1

) radiation-induced gastric or duodenal ulcer or (3) any other gastroduodenal toxicity associated with radiation. Serious gastroduodenal complications were defined as those requiring prompt therapeutic interventions or severe enough to stop or delay the scheduled CCRT protocol. Results: During the median follow-up of 155 days, 62 (52.8%) patients showed endoscopic evidence of radiation-induced gastroduodenal complications after CCRT. Radiation gastric and duodenal ulcers were found in 32 (26.0%) and 20 (16.3%) patients, respectively, while 50 (40.7%) and 42 (34.1%) patients had radiation gastritis and duodenitis, respectively. Bleeding was observed in 13 (10.6%) patients. Eighteen (14.6%) patients experienced serious gastroduodenal complications

Epigenetic Silencing of WIF-1 in Hepatocellular Carcinomas Yun Deng 1 , Zhi-Gang Zhang

Methods: Quantitative real-time PCR analysis was employed to measure mRNA levels of WIF-1 for hepatocellular carcinoma (HCC) cell lines and tissue samples on ABI7300 instrument. Methylation status of WIF-1 was analyzed by bisulfite DNA sequencing and methylation specific PCR. Results: Expression of WIF-1 was repressed due to its promoter hypermethylation and this suppression in HCC cell lines could be restored by demethylating agent. Furthermore, cell growth was inhibited by overexpression of WIF-1 in HCC cells. Conclusions: These results suggest that WIF-1 is frequently inactivated by promoter methylation and it may be a candidate tumor suppressor in HCC. This work was supported by the National Natural Science Foundation of China (30973492), the Shanghai Municipal Program of International Cooperation in Science and Technology (08410700900), and the Programs of

The Effects of Palliative Treatment Modalities of Hepatocellular Carcinoma (HCC) on Patients' Quality of

China Background and aim: Hepatocellular Carcinoma (HCC) is one of the major causes of cancer deaths in Hong Kong. Palliative treatment modalities are commonly used in about 80% of HCC patients. This study aims to assess the quality of life (QOL) issues including functioning and symptoms subscales among advanced HCC patients and identify factors (demographics, treatments, clinical data, and laboratory results) in determining patients' QOL. Methods: A cross-sectional quantitative design was adopted to examine descriptive, comparison and association data related to QOL

Results: The overall mean score of the global health status/ QOL of patients in this study was low (46.7/100). Patients experienced low role and social functioning and high level of insomnia and fatigue. When comparing the four treatment groups of patients: Transarterial Chemoembolization (TACE), Percutaneous Ethanol Injection (PEI), Radiofrequency Ablation (RFA), and Symptomatic Control Treatments, patients underwent Symptomatic Control Treatments had the poorest quality of life level (16.8/100). Patients below age 60 experienced more emotional and sexual problems (P \ 0.001). Elevation of Alpha Feto Protein (AFP) and bilirubin

Median age at presentation was 66 years and 105 patients (91.3%) had clinical evidence of liver cirrhosis. There were multiple tumor in 59 patients (47.2%). Nine patients underwent hepatic resection and 41 patients received TAE. Five patients received staged liver resection after hemostasis by TAE. Other 80 patients were treated conservatively. The 30-, 60-and 90-day survival rate for all the patients were 45.3, 30.2 and 26.2% respectively. Mean survival for the patients after palliative treatment, TAE, surgery, and two-stage approach were 1.1, 6.8, 21.8, and 59.7 months, respectively. Two cases after two-stage approach have been surviving for 77 and 78 months respectively without disease recurrence. Conclusion: Mean survival in patients treated by TAE, surgery, and two-stage approach was better than that of patients who received palliative treatment only. Properly selected patients with ruptured HCC could also achieve good prognosis by two-stage approach. drain independently into IVC

Postoperative Infection in Patients with Hepatoma Undergoing Liver Transplantation Yue Shi 1 , Qing-Yu Zhao 1

Among the identified pathogens, 31.1% were Staphylococci (64.9% was MRSE/MRSA), 6.7% was Monilia albiicans, and 5.9% was Aerobacter cloacae and Pseudomonas maltophilia. 19.3% of the isolated G -bacilli were ESBL producing strains. Infections were frequently observed in the first week after operation. Serum Alb levels decreased gradually after operation, but recovered to be normal in the first month. Serum levels of the immunodepressant FK506 reached its maximum in the first week with its concentrations maintained around 11.6 lg/L in the first month and decreased in the second month. Conclusion: In patients with liver cancer, the incidence of postoperative infection was high in the first month after LT. G + cocci (mainly MRSE/MRSA Staphylococci) and G -bacilli (mainly Aerobacter cloacae and Pseudomonas maltophilia) were the predominant species causing infection. Monilia albicans is the major species of fungal infection

Tobacco Smoking and Long-term Outcome in Liver Transplantation Maggie Ow 1

There was no difference in patient survival between smokers and non-smokers, with 10-year survival of 84 versus 86%, respectively. There was a trend towards better 10-year graft survival in non-smokers compared to active smokers (90 vs. 79% respectively, P value 0.06). The commonest cause of graft failure in smokers was recurrent hepatitis C. There was a trend towards an increased risk of hepatic artery thrombosis in active smokers (8%), versus non-smokers (3%) (P value 0.09). There was no increase in non-skin malignancy or cardiovascular disease in smokers. Conclusion: Pre-transplant smoking status did not influence patient survival post-transplant. There was a trend towards reduced graft survival and this entire course, AST/ALT level never rose above normal range. She succumbed to sepsis 47 days after LTX. Conclusion: The authors consider reduction in immunosuppression level nor WBC transfusion works for acute GvHD after LTX

There was no statistical difference in mean first checkout time between pp65 and IE antigenemia assay (P = 0.769). The level of pp65 and IE antigenemia had a significant correlation (r = 0.828). Moreover, 150 (11.2%) positive samples were detected by PCR.The distribution of gB genotypes was as follows: gBn1, 60% of patients; gBn2, 13.3%; gBn1 and gBn3 mixed infection, 26.7%, gBn4 and other mixed infection were not found. The level of CMV gB DNA detected and the number of CMV positive pp65 cells correlated well (r = 0.641)

MELD Score and Risk of Mortality in Liver Transplant Candidates with Hepatitis B Cirrhosis: A Lesson from the US Experience W. Ray Kim 1 , Rachel Pedersen 1

Methods: From the Organ Procurement and Transplant Network, all adult primary LTx candidates with ESLD or non-acetaminophen acute liver failure (ALF) between 2003 and 2005 were identified and divided into HBV ESLD, non-HBV ESLD and ALF groups. The competing risk analysis and proportional hazards regression analysis were used to assess mortality at 3 months post-listing. Results: The analysis included a total of 21,575 waitlist registrants including HBV ESLD (n = 450), non-HBV ESLD (n = 18,996), and ALF (n = 2,129)

Result: CMV-Leu and CMV-Pro vectors were proved to be exactly correct by DNA sequencing. The amounts of TGFb1 secreted from both CMV-Leu and CMV-Pro transfected cells were higher than pcDNA3.1-transfected cells in both cell lines. CMV-Pro-transfected cells secreted much more TGFb1than CMV-Leu -transfected cells, and the difference was statistically significant (P \ 0.05) in L02 cells but not in HepG2 cells. The apoptotic rates of CMV-Leu or CMV-Pro transfected cells were greater than pcDNA3.1-transfected cells in HepG2 cells. While in L02 cells the apoptotic rates of CMV-Leu or CMV-Pro transfected cells were much lower than pcDNA3.1-transfected cells. The apoptotic rates of CMV-Pro-transfected cells was also lower than that of CMV-Leu -transfected cells in two cells. And the differences between all transfected cells were statistically significant (P \ 0.05). Conclusion: Leu10Pro encoding Pro10 was associated with the augmentation of TGFb1 secretion

In this research, the antitumor effect of vitamin K along and combination of VK2 and Epirubicin was examined in the human hepatocellular carcinoma cell lines HepG2 and PLC/PRF/5. Methods: HCC cell lines were cultured in DMEM containing 10% of fetal bovine serum. Cytoxicity of vitamin K2, K3, K5, Epirubicin and combination of VK2 and Epirubicin was determined using XTT method. DNA of control and treated cells were extracted and were analysed by agarose gel electrophoresis. Results: IC 50 of VK2, VK3 and VK5 for HepG2 were

PP845 Inhibition of 5, 2 0 , 4 0 -Trihydroxy-6, 7, 5 0 -trimethoxyflavone on Proliferarion of Hepatoma HepG-2 Cell Line Xue-Wu Zhang 1

Apoptosis of HepG-2 cells was investigated further by means of inverted microscope, HE staining, transmission electron microscopy and expression of apoptosis-associated gene bcl-2, bax and p53 were determined by immunohistochemical method. The expressions of caspase-3 protein was detected by Western blot. Results: TTF inhibited the proliferation of HepG-2 cell and the effects were in a time-and-concentration dependent manner. TTF can also induced apoptosis, the G 0 /G 1 phase was arrested and the ratio of apoptosis there was significantly difference than that of control group, respectively. The expressions of apoptosis-associated gene bcl-2, bax and p53 had changed significantly, respectively. Western blot disclosed that followed by the increase of TTF, caspase-3 protein expression was step up gradually. Conclusions: TTF induced apoptosis of HepG-2 cells via induction of apoptosis and cell cycle arrest. PP846 Wild and Mutant Hepatitis B Virus X Protein wth Deletion at OOHterminus Induce Cell Cycle Progression via Down-regulating P16 ad RB Pathway in HepG2 Cells Lin Yang 1 , Xue-Bing Yao 1

However, no significant differences was observed among HepG2/GFP-HBxn (67.950 ± 2.128)%, HepG2 and HepG2/ GFPcells. Western blot analysis show that the levels of p16, RB and phosphorylated RB in HepG2/GFP-HBx and HepG2/ GFP-HBxc cells significantly decreased compared with HepG2, HepG2/GFP cells. However, no decreased level of p16, RB and phosphorylated RB was observed in HepG2/GFP-HBxn cell. Conclusions: Wild and the COOH-terminus-deleted HBx promote cell growth and cell cycle progression via down-regulating p16 and RB pathway

After Poly (I:C) injection, CRACC expression on NK cells from all the organs were significantly enhanced, and moreover, all hepatic lymphocytes remarkably increased the CRACC expression when compared with other organs. Conclusion: These results indicate that CRACC may play an important role in this Poly (I:C)-induced NK cells-mediated liver injury. amplified expression of anti-apoptotic gene-survivin, which also displayed a biphasic phenomenon. Moreover, activation of nuclear factor-kappaB (NF-jB) corresponded to the up-regulation of survivin. Inhibitor of NF jB, BAY 11-7082, suppressed the expression of survivin and eliminated the biphasic response. It reveals that the up-regulation of survivin is mediated by the activation of NF-jB. Conclusion: These data suggest that a low dosage of GCDC cause hepatocyte apoptosis to exhibit the biphasic response, which may result from the expression of Survivin and mediation of NF-jB. PP853 Effects of the Inhibition of Hypoxia-Inducible Factor-1A (HIF-1A) and Interleukin 8 (IL-8) on Angiogenesis and Metastasis in Liver Cancer In Vitro Sung Hoon Choi 1

College of medicine, 250 Seongsanno, Sodaemun-gu, Seoul 120-752, Korea, 4 Yonsei Liver Cancer Special Clinic, Yonsei University Health System, 250 Seongsanno, Sodaemun-gu, Seoul 120-752, Korea Objectives: Hypoxia-inducible factor-1a (HIF-1a) is a central transcription factor involved in various aspects of cancer biology, including proliferation, survival, angiogenesis, and extracellular matrix metabolism under hypoxia. Interleukin 8 (IL-8) is a key mediator of angiogenesis during tumor development. This study evaluated whether adenovirus-mediated small hairpin HIF-1a (shHIF-1a) and IL-8 (shIL-8) inhibited angiogenesis and metastasis in the hepatocellular carcinoma (HCC) cell lines; HepG2 and Huh7. Method: HCC cell lines were infected with adenovirus expressing shRNA and cultured under hypoxic conditions (1% O 2 , 24 h). Following infection, the expression levels of HIF-1a, angiogenesis factors, and matrix metalloproteinase were examined through Western blotting, while tumor growth was measured by cell proliferation assay (MTT assay) and angiogenesis was measured by angiogenesis related assay (Tube formation assay, Migration assay, etc). Results: The inhibition of HIF-1a by adenovirus-mediated shRNA led to the repression of tumor growth, while the inhibition of IL-8 did not. When human umbilical vein endothelial cells (HUVECs) were treated with culture medium from HIF-1a and IL-8 knockdown HCC cell lines, we observed reduced tube formation capacity and invasion compared to HUVECs treated with culture medium from wild-type cells. Similar effects were observed ex vivo (aorta ring assay), where the microvessel formation was inhibited significantly. Conclusions: These data suggest that adenovirus-mediated shHIF-1a and shIL-8 inhibit tumor angiogenesis and metastasis in HCC cell lines

With the rising of IFN-a concentration, APOBEC3G mRNA and protein level rise progressively. When IFN-a concentration is 10 4 U/ml, the expression level of APOBEC3G is the highest. Moreover, the expression level of STAT-1 mRNA and protein also rise progressively, which appear with APOBEC3G expression amount parallelly and relevantly. With the extension of stimulation time of IFN-a, APOBEC3G expression level rise obviously, reach highest at the 8 hours, and thereafter drop gradually. When 10 4 U/ml IFN-a stimulates 8 h, the level of HBsAg, HBeAg, HBV DNA in cultural supernatant and the level of HBV mRNA in HepG 2 2.2.15 cells are obviously lower than the cells untreated with IFN-a. Conclusion: IFNa could induce HepG 2 2.2.15 cells express APOBEC3G. Within the certain limits, APOBEC3G expression presents positive correlation with IFN-a dosage and action time. The expression of APOBEC3G induced by IFN-a could be one of antiviral mechanisms of IFN-a. Whether JAK-STAT signal pathway participates in the expression of APOBEC3G induced by IFN-a need further studying

To observe the effects of APOBEC3G on biological behaviors, including cell growth and migration ability, cell cycle, apoptosis, etc., of HepG 2 2.2.15 cells after highly-expressed APOBEC3G. Methods: Eukaryotic expression vector pcDNA3.1-APOBEC3G was transfected into HepG 2 2.2.15 cells. Limiting dilution monoclonal screening was used to establish HepG 2 2.2.15 cells in which APOBEC3G protein was stably expressed. Western-blot was used to identify and screen HepG 2 2.2.15 cells in which APOBEC3G protein was highly expressed. The changes of cell cycle and apoptosis were detected by using flow cytometry (FCM). The proliferation of cells was analyzed by MTT assay. The migration and movement of cells was determined with rowing trace. Results: The changes of cell cycle, apoptosis, cell growth and cell migration

Conclusion: Highly-expressed APOBEC3G has no obvious effects on biolog

Epidemiologic Study on Plasmid-mediated High-level Resistance to Aminoglycosides in Acinetobacter Baumannii Isolates of Zhejiang Due to 16S Rrna Methylation Yun-Feng Pan 1

of 25 hospitals in Zhejiang were analysed. Three 16S rRNA methylase genes, armA, rmtA and rmtB, were detected by PCR-based assays. Agar dilution and Etest were used to determinate the MIC of these strains to 17 antibiotics. The homology of these isolates was analyzed by pulse-field gel electrophoresis (PFGE). PCR was performed to amplify the coding sequence of 16S rRNA methylases

Multidrug-resistance to various aminoglycoside were identified in 264 strains. Overall, 237 armA-positive isolates were identified, with no rmtA-, rmtB-positive be detected. The prevalence rates of armA among 544 isolates were 43.5%. All strains harboring armA gene which was confirmed by PCR and sequencing. PFGE analysis indicate that 544 strains isolated from 25clinical units, were classified into 6 clone mainly .and that clone A and C had widely spread in Zhejiang. No plasmid was extracted and conjugation was not successful, despite multiple attempts. Conclusions: This is the first report of the emergence of 16S rRNA methylases in Acinetobacter baumannii isolates in Zhejiang. Nosocomial spread of 16S rRNA methylases producing Acinetobacter baumannii is quite severe. The acceleration of aminoglycoside resistance among gram-negative bacilli by producing 16S rRNA methylases

China Methods: We analyze EGF +61 polymorphism among 147 cirrhosis patients and 227 healthy controls by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and use chi-square test with SPSS 10.0 to evaluate the distribution frequency of the genotype and allele. Then we compare the distribution frequency of the genotype and allele in different Child-Pugh grades in cirrhosis to evaluate the relationship between EGF +61 polymorphism and the progression of cirrhosis in Chinese patients. Results: The distribution frequency of AA genotype and A allele frequency of EGF +61 polymorphism were both higher in healthy controls than cirrhosis patients, but no significant difference was observed in the distribution (P [ 0.05). And we still did not found significant difference in the distribution frequency of AA genotype and A allele frequency of EGF +61 polymorphism in different Child-Pugh grades of cirrhosis

China Background: MicroRNAs (miRNAs) are small noncoding RNAs with regulatory functions as tumor suppressors and oncogenes. Although single nucleotide polymorphism (SNP) in miRNA regions have been reported to be rare and unlikely to be functionally important, recent studies have implicated that the rs11614913 SNP in miR-196a2 was associated with susceptibility of lung cancer, congenital heart disease, breast cancer and shortened survival time of non-small cell lung cancer. Our aim was to investigate the association between this genetic variant and the risk and/or progression of HBV-related HCC. Methods: A total of 528 patients with chronic HBV infection were periodically enrolled. The patients were divided into two groups: those without (n = 167) and those with HCC (n = 361). DNA was extracted from blood specimens, and miR-196a2 polymorphism was genotyped by polymerase chain reaction-ligation detection reaction (PCR-LDR). Results: The risk of HCC was non-significantly higher with miR-196a2 CC genotype or miR-196a2 C allele containing genotypes compared with those with the TT genotype among all or male patients. Interestingly, in female patients, we observed the opposite trend: carriage of the T allele contributes to the pathogenesis of HBV-related HCC. Furthermore, in a subsequent analysis of the association between this polymorphism and clinicopathological characteristics, there was still no significant difference in both the distribution of genotype or allelic frequency within HCC patients. Conclusions: Our results suggested that miR-196a2 polymorphism was not associated with both an increased risk and progression of HBV-related HCC in Chinese

Background: NKG2D on NK or NKT cells is involved in acute and chronic liver injury, so, inhibiting the expression of NKG2D ligands on hepatocytes should be a potential approach in related liver diseases. Since there are several ligands of NKG2D, such as Rae-1, Mult-1 and H60 in mice, construction of a liver-specific shRNA vector for simultaneously silencing multiple NKG2D ligands will be highly attractive. Methods: Albumin enhancer/promoter and alpha-fetoprotein enhancer/albumin promoter fragments were prepared from plasmid Albumin2335A and pLIVE, respectively. These promoter fragments were cloned into plasmid pEGFP-C1 to replace the CMV promoter and hepatocyte-specific expression vectors were constructed. A liver-specific shRNA vector for simultaneously silencing multiple NKG2D ligands (Rae-1, Mult-1 and H60) was also constructed. Results: The results showed that albumin promoter had a strong transcription activity in hepatoma cell line such as Hep2, HepG2 and Hepa1-6. EGFP was observed under the fluorescence microscope. Albumin enhancer and alphafetoprotein enhancer raised the transcription activity of albumin promoter as EGFP expression level was increased in the transfection experiments. In another study, after hydrodynamic injection of plasmid containing three shRNA sequences (siRae1-siMult1-siH60), we found the decrease of expression of these three ligands on liver (hepatocytes)

To study the dynamic changes of liver regeneration rate, PCNA and BMP2, the rats were sacrificed at different time points after PH (12, 48 and72 h).NC group was used as 0 h observation time point, namely 0 h group. For each time point indicated, eight rats were used in parallel. Results: Liver regeneration rate and PCNA in all groups were significantly increased gradually after PH. Liver regeneration rate and PCNA in NL,NH groups were significantly decreased than NS group at 12 h after PH (P \ 0.01). There were no significantly differences among three groups at 48, 72 h after PH (P [ 0.05). BMP2 in liver homogenate or immunohistochemistry in NS,NL,NH groups were significantly decreased than NC group at 12

Natural Killer Cells are Involved in Impairment of Murine Liver Regeneration via Tumor Necrosis Factor-A after Carbon Tetrachloride Treatment Rui Sun

After exposure to hepatotoxic injection of CCl 4 , we examined the regenerative response to hepatic injury in models treated with or without polyI:C. Results: PolyI:C treatment impaired CCl 4 -induced liver regeneration. In pol-yI:C-treated mice, the impairment was associated with accumulation and activation of NK cells and secretion of a large number of NK cells-derived TNF-a. The impaired capacity of liver regeneration of polyI:C was enhanced in NK-depleted mice and TNF-a neutralized mice

Mouse that was deficient in fumarylacetoacetate hydrolase (Fah -/-) die as neonates as a result of liver dysfunction. NTBC, a chemical, was reported to protect Fah-deficient mice from death. When murine hepatocytes need to be replaced by human hepatocytes, NTBC water feeding Western-blot analysis. CyclinD1 mRNA expression was determined by semiquantitative RT-PCR; The mRNA levels of some phenotypic markers, including AFP, CK-19,ALB, and two hepatic nuclear factors, HNF-4, HNF-6 were determined by RT-PCR. CK-19 and AFP protein expression were examined by Western-blot analysis. Results: We found that Wnt3a promoted proliferation of WB-F344 cells. Stimulation of WB-F344 cells with recombinant Wnt3a resulted in the accumulation of the transcriptional activator b-catenin, together with its translocation into the nucleus, and the upregulation of typical Wnt target genes cyclin D1 After 3 days of Wnt3a treatment in the absence of any serum, these cells retained their bipotential to express several specific hepatocyte and cholangiocyte phenotypic markers

Guangzhou 510630, China PP871 The Significance on NF-Kb Changes for HGF-mediated Proliferation and Differentiation in Human Fetal Hepatic Stem Cells Yan-Hang Gao

Methods: By ELASA, immunocytochemistry staining and RT-PCR methods used, the NF-kB activity changes in hFHSCs were detected for the role of HGF concentrations in different conditions. Results: HGF can promote the proliferation and differentiation of hFHSCs, in the process, and NF-kB activity showed a positive correlation with HGF concentrations; immunocytochemical show that with the increase in HGF dose, the staining degree of cells nucleus and the number of the cells whose nucleus were stained were both gradually increase the. This result intuitively verified ELASA detection results; RT-PCR showed that HGF can promote the expression of hFHSCs P65 and P50mRNA, I jBmRNA expression was reduced, so that the production of I jB was deseased

Differential Effects of Connective Tissue Growth Factor and Transforming Growth Factor-b on Rat Hepatic Progenitor Cells Ai-Ting Yang 1

PR China Background: Rat hepatic progenitor cells (WB-F344 Cells) can differentiate into hepatocytes and bile duct cells. Connective tissue growth factor (CTGF), a downstream mediator of transforming growth factor (TGF)-b, plays an important role in differentiation of cells and extracellular matrix (ECM) synthesis. But it is unknown that the effects of these two cytokines on hepatic progenitor cells. Methods: MTT assay was used to detected cell viability when different concentrations of TGF-b and CTGF were used to stimulate WB-F344 cells incubated serum-free for 24 and 48 h, respectively. Real-Time PCR

Results: Both TGF-b and CTGF could inhibit the proliferation of cultured WB-F344 cells in serum-free medium in a dose-dependent and time-dependent manner. 90 and 50% inhibition concentrations of TGF-b are 0.5 and 5 ng/ml, while CTGF are 1 and 5 ng/ml. RT-PCR and Western blotting results showed that compared with CTGF used to stimulated WB-F344 cells, TGF-b can changed the phenotype of rat hepatic progenitor cells significantly, including albumin, AFP and CK-19. We also find that TIMP-1 expression decreased when

PPC Protects Liver Function and Ameliorates Endotoxic Shock via Inhibition of Acute Infalmmatory Responses Qing-Yu Zhao 1

United States Aim: To test the effects of polyenylphosphatidylcholine (PPC) on liver injury caused by endotoxic shock and to investigate its mechanism with BMDM (bone marrow-derived macrophages)

LPS and PPC+LPS (P+L) groups. 6 mg/kg E.coli endotoxin was injected into abdominal cavity to cause lethal shock and PPC at 1 ml/100 g (232.5 mg/ kg) was given via tail vein 24 and 6 h before endotoxin. After sacrifice in 24 h, liver weight, enzyme and immunohistochemistry staining of ICAM-1 were done. BMDM were from bone marrow of WT mice

PPC 0.1 mM was given 2 h before LPS (100 ng/mL) stimulation for preincubation. After stimulation, cells were prepared for Western blot detection of Erk, IKKa/b, IjBa, their phosphorylation products (15, 30 min

Supernatant (24 h) was used to measure cytokine (TNF-a, IL-6,-10 and -12) levels with ELISA and NOx production with Griess reaction. mRNA (6hrs) of TNFa, IL-6 and IFNc were checked with real-time PCR

Association between Metabolic Syndrome and Chronic Hepatitis B in Taiwan: A Community-based Study

Methods: We conducted a community-based study in Kaohsiung area located on the southern Taiwan. Overnight fasting blood samples were collected in subjects for metabolic profiles and liver function tests and the patients' medical histories by the questionnaire were taken and body mass index (BMI) and waist circumference were measured. MS was diagnosed based on the Adult Treatment Panel (ATP) III (Asian version) criteria. Hepatitis B surface antigen (HBsAg) and anti-hepatitis C virus (HCV) antibody were also tested. The statistical tests including Student's t test

mg/dL, 194.7 ± 38.4 mg/dL, 57.5 ± 14.8 mg/dL, 123.8 ± 34.9 mg/dL and 24.4 ± 4.25 kg/m 2 , respectively and the prevalences of HBsAg, obesity (BMI [ 27 kg/m 2 ), and MS were 17.2, 21.4 and 19.7%, respectively. The 260 subjects with MS have significantly higher proportions of male gender, age [50 years, and obesity and significantly higher mean age, BMI, AST level and ALT level (All Ps \ 0.001) than subjects without MS. Multivariate logistic regression showed that age, BMI, ALT levels (all P \ 0.001), gender and AST level (P = 0.001). HBsAg status was not associated with MS. Conclusion: In the present community-based study, around one-fifth of the subjects have MS. MS is associated with demographic features

Combined Administration of DPA and Selenium Reverses Beryllium Induced Hepatic Dysfunction Sonia Johri 1

No antidote is available for beryllium induced hepatic abnormalities. D-Panicilamine (DPA) along with selenium was evaluated to encounter beryllium induced characteristic biochemical and ultra morphological alterations. Methods: Female albino rats were exposed to beryllium nitrate (1 mg kg -1 , ip) various biochemical and histopathological parameters were performed. Results: Exposure to beryllium increased its concentration in serum, liver and kidney and caused significant alterations in cytochrome P-450, lipid peroxidation and protein. Beryllium administration significantly altered the lactate dehydrogenase, c-glutamyl transpeptidase, bilirubin, creatinine and urea in serum; activity of acid phosphatase, alkaline phosphatase, adenosine triphosphatase, glucose-6-phophatase and succinic dehydrogenase, triglycerides, cholesterol, protein contents, glycogen contents, lipid peroxidation and glutathione level in liver and kidney as well as severe alterations in histopathology and ultra morphology of liver and kidney proving its toxic consequences at cellular level. Monotherapy of DPA and Se recovered some of the studied parameters moderately towards control

Puspanjolo Barat Raya No. 47

Methods: Design of study: descriptive and analytic cross sectional study enrolling 76 cirrhosis and 30 controls in Gastroentero-Hepatology unit, Tugurejo hospital. Diagnosis of cirrhosis was confirmed by two syndromes of hepatocellular failure and portal hypertensive, severity of liver Cirrhosis was graded according to Child-Pugh's classification, in all cases glucose tolerance was diagnosed by 75 gr-oral glucose tolerance test according to WHO criteria. The mean age was 49.5 ± 10.5 years and 58 patients were men

Conclusion: The ratio of hyperglycemia in Cirrhosis by OGTT is 89% with IGT: 35% and DM 54%. There are the significant positive correlation between glucose concentrations and Child-Pugh's degree

Predicted post-transplant survival calculated by the Metroticket model incorporating explant pathological data (n = 80) was 68% at 5 years

Conclusion: The Metroticket calculator incorporating pretransplant radiological staging criteria is an accurate predictor of post-transplant survival in HBVrelated HCC. However, waiting list drop-out remains an important issue

Background: Hypoxia may activate survival signals in hepatocellular carcinoma (HCC) cells. We have previously shown that hexokinase II (HK II) is actively participating in HCC cell survival, especially in a hypoxic environment, and that HK II inhibitor can induce apoptosis by activating mitochondrial signals. Hypoxia may also activate endoplasmic reticulum (ER) stress, which is counteracted by unfolded protein response such as protein disulfide isomerase (PDI) activation. Since HK II inhibitor may as well activate pro-apoptotic JNK, and ER stress can also lead to JNK activation,we hypothesized that PDI inhibition may potentiate HK II inhibitor-induced JNK activation and apoptosis by increasing ER stress in hypoxic HCC cells. Methods: Huh-7 cells were used in this study. Cells were treated with bacitracin,a PDI inhibitor, and/or 3-bromopyruvate(3-BP),a HK II inhibitor. Cellular apoptosis was analyzed by MPT assay, and apoptotic and kinase signaling pathways were explored by immunoblot analysis. Results: The expression of PDI was increased in HCC cells cultured in hypoxia as compared to normoxic cells. Bacitracin treatment alone did not induce HCC cell death. While 3-BP treatment alone did not induce ER stress in hypoxic HCC cells, 3-BP treatment was able to induce ER stress in the presence of bacitracin. Accordingly, a strong 3-BP-induced JNK activation was observed Aim: Prognosis of advanced cholangiocellular carcinoma is extremely poor. We reported the effectiveness of Combination Therapy of 5-Fluorouracil and Interferon-Alpha for Advanced hepatocellular carcinoma (S Obi et al. Cancer 2006). This study aimed for pilot study of combination therapy of 5-fluorouracil and interferon for advanced cholangiocellular carcinoma. Method: Fifteen patients with cholangiocellular carcinoma received peginterferon (90 ng, subcutaneous injection on weekly) and 5-Fluorouracil (500 mg into hepatic artery on Days 1-5 of the first and second week of each 4-week cycle). The therapy was either terminated at the end of the first cycle in cases with progressive disease, or continued when responses to treatment were evaluated by Eastern Cooperative Oncology Group criteria. Result: Four (27%) patients showed partial response (PR) and complete remission (CR) was not observed. Two patients (23%) showed stable disease (SD). The median survival time was 9.0 months. Adverse events were limited to thrombocytopenia (2 patients), leucopenia (2 patient) Conclusion: The combination therapy with 5-Fluorouracil and Interferon was safe, and improved the survival rate among the responders.

Conclusion: We think that radiation therapy is safe and effective treatment for advanced hepatocellular carcinoma with portal vein thrombosis.

Hsp27 Phosphorylation can Enhance Radioresistance of Hepatocarcinoma Cells by Stimulating Akt Activation Alexander Kabakov 1 , Yulia Makarova 1 , Vladimir Kudryavtsev 1 1 Medical Radiology Research Center, 4 Korolev Street, Obninsk, 249036, Russia Background/Aims: The 27 kDa heat shock protein (Hsp27) is a cytoprotective chaperone whose constitutive expression is increased in many human tumors and associated with poor prognosis. Hsp27 is involved in radioprotective responses of tumor cells, while machinery of such Hsp27-involving radioprotection is not quite unraveled. The present study was aimed at elucidation of a role of Hsp27 phosphorylation in the phenomenon of irradiation-responsive Akt activation (phosphorylation) that contributes to the hepatocarcinoma cell radioresistance. Methods: Human hepatocarcinoma cells (HepG2 and HCCLM3 lines) were exposed to gamma-radiation at clinically relevant doses (3-6 Gy). The site-specific phosphorylation of Hsp27 and Akt or Hsp27-Akt interaction was determined by immunoblotting or immunoprecipitation (IP), respectively. The post-irradiation cell death and survival were assessed in TUNEL or annexin V-staining and clonogenic assays. Transient transfection with dominant negative or constitutively activated Akt was used to manipulate the Akt activity in the cells. Results: It was found that human hepatocarcinoma cells, constitutively expressing high Hsp27 levels, respond to gamma-radiation exposure by Hsp27 phosphorylation and subsequent Akt phosphorylation (activation); later, some cell fractions entered apoptosis. Down-regulation of Hsp27 with RNAi as well as inhibition of the Hsp27 phosphorylation with SB203580 or dominant-negative MAPKAP kinase-2 suppressed the Akt activation and enhanced apoptosis. Effects of (co-)transfection with mutant Hsp27 and/or Akt also suggest implication of the Hsp27 phosphorylation in catalyzing the radioprotective Akt activation in the irradiated tumor cells. IP from the cell lysates revealed Hsp27-,Akt-containing complexes which dissociate when Hsp27 is phosphorylated following irradiation. Conclusion: These findings characterize the Hsp27 phosphorylation as an important event in the radioprotective response of hepatocarcinoma cells. Consequently, pharmacological inhibition of the Hsp27 phosphorylation in human hepatocarcinomas may sensitize them to radiotherapy.

Introduction of RFA in Mongolia: Year 1 Treatment Results at National Cancer Center of Mongolia Chinburen Jigjidsuren 1,2 , Sanduijav Ravjir 1,2 , Yumchinserchin Narangerel 1,2 , Bat-Ireedui Purevbaatar 1,2 , Sayabold Batmunkh 1,2 , Amarsanaa Jazag 1, 2 Aim: The objective of this study was to identify among liver transplantation candidates for for HCC with broader criteria, those at low risk of tumor recurrence. Patients and methods: Predictive factors of tumor recurrence were tested by multivariate analysis by Cox model from 35 preoperative variables in 373 patients transplanted for HCC (1988) (1989) (1990) (1991) (1992) (1993) (1994) (1995) (1996) (1997) (1998) in 14 centers [training cohort (TC)]. A prognostic score was derived from this analysis by linear transformation of b coefficients. The score was tested in a validation cohort (RVC) of 175pt and in a validation cohort (ABM VC) of 478pt, prospectively followed. Results: In the TC, the probability of tumor recurrence at 5 years according to AFP were 20.7% (\100 ng/ml), 32.2% (100-1,000 ng/ml) and 53.8% ([1,000 ng/ml) (p \ 0.001). Three independent predictors of recurrence were identified: the diameter of the largest nodule (p \ 0.0001): score: \3 cm = 0, 3-5 cm = 1, [5 cm = 5, the AFP level (p = 0.01) score \100 ng/ml = 0, 100-1,000 = 2, [1,000 = 3, and the number of nodules (p = 0.05) score: B3 = 0, or C 4 = 2. Two prognostic groups were identified in the TC according to the score B2 or C3. The prognostic model was validated in the two validation cohorts. In patients with a score B2, the incidence of recurrence and overall survival at 5 year were similar whether patients comply or not the Milan criteria. In the ABM VC cohort 59 patients were outside Milan with a 5-year probability of recurrence of 10.1 ± 4.3% and overall survival of 74.4 ± 5.7%. Conclusion: AFP level is highly predictive of HCC recurrence. A prognostic score with AFP identifies among patients outside Milan, those with low risk of recurrence. A score B2 could be proposed to rationally select patients. Background: Primary non-hodgkin lymphoma (NHL) of liver is a very rare malignancy. Surgical resection and chemotherapy are the effective treatment, especially using chemotherapy before surgical resection is more efficacious. Ultrasound-guided or CT-guided biopsy is the most important diagnostic method of primary NHL of liver. We aimed to investigate the different clinical and pathological features of primary NHL of the liver between B and T cell original in needle biopsy of liver. Methods: Here, we report three case of Primary NHL of liver. Etiological factors, clinical character, and histological features of three cases of NHL were retrospectively analyzed. Then the difference between B and T cell original were discussed in combination with immunohistochemical stain. Results: Three cases were all male. But all of them were no HBV/HCV infection in serum. With microscopic examination, the tumorous lymphocytes' destructive action was clearly visible. The tumorous lymphocytes infiltrated in portal area predominantly were B cell original, and infiltrated in sinus hepaticus were T cell original. The B cell original NHL were all expressed CD20. Conclusion: The histological features combined with immunophenotype are enough to diagnose, and differentiate between B and T cell original NHL. That was very important for the selection of chemotherapeutical program before surgical operation. (1) MP showed inhibitory effects on the proliferation of both two kinds of cells, which were related with the concentration of MP. (2) MP showed inhibitory effect on the expression of HBV DNA in HepG2.2.5 cells.(3) MP showed induction of cell cycle arrest in the G0/G1 phase, which was more obvious in the HepG2.2.5 cells than HepG2 cells, and the difference was statistically significant. (4) Treated by different concentrations of MP, expression of integrin a5b1 in HepG2.2.15 cells was significantly higher than that of HepG2 cells, and the expression rate increased as the concentration of MP increased. (5) MP reduced the expression of cyclin A, and expression of cyclinA decreased as the concentration of MP increased. Conclusions: MP showed inhibitory effects on proliferation of hepatoma cells and replication of HBV, in which expression of integrin a5b1 and cyclinA played an important role.

Liver Transplantation for Hepatitis B is Associated with Excellent Graft and Patient Survival Ed Gane 1 , Ken Tan 1 1 Auckland City Hospital, Grafton, Auckland, New ZealandBackground: Prior to the introduction of lamivudine and hepatitis B immunoglobulin (HBIG) prophylaxis, liver transplantation for HBV was associated with rapid HBV recurrence and death. The aim of this study is to report the outcome after liver transplantation for HBV at NZLTU following adoption of new prophylaxis regimens. Method: Medical records of all HBV OLT recipients registered with NZLTU from 1998 to 2009 were retrospectively reviewed. Data was collected on demographic, Indication for OLT, Hepatitis B prophylaxis regimen, HBV recurrence, and Survival. Results: 86 patients were included in this study. Mean age was 53 years and duration of follow up 5.5 years. Thirty-three were Maori, 20 Pacific Islanders, 19 Asian, 11 European. Over the study period, the indication for listing has changed from decompensated cirrhosis (80-14%) to HCC (10-71%). Forty-eight received HBIG-LAM and 36 received LAM-ADV. Of those on HBIG-LAM, 26/48 were switched to LAM-ADV, 3 were switched to LAM-Tenofovir. Six patients became HBsAg positive, of whom four also became HBV DNA positive. The remaining two patients remained HBV DNA negative but had recurrent HCC, suggesting HBs gene integration in tumour. Five year cumulative HBV recurrence rate was 4.4%. 10 patients died (8 recurrent HCC). Post-transplant survival at 1, 5 and 10 years was 96, 89 and 84%, similar to non-HBV patients. Pretransplant HCC was associated with reduced survival (see Fig. 1 ). Conclusions: Liver transplantation for HBV is now associated with very low HBV recurrence and excellent long-term outcomes. However, recurrent HCC remains an important problem. (3.38 ± 1.00) g/L; PLT: (10 8 ± 60) 9 10 9 /L vs. (184 ± 108) 9 10 9 /L, all P \ 0.01]. In the first week after operation, elevated PT, APTT, TT and INR levels decreased gradually, APTT was even lower than the preoperative level [(32.05 ± 6.50) s vs. (40.19 ± 4.11) s, P \ 0.01]. These changes appeared usually on 1-2 days after operation. Decreased PLT and Fib regained slowly at the first week after operation when compared with the preoperative levels [Fib: (2. 13 ± 0.53) g/L vs. (3.38 ± 1.00) g/L, P \ 0.01; PLT: (145 ± 90) 9 10 9 /L vs. (184 ± 108) 9 10 9 /L, P \ 0.05], but the values were normal. Stratification analysis indicated more obvious hypocoagulability in patients with moderate/severe cirrhosis and those with Child-Pugh B level than in their counterparts. Conclusion: LALC's coagulation functions shift from hypocoagulatory to hypercoagulatory or normal during perioperative period. Prevention of bleeding should be focused on at anhepatic phase and on 1-2 days after operation while prevention of thrombosis should be focused on after the first week of operation. The degree of liver cirrhosis and Child-Pugh level could help evaluate postoperative coagulation disorder.

The Relationship Between the ATP within CD4+ T Lymphocyte and the Acute Rejection After Liver Transplantation Background: Recently there are increasing interests in the relationship between CD4+ T lymphocyte and acute rejection. Now we would like to explore the role of it after liver transplantation. Methods: This study contains 77 patients that received LT from February to October 2009. They are divided into AR (acute rejection) and NAR (non-acute rejection) group while 56 healthy people being the control group. Blood specimen were collected preoperatively, 1, 2 and 4 week postoperatively. Results: ATP values within CD4+ T lymphocyte were elevated significantly in each group compared with those preoperatively. It reach the peak level in AR group and was apparently higher than those of the contemporary NAR group (P \ 0.05). By ROC curve analysis, the obvious elevation of the ATP value within CD4+ T lymphocyte 1 week postoperatively has better sensitivity and specificity in diagnosing the AR. The ATP sensitivity rate for early AR was 76.9% and specificity rate 81%. 1 week after LT. The ATP value within CD4+ T lymphocyte at the day of AR has positive relationship with the rejection acting index (RAI), with the relative index(r) being 0.876 (P \ 0.05). After the steroid dump treatment, all the ARs were reversed; and the ATP value declined significantly compared with that of the control group and of the day when AR occurred (P \ 0.05). Conclusions: During the postoperative period, the dynamic change of ATP value within CD4+ T lymphocyte had close relationship with acute rejection after liver transplantation. So it might be used as a feasible and noninvasive monitoring item in diagnosing the AR and the effectiveness of the anti-rejection treatment.

Hypophosphatemia May Not Correlated with Hepatic Regeneration after Living Donor Hepatectomy Ding Yuan 1 , Yong-Gang Wei 1 , Bo Li 1 , Lv-Nan Yan 1 , Tian-Fu Wen 1 , Ji-Chun Zhao 1 , Yong Zeng 1 1 West China Hospital, Sichuan University, NO.37, Guoxue Street, Chengdu, Sichuan, ChinaBackground: Hypophosphatemia has been reported a frequent occurrence after hepatectomy. This event has previously been ascribed to amplified phosphate utilization of hepatic regeneration. The purpose of this study was to investigate the correlation between postoperative hypophosphatemia and both donor morbidity and the degree of hepatic resection. Methods: The cases of 112 living liver donors were retrospectively evaluated, who had undergone right (n = 106) and left (n = 6) hemihepatectomy from March 2006 to December 2008 at liver transplantation center of the West China Hospital. We categorized the living donors according to severity of hypophosphatemia as follows: mild (1.5 to 2.5 mg/dL), moderate (1.1 to 1.5 mg/dL), and severe (\1.0 mg/dL). We compared the incidence of complications in these groups and analyzed the possible correlation between the serum phosphorus levels and the variables of hepatic regeneration. Results: 110 of all cases (98.2%) developed hypophosphatemia postoperatively, including mild (n = 60), moderate (n = 28) and severe (n = 22). Postoperative phosphate began to fall on POD2, typically reaching a nadir (1.88 ± 0.71 mg/dL) on POD3, and returned to the normal range on POD5. The incidence of complications was no found significant difference among three hypophosphatemia groups (P = 0.931). The nadir phosphorus level was no correlated with the follows variables, including the remnant liver volume (P = 0.478), the ratio of graft weight to donor body weight (P = 0.263), the postoperative increase of ALP (P = 0.460) and the serum nadir uric acid level (P = 0.657). However, the phosphorus decrease positively correlated with the uric acid decrease between POD1 and POD3 (R = 0.317, P = 0.038), but the increases of two variables were not significantly correlated each other between POD3 and POD7 (P = 0.895).Conclusion: Hypophosphatemia after hepatectomy was a frequent occurrence in healthy living donors. However, the phenomenon was transient and selflimited with hepatic function recovery, and unlikely to induce serious postoperative outcome. Furthermore, we presumed that the minor plasma phosphorus possibly influxes into hepatocytes so as to sustain salvage nucleotide synthesis after partial hepatectomy, but the event is no responsible for hepatic regeneration.

Ductular Reaction in Hepatitis C Recurrence Post Liver Transplantation Emilia Prakoso 1,2 , James Kench 2 , Andrew Clouston 3 , David Bowen 1,2 , Geoffrey McCaughan 1,2 , Nicholas Shackel 1,2 1 Centenary Institute Sydney, NSW -Australia, 2 Royal Prince Alfred Hospital Sydney, NSW -Australia, 3 Envoi Pathology Brisbane, Queensland -Australia Background: Ductular reaction (DR) is a reaction of ductular phenotype associated with liver injury and fibrosis. The appearance of DR in the setting of hepatocellular/cholangiocyte injury suggests there are regenerating hepatic progenitor cells (HPCs). In HCV recurrence post-liver transplantation (LTx), the pathogenesis of progressive fibrosis is unclear and the role of DR in this setting is unknown. We aimed to characterise DR, HPC, replicative arrest and intrahepatic proliferation in patients with HCV recurrence post-LTx and to correlate these changes with liver fibrosis and steatosis. Methods: Immunohistochemistry was performed on liver biopsies with HCV recurrence post-LTx. Donor livers (n = 15) served as controls. DR (ck7) and a-SMA (activated stellate cells and myofibroblasts) were quantified by image analysis. HPC was calculated by counting isolated ck7-positive cells periportally. Hepatocellular proliferation (Ki-67) and replicative arrest (p21) were assessed by counting positive hepatocyte nuclei. Fibrosis was staged (F0-4) according to Scheuer. Results: There were 105 patients with 194 liver biopsies. DR, HPC, a-SMA, replicative arrest, proliferative index and replicative arrest ratio were higher in HCV recurrence patients vs control donors (P B 0.020). There was significant positive correlation between DR and HPC (r s = 0.629, P \ 0.001). There were also significant positive correlations (P \ 0.001) between DR and HPC with a-SMA, fibrosis, portal and total Scheuer's scores. DR also positively correlated to proliferative index (P \ 0.001). Steatosis (P = 0.018) and lobular score (P = 0.003) only correlated to HPC, but not DR plateau and HPC peaked at F3. Conclusions: The DR and HPC responses correlate with the extent of fibrosis in early stage liver injury. The lack of correlations in the latter stages of injury is probably related to the diminished liver regeneration in advanced fibrosis. Further studies are warranted to investigate the role of the DR in liver injury and regenerative responses in HCV recurrence post-LTx. Background: Survival of the partial graft after living donor liver transplantation owes much to its tremendous regenerative ability. With excellent venous outflow capacity, a graft within a wide range of graft-to-standard-liver-volume ratio can cope with portal hypertension that is common in liver transplant recipients. Methods: However, when the ratio range is exceeded, modulation of graft vascular inflow becomes necessary for graft survival. The interplay between graft-to-standard-liver-volume ratio and portal pressure, in the presence of portosystemic shunt or otherwise, requires individualized modulation of graft portal and arterial inflows. Portal inflow boosting by shunt ligation can be guided by transonic flowmetry, whereas portal inflow muting by splenic artery ligation can be monitored by portal electronic manometry. Results: Here we describe four cases to illustrate this. One patient had hepatic artery thrombosis resulted from splenic artery steal syndrome which was the sequela of small-for-size syndrome. Emergency splenic artery ligation and reanastomosis of the hepatic artery successfully muted the portal inflow and boosted the hepatic arterial inflow. Another patient with portal vein thrombosis underwent thrombendvenectomy. Portal inflow was boosted with ligation of portosytemic shunt, which is often present in these patients with portal hypertension. The coexistence of splenic aneurysm and splenorenal shunt required ligation of both in the third patient. The fourth patient, with portal pressure and flow monitoring, avoided ligation of a coronary vein which became a main portal inflow after portal thrombendvenectomy. Conclusion: A management scheme of graft inflow modulation guided selectively by transonic flowmetry or portal manometry was formulated. Aim: To retrospectively investigate the experience of microsurgical hepatic artery reconstruction and management of hepatic arterial thrombosis (HAT) in adult-to-adult liver transplantation (ALDLT). Methods: From 1/2001 to 9/2009, 182 recipients, 10 of which received dual grafts, with end-stage liver disease underwent ALDLT. Mean age was 42 years (range 18-68 years), male-to-female ratio was 157:25. Microsurgical techniques and running sutures with back-wall first techniques were performed in all arterial reconstructions under surgical lope (3.59). Direct end-to-end anastomosis was applied between the recipient hepatic arteries (HA), including 62 right HAs, 20 left HAs, 90 proper HAs, 11 common HAs, and 2 aberrant right HAs arising from superior mesenteric artery, and the graft HAs. Intimal dissections were handled by interposition of great saphenous vein (GSV) between donor right HA and recipient common HA (3 cases) or abdominal aorta (AA) (2 cases), by interposition of cryopreserved iliac vessels between donor right HA and recipient AA (2 cases). Results: In incipient 48 patients of this series, HAT was encountered in 4 patients, and thereafter did not occur in consecutive 124 cases. Overall incidence of HAT was 2.20%. All HATs were suspected by color Doppler ultrasonography and confirmed by hepatic angiography. Two of these HATs, which occurred on 1st and 7th days following ALDLT, were immediately revascularized with GSV between graft and recipient AA, one of these HATs occurred on 46th days after ALDLT with no symptom, remain one of these HATs, which occurred on 3rd days following ALDLT, was cured by thrombolytic therapy combine with anticoagulant, and died of multiorgan failure on 36th days after ALDLT. No death related to HAT. Conclusion: Applying microsurgical techniques and running suture with backwall first technique for HA reconstruction can reduce the high risk of HAT following ALDLT. Background: The CMV pp65 antigenemia assay has been widely used to assess viral load of CMV infections in transplant patients and is often considered as a golden standard. The Quantitative real-time PCR for CMV DNA provides sensitive and specific data for detecting CMV as well as monitoring the infection. However, each laboratory uses its own standard threshold. In this study, we have described the real-time PCR and compared it with CMV pp65antigenemia assay in Chinese haematopoietic stem cell (HSC) and solid organ transplant (SOT) patients. Methods: 73 patients, 56 men and 17 women who underwent HSC transplantation and 181 recipients, 128 men and 53 women who underwent SOT were included in this study. Blood specimens were collected and used for detecting the CMV-DNA loading in plasma and the CMV pp65 expression in leukocytes, respectively. Eight healthy men distributed to the control group. Results: CMV DNA positive in HSC transplant recipients and in SOT recipients was 52.1% (0-8.14 9 10 6 copies/ml), 57.9% (0-6.08 9 10 6 copies/ml), respectively. The positive antigenemia in HSC transplant recipients and in SOT recipients was 49.3% (positive cell number 1-30 per 5 9 10 4 WBC), 53.9% (positive cell number 1-37 per 5 9 10 4 WBC), respectively. The positive rate by both assays was 43.8% (32/73) and 45.9% (83/181), respectively. The negative rate in both assays was 41.1% (30/73) and 40.3% (73/181), respectively. The coincidence was 86.5 % in post-HSC transplant patients and 87.5% in SOT transplant patients. X 2 = 0.12, P [ 0.05. Conclusion: The real-time PCR assay was able to detect CMV reactivation in HSC transplant patients and SOT patients as well as pp65 antigenemia assay. Background: The reported recurrence rate of hepatocellular carcinoma (HCC) within the Milan criteria after liver transplantation is very low, and the prognosis is extremely favorable. We experienced two rare recurrences of HCC within the Milan criteria. We report these cases with a consideration of the literature. Case 1: A 47-year-old man had three HCCs each about 2.5 cm in diameter. The postoperative pathological diagnosis was one moderately differentiated HCC and a multicentric outbreak of suspected multiple well-differentiated HCC 3-4 mm in diameter. It recurred as a stalked polyp in the hypopharynx 22 months postoperatively. A polypectomy was performed, but 2 years after that procedure, it metastasized to the cervical lymph nodes. A cervical lymph node resection was performed, and he was alive without further recurrence 12 months postoperatively. Case 2: A 61-year-old woman had a one 4.0-cm-diameter HCC. All tumor makers were negative including CEA, CA19-9, PIVKA-2 and AFP. The HCC was hypovascular on dynamic computed tomography (CT) and positive on fluorodeoxyglucose-positron emission tomography/CT (FDG-PET/ CT; SUV 5.66). The postoperative pathological diagnosis was undifferentiated HCC accompanied by sarcomatoid alteration. It recurred in the retroperitoneum, mesenteric lymph nodes, and lumbar spine in early postoperative period. Conclusion: In follow of the patient, we have to be careful for not only the graft, lung, and bone, but also the head and cervical region via the portalvertebral venous system. Some transplantation centers reported that in addition to tumor size and number, tumor differentiation and pathological vascular invasion may contribute to a recurrence. But these factors are not clear preoperatively. If atypical HCC is recognized that is hypovascular, strongly positive on FDG-PET, and negative for tumor markers, it may be necessary to exclude in from transplantation within the Milan criteria. Results: Ten types of complications were diagnosed out. The incidence of 7 out of 10 complications was 60.2-100% within the first 3 months after OLT. According to the diagnostic frequency, 10 complications were divided into 3 stages. Stage I (early stage:\1 month): Mainly seen as small-for-size liver syndrome, ischemia reperfusion injury, acute cellular rejection and borderline rejection, the incidences accounting for total frequency of each lesion in this stage was 100, 97.2, 64.7 and 60.2%, respectively. Stage II (middle stage: 1-3 month): Mainly seen as vascular complications, cytomegalovirus infection and drug-induced liver injury, the incidences accounting for total frequency of each lesion in this stage was 64, 62.5 and 60.6%, respectively. Stage III (late stage: [3 month): Mainly seen as chronic rejection, primary liver disease recurrence and bile duct complications, the incidences accounting for total frequency of each lesion in this stage was 100, 91.3 and 68.5%, respectively. Percent 59.5 of central perivenulitis (CP) type ACR and 82.1% of late ACR (LACR, over 6 months after OLT) had more than twice ACR attacks, which were significantly higher than that of portal triad (PT) type ACR (30.6%, P B 0.001). Conclusion: The first 3 months after OLT is a high-incidence period of complication occurrence. CP type ACR is predisposed to repeated attack. Frequent attacks of ACR, or the first attack of ACR is CP pattern, might be two high-risk factors for LACR. The present staging provides a useful reference for determining the scope of diagnosis and differential diagnosis of OLT complications.

Yosuke Osawa 1,2 , Masahito Nagaki 2 , Yoichi Yasuda 2 , Hisataka Moriwaki 2 , Mitsuru Seishima 1 1 Department of Informative Clinical Medicine, Gifu University Graduate School of Medicine, 1-1 Yanagido Gifu Japan, 2 Department of Gastroenterology, Gifu University Graduate School of Medicine, 1-1 Yanagido Gifu Japan Background and aims: Kupffer cells, resident tissue macrophages of the liver, play a key role in the regulation of hepatic inflammation, hepatocyte death, and fibrosis that characterize liver diseases. However, it is controversial whether Kupffer cell promotes or protects from liver injury. To explore this issue, we examined the role of Kupffer cell in liver injury, cell death, regeneration, and fibrosis on cholestatic liver injury. Methods: A model of partial bile duct ligation (PBDL) was developed. The left hepatic duct of wild-type C57BL/6 male mice was ligated with 6-0 silk, and the animals were sacrificed 10 days after the surgery. Liver injury, regeneration, and fibrosis were compared between the ligated left and non-ligated right lobes. Hepatocyte apoptosis was induced by administration of D-galactosamine and TNF-alpha to the PBDL mice. The effects of Kupffer cell depletion by alendronate liposomes, or inhibition of AKT by adenovirus expressing dominant-negative AKT were investigated. Results: In the cholestatic liver injury, remaining viable cells represented tolerance for tumor necrosis factor (TNF)-alpha-induced hepatocyte apoptosis and regenerative features along with AKT activation. Inhibition of AKT abolished the survival and regenerative properties in hepatocytes. Moreover, Kupffer cell depletion increased hepatocyte damage and the sensitivity of TNF-alpha-induced hepatocyte apoptosis in ligated lobes. Kupffer cell depletion decreased hepatocyte regeneration and liver fibrosis with reduced AKT activation. Conclusion: Kupffer cell has a protective role for hepatocyte damage, and promotes cell survival, liver regeneration, and fibrosis in cholestatic liver disease. Kupffer cell is crucial for AKT activation of hepatocytes that is required for the survival and regenerative responses. Background/Aims: The incidence of hepatocellular carcinoma (HCC) is rising due to alcohol drinking, hepatitis C viral infection, and metabolic syndrome. Differential expression of CYP2E1 may play a pleiotropic role in the multistep process of liver carcinogenesis. The antitumor effect of trichostatin (TSA) as well as the apoptotic effect of CYP2E1 has received a lot of attention. However, very little is known regarding the effect of TSA on CYP2E1 gene regulation and the mechanism by which TSA induces apoptosis. The current study examines the mechanism by which TSA regulates the expression of CYP2E1 and the role of CYP2E1 in TSA-mediated apoptosis of HepG2 cells. Methods: HCC cells and normal primary human hepatocytes were treated with TSA (10 lM). Using real-time PCR, ChIP assay, siRNA and confocal microscopy, we studied the effect of TSA on CYP2E1 expression and the function of CYP2E1 in TSA-induced apoptosis of HepG2 cells.Background: Epidermal growth factor (EGF) is a kind of single-chain polypeptide consisting of 53 amino acid which widely exist in the tissues and extracellular fluid of body. Studies have found that EGF +61 polymorphism in 5 0 untranslated region (5 0 UTR) play an important role on liver including promoting the synthesis of DNA in liver, stimulating the regeneration of liver cell and protecting the liver. Now we study the EGF +61 polymorphism in cirrhosis patients and healthy people to explore the relationship between EGF +61 polymorphism and the occurrence as well as progression of cirrhosis in Chinese patients. will be withdrawn. Since NTBC withdrawn will induce liver inflammation, the role of hepatic lymphocytes in the damage is studied. Methods: We examined the progressive changes in body weight, survival rate, hepatocytes and liver inflammation when the protective drug is withdrawn. Results: After removing of NTBC, we found that the body weight of Fahdeficient mice reduced gradually and almost all the mice died finally when the body weight largely reduced (low to 70%). The survival rate of the NTBCwithdrawn mice was only 10% compared to 100% of the NTBC-protected mice. At the same time, the liver in the drug-withdrawn mice became smaller and the serum level of ALT was increased week after week. Interestingly, the frequency of T cells in the liver of NTBC-withdrawn mice increased gradually while the frequency of NK cells decreased. Furthermore, histological examination showed that the lymphocyte infiltration in the liver was more and more serious with the extension of the time after NTBC withdrawn. Conclusion: These results indicated that the inappropriate liver T cell response may be responsible for the death of NTBC-withdrawn Fah -/mice. Objective: To explore the biological effects of human hepatocyte growth factor (HGF) gene expression on CCl4 injured human hepatocyte and rat hepatic stellate cell line and its protective effect of HGF to CCl4 injured human hepatocyte line (HL-7702), and also the effect of HGF to CCl4 injured rat hepatic stellate cell line (CFSC-2G cells) apoptosis, further explore the mechanism of apoptosis produced. Methods: The proliferative activity of cells was determined by MTT method, the expression of caspase-3, -8, and -9 were determined by Western Blot method. Results: The normal human liver cells (HL-7702) transfected with PCI-neo-HGF and cultured for 48 h, the cell proliferation activity was significantly higher than the PCI-neo, liposome and non-transfected HL-7702 cells (P \ 0.01), and cell proliferation activity had increase trend with the increase of transfected dose of PCI-neo-HGF, but there did not have significant difference between different dose groups. The expression of caspase-3 protein decreased significantly in PCI-neo-HGF-transfected HL-7702 cells compared with CCl4 injured HL-7702 cells and PCI-neo-transfected HL-7702 cells. The expressions of caspase-8 and caspase-9 protein did not be detected in PCI-neo-HGF-transfected HL-7702 cells. The expression of caspase-3 protein increased in PCI-neo-HGF-transfected CFSC-2G cell compared with CCl4-induced CFSC-2G cell, and the expression of caspase-9 protein was also positive. Conclusion: The expression of PCI-neo-HGF gene can promote the growth and proliferation of HL-7702 in vitro, and inhibit the apoptosis of CCl4 injured HL-7702, and promote the apoptosis of rat CFSC-2G cell, the possible mechanism of promoting apoptosis in rat CFSC-2G cell may be due to the increase the expression of caspase-3 and -9 protein. (20, 40, 80, 160, 200 ng/ml) in the serum-free medium for 24 h. Cell proliferation was measured by Brdu incorporation analysis, un-treated WB-F344 cells were taken as control. After treatment with Wnt3a (160 ng/ml) for 24 h, the subcellular localization and protein expression of b-catenin in Wnt3a treated and untreated WB-F344 cells were examined by Immunofluorescence Staining and Objective: The hepatocyte-like cells induced from bone marrow mesenchymal stem cells (BMSCs) recover liver function in animal and human with liver failure. Our initial findings revealed that human BMSCs improved liver function in hepatitis B patients with end stage liver disease. However, the susceptibility of BMSCs to HBV infection during induction toward hepatocytes remains unknown. The aim of this study is to investigate the susceptibility of BMSCs to HBV infection during induction toward hepatocyte and the role of asialoglycoprotein receptor (ASGPR) in HBV infection. Methods: BMSC obtained from hepatitis B patients were cultured with HBV infectious serum in vitro and induced to differentiate into hepatocyte through exposure to HGF, FGF-4, and EGF. Subsequently these cells were assayed for the presence of HBeAg, HBsAg, HBV DNA and ASGPR. Results: After 6 days of exposure, BMSC-derived hepatocyte-like cells showed hepatic special genes and proteins, that were AFP, CK18, ALB and got hepatocyte functions. The BMSC was resistant to HBV infection in HBV patients and during induced to hepatocyte in vitro. ASGPR was low expressed in BMSC, but increased on the induced BMSC. Conclusion: BMSC proved resistant to HBV infection, both in vivo and during differentiation into hepatocytes in vitro. The expression of ASGPR increased during BMSC was induced to hepatocytes. ASGPR may play an important role in HBV infection. Background; Engelbreth-Swarm (EHS) gel has been reported to maintain the mature hepatocyte phenotypes in primary cultured hepatocytes. We investigated the effects of EHS gel on the differentiation of fetal liver cells, which contain stem/progenitor cells. Methods; Fetal liver cells were isolated from embryonic day 14.5 livers of mice. After deletion of hematopoietic cells, the cells were plated on either normal plastic, EHS gel coated, or type 1 collagen coated dishes and cultured for 4 days in appropreate medium. Expression of albumin and CK19 were examined by double immunofluorescent staining. Liver specific protein and mRNA expression in the cultured fetal liver cells were examined by Western blot analysis and quantitative real-time RT-PCR. Then we transplanted the fetal liver cells with or without EHS gel to BALB/c nude mice subcutaneously to investigate cell survival and functions in vivo. Result: The isolated fetal liver cells cultured on EHS gel formed a spherical shape and increased liver-specific gene expressions compared with cells cultured on normal or collagen coated dishes. The hepatic progenitor cells that were transplanted subcutanoeusly to BALB/c nude mice could survive and express hepatocyte marker alpha-hetoprotein when the cells were transplanted with EHS gel. Conclusion; These findings demonstrate that EHS gel supports cytodifferentiation from immature progenitor cells to mature hepatocytes and maintain its differentiated phenotypes in vitro and in vivo. Background: Stem cells located in bone marrow (BM) have been shown to mobilize and migrate to the liver in response to injury and thereby contributing to liver regeneration. In this study, we investigated the resident stem cell population in adult human liver and their mobilization during liver transplantation. Methods: Liver tissue samples and perfused graft preservation fluids were collected for stem cell isolation at the time of liver transplantation. Phenotypic and functional characterization was extensively investigated in vitro, in NOD/SCID mice and in transplant patients. Results: Culturing of perfusates and liver biopsies, resulted in the expansion of mesenchymal stem cells (MSC). Liver-derived MSC were equivalent to BM MSC in adipogenic and osteogenic differentiation and Wnt-stimulated proliferative responses. Moreover, the genome-wide gene expression was highly similar, with a two or more-fold difference found in only 82 of the 32,321 genes (0.25%). Liver MSC differentiation into hepatocyte-lineage was demonstrated in vitro, by acquiring the ability to support HCV infection. After engraftment in mice, a majority of MSC survived and maintained stem cell characteristics, indicating selfrenewal, and a subpopulation differentiated into hepato-like cells. Like BM MSC, liver MSC were found to be immunosuppressive shown by significant inhibition of T cell proliferation. Meanwhile, we found the mobilization of hematopoietic stem cells (HSC) during graft perfusion, which were characterized by surface marker expression and colony forming capacity in vitro and after engraftment in mice. After liver transplantation, donor HSC continue to migrate from the graft and were detectable in recipient circulation, with approx. 2-8% of peripheral blood CD34 + cells being of donor origin. Conclusions: Adult human liver contains resident population of MSC and HSC. Graft cold-storage associated with hypoxia results in mobilization of hepatic MSC and HSC, suggesting responsiveness to tissue injury. Their continuous migration after transplantation may contribute to hematopoietic chimerism and organ tolerance in patients.

Background: MSCs are characterized by self-renewal and multi-directional differentiation. Autogenetic MSCs do not cause immunological rejection and have been clinically used in France, Germany, Japan, Italy and China. We have applied autogenetic MSCs in the treatment of hepatitis B (HB) patients and satisfactory therapeutic effects were achieved. To improve the effectiveness of autoplastic transplantation and investigate the mechanism, biological characteristics of MSCs from HB patients were explored in our previous studies. This study aimed to investigate and compare differentiation of MSCs from HB patients with liver failure into hepatocytes-like cells with different induction systems in vitro. Methods: The differentiation of MSCs from HB patients with liver failure were induced in vitro into hepatocytes-like cells by three culture media [serum-free medium (group 1), auto serum-containing medium (group 2) and medium supplemented with fetal bovine serum (FBS) (group 3)]. And the cell morphology, cell growth curve, amount of urea and glycogen and mRNA expressions of ALB, CK18 and AFP were detected and compared.Result: Morphological changes in group 1 and 2 were more evident than that in group 3, and cell growth in group 3 was faster than the other two groups. The amount of urea and glycogen in group 1 and 2 was significantly elevated when compared with that in group 3 after 6 days of culture. RT-PCR analysis indicated the mRNA expressions of ALB, CK18 and AFP in group 1 and 2 were markedly increased as compared to that in group 3. Conclusion: The differentiation of MSCs from HB patients with liver failure into hepatocytes-like cells can be induced by three different cell culture media and the inductive effects were more profound in cells grown in the serum-free medium and auto serum-containing medium.Conclusion: Acute pancreatitis is an acute inflammation of the pancreatic gland, the evolution of the disease is rapid and the extend of the inflammation reaction is violence, so the level of TNFa increase highly and rapidly notably in the first 24 h of the AP; therefore the dosage of TNFa is very useful and important in the diagnosis and prognosis of AP.

Pre Hepatic and Post Hepatic Portal Hypertension: Hospital Selayang Experience Saravana Kumar Karunanathy 1,2,3 , Soek Siam Tan 1,2,3 , Ooi Keat Tan 1,2,3 , M. R. Syed 1,2,3 , A. I. Shamsul 1,2,3 , S. Sachitanandan 1,2,3 , D. Khoo 1,2,3 , Y. M. Chan 1,2,3 1 Hepatology Department, Hospital Selayang, 2 Lebuhraya Selayang-Kepong, 68100 Batu Caves, 3 Selangor, MalaysiaBackground: To assess the aetiology, mode of presentation and spectrum of patients with pre and post hepatic portal hypertension. Methods: Nine cases of pre and post hepatic portal hypertension who presented to Hospital Selayang from 2001 till May 2009 were reviewed. Results: The mean age group at presentation was 29 years of age. Four of the cases were female and five were male. Seven cases presented with upper gastrointestinal bleed due to esophageal varices, one case with ascites and one case with acute liver failure. The chart below summarizes the laboratory results of the cases on presentation. All patients had an ultrasound and a subsequent CT abdomen which revealed a non cirrhotic liver with evidence of portal hypertension. The reason for portal hypertension was portal vein thrombosis in six cases, splenic vein thrombosis in one case and Budd Chiari syndrome in two cases. The underlying aetiology for the thrombotic tendencies were protein C and S deficiencies in two cases, previous umbilical catheterization in one case, polycythaemia in one case, chronic pancreatitis in one case with the remaining four cases pending further haematological workout. Conclusion: In young patients who present with complications of portal hypertension with an essentially normal liver imaging and biochemistry, a pre or a post hepatic cause should be considered. The underlying reason for the thrombotic tendencies should be assessed as systemic diseases can present in this manner to the hepatology unit.

The Initial Screening of Epitope in HCMV gB Jun Fan 1 , Xuan Zhang 1 , Jun-Ya Shen 2 , Min-Huan Li 1 , Ya-Dan Ma 1 Background: HCMV is still an important cause of morbidity and occasional mortality in transplant recipients. Envelope glycoprotein B(gB) of HCMV plays an important role in HCMV infection and induction of immune response. gBn1 is a major pathogenic genotype,and holds an obvious quantitative advantage in HCMV infected Chinese. Methods: Based on the GenBank sequence (M60929), amino acid sequence of gB antigen peptide was selected and decided. After the peptide gB conjugated Keyhole Limpet Hemocyanin immunized New Zealand rabbits (including the initial vaccination and three times of enhanced immunization), serum of rabbits were obtained and purified. ELISA, immunoprecipitation and immunocytochemical staining were used to identify antibodies and the titer of antibody. Results: Titer of the serum could reach 1:64,000.Through ELISA assay, the serum could specificly link to gB peptide, and it could both interact with HCMV AD169 and Towne strain. Furthermore, the serum could interact with HCMV AD 169 and Towne strain with immunoprecipitation and Immunofluorescence technique. With the serum, some plasma from Chinese could be verified that it infect HCMV in immunocytochemical staining. Conclusions: The synthetical peptide has a favourable immunogenicity. In the future, more research would be dedicated to screening epitope in the synthetical peptide, which could help us to know HCMV, understand neutralizing antibody of HCMV, prepare HCMV vaccine and antivirus drug.

Bleeding Control in the Laparoscopic Partial Liver Resection Special Reference to Clinicopathological Examination about RFA Precoagulation Fumiki Kushihata 1 , Jota Watanabe 1 , Tetsuya Mizumoto 1 , Yoshikuni Yonenaga 1 , Akifumi Miyoshi 1 , Taiji Toyama 1 , Kazuo Honda 1 , Yasutsugu Takada 1 1 Surgery, Ehime University, Toon city, Ehime prefecture, JapanBackground: Laparoscopic partial hepatectomy (LPH) for liver tumors is the most common procedure in laparoscopic liver resection. But owing to its easily bleeding tendency, it is very important to regulate the control of it during hepatic resection. We have done mainly with RFA precoagulation, but there is little fundamental examination. Purpose: We perform a clinicopathological examination of a RFA precoagulation and examine an optimum precoagulation method for LPH. Object and method: LPH enforcement 12 examples in our department concerned. RFA using cool tip TM needle and, for in principle left liver 60 W/ 1 min, performed it for right liver 80 W/1 min. We examined cauterization depth by the pathological examination of the sample, weight, bleeding, postoperative liver function, CRP and postoperative complications including bile leakage. Result: The left liver was cauterized in depth than the right liver, and there were few amounts of bleeding. We did not recognize the sample weight and the cauterization depth and the relevance with the postoperative liver functional impairment and complication. Conclusion: RFA precoagulation is feasible and easy to dissect the hepatic vein and parenchyma. We may simplify the RFA precoagulation of left liver LPH.

Is There a Benefit of Anticoagulant Therapy in the Survival? Filiz Akyuz 1 , Yilmaz Cakaloglu 1 , Binnur Pinarbasi 1 , Kadir Demir 1 , Sadakat Ozdil 1 , Fatih Besisik 1 , Gungor Boztas 1 , Zeynel Mungan 1 , Sabahattin Kaymakoglu 1 1 Istanbul Medical Faculty, Department of Gastroenterohepatology, Capa, Aim: To determine the factors that effect survival and prognosis especially the anticoagulant treatments' role in the patients diagnosed with BCS. Material/Methods: Between 1995 and 2004, 30 patients (60% female) diagnosed with BCS based on the clinical, biochemical, imaging and histological findings were evaluated retrospectively. Clinical status, etiologic factors, laboratory and imaging findings at the admission and development of complications and survival during the follow-up period were determined. Results: Mean ages of patients were 36 ± 12 years [Median diagnosis period 5 months, mean follow-up time 48 ± 40 months (6-120)]. 7% of patients were diagnosed acute, 40% subacute and 53% chronic BCS according to disease duration. Centrilobular necrosis was found in 9 of 17 (56.7%) biopsy performed patients. Etiologic distribution were as follows: 5 APS, 6 factor V Leiden mutation (3 heterozygote), 5 myeloproliferative disease, 2 IVC stenosis, 1 tumour infiltration, 2 protein C and 1 protein S deficiency, 1 PNH and 1 Behcet disease. There were multiple etiologic factors in 5, where etiologic factors were not detected in 37% of patients. Twelve of them received anticoagulant therapy, whereas 16 were followed-up without treatment. Mesoatrial shunt surgery was performed in four cases; only one patient had Denver shunt operation. Five patients died after mean 14.5 ± 9 months follow-up period. Although, there was a positive correlation between survival and centrilobular necrosis (r = 0.683, P = 0.014), negative correlation was observed between albumin levels and survival (r = -0.0586, P = 0.008). These factors were not effective over survival in Cox regression analysis. The mean survival periods were 94.5 months (%95 CI 63-125 months) and 96.4 months (%95 CI 72-120 months) in anticoagulant treated and untreated patients, respectively (log rank analysis P [ 0.05). Conclusion: Most of patients with BCS apply to hospital in subacute and chronic stage and approximately half of them have underlying thrombotic risk factor. In our study, no beneficial effect of anticoagulant therapy was observed on the survival. Aim: The ''Metroticket'' prognostic model for survival post liver transplant for HCC was developed by (Lancet 2009) from a European cohort of predominantly alcoholic liver disease and hepatitis C-related HCC. The aim of this study was to evaluate the prognostic value of this Metroticket model in an independent cohort of predominantly HBV-related HCC in an Asia-Pacific transplant programme. Methods: All patients listed for HCC at NZLTU between January 1998 and December 2008 were included. For each patient, a Metroticket predicted 3 and 5 year survival score was calculated using the pre-transplant measurements of tumour size and number (http://www.hcc-olt-metroticket.org/calculator/ index.php). The primary outcome was the comparison between observed 3 and 5 year survival with predicted survival calculated by the Metroticket model incorporating pre-transplant radiological measurements of tumour size and number. The secondary outcome was the comparison with predicted survival calculated by the Metroticket model incorporating explant pathological measurements of tumour size and number. Results: 84 patients were listed for HCC-71 were transplanted and 13 delisted for disease progression. Another nine patients had incidental HCC in the explant. On intent to treat analysis, survival after listing was 72% (62-82%) at 3 years and 68% (58-80%) at 5 years. Predicted post-transplant survival calculated by the Metroticket model incorporating pretransplant radiological data (n = 71) was 76% at 3 years and 70% at 5 years. Observed post-transplant survival was 83% (49/59) at 3 years and 74% (35/47) at 5 years. Actuarial 3 and