key: cord-0034120-iqdi99pd
authors: nan
title: 25th Annual Meeting of the Austrian Society of Transplantation, Transfusion and Genetics Graz, October 19–21, 2011
date: 2011
journal: Eur Surg
DOI: 10.1007/s10353-011-0041-z
sha: 5fcaa5b826f3255958984041b5e25f3549b12ae7
doc_id: 34120
cord_uid: iqdi99pd

nan

Grundlagen. Parvovirus B19 (PVB19) als Ursache einer schweren aplastischen Anämie (SAA) oder eines myelodysplastischen Syndroms (MDS) ist bisher erst selten beschrieben. Es existieren auch kaum Berichte über Stammzell-Transplantationen (SCT) bei diesen Krankheitsbildern.

Methodik. Bei 6 Patienten mit SAA (n ¼ 4) oder MDS (n ¼ 2) wurde bei Diagnosestellung eine frische Infektion mit PVB19 mittels positivem Nachweis von IgM, IgG und PCR nachgewiesen. Alle Patienten erhielten eine SCT unter dem Schutz repetitiver intravenöser Immunglobulin (IVIG)-Gaben. Das virologische Monitoring mittels PCR erfolgte in 5 von 6 Fällen in der Akutphase wöchentlich, danach monatlich bis zur hämatologischen Rekonstituierung.

Ergebnisse. Die 4 Patienten mit SAA erhielten Knochenmark eines HLA-identen Geschwister-Spenders (n ¼ 3) oder HLA-idente hochgereinigte, periphere Stammzellen eines unverwandten Spenders (n ¼ 1). Ein Patient entwickelte nach SCT eine chronische, hypoplastische Anämie, die bis zur vollständigen Blutbild-Regeneration am Tag 186 andauerte. Der Posttransplantations-Verlauf der ü brigen 3 Patienten war komplikationslos mit einem Leukozyten-Engraftment nach 8-31 Tagen und einer hämatologischen Rekonstituierung nach 36-112 Tagen. Die 2 Patienten mit MDS wurden nach Konditionierung mit Thiotepa, Fludarabin und ATG einer SCT mit unverwandten Spendern (1x Knochenmark, 1x periphere hochgereinigte Stammzellen) unterzogen. Nach zögerlichem Leukozyten-Engraftment dauerte es 1260 bzw. 119 Tage bis zur Blutbild-Normalisierung, PVB19 wurde in der PCR bis zum Tag 686 bzw. 139 nachgewiesen.

Schlussfolgerungen. Eine erfolgreiche SCT bei diesen Patienten ist möglich, sofern eine engmaschige Monitorisierung der PVB19-PCR und eine konsequente repetitive IVIG-Therapie erfolgen. Der Transplantationsverlauf kann durch ein zögerliches Engraftment und eine prolongierte "poor graft function" kompliziert werden.

Retrospective study to test ferritin serum levels as biomarker for graft-versus-host disease-associated non-infectious inflammatory reaction in 117 children after hematopoietic stem cell transplantation Background. Myelodysplastic syndromes (MDS) are a heterogeneous group of stem cell disorders characterized by bone marrow dysplasia, peripheral cytopenia, and an enhanced risk to transform to acute myeloid leukemia (AML). In most patients, treatment options are limited to supportive care and palliative cytoreduction. However, in a group of patients, intensive therapy can be offered. The only established curative treatment approach for these patients is haematopoietic stem cell transplantation (SCT) .

Methods. In the present study, we retrospectively analyzed a cohort of 60 adult patients (33 males, 27 females) with MDS (n ¼ 28) or MDS transforming into secondary AML (n ¼ 33), who underwent SCT at our institution between 1988 and 2010. Fifty-one patients had an HLA-identical related transplant donor, and 9 had an HLA-matched unrelated donor. The median age at time of SCT was 44 years (range: 18 to 68 years). According to the WHO classification, 4 patients had RA, 1 RARS, 3 RCMD, 1 RCMD-RS, 6 RAEB-1, 12 RAEB-2, 1 CMML, and 32 had AML following RAEB at SCT. Conditioning consisted of chemotherapy plus total body irradiation (55/60 patients) or chemotherapy alone (5/60 patients). Graft versus host disease (GvHD) prophylaxis consisted of a combination of low-dose methotrexate and cyclosporine A (37/60 patients) or cyclosporine A plus mycophenolat mofetil (23/60 patients).

Results. Patients were followed up with a median observation time of 16 months (range: 1-218). Currently, 34 patients (57%) are alive. Of the 26 patients who died, post-transplantation relapse occurred in 12 patients, and 14 patients died of treatment-related causes (multi-organ failure, n ¼ 7; sepsis, n ¼ 4; haemorrhage, n ¼ 3). In multivariate analysis we identified pre-transplantation ferritin as a significant adverse prognostic variable for survival in our MDS patients. In contrast, the overall outcome after SCT was independent of IPSS risk categories or the WHO classification.

Conclusions. Based on these data and similar published data we recommend to select patients with MDS or sAML for SCT who are considered candidates for SCT according to the presence of pre-transplantation iron-overload.

Significantly worse survival of patients with chronic graft-versus-host disease (cGVHD) defined according to the National Institutes of Health (NIH) consensus criteria and thrombocytopenia data and expert opinion. For both acute as well as cGVHD new definitions were proposed. We performed a prospective study on all consecutive patients undergoing allogeneic hematopoietic stem cell transplantation (HCT) since 2005 to assess the prognostic impact of the new cGVHD staging criteria.

Methods. One hundred seventy-eight patients (85 males, 93 females) with a median age of 40 years alive on day þ 100 after HCT with myeloablative (n ¼ 110) or reduced-intensity (n ¼ 68) conditioning and a related (n ¼ 37) or unrelated (n ¼ 141) stem cell donor were enrolled into the study. Starting on day þ 100 after HCT all patients were assessed clinically every 3 months in the outpatient clinic for cGVHD activity according to the NIH consensus criteria (Filipovich AH et al, BBMT 2005; 11:945-956) .

Results. One hundred twenty-six (71%) patients experienced acute GVHD grades I to IV including 11 with recurrent, 11 with persistent, and 10 with late-onset acute GVHD after a median of 18 (range, 9-120) days after HCT. One hundred-fifteen patients (65%) experienced cGVHD after a median of 151 (range, 82-510) days after HCT. Eighty-nine patients (77%) had classic cGVHD and 26 (23%) overlap syndrome. Probability of overall survival (OS) at 3 years for late-acute GVHD, chronic classic and overlap cGVHD were 69%, 83%, and 73%. Three-year OS for mild, moderate and severe cGVHD at onset were 93%, 79%, and 62.5% and significantly different between mild and severe (p ¼ 0.007). Patients with progressive onset type of cGVHD had significantly worse three-year survival compared to de novo and quiescent (54.5% vs. 89.5%, 84%, p < 0.01). Three-year OS was also significantly worse in patients with platelet counts below 100 G/l at onset of cGVHD (35% vs. 86%, p < 0.0001).

Conclusions. This prospective analysis supports the importance of distinguishing late-acute GVHD from cGVHD and indicates a prognostic value of thrombocytopenia and progressive onset type of cGVHD for HCT outcomes. Current concepts suggest a major role for innate immunity in the initiation of acute graft-vs.-host-disease (aGvHD) as well as in Crohn's disease. Identical single nucleotide polymorphisms (SNP) have been described as risk factors for both diseases. Recently, the noncoding C/T polymorphism rs2274910 in intron 3 of the intelectin 1 (ITLN1) gene (human lactoferrin receptor) has been associated with Crohn's disease. We retrospectively typed this polymorphism in AML patients and their donors. A pilot study of 19 patients in CR1 transplanted from HLA identical sibling donors after myeloablative conditioning was confirmed in a second cohort including 40 other AML patients. A total of 59 consecutive patients (median age 43 yrs (18-63); 25/34 CR1/advanced disease; 40 myeloablative and 19 reduced intensity conditioning; 54 peripheral stem cells) were tested. T-alleles were found at a frequency of 28.8% in recipients and 29.7% in donors. 50.8% of patients had a CC genotype. In the pilot study, 2 out of 12 patients with a CC genotype versus 5 out of 7 patients with a T-allele had acute intestinal aGvHD (p ¼ 0.045). These results were confirmed in a second cohort. Acute intestinal GvHD was found in 3/18 patients (16.7%) with a CC genotype and 12/ 22 patients (54.5%) with a T-allele (p ¼ 0.014). In a combined analysis (n ¼ 59) we found intestinal aGvHD in 58.6% of patients with a T-allele versus 16.7% of patients with a CC genotype (p < 0.001). The lower incidence of aGvHD grades II-IV in patients with a CC genotype (p ¼ 0.019) was only due to less intestinal aGvHD while no difference was found with regard to skin or liver aGvHD. We did not see any association between donor genotypes and aGvHD. The strong association (RR 3.52; 95% CI 1.5-8.2) between this SNP and the incidence of intestinal aGvHD is in accordance with reports about SNPs in other genes associated with both aGvHD and Crohn's disease. Our results further support the concept of the importance of the gut associated innate immune system in the initiation of aGvHD. Grundlagen. Klinische Studien zeigen, dass sich Minorhistokompatibilitätsantigen (mHag)-spezifische Spender T-Zellen positiv in der Therapie eines leukämischen Relapses nach HLAidenter hämatopoetischer Stammzelltransplantation (SZT) auswirken. Der prädiktive Wert der mHag-Typisierung für das Auftreten eines Graft-versus-Leukämie Effektes oder von Komplikationen nach SZT ist jedoch nach wie vor umstritten. Auch Langzeitstudien über den Einfluss von mHag-Mismatches auf das Ü berleben der Patienten fehlen. Deshalb haben wir in der vorliegenden Studie Unterschiede in zehn verschiedenen mHag in insgesamt 217 Patienten und ihren HLA-identen Familienoder Fremdspendern untersucht und sie mit Ü berleben und Komplikationen nach allogener SZT korreliert.

Methodik. Die Bestimmung der mHag wurde mittels Allelspezifischer Polymerase-Kettenreaktion mit Hilfe eines kommerziell erhältlichen mHag-Typisierungskits durchgeführt. Die Wahrscheinlichkeit des Ü berlebens (bis zu 22 Jahre nach SZT) wurde mittels Kaplan-Meier Ü berlebensanalyse, die Komplikationsraten mittels Cumulativer Inzidenz berechnet.

Ergebnisse. Von den 217 Patienten/Spenderkombinationen wiesen 119 keinen mHag-Mismatch, 67 einen mHag-Mismatch und 31 zwei oder mehrere mHag-Mismatches auf. Sämtliche Patienten mit einem HA-1-, HA-2-, HA-8-(HLA-A2-restringiert) oder HA-3-(HLA-A1-restringiert) Mismatch zeigten eine geringere Rezidivrate als Patienten ohne den entsprechenden mHag-Mismatch. Ein HA-2-und ein HA-3-Mismatch waren jedoch mit einer erhöhten Inzidenz an akuter GvHD und daher mit einer geringeren Ü berlebensrate verbunden. Patienten/Spenderkombinationen mit einem HY-Mismatch profitierten mit einem leichten Ü berlebensvorteil (48 % vs. 40 %). Interessanterweise wirkten sich mehrere mHag-Mismatches positiv auf das Langzeitüberleben aus. Vier Jahre nach SZT betrug das Gesamtüberleben von Patienten ohne Mismatch 44 %, von Patienten mit einem Mismatch 43 %, von Patienten mit 2 Mismatches 55 % und von Patienten mit 3 Mismatches 75 % (allerdings nur 8 Patienten). In Ü bereinstimmung mit diesem Ergebnis betrug die Inzidenz der Nonrelapse-Mortalität von Patienten ohne Mismatch 50 %, von Patienten mit einem Mismatch 54 % und von Patienten mit zwei bzw. drei Mismatches 36 % bzw. 14 %. Eine Erklärung für dieses Ergebnis könnte der Graft-versus-Leukämie Effekt sein, der durch die erhöhte Anzahl an Spender T-Zellen induziert wird, die verschiedene hämatopoetisch-restringierte mHag auf den residuellen Leukämiezellen des Patienten erkennen und diese zerstören.

Schlussfolgerungen. Die Bestimmung von mHag in Patient und Spender vor der SZT wäre eine sinnvolle Maßnahme zur zusätzlichen Risikoabschätzung von Komplikationen, vor allem aber Ü berleben nach SZT.

"Stuhltransplantation" als erfolgreiche Therapie bei schwerer idiopathischer Enterocolitis Grundlagen. Die normale humane Darmflora umfasst etwa 100 Billionen Bakterien, von denen der Großteil mit konventionellen Methoden nicht kultivierbar ist. Störungen in der Zusammensetzung der Darmflora scheinen bei der Entstehung von Darmentzündungen und Infektionen eine wichtige Rolle zu spielen. Die fäkale Bakterientherapie (Stuhltransplantation) zielt auf die Wiederherstellung einer gestörten Darmflora durch Ü bertragung von Stuhl eines Gesunden in den Darm eines Erkrankten. Diese Therapie wird erfolgreich bei therapierefraktären Clostridium difficile Infektionen eingesetzt.

Fallbericht und Methodik. Eine 16-jährige Patientin wurde aufgrund eines Schädelhirntraumas intensivmedizinisch behandelt. Unter einer mehrfachen, antibiotischen Therapie in Kombination mit Steroiden entwickelte die Patientin eine schwere Enterocolitis mit ausgeprägter Diarrhoe (bis zu 7 Litern Stuhl pro Tag). Die histologischen und endoskopischen Befunde der Darmentzündung zeigte das Bild einer Graft versus Host Disease (GvHD) artigen Entzündung. Eine Abklärung auf mögliche Ursachen insbesondere auf infektiöse Erreger blieb negativ. Im Verlauf nach 8 Wochen kam es zu einer spontanen Besserung der Entzündung im Dünndarm, im Dickdarm persistierte jedoch die schwere Colitis. In den histologischen Proben aus dem Colon zeigte sich ein fast kompletter Verlust des Epithels. Als Ursache der Colitis wurde eine Störung der Darmflora durch die antibiotische Therapie angenommen und deshalb bei der Patientin eine Stuhltransplantation durchgeführt. Als Stuhlspender wurde die gesunde Mutter der Patientin herangezogen, diese wurde zuvor auf übertragbare Infektionskrankheiten getestet. Der Stuhl wurde mit Kochsalzlösung verflüssigt und filtriert. Im Anschluss wurde bei der Patientin eine Coloskopie durchgeführt und insgesamt 400 ml des flüssigen Spenderstuhls in das Ileum und Colon appliziert.

Ergebnisse. Die Stuhltransplantation führte bei der Patientin in der endoskopischen Kontrolle nach 1 Woche zu einer rapiden Besserung vor allem des histologischen Befundes. Ebenso kam es zu einem schrittweisen Rückgang des Stuhlvolumens mit Normalisierung der Stuhlfrequenz nach 6 Wochen. In der Kontrolle nach 2 Monaten war der endoskopische und histologische Befund im Colon normal. Im weiteren Beobachtungszeitraum nach 5 Monaten ist es zu keiner neuerlichen Reaktivierung der idiopathischen Enterocolitis gekommen.

Schlussfolgerungen. Die GvHD-artige idiopathische Enterocolitis nach Antibiotikatherapie scheint durch eine persistierende Störung der Darmflora mitbedingt zu sein. Die Stuhltransplantation war bei der beschriebenen Patientin mit diesem schweren Krankheitsbild eine sehr effektive Therapiemaßnahme.

Tissue specific epigenetic memory: differentiation capacity of mesenchymal stem cell is restricted to their tissue of origin Human mesenchymal stem and progenitor cells (MSPCs) are currently evaluated in clinical trials for bone and marrow regeneration and their immune modulation potential. MSPCs from virtually all tissues appear indistinguishable regarding immunephenotype and multipotent differentiation capacity in vitro. Improvements of limited clinical efficiency are hampered by a lack of understanding MSPC functionality in vivo. Here we demonstrate that the capacity of in vivo endochondral bone formation followed by infiltration of hematopoietic components can be used as a surrogate to determine in vivo MSPC-multipotentiality. MSPCs from bone marrow (BM), adipose tissue (AT) and umbilical cord (UC) have been isolated. Comparative analysis of immune-phenotype, adipo-, chondro-and osteogenic differentiation potential in vitro as well as expression analysis of key mesenchymal lineage genes were performed. Epigenetic profiling of MSPCs was done using a methylation array. In vivo differentiation capacity was tested by transplanting MSPCs subcutaneously into immune-compromised mice. The developmental sequence of chondrogenic and osteogenic as compared to perivascular mesenchymal tissue formation was analyzed. Formation of a marrow niche with establishment of the complete host hematopoiesis was studied. Secondary transplants of MSPCs were performed and analyzed equally. MSPCs from all tissues analyzed show an almost identical immune-phenotype using a MSPC marker profile. Osteo-and adipogenic differentiation potential in vitro as well as gene expression can not distinguish tissuespecific MSPCs. MSPCs from all tissues except BM lack in vitro chondrogenic differentiation potential using stringent 3D-chon-drogenesis assays followed by appropriate histological staining. In vivo studies could strengthen these findings, because only BMderived MSPCs were able to generate bone through an endochondral ossification process with subsequent invasion of mouse marrow. These results correlate with the epigenetic status of the cells since BM-derived MSPCs cluster separately in principalcomponent-analysis (PCA), whereas MSPCs from all other tissues cluster together. BM was the only tissue containing MSPCs with multipotent differentiation capacity. This is reflected by a BMspecific epigenetic profile that differs from all other tissues analyzed. Since cartilage formation is an essential developmental process important for bone generation and hematopoiesis attraction, an epigenetic predisposition of BM-MSPCs to undergo endochondral ossification makes these cells unique for bone and marrow regeneration purposes. Grundlagen. Die Hemmung des Renin-Angiotensin-Systems (RAS) entweder mit ACE-Hemmern oder mit Angiotensin-Rezeptor-Blockern (ARB) verlangsamt die Progression unterschiedlicher chronischer Nierenerkrankungen und der chronischen Transplantatdysfunktion. RAS-Blockierung kann auch durch eine direkte Renin-Inhibierung erreicht werden, die aber infolge einer verminderten Produktion der protektiven Angiotensin II-Spaltprodukte wie z. B. Angiotensin (Ang)(1-7) zusätzliche Effekte haben kann.

Methodik. Im Fischer-Lewis Nierentransplantationsmodell der Ratte wurde der Effekt von Aliskiren (10 mg/kg/Tag) auf die Entstehung der chronischen Transplantatdysfunktion im Vergleich zu einer Behandlung mit Vehikel oder mit dem ARB Candesartan als bekannter Hemmer der chronischen Transplantatdysfunktion untersucht. Analysiert wurden die Histologie der Niere und der Verlauf der Proteinurie, der Serum-Spiegel von Ang (1-7) und der Spiegel von Angiotensinogen im Harn als Indikator für die intrarenale RAS-Aktivität.

Ergebnisse. Im Gegensatz zu Candesartan verminderte Aliskiren weder die klinischen (Proteinurie, Kreatinin-Clearence) noch die histologischen Zeichen (Glomerulosclerose, interstitielle Fibrose, Makrophagen-Infiltration) der chronischen Transplantatdysfunktion. Candesartan verbesserte im Vergleich zu Vehikel-und Aliskiren-behandelten Gruppen sowohl die Proteinurie als auch den Histologieschaden. Die mit Aliskiren behandelten Ratten zeigten einen verminderten Serum-Spiegel des protektiven Ang(1-7) und einen höheren Harn-Angiotensinogen-Spiegel im Vergleich zu den mit Candesartan behandelten Tieren.

Schlussfolgerungen. Der Renin-Blocker Aliskiren verminderte das Fortschreiten der chronischen Transplantatdysfunktion nicht. Der fehlende Schutzeffekt hängt wahrscheinlich entweder mit der verminderten Produktion des protektiven Ang(1-7) oder mit einer ineffektiven intrarenalen RAS-Blockierung zusammen.

Kein Unterschied im Gehalt an energiereichen Phosphaten in Organen von Lebendspendern, hirntoten Spendern und "non-heartbeating" Spendern im Tiermodell Schlussfolgerungen. Zusammenfassend sind unterschiedliche miRNA-Signaturen mit zellulärer oder humoraler Abstoßung sowie DGF assoziiert. Die identifizierten miRNAs und Target-Gene werden neue Sichtweisen auf die molekulare Regulation von Transplantatabstoßung und Transplantatversagen erlauben und so den Weg für neue Therapieansätze ebnen. Darüber hinaus könnten die identifizierten miRNAs und Target-Proteine in Zukunft als neue diagnostische Marker eingesetzt werden.

Correlation of recipient factors with the course of lymphocytes after alemtuzumab induction in renal transplantation Background. Alemtuzumab, an anti-CD52 T-cell and B-cell depleting monoclonal antibody, used as induction therapy in renal transplantation (KTs). The recovery of lymphocytes after alemtuzumab induction has been investigated in a number of trials, however, the clinical course after KTx has not been correlated with lymphocyte recovery. Herein, we correlate the outcome as well as recipient factors with lymphocyte recovery after induction with alemtuzumab.

Methods. Single center retrospective analysis of 225 patients/consecutive kidney transplantations between 01/ 2004 and 12/2010 which received alemtuzumab as induction agent (one dose of 30mg). Patients were devided into 4 groups according to lymphocyte recovery at 4 points of time (pre-Tx, 1-3 weeks post-Tx, 3 weeks-3 months post-Tx and 3-6 months after KTx). The relevance of recipient-characteristics was analyzed. Delayed kidney graft function (DGF) was defined as requirement for more than one dialysis within the first week after KTx. Statistical analysis was performed with SPSS 17.0 software (SPSS Inc., Chicago, IL, USA). Analysis of variance for repeated measurements with measurement time as withinsubject factor and with age, CMV status, DGF status as between subject factors were performed.

Results. Median age of all recipients was 49.63 years, 65.33% were male. Mean lymphocyte counts were 22.8 AE 9.41% pre-Tx, 2.61 AE 3.11% between week 1 and 3, 6.98 AE 6.7% between 3 weeks and 3 months after Tx and 18.20 AE 11.48% at later time points. Among all factors analyzed, DGF, CMV status and age showed a significant correlation with lymphocyte counts. DGF occurred in 27.56% of the recipients. The lymphocyte-counts in the DGFgroup were significantly higher, 10.7% vs. 13.13% (p ¼ 0.036) post-Tx. CMV-status of the recipient influences the quantity of lymphocytes pre-Tx significantly (p ¼ 0.009). Age showed a significantly influence on the lymphocyte count 3 months post-Tx (p ¼ 0.032).

Conclusions. CMV-status and age have a significant impact on lymphocyte recovery after alemtuzumab induction therapy. Lymphocyte counts early after transplantation represent a prognostic factor for kidney function early after transplantation. A detailed analysis of phenotype and function of lymphocytes after alemtuzumab induction together with a correlation with the clinical course is warranted.

Belatacept-based immunosuppression in de novo liver transplant recipients: 1-year experience from a phase II study Background. Calcineurin inhibitors contribute to substantial toxicities in liver transplant (LT) patients that can lead to renal dysfunction/failure and cardiovascular (CV) disease. This phase II study evaluated belatacept versus tacrolimus (Tac) in de novo LT recipients.

Methods. 250 LT recipients were randomized to belataceptbased (high dose [HD] or low dose [LD]) or Tac-based regimens. All patients received steroids for the first 3 months.

Results. Demographic characteristics were similar among groups; 46% of patients were HCV positive. The primary endpoint (composite of acute rejection [AR], graft loss [GL] , and death by month 6) occurred more frequently in belatacept groups. By year 1, more deaths and GL occurred with belatacept LD vs. other groups. Causes of early (by 6 weeks) death and GL were mostly due to postoperative complications; thereafter causes included sepsis, PTLD, and multi-organ failure. AR was more common with belatacept-based regimens compared to Tac þ MMF. By 1 year, mean cGFR was 15-34 mL/min higher in belatacept vs. Tac groups. Lower blood pressure and less neurotoxicity were observed with belatacept treatment. 2 PTLD cases and 1 PML case (HD group) occurred in the belatacept groups.

Conclusions. Belatacept HD group had comparable efficacy relative to Tac alone but was less effective compared to Tac þ MMF. Belatacept LD group was less effective with more deaths and GL observed compared to both Tac-based groups. Belatacept provided improved renal function with less CV/metabolic and neurotoxicities vs. Tac-based regimens. PTLD and PML were observed with belatacept. The optimal dose/regimen of belatacept in LT is yet to be determined.

CMV late phase-induced mTOR activation is essential for efficient virus replication in human M2-polarized macrophages Human Cytomegalovirus (CMV) remains one of the most important pathogens following solid-organ transplantation, potentially leading to CMV disease, allograft dysfunction, acute and chronic rejection and opportunistic infections. Mounting evidence indicates that mammalian target of rapamycin (mTOR) inhibitors may decrease the incidence of CMV infection in renal transplant recipients. Here we aimed to elucidate the molecular mechanisms of this effect by employing a human CMV (HCMV) infection model in human macrophages, since myeloid cells are the principal in vivo targets of HCMV and the major viral source during early CMV disease. Here we demonstrate that polarization of macrophages into M1 and M2 macrophages resulted in highly divergent host cell permissiveness for HCMV with optimal infection susceptibility in M2 versus M1 macrophages. Employing an ultra-high purified HCMV stock (TB-40/E) selective rapamycinindependent induction of IFN-transcripts, but no proinflammatory cytokines or early signalling events including MAPK and mTOR signalling could be detected. Assessment of viral gene expression and mTORC1 activity during the course of macrophage infection revealed that rapamycin significantly suppressed CMV replication 3 and 5 days post infection, while CMV proliferation in fibroblasts was largely unaffected by mTOR-blockade. Analysis of mTORC1 activation and late phase viral proteins including pUL-44 and pp65 signified an exquisite mTORC1 dependency of protein synthesis during the late phases of viral replication. Collectively, these data indicate that mTORC1 is essential for virus replication during late phases of the viral cycle in myeloid cells which might explain the potent anti-CMV effects of mTOR inhibitors after organ transplantation.

Background. Brain death (BD) triggers inflammatory signals and graft injury. Clinically, reduced organ quality as a result of brain death contributes to inferior graft and patient survival.

Regulatory T-cells are primarily known for their control of the immune response. We analyzed CD25þ/FoxP3þ T-cell subpopulations in recipients of either brain dead and native donor hearts. To dissect the role of donor immune competent cells in communicating inflammatory signals following BD we performed syngeneic cardiac transplants from wildtype and immune deficient donors.

Methods. Hearts from brain dead wild type (BD/WT) C57BL6 mice and Rag2/double knockout (BD/KO) mice were procured 3 hrs following brain death induction and transplanted into WT recipients. Hearts originating from native C57BL6 mice served as controls. By 3 days, T-lymphocyte subpopulations (CD4, CD8, CD4/CD25/FoxP3) were assessed by flow cytometry and graft-specific changes were assessed by IHC and RT-PCR.

Results. Blood pressure remained stable following BD induction and intragraft cell infiltration was comparable in all groups (p ¼ n.s.). Cardiac isografts from BD/WT donors demonstrated pronounced cellular infiltrates; only few cells infiltrated hearts from living WT donors (p ¼ 0.025). Of note, CD4/CD25/FoxP3 regulatory T-cells counts were significantly elevated in recipients of BD/WT grafts (p ¼ 0.04). Recipients of BD/KO hearts featured reduced CD4 þ T-cell infiltrates, but comparable FoxP3 levels (p ¼ n.s.). In contrast, BD/KO heart recipients showed significantly lower IL-6, IFN-, and TNF--levels (p ¼ 0.03, 0.005, 0.01).

Conclusions. Inflammatory events after brain death are counterbalanced by significantly elevated rates of CD4/CD25/ FoxP3 reg. T-cells. Less pronounced consequences of brain death in hearts originating from immune deficient donors suggest a critical role of immune competent cells in communicating inflammatory signals.

Lipocalin-2 as a therapeutic agent in chemotaxis during ischemia and reperfusion injury following solid organ transplantation Background. Neutrophil gelatinase-associated lipocalin (NGAL/Lcn-2) expression is associated with ischemia/reperfusion injury (IRI) following transplantation and correlates with polymorphonuclear cell infiltration. To get insight into the regulatory role of Lcn-2 during IRI the expression of different chemokines and adhesion molecules were analyzed in a murine heart transplantation model.

Methods. The murine heterotopic heart transplant model also implying exogenous i.p. application of Lcn-2 was used for in vivo experiments. The mRNA expression of the chemokines MIP-2, LIX, KC, MCP-1, IL-6 and CCL-6 and their receptors CXCR2 and CCR2 as well as ICAM-1 was analyzed by qPCR. Immunohistochemistry was performed in heart sections and correlated with neutrophil infiltration at various time points (2, 12, 24 and 48 h).

Results. Significant lower granulocyte infiltration and serum creatinine kinase levels during IRI were observed in the Lcn-2-/transplants correlating with a stable ICAM-1 expression compared to the Lcn-2 wt setting (>5fold expression at 2 h of reperfusion). In the early phase of reperfusion (2 h) MCP-1, KC, LIX and MIP-2 showed a lower expression pattern in the Lcn-2-/transplants with delayed upregulation at 12 h (LIX, MIP-2). After i.p. Lcn-2 application no significant difference in apoptosis was observed. The number of infiltrating granulocytes was reduced after application of the iron-loaded Lcn-2 compared to iron-free Lcn-2.

Conclusions. Our data point to a possible chemotactic role of Lcn-2 which may also affect the expression of particular chemokines in the early phase of IRI. The role of the iron binding capacity of Lcn-2 in chemotaxis during IRI is still unknown. Understanding these regulatory mechanisms will be crucial to establish treatment strategies for IRI during solid organ transplantation.

Glomeruläre Effekte des mTOR-Inhibitors Rapamycin in der nephrotoxischen Serumnephritis

Grundlagen. Der mTOR Inihibitor Rapamycin ist ein Immunosuppressivum, welches klinische Anwendung zur Prävention von Abstossungsreaktionen bei nierentransplantierten Patienten gefunden hat. Einer der limitierenden Faktoren bei der klinischen Anwendung ist das de novo Auftreten von Proteinurie. Ziel der Arbeit war es, die glomerulären Veränderungen der Rapamycininduzierten Proteinurie in einem experimentellen Modell der nephrotoxischen Serumnephritis (NTS) näher zu untersuchen.

Methodik. NTS wurde in C57BL/6 Mäusen induziert und die Gabe von Rapamycin in einer Dosis von 0,5 mg/kg 14 Tage nach Induktion begonnen. Die Mäuse wurden am Tag 35 sakrifiziert und Glomeruli zur Genexpressionsanalyse isoliert. Außerdem wurde mittels Immunhistochemie die glomeruläre Expression von T-Zell-(CD4), Makrophagen-(F4/80, CD68), Podocyten-(Nephrin), sowie Endothelzellmarkern (CD31) untersucht.

Ergebnisse. Mäuse, die mit Rapamycin ab Tag 14 behandelt wurden, entwickelten eine signifikant erhöhte Proteinurie im Vergleich zu den Kontrolltieren. Dies war mit einer verstärkten glomerulären Infiltration von CD4-positiven (mittlere intraglomeruläre positive Zellen/50 Glomeruli 4,43 AE 0,68 vs 2,23 AE 0,57; p < 0,05), F4/80-positiven (8,2 AE 1,58 vs. 3,64 AE 0,74; p < 0,05) und CD68-positiven Zellen (18,80 AE 3,05 vs. 9,15AE 1,61; p < 0,05; n ¼ 13 Rapamycin, n ¼ 12 Vehikel) assoziiert. Keine Unterschiede fanden sich in der glomerulären Färbung von Nephrin und CD31. Real-time PCR der aus Glomeruli gewonnenen RNA zeigte, dass die Rapamycin-Behandlung zu einer verstärkten Expression der proinflammatorischen Zytokine IL-6 (12,0 AE 5,2 vs. 1,0 AE 0,1; p < 0,05) und TNF-(3,4 AE 1,0 vs. 1,0AE 0,1; p < 0,05) führte, während sich hierbei keine Hinweise auf eine Schädigung der Endothelzellen ergab. Außerdem fand sich eine gesteigerte glomeruläre Expression des Transkriptionsfaktors Foxp3. Bei TX-Patienten/innen findet man eine bis zu 200-fache Erhöhung der sog. ,non-melanoma skin cancer'-Gruppe und eine 5-fache Steigerung der Melanom-Inzidenz im Vergleich zur Normalbevölkerung. Das klinische Bild dieser Neoplasien ist nicht selten atypisch, das Verhalten aggressiv und die Prognose ungünstig. Die Intensität bzw. Dauer der Immunsuppression und die Wahl der immunsuppressiven Medikamente in Verbindung mit der aktuellen und zurückliegenden UV-Exposition korreliert dabei mit der Tumorentstehung. Besonders wichtig sind regelmäßige Nachsorge-Untersuchungen post Transplantation, welche in speziellen dermatologischen Ambulanzen durchgeführt werden sollten. Hierbei stehen heute moderne Hauttumor-Therapieformen (,targeted' therapy und Immunmodulatoren) zur Verfügung. Background. We compared steady state pharmacokinetics of mycophenolate mofetil -Myfenax + (Teva) and CellCept + (Roche) -in stable kidney transplant recipients (KTR).

Methods. This was an international, multi-centre, randomized, open-label, two-treatment, two-sequence crossover study with a 3-months follow-up. We included KTRs at least 12 months post-transplantation with stable renal graft function for at least 3 months. The maintenance treatment consisted of MMF in combination with tacrolimus with or without steroids. At the end of the two treatment periods 6-hour or 12-hour pharmacokinetic studies of mycophenolic acid (MPA) were performed.

Results (0-6h) , and Cmin of MPA were within the bioequivalence margins and Cmax was marginally outside of these margins in this set of patients. The numbers and types of adverse events were not different between the two treatments.

Conclusions. The steady state pharmacokinetics of MPA as well as adverse events are comparable for Myfenax and CellCept in Tacrolimus treated stable kidney transplant recipients.

Belatacept in de novo kidney transplant recipients -10-year experience in a single center Background. Belatacept is a co-stimulation blocker which has recently been approved as immunosuppressive therapy in renal transplant recipients. Here we assess the outcome of patients that have been treated with belatacept for 10 years after kidney transplantation.

Methods. In our center, 22 patients were enrolled in the phase II multi-center belatacept trial that started in 2001. Patients were randomly assigned to belatacept-(n ¼ 14) or CyA-(n ¼ 8) based immunosuppression (all patients received basiliximab, MMF and steroids). In this retrospective analysis we report the outcome of the belatacept group as of June 2011. Patient and graft survival, the incidence of acute rejection, kidney function (calculated GFR; MDRD [ml/min/1.73 m 2 ]) and cardiovascular risk profiles (triglycerides and cholesterol) are presented.

Results. At an average of 9.2 (8.5-10.1) years after kidney transplantation 9 out of 14 belatacept (64%) patients are still on therapy. Five patients discontinued the study: 1 due to lack of efficacy (ATG-resistant rejection), 1 due to PTLD, 1 due to pneumocystis carinii pneumonia and 2 patients withdrew consent (1 and 3.5 years after transplantation with a functioning graft; GFR: 65.4 and 46.05 ml/min/1.73 2 ). Three patients died: the patient with PTLD and the patient with pneumocystis carinii pneumonia shortly after discontinuation and the patient with lack of efficacy 6 years later due to cardiac arrest. Two patients developed biopsy-proven acute rejection (2/14; 14.3%) (BANFF IIa, BANFF IIb). In 8 out of 9 patients kidney function remained stable over time: mean GFR 12 months after kidney transplantation 59.64 (SD 10.6), mean eGFR at the time of the last visit: 59.32 (SD 11.2). One patient had an impaired kidney function with a GFR of 45.03 at the last visit (GFR at month 12: 68). Mean serum triglycerides and cholesterol at 12 months after transplantation were 167 (SD 70) and 207 (SD 34) respectively, and changed little over time (mean triglycerides: 159 (SD 44) and cholesterol: 201 (SD 44) at the last visit).

Conclusions. In this selected group of patients enrolled in a phase II trial, continuous use of belatacept therapy for 10 years is associated with stable long-term graft function in a high percentage of patients.

The Innsbruck Hand Transplant Program: update at 11 years after the first transplant Background. We describe here the outcome after two bilateral hand, one bilateral forearm and one unilateral hand transplantation at 11/8/5 and 2 years after transplantation.

Methods. Four patients received a bilateral hand (n ¼ 2), a bilateral forearm (n ¼ 1) or a unilateral hand transplant between March 2000 and July 2009. Induction therapy with ATG (n ¼ 2) or alemtuzumab (n ¼ 2) was followed by tacrolimus, prednisolone, MMF (n ¼ 3) or tacrolimus and MMF (n ¼ 1) maintenance IS. Later, sirolimus/everolimus was added under simultaneous withdrawal (n ¼ 2) or dose reduction (n ¼ 1) of tacrolimus (n ¼ 1) or MMF (n ¼ 1). Steroids were avoided in one and withdrawn in two patients.

Results. Range of motion reached up to 70% of normal with a grip strength of 2-10 kg. Hand function correlated well with time after transplant and amputation level. Intrinsic hand muscle function recovery and discriminative sensation were observed in all patients. Complications included CMV infection, fungal infection, hypertension, hyperglycemia, transient creatinine increase and headache. Three, six, four, and one rejection episode were successfully treated with steroids, anti-CD25, anti-CD52 antibodies and/or intensified maintenance IS. Skin histology at current shows no or mild perivascular lymphocytic infiltrates without signs of progression. Vessels are patent without signs for luminal narrowing or intimal proliferation.

Conclusions. The overall functional outcome and patient satisfaction after bilateral hand, bilateral forearm and unilateral hand transplantation are highly encouraging. All patients are now free of rejection with moderate levels of IS.

Renal function efficacy measures following conversion from calcineurin inhibitors to sirolimus after cardiac transplantation in patients with renal insufficiency Methods. This was a randomized (1:1), comparative, multinational, phase 4 study of patients 1-8 years posttransplant. Patients on CNI therapy with GFR 40-90 mL/min/1.73 m 2 were eligible for inclusion. Exclusion criteria included acute rejection (AR) within 3 mos or proteinuria >500 mg/d. Primary end point was 1-yr change from baseline in creatinine clearance (CLcr).

Results. A total of 114 patients were randomized and treated (SRL 57; CNI 57); mean baseline GFR was $57 mL/ min/1.73 m 2 . In the ITT LOCF analysis, 1-yr adjusted mean change from baseline in CLcr was þ 4.0 and À1.2 mL/min/ 1.73 m 2 for SRL vs. CNI, respectively (p < 0.001); for the OT analysis, values were þ4.7 and À2.1, respectively (p < 0.001). AR rates were numerically higher in the SRL group; there was 1 AR with hemodynamic compromise in each group. A significantly higher (33.3% vs. 0%; p < 0.001) treatment discontinuation rate due to AEs was seen in the SRL group. Most common treatment-emergent AEs that were significantly higher in the SRL group were diarrhea (28.1%), rash (28.1%) and infections (47.4%).

Conclusions. Conversion to SRL from CNI therapy resulted in improved renal function in cardiac transplant recipients with renal insufficiency, but was associated with an attendant AR risk and higher discontinuation rate due to AEs. Background. ATG-induction therapy after heart transplantation is still controversial and used only by less than 50% of centers. Moreover, there exist no data about the optimal dosage of ATG-induction. The aim of this study is to compare different doses of ATG-induction.

Methods. Between 1996 and 2009 586 cardiac transplants were performed in our center. 523 (89%) patients with full data sets were included in the analysis. The median age was 56 years, 21% (n ¼ 112) were female. The median follow-up-time was 98 months. Patients were divided into 3 different groups according to total ATG dose: Group A: ¼ 4.5 mg/kg vs. Group B: 4.5-7.5 mg/ kg vs. Group C: >7.5 mg/kg. Survival, incidence of rejection, infection, graft vasculopathy and cancer were compared by Kaplan-Meier-analyses (log rank test).

Results. There was better early (12 m) and late (150 m) survival in Group B (A: 80%, 43%; B: 90%, 65%; C: 88%, 58%; p ¼ n.s.), however, the difference was not significant. Freedom from treated acute rejection was better in group B (88%) compared to A and C (79%, 80%, p ¼ 0.08). Signs of histological rejection were significantly different between the groups (A: 25%, B: 18%, C: 33%; p ¼ 0.03). Group B had the lowest incidence of severe infection (A: 37%, B: 21%, C: 51%; p < 0.01). CMV infection incidence was higher in group C (35%) compared to groups A, B (20%, 23%; p < 0.01). There was no significant difference in freedom from graft vasculopathy between the groups (A: 91%, B: 85%, C: 79%; p ¼ n.s.). The incidence of cancer was similar in all ATG groups (A: 3%, B: 7%, C: 11%; p ¼ n.s.).

Conclusions. Different doses of ATG-induction seem to have a significant impact on the outcome of heart transplantation. There is a strong need for more studies on optimization of ATG therapy.

Nighttime kidney transplantationa risk or a need?

Background. Kidney transplantation is performed as emergency surgery also as a nighttime procedure to reduce cold ischemia time und therefore improve outcomes after kidney transplantation. However, surgical procedures performed during nighttime in the context of 24 hour shifts and sleep deprivation have been associated with a higher rate of complications than elective surgery. The aim of this retrospective analysis is to determine the impact of nighttime surgery on the outcome after kidney transplantation.

Methods. All kidney transplants from cadaveric donors performed at our center from January 2000 through January 2010 (n ¼ 873) were included in this retrospective study and grouped according to the time of surgery: daytime (from 8 a.m. to 8 p.m., n ¼ 608) versus nighttime (from 8 p.m. to 8 a.m., n ¼ 265). Statistical analysis compared patient and graft survival in the two groups, rate of delayed graft function and acute rejection as well as surgical complications.

Results. 5-year patient survival rates of 87% and 85% and 10-year patient survival rates of 73% and 69% in daytime and nighttime kidney transplants did not show any significant difference. Also, graft survival at 5 and 10 years did not differ significantly (85% versus 85% and 69% versus 60%) in the two groups. Delayed graft function occurred in 31% of daytime transplants compared to 37% of nighttime procedures (p ¼ 0.06). Acute allograft rejection was observed in 23% of daytime graft recipients compared to 18% of nighttime graft recipients (p ¼ 0.13). Furthermore, nighttime procedures were associated with a 23% risk of surgical complications which was not significantly different from daytime surgery (22%, p ¼ 0.7).

Conclusions. Nighttime kidney transplants are not associated with a higher surgical complication rate than daytime procedures. Nighttime kidney transplant procedures have the same outcome as daytime transplants and are necessary to keep cold ischemia time as short as possible.

Ursolic acid a constituent of Dwarf Elder (Sambucus ebulus) inhibits surface expression of endothelial adhesion molecules and prevents intimal hyperplasia in an in vivo model for bypass surgery Background. In a recent study for the identification of compounds capable of inhibiting VCAM-1 expression we isolated ursolic acid (UA) from extracts of Dwarf Elder. Herein, we analyse the mechanism of action and the in vivo applicability of the compound in a setting for venous bypass graft intimal hyperplasia.

Methods and Results. Our analyses indicate that UA does not interfere with the adhesion molecule expression pathway at or upstream of NFK-B, as NFK-B translocation and NFK-B mediated transcription were not affected by UA. However, UA inhibited adhesion molecule protein synthesis suggesting an inference with the translation of adhesion molecules. In an in-vivo rat model for vein graft disease, UA-containing Sambucus extracts inhibited endothelial VCAM-1 expression, induced cell death in smooth muscle cells, and inhibited intimal hyperplasia in a rat model for venous bypass graft disease.

Conclusions. Our results suggest that the inhibition of VCAM-1 expression constitutes a valuable target for drug discovery in the field of cardiovascular research. UA may be an interesting agent in the prevention of vein graft disease and graft failure.

The killer-cell immunoglobulin-like receptor (KIR) genotype correlates with acute kidney failure in the early post-liver transplantation period Acute renal failure (ARF) is a major complication following liver transplantation (LT) leading up to chronic end-stage renal disease. The etiology of post-LT impairment is multifactorial, but it is suggested that e.g. during ischemia initial insults provoke morphological and functional changes within the vascular and tubular epithelium. Because it has been demonstrated that ischemic ARF can occur in the absence of classical T cell function and that natural killer (NK) cells can kill syngeneic tubular epithelial cells in vitro, we aimed to elucidate the role of NK cells and their receptors in the context of early post-liver transplant ARI and ARF more precisely. Patients with impaired kidney function (serum creatinine >1.2 mg/dl, n ¼ 13) illustrated heightened peripheral NK cell frequencies prior to LT compared with patients showing stable renal function (n ¼ 9) (17.22% AE 10.56 versus 12.98% AE 9.09). We further tested retrospectively 89 liver transplant recipients for their killer-cell immunoglobulin-like receptor (KIR) genotype and the risk of ARI/ARF. During the first week post-LT, ARI occurred in 12% and ARF in 22% of the patients. ARI was a significant risk factor for acute rejection (p ¼ 0.0009) and ARF led to elevated serum creatinine levels (>1.2 mg/dl) at the time of hospital discharge (p ¼ 0.008). Interestingly, significantly less patients having a homozygous KIR haplotype A/A (char-acterized by the presence of only one activating KIR gene) displayed a stable early postoperative kidney function, compared to patients with a KIR haplotype B/x (more than one activating receptor) (p ¼ 0.025, odds ratio 2.3, CI ¼ 1.3-3.9). Moreover, the absence of KIR2DL2/DS2 genes significantly influenced the risk of ARF (p ¼ 0.05). A multivariate regression model of clinical and genomic risk factors for ARI/ARF confirmed a link between the KIR haplotype A/A and post-LT acute renal failure (p ¼ 0.04). In summary, we observed a higher percentage of NK cells prior to LT in patients with impaired renal function and identified the KIR haplotype A/ A as an independent genetic risk factor for ARF within the first postoperative week. Our data provide new aspects of an innate immune response within the setting of post-LT kidney injury and failure. Background. New-onset diabetes after transplantation (NODAT) is a serious complication after kidney transplantation affecting graft and patient survival. Currently, no guidelines exist about the ideal management of renal transplant patients with impaired glucose tolerance (IGT, blood glucose >140 and < 200 mg/dl) who are at highest risk for developing manifest diabetes. Here, we studied the efficacy and safety of vildagliptin and pioglitazone in patients with IGT after renal transplantation.

Methods. After routine oral glucose tolerance testing (OGTT) in our center, 48 stable renal transplant patients with IGT were recruited in this double blind, prospective study and randomized 1 : 1 : 1 to receive either 50 mg vildagliptin, 30 mg pioglitazone or placebo once daily for 3 months. After treatment OGTTs were performed and HbA1c levels along with body mass index (BMI), metabolic and safety parameters were evaluated.

Results. So far 45 patients have finished the study. 2 hour glucose values improved in all three groups with the greatest mean improvement in the pioglitazone group. Fasting glucose values also showed the strongest improvement in the pioglitazone group as well as HbA1c (vildagliptin: 0.079% AE 0.249%, pioglitazone: À0.153% AE 0.318%, placebo: þ0.056% AE 0.294%). BMI increased in the pioglitazone group and decreased in the vildagliptin and placebo group. The difference in HbA1c between pioglitazone and placebo reached marginal statistical significance at p ¼ 0.05. Safety analysis showed no significant influences on glomerular filtration rate (GFR).

Pioglitazone led to significant reductions in GPT and Gamma-GT as well as increases in HDL. Adverse events occurred at similar rate in all three groups and were generally mild and reversible.

Conclusions. Vildagliptin and pioglitazone are both safe and effective in patients after renal transplantation. However, only pioglitazone led to significant improvements in glycemic control. Furthermore, pioglitazone led to improvements in GFR and lipid metabolism besides causing the largest weight gain. While further studies with longer observation periods are urgently required, pioglitazone seems to possess efficient beta-cell protective potency.

Clinical relevance of preformed complement-and non-complement-fixing HLA alloreactivity in cardiac transplantation S. Mahr 1 , M. Wahrmann 2 , A. Zuckermann 1 , G. Böhmig 2

Background. There is increasing evidence for a role of alloantibody-mediated rejection in organ transplantation. Solid phase HLA antibody detection using bead array technology may help identify patients at risk of rejection and graft loss.

Methods. In this retrospective monocentric cohort study we evaluated 229 consecutive heart transplant recipients (transplantation between 2000 and 2006; immunosuppression: ATG induction and calcineurin inhibitor-based maintenance therapy) for the presence of preformed (complement-and non-complement-fixing) HLA alloantibodies. Sera obtained immediately before transplantation were screened by FlowPRA, and test-positive sera were subjected to Luminex-based single antigen testing including a test modification for detection of in vitro C4d deposition.

Results. Seventeen recipients (7.3%) were found to have preformed IgG type donor-specific alloantibodies (DSA), five of them with C4d-fixing capability. The presence of DSA was related to retransplantation and previous pregnancies, but not associated with prior implantation of a ventricular assist device. Evaluating clinical endpoints, we found an association between DSA and acute rejection (>grade 1A according to the ISHLT 1990 grading system; no DSA: 17%; [IgG]DSA: 33%; [IgG/C4d] DSA: 60%). However, in our study cohort, sensitization had no effect on long-term survival rates (5-year transplant survival: 72% vs. 92% vs. 80%) or rates of chronic transplant vasculopathy (20% vs. 18% vs. 20%). Moreover, none of the DSA-positive patients was in need of extracorporeal membrane oxygenation, and the duration of post-transplant intensive care did not differ between groups.

Conclusions. In conclusion, our data point to a relationship between preformed donor-specific alloreactivity and acute rejection. However, possibly as a result of our local immunosuppressive regimen, which also includes induction therapy with a depleting anti-lymphocyte antibody, such reactivity did not influence long-term allograft outcomes. Background. So far, no data is available on the use of everolimus in pediatric cardiac transplantation. We present the first results of everolimus in pediatric heart transplantation.

Methods. From 2000 to 2008, HTx was performed in 86 children aged below 17 years. 52 of these children received continuously either mycophenolat mofetil (MMF) (Group A, n ¼ 28) with standard cyclosporine A (CsA) exposure or were randomized to receive everolimus (EVE) (Group B, n ¼ 24) with reduced CsA exposure in combination with steroids. During a follow-up period of 24 months, efficacy with regard to acute rejection, allograft vasculopathy and survival as well as safety with regard to myelosuppression, infection, tumors, lipids and body growth was studied.

Results. Mean post Htx CsA exposure was significantly lower in EVE treated children compared to MMF at month 6 (167 ng/ml versus 284 ng/ml), month 12 (mean 134 ng/ml versus 235 ng/ml) and month 24 (79 ng/ml versus 76 ng/ml). Mean daily dose of EVE was 1.16 AE 0.56 mg/m 2 body surface area resulting in trough levels of 4.8 AE 1.3 ng/ml by 24 months of follow-up. Two cases in each group encountered biopsyproven acute rejection >grade 2R (7% versus 8%; p ¼ 0.043). Cardiac allograft vasculopathy at month 12 was detected in 4 children (16%) with EVE compared to 18% in MMF. Kidney function deteriorated in both groups during the first month, recovering after 24 months to a glomerular filtration rate of 95.1 AE 0.3 ml/min/sqm for EVE versus 88.9 AE 5.2 ml/min/sqm for MMF. Rates of myelosuppressive disorders (anaemia, leukopenia, thrombocytopenia), infections and hyperlipidaemia was low and comparable in both groups. Children with EVE did not encounter inferiority in body growth. 1 child with EVE died following intractable ventricular fibrillation during myocardial biopsy compared to 5 children with MMF (2 deaths due to allograft rejection).

Conclusions. Everolimus with reduced CsA combined with steroids in paedriatic cardiac transplant recipients enables high efficacy and is comparatively safe to immunosuppression with MMF and standard exposure of CsA.

Transplantation of a minor-mismatched skin graft elicits a rapid humoral response including the induction of antigen-specific IgE Background. Sensitization to major and minor antigens is a critical problem in transplantation medicine with antibodies of IgM, IgA and IgG isotypes have been demonstrated towards major (i.e. MHC) and minor (i.e. non-MHC) antigens. In a new transgenic mouse model we investigated humoral antigenspecific responses towards a highly immunogenic non-MHC antigen. A transgenic mouse expressing the well-characterized major grass pollen allergen Phl p 5 ubiquitously on the cell surface was generated.

Methods. A Phl p 5-transgenic Balb/c mouse was generated by pronuclear injection integrating Phl p 5 (and GFP) fused to a leader peptide and a transmembrane domain. Splenocytes (2.5Â106 per mouse), cell extracts containing mainly membrane proteins and recombinant (r) Phl p 5 [5 mg AE Al(OH)3 per mouse] were injected subcutaneously into naïve Balb/c mice (n ¼ 8 per group). Furthermore tail skin of Phl p 5-transgenic mice was grafted onto naïve Balb/c (n ¼ 13 in two independent experiments). The Phl p 5-specific humoral response was determined in sera by ELISAs and T-cell proliferation assays were assessed.

Results. Surprisingly, a prompt rejection of Phl p 5-transgenic skin was elicited (within 8-10 days), accompanied by a strong Phl p 5 specific antibody response including Phl p 5-specific IgE. Additionally to the skin grafted group, mice receiving splenocytes or rPhl p 5 plus adjuvant showed a comparable response in terms of Phl p 5-specific IgE throughout the whole follow-up (week 1, 2, 3, 5, 7, 9, 11) suggesting an unusually strong immune response to cell or tissue-bound Phl p 5. Furthermore Phl p 5-specific IgG isotypes, IgA and IgM were induced. Besides splenocyte-proliferation assays showed Phl p 5 specific T-cell responses in all groups of mice that showed strong humoral responses.

Conclusions. The high immunogenicity of tissue-bound Phl p 5 may represent a new stringent model for studying humoral responses towards non-MHC antigens. Notably, the immune reaction included a rapid IgE response in this model.

The influence of recipient age on chimerism-based tolerance induction K. Hock 1,2 , R. Oberhuber 2,3 , Y.-L. Lee 2 , T. Wekerle 1 , S. G. Tullius 2 Background. Immune senescence substantially alters alloreactivity. Higher frequencies of memory T cells (Tmem) found in older recipients are considered a substantial barrier to tolerance induction. Tolerance induction through mixed chimerism holds promise for clinical translation but has only been investigated in young recipients so far. As the average recipient age has increased substantially in clinical organ transplantation, we investigated the consequences of recipient age on the outcome of costimulation blockade based allogeneic bone marrow transplantation (BMT) for the purpose of mixed chimerism and tolerance induction.

Methods. Young (2 months; weighing approx. 20 grams) and old (12 months; 25 g) recipients (C57BL/6) were treated with 3 or 1 Gy total body irradiation (TBI, d-1) and received adjusted to the body weight 20Â10 6 and 25Â10 6 un-separated Balb/c BM cells (d0) and co-stimulatory blockade with anti-CD154 mAb (d0) and CTLA4Ig (d þ 2) was administered. Lymphocyte subsets and cytokine production were compared between young and old naïve mice and multilineage chimerism was followed by flow cytometry.

Results. Old mice contain significantly more CD4 (p < 0.05) and CD8 (p < 0.001) memory T cells (CD44highCD62Llow), early activated CD4 T cells (CD4 þ CD69 þ ; p < 0.01), less CD4 and CD8 T cells and comparable amounts of regulatory T cells (Tregs; CD4 þ CD25 þ FoxP3þ; p ¼ n.s. vs. young animals). Moreover, older CD4 and CD8 T cells release more IFN-(CD4: p < 0.05), IL-2 (not CD8 T cells), IL-6 (CD4: p < 0.05), IL-10 and TNF-. Chimerism developed earlier in old recipients: within one week most older recipients became chimeric following an irradiation with 3 Gy TBI and co-stimulatory blockade (17/18 vs. 9/18 chimeras, d þ 7 and 16/17 vs. 12/17, d þ 14). Old recipients became even chimeric with a reduction of the total body irradiation to 1 Gy. In sharp contrast, none of the young recipients became chimeric under those conditions (4/8 vs. 0/8 chimeras in young recipient, p < 0.05, d þ 30).

Conclusions. Recipient age is linked to a faster donor BM engraftment and chimerism. Moreover, chimerism is attainable with a lower dose of irradiation. Those results support the clinical relevance of the chimerism strategy for tolerance induction. Background. In allogeneic bone marrow transplantation (BMT) donor T cells have pleiotropic effects. Surprisingly, cotransplanting high doses of donor T cells with donor BM causes rejection of donor BM despite costimulation blockade. Therefore, in the present study we investigate the molecular mechanisms responsible for this seemingly paradoxical phenomenon.

Methods. Recipients (C57BL/6) were treated with 3 Gy TBI (d-1) and received approximately 20Â106 unseparated Balb/c BM cells (d0) and costimulation blockers anti-CD154 mAb (d0) and CTLA4Ig (d þ 2). 30Â10 6 Balb/c, CB6F1 (Balb/cÂB6), irradiated Balb/c or C3H CD4 T cells (isolated by MACS) were co-transplanted in addition. Groups either received anti-IL-6, anti-IFN-, anti-LFA1 mAb or rapamycin. Multilineage chimerism was followed by flow cytometry and cytokine release was analyzed.

Results. Co-transplantation of 30Â10 6 CD4 T cells but not CD8 T cells triggered rapid BM rejection of donor BM under costimulation blockade within one week in an otherwise successful protocol (0/13 vs. 17/20 chimeras, p < 0.001). The levels of IL-6, IFN-, IL-17A (p < 0.05) and TGF-were found to be higher in mice treated with additional donor T cells. The neutralization of IL-6, but not of IFN-resulted in a significantly higher rate of chimerism induction compared to controls (5/7 vs. 0/5 chimeras; p < 0.05). The injection of CB6F1 or irradiated Balb/c CD4 T cells did not abrogate chimerism (5/6 and 4/5 vs. 0/4 chimeras with Balb/c T cells; p < 0.05), whereas C3H CD4 T cells induced BM rejection (0/5 vs. 9/9 chimeras BMT, p < 0.001). The additional treatment with rapamycin or anti-LFA1 overcame the negative effect of donor T cell injection (5/5 and 6/6 vs. 0/4 chimeras; p < 0.01).

Conclusions. The abrogation of BM engraftment through cotransplantation of donor CD4 T cells involves IL-6, requires proliferative capacity of the co-transplanted T cells and needs to recognize the recipient as allogeneic. Neutralisation of IL-6, rapamycin and anti-LFA1 overcome the effect of co-transplanted donor CD4 T cells and offer potential targets for therapeutic intervention in costimulation blockade-resistant rejection. 

Background. Mixed chimerism is an effective strategy for the induction of transplantation tolerance, however, translation from murine models to the clinical setting is challenging. One hurdle is the high frequency of alloreactive memory T-cells (Tmem) found in the (pre)clinical setting, which is not present to a similar degree in mouse models. To better model this clinical reality, we have developed a murine model in which the transfer of 3Â10 7 T-cells from sensitized mice inhibits the induction of chimerism and tolerance in a well-characterized costimulation blockade-based bone marrow transplantation (BMT) model. Here, we evaluated whether treatment with clinically approved drugs (rapamycin, anti-LFA-1mAb) can overcome costimulationresistant rejection triggered by memory cells and tried to gain insight into the underlying mechanisms.

Methods. T-cells sensitized towards donor (3Â10 7 Tmemenriched T-cells) were adoptively transferred into groups of naïve B6 mice prior to BMT (d-7). BMT recipients received 1Gy total body irradiation (d-1), 20Â10 6 fully mismatched Balb/c bone marrow cells (d0) and costimulation-blockade consisting of anti-CD154mAb (1 mg, d0) and CTLA4Ig (0.5 mg, d þ 2). Groups additionally received rapamycin (0.1 mg/d, d-1/0/2) or anti-LFA-1mAb (0.1 mg/d, d-1/2). Multilineage chimerism and deletion of donor-reactive T-cells were followed by flow-cytometry and in vitro tolerance was assessed by MLR. Transferred T-cells were followed in vivo using CD45.1/2 alleles as congeneic markers in peripheral blood (d-2/12) and lymphoid organs (d7). IFNgamma-production upon stimulation with donor antigen was measured by ELISPOT and intracellular flow-cytometry.

Results. Rapamycin and anti-LFA-1 were able to overcome costimulation-blockade resistant rejection and induced stable multi-lineage chimerism in pre-sensitized recipients (0/7 control vs. 7/7 rapa, p < 0.001; 3/7 anti-LFA-1, p ¼ 0.2; representative for multiple experiments). Chimeras showed central and peripheral deletion of donor-reactive T-cells and donor-specific hyporesponsiveness in vitro. The frequency of transferred T-cells decreased over time, but less so in the rapamycin-treated group. IFNgamma-production was reduced by rapa or anti-LFA-1 treatment.

Conclusions. Rapamycin and anti-LFA-1 overcome the additional T-memory-cell barrier for tolerance induction, although rapamycin seems to be more potent in Tmem tolerization in this model. The trend that elimination of Tmem is lower in the rapamycin-treated group and central deletion is more pronounced early after BMT suggests different mechanisms for Tmem tolerization.

Lunchsymposium Virusinfektionen in der Organ-und Stammzelltransplantation 035 Diagnostik von Virusinfektionen in Transplantationspatienten

Patienten nach Organ-oder Knochenmarkstransplantationen werden immunsuppressiv behandelt und weisen daher ein anderes Spektrum von potentiell gefährlichen Virusinfektionen auf als immunkompetente Personen. Vor allem Viren, die im normalen Wirt nur leichte oder asymptomatische Infektionen verursachen und latent im Organismus verbleiben, wie Herpesviren (Zytomegalievirus, Epstein-Barr Virus etc.) oder auch Polyomaviren (BK-, JC-Virus), können unter Immunsuppression hoch replizieren und zu schweren und potentiell tödlichen Infektionen führen.

Um schwere Infektionen oder Reaktivierungen durch verschiedene Viren zu verhindern, wird heute routinemäßig in regelmäßigen Abständen im Verlauf nach Transplantation der direkte und quantitative Nachweis verschiedener Viren (vor allem von Zytomegalievirus) mittels PCR im Blut, aber auch in verschiedenen anderen Patientenmaterialien durchgeführt. Ziel der Virusdiagnostik ist es hier nicht so sehr eine klinisch relevante Virusinfektion nachzuweisen, sondern vielmehr das Vorhanden-sein einer signifikanten Virusreplikation frühzeitig, noch vor Krankheitsbeginn zu erkennen. Das ist dann auch die Basis für die "präemptive" antivirale Therapie, die gegeben wird, wenn die Viruslast eine bestimmte Höhe überschreitet, aber noch bevor klinische Symptome auftreten.

Die Festsetzung der virologischen Grenzwerte fü r den Einsatz der präemptive Therapie ist aber eine große Herausforderung, da vor allem Herpesviren wie Zytomegalievirus (CMV) oder Epstein-Barr Virus (EBV), aber auch Polyomaviren auch ohne jede Krankheitsrelevanz im Organismus nachgewiesen werden können. Neben den quantitativen PCR Ergebnissen hängt der Einsatz einer präemptiven Therapie auch von bestimmten weiteren Aspekten ab, wie vom Donor/Recipient Virus-serostatus, dem transplantierten Organ oder vom Material in dem man die Virusnukleinsäure nachweist. Daher ist ein enges Zusammenspiel von virologischer Befunderstellung, Interpretation und der Kenntnis der klinischen Aspekte notwendig, um eine optimale virologische Kontrolle der einzelnen Patienten zu erreichen.

Antivirale Therapie bei Patienten nach Organ-oder hämatopoetischer Stammzelltransplantation

Klinische Abteilung für Pädiatrische Hämato-/Onkologie, Universitätsklinik für Kinder-und Jugendheilkunde, Medizinische Universität Graz, Graz, Österreich Zahlreiche humanpathogene Viren persistieren nach Primärinfektion im Körper und können unter geschwächter Abwehrlage reaktiviert werden. Zu diesen zählen die Herpesviren Herpes simples 1 und 2 (HSV 1, 2), Cytomegalievirus (CMV), Epstein-Barr-Virus (EBV), Varicella Zoster Virus (VZV) und HHV-6 sowie das Parvovirus B 19 (PVB19), das Adenovirus (ADV), die Hepatitisviren B (HBV) und C (HCV) und die Polyomaviren BK-Virus (BKV) und JC-Virus (JCV). Diese und andere Viren (z. B. Enteroviren) können bei Empfängern von Organoder Stammzelltransplantationen durch Primärinfektion oder Reaktivierung zu schweren Krankheitsverläufen ("Virus-Sepsis") und immunologischen Prozessen wie Transplantatabstoßung oder Graft-versus-Host Disease führen.

Neben unspezifischen Maßnahmen wie Reduktion der Immunsuppression, Gabe von Immunglobulinen und Immunmodulation mittels Interferon, wurde in den letzten Jahren eine zunehmende Zahl antiviraler Substanzen verfügbar. Deren antiviraler Effekt beruht auf unterschiedlichen Mechanismen: Hemmung der Aufnahme der Viren in die Wirtszelle (Amantadin, Pleconaril), Inhibierung der Replikation des viralen Genoms (Aciclovir, Ganciclovir, Cidofovir, Foscarnet, Ribavirin), Unterdrückung der Freisetzung von Viruspartikeln aus der Wirtszelle (Oseltamivir, Zanamivir).

All diese Substanzen wirken virustatisch und nicht viruzid, können also die Virusvermehrung hemmen, nicht jedoch die Viren gänzlich eliminieren.

Während einige Substanzen relativ spezifisch in die Replikationsmechanismen einzelner Viren eingreifen (Aciclovir, Ribavirin, Oseltamivir, Zanamivir), zeigen andere Substanzenmeist jene mit breiterem Wirkspektrum (Cidofovir, Foscarnet) -auch eine vermehrte Aktivität innerhalb humaner, molekularer Mechanismen, was zu einer beträchtlichen Zahl unerwü nschter Arzneimittelwirkungen fü hrt (v. a. Nephro-, Myelo-, Neurotoxizität).

Handelsname DNA-Viren RNA-Viren

Ciprofloxacin** Ciproxin þ (in vitro) Wirkspektrum ausgewählter Virustatika (ohne anti-retrovirale Substanzen) Wirksamkeit entspricht nicht den zugelassenen Indikationen * experimentell, nicht verfügbar ** experimentell Einige Substanzen haben im Tierversuch ein carcinogenes (Ganciclovir, Cidofovir) und/oder teratogenes (Ganciclovir, Cidofovir, Ribavirin) Potenzial gezeigt.

Unter zu geringer Dosierung und/oder mangelhafter Medikamenteneinnahme kann es zu viralen Mutationen und -daraus resultierend -zur Resistenzentwicklung kommen.

Tabelle 1 gibt einen Ü berblick über die wichtigsten Virustatika und deren Wirkspektrum.

Viele dieser Substanzen sind jedoch für die Anwendung im "Transplantations-Setting" nicht offiziell zugelassen.

In Zukunft sind weitere, teilweise dzt. noch experimentell eingesetzte Therapieoptionen zu erwarten: so scheint Ciprofloxacin als Gyrasehemmer auch gegen Polyomaviren wirksam zu sein, mit CMX001 wird eine wirksamere und nebenwirkungsärmere Weiterentwicklung von Cidofovir derzeit klinischen Tests unterzogen.

Reaktivierungen von persistierenden Virusinfektionen sind häufige Komplikationen nach Transplantation. Je nach Risikokonstellation reicht das klinische Spektrum von einem harmlosen Infekt bis zu einem lebensbedrohlichen Infektionsverlauf. Die häufigsten Pathogene sind Cytomegalievirus (CMV), Epstein-Barr Virus (EBV) und Adenoviren (ADV). Die Möglichkeiten der virostatischen Pharmakotherapie sind meist limitiert. Alle drei Virusinfektionen haben gemeinsam, dass fü r eine Elimination eine suffiziente T-Zellimmunität notwendig ist. Darauf baut die Rationale eines adoptiven Immuntransfers auf, bei dem Ag-spezifische T-Zellen von einem gesunden Spender in einen erkrankten Empfänger infundiert werden. Hierbei ist das Therapieziel eine adaptive Immunität im Empfänger zu induzieren. Im Rahmen einer risikoadaptierten Therapie wurde bei refraktären Virusreaktivierungen nach Transplantation dieses Therapieverfahren angewendet. Eine Ag-spezifische T-Zellantwort konnte im Empfänger gegen immundominante Epitope (pp65 fü r CMV, Hexonprotein fü r ADV und EBNA1 fü r EBV) induziert werden um den Empfänger vor Virus-assoziierten Komplikationen zu schü tzen. Background. We investigated the safety and efficacy of T and B cell depleted peripheral stem cells from full haplotype mismatched parental donors in pediatric patients.

Methods. Use of the Clinimacs system and CD3/CD19 coated magnetic microbeads resulted in a 4 log depletion of T cells and allowed to cotransfuse high numbers of donor NK cells (median: 107Â10 6 /kg). TBI or busulfan based myeloablative regimens or a melphalan based intensity reduced regimen were used. All patients underwent intensive pretreatment according to current study protocols; 41/106 already received previous allogeneic transplantations. The diagnoses were: acute leukemias and MDS (n ¼ 60), solid tumors (n ¼ 32) and nonmalignant diseases (n ¼ 14).

Results. Primary engraftment occurred in 89% of patients. After TLI based reconditioning and second haploidentical stem cell donation, final engraftment was achieved in 100%. Median time to reach >500 neutrophiles/ml and independence from platelet substitution was 10 (8-15) and 9 (5-59) days respectively. 35% of patients had no GvHD, 36% had grade I, 23% had grade II and 4% had grade III. Chronic limited and extensive GvHD was observed in 8 and 11%. Transplant related mortality was 0% at day 100 and 8% at one year. Event free survival at 3 years was 66% for patients with leukemias in any CR and 80% for patients with nonmalignant diseases. Over all survival at 2 years was 20% for patients with solid tumors. Relapse or progression were the major causes of death. Thus, pilot studies with IL-15 stimulated grafts and posttransplant donor-NK cell infusions were initiated and are currently ongoing.

Conclusions. Transplantation of CD3/CD19 depleted haploidentical stem cells resulted in a fast recovery of neutrophiles and platelets. Engraftment rates similar to that of patients with myeloablative standard conditioning and positive selected stem cells could be achieved, possibly due to a graft facilitating effect of cotransfused NK cells. The regimen helped to minimize TRM, despite intensive pretreatment (including previous transplantation). However, relapse remains a major problem and further immunotherapeutic elements have to be evaluated.

Antibody based immunotherapy combined with haploidentical stem cell transplantation for high risk neuroblastoma Background. Pediatric patients with relapsed metastatic neuroblastomas have a poor prognosis and additional therapeutic strategies are warranted. We present preliminary results with haploidentical stem cell transplantation and posttransplant immunotherapy with an anti-GD2 monoclonal antibody (CH14.18 CHO).

Methods. T and B cell depleted haploidentical stem cells in combination with a toxicity reduced conditioning regime (melphalan, thiotepa, fludarabine and OKT3, now substituted by low dose ATG + Fresenius) were used for transplantation. In a subsequent clinical trial, we evaluate the feasibility and safety of antibody infusions against the neuroblastoma antigen GD2: 6 cycles of mAb CH14.18/CHO (20 mg/m 2 infusion for 5 days; in cycles 4-6, 1Â10 6 units/m 2 interleukin 2 (IL-2) is given additionally on days 6, 8 and 10).

Results. Haplotransplantation without subsequent antibody infusions resulted in a 2-year over all survival of 28%. TRM at day 100 was 0%, single cause of death was relapse/ progression. Thus, transplantation itself seems to be not sufficient for most patients but may be a platform for further immunotherapies on the basis of antibody dependent NK cell mediated cytotoxicity and complement lysis. 12 patients have been enrolled in our CH14.18 study and received a total of 42 antibody cycles. Side effects were: inflammation symptoms (pain 42/42, fever 36/42 and CRP elevation 35/42 cycles), which decreased or disappeared by increasing cycle numbers; accommodation disturbance (n ¼ 3), seizures (n ¼ 2), loss of weight (n ¼ 2). During antibody infusions, endogenous secretion of IL-2 in the patients was significantly increased (880 U/ ml prior vs. 1580 U/ml post), which resulted in an also significantly increased stimulation of natural killer (NK) cells in peripheral blood (measured by CD69 expression). Predominantly ADCC exerting NK cells (with expression of the Fc-gamma receptor CD16) were stimulated. In 3 out of 5 investigated patients, NK cell and complement mediated anti-tumor effects against neuroblastoma cells were detectable. 5 patients finished the protocol so far and were evaluable after 6 cycles. 3/5 patients responded and reached a complete remission or improved their partial remission, 2 patients progressed.

Conclusions. Preliminary results of our ongoing study suggest an anti-tumor effect of the donor derived immune system in vitro and in vivo. SCT from a matched sibling donor (MSD) is the treatment of choice for children with SAA, however limited by donor availability. Immunosuppression (IS) as the other option has the disadvantage of high treatment failures. 7 patients with SAA without a MSD and refractory to previous IS, median age 11 years (5-16), median interval from diagnosis to SCT 12 months (3-155) and median number of transfusions before SCT 55 (18-116) underwent alternative donor SCT. Donors were matched unrelated (MUD) (n ¼ 4), mismatched unrelated (n ¼ 2) and haploidentical (n ¼ 1). Conditioning regimens contained cyclophosphamide and either muromonab-CD3 or anti-thymocyte globulin (ATG) in 7/7, thiotepa (THT) in 6/7, fludarabine (FLU) in 5/7 and total lymphoid irradiation in 2/7 patients. Grafts were either CD34þ selected and/or CD3/19 depleted using the Miltenyi CliniMACS device. Median yield of purified CD34þ cells was 10.17Â10 6 /kg (7.85-24.3) and median CD3þ number was 5.5Â10 4 /kg (0.84-10). All patients had WBC-engraftment on median day 10 (8-12). There was no GVHD prophylaxis in 2 patients and either cyclosporine-A or mycophenolate mofetil up to day þ 60 in 5 patients. 1 MUD-HSCT recipient who had 6 IS courses and the longest interval to HSCT (13 yrs) and who received the highest CD 3þ dose (10Â10 4 /kg) developed GVHD (grade II) on day þ 125 with progressive kidney failure due to microangiopathic hemolytic anemia. All children are alive with a median follow up of 20 months (8-149) with stable complete engraftment and stable chimerism of median 98.5% (90.65-100).

Conclusions. Long interval to transplant, multiple transfusions and long term immunosuppression before SCT are associated with poor alternative donor SCT-outcomes in SAA. Cyclophosphamide conditioning including FLU, THT and ATG but without TLI, high doses of purified CD34þ cells and/or CD3/19 depletion may prevent graft rejection and GVHD and hasten engraftment. This may facilitate the decision for the earlier use of unrelated stem cells preventing complications from prolonged and multiple immunosuppressive therapies and presumably increasing the survival of patients with SAA without a matched sibling donor.

Extracorporeal photopheresis in patients with high bleeding risks Background. For anticoagulation in extracorporeal photopheresis (ECP) to prevent clotting in the extracorporeal circuit, the manufacturer's recommendation is the use of heparin (normally 10,000 Units). However, patients with acute graft-versushost disease (aGvHD) after allogeneic hematopoietic stem cell transplantation are at high risk for bleeding complications due to low platelet counts, intestinal lesions from aGvHD or impaired hepatic function. For these patients, alternative anticoagulation using acid citrate dextrose (ACD-A) has been infrequently used in small patient cohorts.

Methods. We investigated the safety and efficacy of this approach in 94 consecutive patients (43 male, 51 female) with aGvHD (45% with gut GvHD) undergoing ECP with ACD-A anticoagulation at a single institution 2 to 3 times per week on a weekly basis until complete resolution of aGvHD. A total of 1242 ECP procedures were analyzed with respect to side effects and changes in haemoglobin and platelet levels. Moreover, in a proportion of ECP treatments activated partial thromboplastin time (aPTT) was monitored. Priming of the separator was performed with 1,000 Units of heparin followed by anticoagulation with ACD-A at a ratio of 1:10 during the procedure.

Results. ECP was tolerated well by all patients. In only 0.2% of procedures mild citrate reactions seen as transient paresthesias were observed but resolved without the need for calcium substitution. In no case bleeding complications were noted during or after citrate anticoagulation. During ECP, aPTT levels increased marginally from a median of 32.1 sec. (range, 25.3 to 44.5 sec) before to a medium of 35.4 sec. (range, 25.4 to 54.7 sec.) after ECP, respectively. At start of ECP 51% of patients had platelet counts <100 G/l and 26% <50 G/L, respectively. Haemoglobin and platelet counts decreased by 11% and 14%, respectively.

Conclusions. In conclusion, citrate anticoagulation during ECP is a feasible and safe alternative for patients with high risk for bleeding complications, especially for those after allogeneic hematopoietic stem cell transplantation.

Allogeneic stem cell transplantation with reduced intensity conditioning in patients with therapy-related myeloid neoplasms Background. Therapy-related myeloid neoplasms (t-MNs) occur as late complication after cytotoxic therapy for malignant and non-malignant disorders. In most patients with t-MNs, allogeneic haematopoietic stem cell transplantation (HSCT) is the only potentially curative approach. Performed with myeloablative conditioning this option is associated with a high transplantrelated mortality (TRM). Here we report our results in using reduced intensity conditioning (RIC) in these patients.

Methods. Between July 2000 and February 2011, 18 patients (male: 5; female: 13; median age: 48.5 years; range: 28-66) with t-MNs underwent RIC HSCT either from a matched sibling (n ¼ 8) or a matched unrelated (n ¼ 7) or one antigen mismatched unrelated donor (n ¼ 3). Primary disorders were solid tumors and haematologic malignancies in 9 patients each. Patients presented with t-MDS (n ¼ 4), t-AML (n ¼ 13) and t-MDS/MPN (n ¼ 1). Cytogenetic analysis revealed clonal aberrations in 15/18 patients. Nine patients were transplanted in 1st CR, one in 2nd CR, five in PR and three in relapsed/refractory disease. Furthermore, five patients showed persistant primary disease. Conditioning regimen consisted of fludarabine/melphalan (n ¼ 15) or fludarabine/ low-dose TBI (n ¼ 3). GvHD prophylaxis consisted of cyclosporine and mycophenolate mofetil and additionally anti-thymocyte globulin in unrelated transplantations.

Results. After a median follow-up of 31 months (range: 3-97) 9 patients are alive and in CR. Causes of death were: relapse (n ¼ 4), sepsis/multiorgan failure (n ¼ 4) and GvHD (n ¼ 1). Acute and chronic GvHD was observed in 6 and 5 patients, respectively. The estimated OS is 53.8% at one and 47.9% at three years; the DFS is 48.1% at one and three years, respectively. Of 5 patients transplanted with active primary neoplasms, 2 are alive at 1 year showing CR of both primary and secondary neoplasms which was associated with the development of chronic GvHD.

Conclusions. The results of RIC HSCT in these patients with poor risk disease are encouraging with respect to TRM, relapse rate and overall survival. Our data show a more favourable outcome compared with previous reports using a myeloablative conditioning. Furthermore, allogeneic HSCT should be considered as a potentially curative strategy for patients with t-MNs and persistant primary malignancy.

The bone component of CTA gives rise to donor HSCs which migrate to recipient thymus and differentiate to mature T cells Background. Composite Tissue Allotransplantation (CTA) is immunologically unique in that it represents the only type of graft to include a vascularized functional bone marrow component. Here we studied if the bone component of a composite tissue graft represents the source of HSCs that differentiate in the thymus and thereby reconstitute a functional immune system (CD3þ T cells in peripheral blood/lymphoid organs) in immuno-deficient B6/SCID recipients rather than originate from donor mature passenger T cells that expand in the host.

Methods. B6 (WT/nude) murine composite tissue grafts (osteomyocutaneous or myocutaneous) were transplanted heterotopically to B6 (WT/scid) recipients using a non-suture cuff technique for revascularization. Flow cytometry of peripheral blood (CD3þ, CD19þ) was performed at pod 7, 14, 21, 28, 56. In addition, histopathology (H&E) and immunohistochemistry of tissues was performed at indicated time points. To assess immunocompetence, allogeneic skin grafts (Balb/c) were transplanted to either naïve B6/nude, naïve B6/scid or B6/scid mice that prior received a B6/nude CTA.

Results. The surgical success rate was 85% in all groups. As expected no CD3þ cells and no rejection of skin allografts were detected in B6/nude and B6/scid controls. B6/scid mice that received B6/nude osteomyocutaneous flaps demonstrated B and T cell immunity from pod 7 and 21 respectively. The percentage of CD3þ and CD19þ cells within peripheral blood mononuclear cells steadily increased to 57.7% and 17.1% respectively at pod 56. Allogeneic skin allografts were rejected 2 weeks after transplantation. However, no B and T cell reconstitution was observed in B6/scid mice receiving B6/nude myocutaneous flaps (without bone component).

Conclusions. The vascularized bone marrow component of CTA provides an effective source of HSCs to restore immunocompetence in T-and B-cell deficient mice. This might also contribute to chimerism induction and maintenance after CTA and facilitate the clinically observed immunoprivilege of CTAs.

Early in vivo signaling events involved in neo-vasculogenesis via stem cell transplantation using proteomic profiling Background. The precise mechanisms regulating human neo-vasculogenesis and organ regeneration are still unclear. It has been shown that neo-vasculogenesis can be induced in immunocompromised NOD/scid/IL-2-receptor gamma-knockout (NSG) mice in vivo by transplantation of human bone marrowderived mesenchymal stem and progenitor cells (hMSPCs) and umbilical cord-derived endothelial colony-forming progenitor cells (hECFCs), whereas transplantation of pure hMSPC or hECFC cells lack the capacity to form stable functional vessels (Reinisch et al., Blood, 2009) . Understanding the activity of the mediators of neo-vasculogenesis would provide us with tools to develop strategies for therapeutic intervention as well as angiogenesis and regenerative applications.

Methods. Autologous pairs of MSPCs and ECFCs were transplanted subcutaneously in Matrigel plugs at a ratio of 20:80 into NSG mice and implants were harvested 1 day post transplantation to investigate proteomic profiling using KAM 1.3 Antibody Microarray (www.kinexus.ca), testing over 800 signaling and phospho-proteins. The state of vessel formation and stability of the transplants were verified by immunohistochemistry of the explants 2 and 8 weeks post transplantation.

Results. Protein microarray data analysis revealed significant alteration in the expression and activity of components 1) regulating apoptosis, mitotic check point control, and centrosome structure; 2) modulating glycolysis and the coordinated expression of cyto-protective genes; and 3) mediating cell adhesion, migration and tissue invasion. Selected targets are currently being validated by Western blotting to allow for the development of novel therapeutic intervention strategies. A detailed expression and interactive network analysis of targets will be presented and discussed.

Conclusions. Our data confirm that more than one purified cell type is needed for tissue engineering in vivo and suggest that composite cellular transplantation may be useful for future therapeutic strategies. Proteomic profiling unraveled at least three distinct but partially overlapping signaling networks involved in the complex process of vascular regeneration. Understanding the origin and activity of the mediators of vessel formation and repair would provide us with tools to develop and further optimize novel cell transplantation strategies.

Oxygen sensing of mesenchymal stem and progenitor cells promotes neo-vasculogenesis in vivo Background. Vascular regeneration requires a stringent interaction of somatic endothelial colony-forming progenitor cells (ECFCs) with mesenchymal stem and progenitor cells (MSPCs). Hypoxia in ischemic tissue is a key factor driving the revascularization machinery. Because ECFCs, despite hypoxic stimulation, only form patent vessels in vivo in the presence of MSPCs, we hypothesized that MSPCs play a decisive role in oxygen sensing during vasculogenesis.

Methods. Adult human ECFCs were isolated directly from whole venous blood and MSPCs from human bone marrow aspirates. Pooled human platelet lysate entirely replaced fetal bovine serum during cell culture. Progenitor cell phenotype, long-term proliferation, molecular cellular response, wound repair as well as migratory and vasculogenic functions were monitored under reduced oxygen levels (5% O 2 ), severe hypoxia (1% O 2 ) and standard culture conditions (20% O 2 ). ECFC and MSPC interaction in vivo and the influence of protein synthesis were studied in immune-deficient NSG mice after subcutaneous injection with matrigel. Immune histochemistry and TUNEL assays were performed on explants in the time course after transplantation.

Results. In vitro ECFC and MSPC proliferation was reduced with declining oxygen levels, while the absolute colony number remained unchanged. ECFC vascular wound repair and vascular-like network formation in vitro improved with escalating oxygen supply. ECFCs stabilize hypoxia-inducing factor-1 (HIF-1) only under 1% O 2 , while MSPCs stabilize HIF-1 under 1% O 2 as well as 5% O 2 conditions. In a mouse model, injected ECFCs underwent apoptosis after 1 day and attracted mouse leucocytes. In vivo co-cultured ECFCs and MSPCs formed perfused human vessels 7 days after transplantation. Perivascular cells, but not ECFCs, in vivo were positive for HIF-1. Inhibition of MSPC, but not ECFC, protein synthesis and HIF-1 stabilization prior to co-implantation blocked vessel formation.

Conclusions. These data indicate that hypoxic ECFCs alone are not able to function in vitro and form patent vessels in vivo. MSPCs react to the low oxygen environment and promote ECFCs to form vessels at least in part by rescuing ECFCs from hypoxia-induced apoptosis. This suggests that oxygen sensing MSPCs are a key factor in stem cell transplantation and regenerative medicine. Background. Low sensitivity and specificity rates for the detection of invasive pulmonary mycosis result in a major cause of mortality among immunosuppressed children. We sought to determine the accuracy of percutaneous computed tomographyguided biopsy in children with cancer and hematopoetic stem cell transplantion.

Methods. We retrospectively reviewed 17 imaging-guided percutaneous biopsies of 17 children for suspicious lesions detected by computed tomography. Thirteen were being treated for hematologic malignancies, three for solid tumors, one for immunodeficiency; 47% had received allogeneic bone marrow or peripheral stem cell transplants. The accuracy of the percutaneous lung biopsy was determined by subsequent surgical resection, autopsy, or clinical course.

Results. Histopathological studies showed 11 biopsy specimens with septate hyphae, indicating a mold, including 6 with Aspergillus, 4 with Mycoraceae, 1 with Aspergillus and Mycoraceae colonies in culture; 2 specimens revealed Candida, 1 Saccharomyces. The remaining 3 biopsies revealed bronchiolits obliterans pneumoniae. Invasive pulmonary mycosis was detected by percutaneous biopsy with 100% sensitivity and 100% (14/14) specificity. Percutaneous biopsy results influenced the surgical decision in 21% (3 of 14) and changed the treatment option in 78% of the cases. Pneumothorax complicated the biopsy in one patient.

Conclusions. Percutaneous computed tomography-guided biopsy is an accurate technique for the diagnosis of invasive pulmonary mycosis in children. It reveals the local epidemiology, correctly determines the therapeutic anti-mycotic agent and influences the choice of prophylaxis for invasive mycosis. Methods. We now report for the first time a modified implantation technique by cannulating the right atrium in order to reach better hemodynamics for right side support. To place the HeartWare system in the area of the right atrium we have created a cavity (8Â8 cm) in the right-sided pericardium using a PTFE membrane.

Results. The design of the HeartWare device enables a quick and less invasive implantation. The small size of the pump allows for intrapericardial placement even in biventricular support. To prevent compromise of the right atrium it is necessary to build a cavity in the pericardium for pump placement.

Conclusions. Adequate pump flow was observed during total support time. The presented implantation technique allows a safe and elegant bridging-to-transplant in Htx candidates representing biventricular failure.

Use of cardiopulmonary bypass for lung transplantation: institutional experience Background. The use of cardiopulmonary bypass (CPB) for lung transplantation is still judged controversial. However, in 30% of lung transplantations CPB support during the surgical procedure is required for hemodynamic and/or respiratory instability or for repair of intracardiac pathologies. This study aimed to determine if the use of CPB has an effect on survival.

Methods. A retrospective review of 190 lung transplants in 180 patients (mean age: 59 years ) with different lung pathologies who underwent bilateral or single lung transplantation between November 1993 and June 2011 was performed. 75 patients (39.4%) in whom elective as well as emergent CPB was necessary were compared with 115 cases (60.6%) without CPB.

Results. The indications for lung transplantation were chronic obstructive pulmonary disease (COPD, n ¼ 111), fibrosis (n ¼ 32), alpha-1 antitrypsin deficiency emphysema (n ¼ 13) and others (n ¼ 34). The follow up period ranged from 2 days to 14 years. Total mean survival was 980 days (range 0 to 5276 days). There was no significant difference in mortality between the CPB and the control group (p ¼ 0.723). The comparison between COPD patients and other indications for lung transplantation also revealed no significant difference in survival (p ¼ 0.11). There was also no difference between COPD patients in whom surgery was performed with CPB or without (p ¼ 0.676). Survival rates in patients over 60 years were similar to patients under 60 independent from the use of CPB (p ¼ 0.676). There was also no significant difference in the usage of CPB between single versus bilateral lung transplantation (p ¼ 0.44).

Conclusions. This study revealed no increase in mortality after lung transplantation when cardiopulmonary bypass was required. When CPB appears necessary it should be employed right away to avoid hemodynamic compromise or severe reperfusion edema due to unacceptable pulmonary arterial pressures.

Erfahrungsbericht aus dem Deutschen Herzzentrum Berlin 

Successful bridging to heart transplantation using the Levitronix CentriMag system and DuraHeart LVAD in a patient with resuscitation related liver injury N. Reiss 1 , L. Kizner 2 , U. Schulz 2 1 Department of Cardiac Surgery Heidelberg, Universitätsklinikum Heidelberg, Germany; 2 Heart Center North Rhine-Westphalia, Ruhr-University of Bochum, Bad Oeynhausen, Germany Background. Mortality rates from cardiogenic shock after acute myocardial infarction remain extremely high. Efforts have been made to develop ventricular assist devices capable of providing complete hemodynamic support in this situation. Mechanical circulatory assistance represents an evident problem when bleeding complications occur. We report a very rare case of successful bridging to heart transplantation despite severe resuscitation related liver injury.

Methods. A 54-year-old male patient underwent failed percutaneous coronary intervention of LAD with consecutive prolonged resuscitation. A Levitronix CentriMag system was implanted via femoral vessels for rapid hemodynamic stabilization. An acute laparotomy was necessary because of severe injury of the left lobe of the liver. During laparotomy the abdominal cave was tamponated by multiple compresses. Three times relaparotomy was necessary to achieve final hemostasis. After four weeks of Levitronix CentriMag support the system was switched to DuraHeart LVAD for long-term assistance as a bridge-to-transplant.

Results. Successful heart transplantation was performed after complete recovery and mobilization of the patient 320 days after the disastrous and hopeless initial situation.

Conclusions. The present case demonstrates that successful bridging to heart transplantation is possible even in cases of severe bleeding complications. Special attention is given to the thin line between bleeding complication and necessary anticoagulation because of mechanical circulatory assist. Background. Advagraf (tacrolimus extended-release capsules) has been tested in de-novo liver and renal transplant recipients. In cardiac transplantation there exists only data on conversion of tacrolimus to advagraf. The aim of this analysis was to evaluate efficacy and safety of de novo use of advagraf in cardiac transplant patients.

Methods. 10 patients received advagraf after ATG induction therapy, 8.1 AE 3.3 days after cardiac transplantation. Mean patient age was 40.1 AE 16.4 years. All patients received also mycophenolate and steroids. Follow-up was 4.4 AE 1.7 months. Dosages, tacrolimus drug levels (tac) and creatinine (crea) levels were recorded 1, 2 weeks and 1 to six months post transplant. Clinical events were defined as acute rejection, infection type, new onset diabetes mellitus (NODM) and renal dysfunction.

Results. Six-month survival was 100%. Two patients were converted to cyclosporine due to NODM 1 and 3 months post transplant. No rejection episodes were recorded during follow-up. Two infections were documented 6 months after transplant (bacterial pneumonia and CMV infection). Advagraf starting dose was 6.2 AE 1.9 mg/d. Crea before start was 1.14 AE 0.34 mg/dl. First measured tac levels at steady state (5 days post drug start) were 7.8 AE 3.7 ng/ml. Until the end of the first month advagraf was slowly increased to 10.1 AE 1.9 mg/d with tac levels of 10.7 AE 2.8 ng/ml and crea of 1.22 AE 0.31 mg/dl. Three and six month drug doses were 9.3 AE 2.11 and 8.2 AE 0.8 mg with corresponding tac levels of 11.2 AE 2.9 and 11.2 AE 1.3 ng/ml. Crea was 1.47 AE 0.22 and 1.49 AE 0.32 mg/dl. There were no events of renal failure.

Conclusions. Advagraf de-novo therapy shows acceptable efficacy and safety early after cardiac transplantation. ATGinduction therapy might be responsible for a low risk of acute rejection despite lower early tac levels.

Usage of If-inhibitors in regulation of heart rate after heart transplantation Background. Due to denervation of the heart from N. vagus, patients physiologically have a heart rate of 90-100 beats per minute (bpm) after heart transplantation. Inhibiting the funnychannels (If) of the pacing-cells of the sinus node reduces ionic-influx and spontaneous depolarization and hereby decreases the heart rate. To improve myocardial oxygenation by extending diastolic time, reducing the heart rate is an important goal for these patients. Objective of the current study is to show the efficacy of the If-inhibitor ivabradine (Procoralan + ) on the transplanted heart.

Methods. This is a single-center retrospective study including 143 patients, who underwent heart transplantation from 1985 to 2011 at our center in Vienna and who were in need of a heart rate regulating therapy after transplantation. Only patients in stable conditions and with sinus rhythm were treated with ivabradine. An average dose of 10 mg per day was used to control the heart rate. Analyzed data were collected during routine checkups in our long-term outpatient-clinic and during ergometry examination by periodical measurements of heart-rate, bloodpressure and ECG.

Results. Mean age at the time of transplantation was 51.3 AE 11.9 years, 117 patients (81.8%) were male (age 16-71 years) and 26 patients (18.2%) were female (age 14-68 years). Before establishing the Procoralan + therapy, patients had an average heart rate of 96.2 AE 11.9 bpm. With the Procoralan + therapy, patients had an average heart rate of 83.1 AE 9.5 bpm. After an average time of 1.8 AE 1.4 years, usage of ivabradine has reduced the heart rate by 14.9 AE 9.4%. In 7 cases (4.8%) we had to terminate therapy because of gastro-intestinal side effects and in 4 cases (2.7%) even under lowest ivabradine dosage patient developed bradycardia.

Conclusions. Using ivabradine after heart transplantation is a safe way to lower heart rate and could become a new indication for If-inhibitors. The individual dosage for each patient has to be found in augmenting the initial daily dosage of 5 mg slowly to the optimum dosage of 10 mg per day.

Comparison between referral and explant diagnoses in lung transplant recipients: discrepancies and additional findings Background. Lung transplantation is a widely accepted therapeutic option for a range of pulmonary conditions in which the diagnosis is often based on clinical data or on limited biopsy material. Posttransplantation complications and recurrence of underlying disease may be related to the primary disease, and an accurate diagnosis is therefore essential.

Methods. A pathologic review was performed on 1056 primary lung transplantations over a period of 22 years (1998 to 2010). Diagnoses of native lungs were compared with referral diagnoses to assess the presence of discrepancies or expanded results like malignancies or infections.

Results. 73 (7%) cases presented a different or expanded diagnosis. Discrepancies between referral diagnosis and histopathology were found in 34 of 1054 cases (3%). The highest percentage of discordance was depicted in chronic obstructive lung diseases (12 of 344), with the final diagnosis of UIP (n ¼ 4), chronic interstitial fibrosis (n ¼ 4), silicosis (n ¼ 2), LAM (n ¼ 1) and sarcoidosis (n ¼ 1). 16 patients who were referred with the diagnosis of an interstitial lung disease had predominantly emphysema (n ¼ 12), bronchiectasis (n ¼ 2) and histiocytosis X (n ¼ 2). Expanded results included Aspergillus (n ¼ 11) and mycobacterial (n ¼ 16) infections, carcinomas (n ¼ 10), cystic adenomatoid dysplasia (n ¼ 1) and carcinoid (n ¼ 1). However, short-and long-term survival was not different in patients with different diagnoses, malignancies or implanted infections. Interestingly all mycobacterial infections and all malignancies occurred in patients with COPD.

Conclusions. On account of this high rate of discrepancies and its possible influence on survival, frequently repeated clinicopathologic investigations should be performed during the waiting list period. 

Background. Induction therapy with alemtuzumab, followed by lower than conventional intensity post-transplant immunosuppression, has been associated with reduced morbidity and mortality in abdominal and heart transplantation. We performed a prospective randomized trial in lung recipients receiving alemtuzumab in conjunction with reduced immunosuppression compared with patients receiving thymoglobulin in association with routinely dosed immunosuppression.

Methods. 60 lung TX recipients were prospectively randomized in two groups: group A received alemtuzumab in conjunction with early lower-dose tacrolimus, lower-dose steroids, and half-dose mycophenolate mofetil, compared with group B receiving thymoglobulin in association with routinely dosed immunosuppression. Survival analyses examined patient and graft survival, freedom from acute cellular rejection (ACR), lymphocytic bronchiolitis, bronchiolitis obliterans syndrome (BOS), kidney function, infectious complications and post-transplant lymphoproliferative disorder (PTLD).

Results. There were no significant differences in 6-and 12month survival (alemtuzumab 96% vs. ATG 93% and 93% vs. 96%, respectively, p ¼ NS). Acute cellular rejection episodes ¼ A2 within the first TX year were significant lower in group A (alemtuzumab 0 vs. ATG 0,33; p ¼ 0.019), lymphocytic bronchiolitis was not different between the groups (cumulative B scores group A 2.9 AE 2.7 vs. group B 3.2 AE 2.3 per patient, p ¼ 0.74). There were no significant differences in bacterial (group A 2. Conclusions. Alemtuzumab induction in conjunction with reduced immunosuppression significantly reduces higher grade rejection rates with comparable survival results and infectious complications to high-dose standard immunosuppression. The incidence of early BOS was higher after alemtuzumab-induction. Background. Cardiac allograft vasculopathy (CAV) is a major cause of allograft failure in long-term heart transplant patients. Even without typical angina symptoms and diffuse lesions, CAV resembles atherosclerosis, but shows more concentric rather than eccentric intimal proliferation. Both, proximal and distal portions of the coronary tree are involved. Cardiac angiographies are periodically performed in the routine long-term follow up after heart transplantation.

Methods. Cardiac angiographies and interventions have been performed on 19 patients (17 male and 2 female) without any complications and substantial advantage in this cohort of patients. 6 Patients already had previous interventions. Mean time after transplantation was 11.4 AE 5.8 years.

Results. Follow-up angiography showed no in-stent restenosis in the patients with previous interventions after a mean time of 36 AE 18 months. 3 patients were in need of interventions, 2 of them in LADp3 with endeavour stents and 1 patient in RCA1 with cypher stent. No complications have been observed.

Conclusions. Radial angiography is a safer method for routine follow-up for patients after heart transplantation, especially in long-term follow-up, as femoral approach often leads to complications.

tation and repeated antigen immunoadsorption (Glycosor-bABO) until the isoagglutinine titre was less than 1:8. In addition intravenous immunoglobulin (Intratect, Biotest Pharma) was administrated at a dose of 1.0 g/kg body weight after each immunoadsorption. A standard immunosuppressive regimen (tacrolimus with a target level of 10 to 12 ng/ml, mycophenolate mofetil at a dose of 1 g twice a day and prednisolone) with IL-2 antibody induction was started approximately two weeks prior to kidney transplantation.

Results. A total of seven ABO incompatible kidney transplantations were performed. Patients' characteristics are summarised in Table 1 . All patients showed an immediate graft function and their serum creatinine decreased to a median level of 1.69 mg/dl (range: 1.09-3.39 mg/dl) on day 7. In one patient there was an early loss of graft function due to thrombotic microangiopathy despite rescue treatment with bortezomib and eculizumab leading to graft loss on day 21. After a median follow up of 3.7 months, serum creatinine averaged 1.7 mg/dl (range: 1.25-2.27 mg/dl). In the performed renal biopsies there was no evidence of relevant acute or chronic graft injury, all allografts showed C4d positive immunostaining.

Conclusions. In conclusion, our small case series demonstrated that perioperative isoagglutinine removal in combination with rituximab is an effective and safe protocol for ABO incompatible transplantation. Therefore we offer ABO incompatible kidney transplantation to patients as alternative to cadaveric kidney transplantation to shorten the waiting time on dialysis. Background. Amyloidosis is a protein misfolding disorder where conformationally changed proteins are pathologically deposited as abnormal insoluble fibrils in distinct tissues potentially leading to morphological and functional disintegration. Heredi-tary amyloidosis is a rare autosomal-dominant disorder arising from mutations in several genes like in the fibrinogen A-chain that affect various organs. Mutations in the fibrinogen A-chain have been discussed to lead to a predominant renal deposition of amyloid generally leading to ESRD.

Methods. Here, we describe a large Spanish family with 101 members affected by the rare fibrinogen R554L mutation and isolated renal amyloidosis.

Results. Most patients presented with late-onset gross proteinuria and a variable course of decline of renal function. In contrast to other reported fibrinogen A mutations, extrarenal involvement or accelerated arteriosclerosis employing cardiac magnetic resonance imaging, carotid ultrasound and tissue biopsy was not observed indicating a highly selective process of renal amyloid deposition. Histological analysis revealed almost complete obliteration of the glomerular and specifically the mesangial architecture despite moderately reduced kidney function. Treatment with angiotensin-converting enzyme and subsequently angiotensin-II receptor inhibitors did not significantly affect proteinuria. Two affected members underwent renal transplantation with the longest surviving graft exhibiting 13 years of survival and graft failure due to surgical and infectious problems, but recurrence of amyloidosis.

Conclusions. Collectively, these results introduce a new perspective on this atypical fibrinogen mutation with significant pathophysiological and clinical impact also with regard to novel therapeutical approaches. Methods. 217 enteric drained whole pancreas transplants (PTx) in 208 patients performed at our center during a seven year period were retrospectively analysed. Prophylactic immunosup- Type II DM was defined as requirement of insulin or oral antidiabetics in the presence of C-peptide production to maintain blood glucose levels <200 mg/dl. Results. Actuarial patient, pancreas and kidney graft survival at one year were 96.4%, 88.5% and 94.8%. At last follow up, two patients were lost, 30 patients died, 26 pancreatic grafts and 16 renal allografts were lost. A total of 17 patients were C-peptide positive at the time of PTx; three patients were excluded from analysis due to early graft loss (n ¼ 2) and DM type II due to partial insufficiency of a previous pancreatic allograft (n ¼ 1). Of the 14 study patients (13 men/1 woman with a median age of 56 [range 41-62] years, two died [invasive fungal infection 1, intracerebral bleeding 1]), another two lost their pancreatic graft [hyperacute rejection 1, chronic rejection 1]) but none their renal graft. Rejection rate was 28.6%, infection rate 64% and surgical complication rate 50%. Median serum creatinine at 5 years follow up was 1.3 (range 0.9-3.6) mg/dl and HbA1c was median 3.2 (range 0.9-6.1)%.

Conclusions. Pancreas transplantation seems to be a good therapeutic option of selected patients with type II DM. Due to the fact that these patients are older than their counterparts with type I DM and the fact that we found these patients to have a high infection rate, less intensive immunosuppressive regimens are recommended. Ergebnisse. Von allen 1064 Nierentransplantierten waren 132 Patienten (12 %) Typ-1-oder Typ-2-Diabetiker und hatten 113 Patienten (11 %) bereits PTDM. Aus n ¼ 307 OGTTs ergab sich PTDM bei 29 Patienten (9 %) und IGT bei Patienten 62 (20 %). Signifikante Prädiktoren für PTDM und/oder IGT beinhalteten höheres Lebensalter, Immunsuppression mit Tacrolimus, höheres C-reaktives Protein und niedrigeres Serum-Albumin. OGTTs mit Bestimmung von Insulin und C-Peptid (n ¼ 105) zeigten im Vergleich mit 1357 OGTTs nicht-transplantierter Patienten (Datensatz G. Pacini) eine erhöhte Insulinsensitivität (OGIS: 414AE 67 vs. 316AE 53 mLÂmin -1 Âm -2 ; ISIcomp: 6,6 AE 4,5 vs. 3,9AE 2,4, beide p < 0,001). Diese Ergebnisse waren in Sensitivitäts-Analysen gematchter OGTTs (nach Alter, BMI, Geschlecht, 2h-Blutzucker im OGTT) konsistent, des weiteren hatten Nierentransplantierte mit Normoglykämie (2h-Blutzucker im OGTT < 140 mg/dL) ebenso wie mit Hyperglykämie (2h-Blutzucker im OGTT 140-199 mg/dl bzw. >200 mg/ dL) signifikant bessere Insulinsensitivität als die Normalbevölkerung ohne oder mit IGT bzw. Typ-2-Diabetes. Die Regressions-Kurven der Insulin-Ausschüttung und des insulinogenen Index, aufgetragen gegen den 2h-Blutzucker im OGTT, wiesen zwischen Transplantierten und Nicht-Transplantierten eindeutige Unterschiede auf, mit besonders starkem Abfall der Insulinsekretion bei Nierentransplantierten mit 2h-Blutzucker-Werten >200 mg/dL (Insulin-area under the curve (AUC) 3,4 AE 1,9 vs. 6,1 AE 3,4 mU/mL 2h, p ¼ 0,002). Eine Validierung aller verwendeter OGTT-Indizes erfolgte anhand 23 euglykämisch-hyperinsulinämischer Clamps.

Schlussfolgerungen. Insulin-Sekretionsverlust könnte das dominante Problem bei der Entwicklung eines PTDM nach Nierentransplantation sein und verlangt nach therapeutischen Strategien zum Schutz der Betazellen.

Hämoglobinvariabilität ist assoziiert mit Mortalität nach Nierentransplantation Ergebnisse. Die Hazard Ratio (HR) fü r Mortalität und Transplantatversagen stieg signifikant mit der Hämoglobinvariabilität an. Im linearen Modell betrug der Anstieg fü r die Mortalität 2,35 (95 % Konfidenzintervall 1,75-3,17; p < 0,001), fü r das funktionelle Transplantatversagen 2,45 (1,76-3,40; p < 0,001). In einem klinischen Expertenmodell, das fü r die Verwendung von ESA, Hämoglobin, Alter bei der Transplantation, Diabetes, Tage an der Dialyse, glomeruläre Filtrationsrate, Abstoßung und Jahr der Transplantation adjustiert wurde, war die Hämoglobinvariabilität mit der Mortalität assoziiert (HR: 2,11; 1,51-2,94; p < 0,001), jedoch nicht fü r das funktionelle Transplantatversagen (HR: 1,34; 0,93-1,93; p ¼ 0,121). Ein ähnliches Ergebnis erhielten wir beim Purposeful Selection Modell: Die Hämoglobinvariabilität war signifikant assoziiert mit Mortalität (HR: 2,63; 1,70-4,08; p < 0,001) und nicht signifikant mit dem funktionellen Transplantatversagen (HR: 1,27; 0,63-2,54; p ¼ 0,509).

Schlussfolgerungen. Hämoglobinvariabilität unter ESA Therapie nach Nierentransplantation ist mit erhöhter Mortalität assoziiert.

Influence of recipient CYP3A5 genotypes on the pharmacokinetics of tacrolimus in liver transplant recipients Background. Tacrolimus, which is widely used in liver transplant recipients, shows high interindividual variability in pharmacokinetics. It is primarily metabolized by hepatic cytochrome P450 3A4 and 3A5 (CYP). CYP3A5 is also expressed in the kidney and in the intestine. In this study, we evaluated the influence of recipient CYP3A5 genotypes on the pharmacokinetics of tacrolimus in patients after liver transplantation.

Methods. Patients after liver transplantation with tacrolimus maintenance therapy were included into the study. CYP3A5 genotypes of the recipients were established. Clinical and laboratory data at various time points (1 month, 3 months, 1 year and 3 years after start of medication with tacrolimus) were evaluated retrospectively. Doses and trough levels of tacrolimus were noted and a screening of all concomitant medications with possible CYP3A5 interaction was performed. SPSS for windows (release 14.0; SPSS, Inc) was used for statistical analyses. Continuous variables were analyzed by ANOVA-test and presented as means AE standard deviation, variables deviating from normal distribution were analyzed by rank sum test and presented as median and range. Categorical variables are presented as percentages and were compared by chi-square test. The criterion for statistical significance was p < 0.05.

Results. Ninety-two liver transplant recipients participated in the study. CYP3A5 genotypes were successfully determined in all subjects and did not deviate from the Hardy-Weinberg equilibirum. 86% of the patients carried the CYP3A5 *3/*3 genotype and were thus classified as CYP3A5 non-expressors. 14% carried the CYP3A5 *1/*3 genotype and were classified as heterozygous expressors. No homozygous CYP3A5 expressor (*1/*1) was found. Neither tacrolimus dose nor levels were significantly different between CYP3A5 expressors and non-expressors at any point of time

Conclusions. The pharmacokinetics of tacrolimus in patients after liver transplantation is not influenced by the recipient CYP3A5 genotype. It is mostly the hepatic metabolism that contributes to the excretion of tacrolimus. The donor CYP3A5 genotype might be useful to predict the tacrolimus pharmacokinetics.

Background. Due to predisposing factors like massive ascites or kachexia incisional hernias occur in about 30% of patients after LTX. Following LTX the immunosuppressive regimen has been associated with the development of incisional hernias. Appropriate closure techniques are still controversially discussed. The usage of Mesh grafts is feasible in those patients as direct closure often results in hernia reccurrence.

Methods and Results. We would like to present a patient who suffered from an incisional hernia 6 months following LTX. The hernia was repaired using a Mesh graft (Permacol + ) which was placed in a sublay technique. The patient primarily recovered and was dismissed on day 7 following hernial repair. After 14 days he presented with fever and general illness. An abdominal CT scan revealed a fluid formation above the Mesh graft. Open drainage had to be performed. The healing of the wound was impaired with the Permacol + graft completely visible in the wound. VAC was applied directly on the Permacol + patch. The wound closed completely without any sings of reinfection or peritonitis until now. An abdominal CT scan 4 months showed no pathologies.

Conclusions. The usage of a Permacol + patch seems likely to repair incisional hernias after LTX whereas severe wound infections might occur after the placement of biological grafts due to immunosuppression. In our patient a VAC treatment was applied and managed the developed infection well even if it had to be placed directly on the patch. Grundlagen. Empfänger-Desensibilisierung mittels Blutgruppen-spezifischer Immunadsorption (ABO-IA) ist derzeit das Mittel der Wahl, um eine erfolgreiche Transplantation über die ABO-Blutgruppen-Barriere zu ermöglichen. Zu diesem Zweck wäre der Einsatz von regenerierbaren, nicht-Antigen-spezifischen (semiselektiven) Adsorbern ebenfalls denkbar, aber die Effizienz dieser Immunadsorptionsmaterialien wurde in diesem Zusammenhang noch nicht getestet.

Methodik. Acht mit ABO-IA behandelte Transplantationskandidaten und 39 Patienten, die mit verschiedenen Indikationen außerhalb der ABO-inkompatiblen Transplantation mit semiselektiver Immunadsorption behandelt wurden, wurden in die Studie inkludiert. Die semiselektive Behandlung basierte auf IA mit Protein A, Peptid-Ligand und Anti-human Immunglobulin Antikörpern. Antikörper-Muster (IgG, IgG1-4 Subklassen, IgM und Komplement-fixierende Reaktivitäten) wurden mit Durchflusszytometrie und konventionellen Agglutinationstests analysiert.

Ergebnisse. Die sensitive Durchflusszytometrie-Analyse der mit ABO-IA desensibilisierten Transplantationskandidaten ergab eine profunde, aber auch oft unvollständige Reduktion der ABO-Blutgruppen-spezifischen Antikörper (anti-A/ B). Persistierende Komplement-oder auch Nicht-Komplement-fixierende Reaktivitäten konnten jedoch nicht mit Abstoßung oder mit C4d-Ablagerung in Kapillaren von Protokoll-Biopsien assoziiert werden. Beim Vergleich zwischen semiselektiver und selektiver IA im Rahmen einer einmaligen Behandlungssitzung stellten sich die semiselektiven Adsorber als effizienter in der Reduktion von anti-A/B IgG heraus (Reduktion der Mediane auf 28 % der Ausgangswerte versus 59 % bei ABO-IA, p < 0,001). Im Gegensatz dazu wurden Blutgruppen-spezifisches IgM (74 % versus 30 %, p < 0,001) und IgG3 (72 % versus 42 %, p < 0,05) weniger gut dezimiert, ohne erkennbare Unterschiede zwischen den getesteten semiselektiven Adsorber-Typen. Eine Analyse nach vier konsekutiven IA-Sitzungen zeigte, dass die unterlegene Effizienz nicht durch wiederholte Behandlungen wettgemacht werden konnte.

Schlussfolgerungen. Unsere Beobachtung einer limitierten Adsorptionskapazität bezü glich bestimmter Blutgruppenspezifischer Immunglobulin-(Sub-)Klassen lässt zu erhöhter Vorsicht bei der Verwendung von semiselektiven Immunadsorptionstechniken bei ABO-inkompatibler Nierentransplantation raten.

New immunosuppressive strategies for preserving renal function and improving outcome J. M. G. Boira

The prevention of acute rejection (AR) has been the main goal of new immunosuppressants (IS) introduced in renal transplantation in the last decades. Nowadays -with the use of calcineurin inhibitors as mainstay IS -the rejection rates are low. However the nephrotoxicity of these agents contribute to the chronic allograft injury which may result in graft loss in the long term. New IS agents should effectively prevent AR and preserve renal function. New biologic and small molecule agents are devoid of nephrotoxicity and interfere with crucial mechanism of allo-immune response. During the presentation recent data on new agents and regimes under development in the prevention of AR will be discussed. Das Ziel ist es daher, zum einen die Empfänger T-Lymphozyten zu eliminieren oder zu tolerisieren und zum anderen die Spender T-Lymphozyten zu tolerisieren. Eine Methode, die sich zur Eliminierung und Tolerisierung der Empfänger T-Lymphozyten in der Knochenmarktransplantation bewährt hat, ist, neben der medikamentösen Therapie, die total nodale Lymphknotenbestrahlung. Damit können Abstoßungsreaktionen selbst bei HLA-nichtidenten Knochenmarktransplantationen weitgehend verhindert werden. Zur Behandlung der GvHD hat sich in der allogenen Knochenmarktransplantation die extracorporale Photopherese (ECP) bewährt, die durch im einzelnen nicht vollkommen aufgeklärte Mechanismen eine Toleranz induziert und somit in der Lage ist, die GvHD ohne intensive Immunsuppression zu mildern oder auch ganz zu eliminieren.

Anhand von Fallbeispielen wird diskutiert werden, dass die total nodale Lymphknotenbestrahlung und ECP auch in der Organtransplantation eine Rolle spielen könnten, da die Mechanismen der Toleranz oder Abstoßung sich prinzipiell nicht von einer Knochenmarktransplantation unterscheiden.

Plenarsitzung IV: Immun-und Organtoleranz/Toleranzinduktion II 067 Approaches to tolerance: mouse to man

Harvard Medical School, Division of Transplantation, Massachusetts General Hospital, Boston, MA, USA Over the past two decades, non-specific immunosuppression (I.S.) incorporating polyclonal and monoclonal antibodies, calcineurin inhibitors (CNI), mTOR inhibitors, anti-metabolites and steroids have greatly improved early allograft survival rates. However, the rate of chronic allograft loss has decreased only modestly over that observed earlier. Following kidney transplantation, for example, the rate of chronic attrition is discouragingly similar to that observed two decades ago, generally resulting from "chronic rejection" or, perhaps more accurately, "chronic allograft nephropathy". The relentless progression of similar histopathologic processes is manifested as vasculopathy in the coronary arteries of heart allografts (CAV) and in lung transplants as obliterative bronchiolitis (OB) despite the long-term administration of currently available I.S.

Thus, the ultimate goal of I.S. for transplant recipients is the development of long-term donor-specific nonresponsiveness which leaves responses to other foreign stimuli intact. Successful induction of "transplant tolerance" would avoid the complications of chronically administered I.S. and reduce the risks of chronic rejection. Numerous approaches employing both peripheral and central tolerance mechanistic strategies have been successfully developed in rodent models. Currently, several of these approaches appear to be clinically applicable, having now proved to be effective in large animal models and preliminary clinical trials. The most successful to date has been induction of hematopoietic mixed chimerism.

The mixed chimerism approach, an initially presumed central tolerance strategy, has been extensively studied in our laboratories, first in rodent, non-human primate, and porcine models, and now in human recipients of renal allografts. Using a non-myeloablative conditioning regimen followed by donor bone marrow (DBM) transplantation, we observed successful production of mixed chimerism in non-human primate renal allograft recipients, followed by normal kidney function with no evidence of chronic rejection for periods as long as thirteen years after discontinuing all I.S. In vitro studies of these recipients as well as of humans suggest that both central (clonal deletion) and peripheral (regulatory cells) mechanisms are involved.

Encouraged by the pre-clinical studies, we have to date extended this approach to 20 human renal allograft recipients. Ten individuals with ESRD secondary to multiple myeloma received a nonmyeloablative preparative regimen consisting of perioperative cyclophosphamide, equine ATG, CsA, and thymic irradiation. The recipients underwent HLA-matched kidney transplantation followed by iv infusion of DBM and subsequent donor lymphocyte infusions (DLI) to enhance the anti-myeloma effect. Transient lymphohematopoietic chimerism was achieved in all patients and only two reversible rejection episodes have occurred with a follow-up of 1 to 13 years after discontinuing I.S. Longterm control of the underlying malignancy was better than with any other currently available treatment, but 4 of these individuals required additional therapy for recurrent myeloma.

Ten patients with ESRD and no myeloma received HLA-mismatched kidneys after a similar conditioning regimen except that MEDI-507, an anti-CD-2 mAb, was used in place of ATG and DLI was not used. Transient mixed chimerism and apparent allograft tolerance has been achieved in 7 of these recipients. These 7 patients have received no I.S. for periods of 2-9 years. Two allografts were lost: acute humoral rejection (1); thrombotic micro angiopathy (1) and the last recipient has been returned to chronic I.S. after developing acute rejection when I.S. was withdrawn.

These observations plus similarly encouraging observations in several clinical trials elsewhere suggest that tolerance induction may become a more widely applicable clinical reality.

Experimental rationale for using hematopoietic stem cell transplantation for tolerance induction

It was recognized a long time ago that an individual that shares hematopoietic cells with another, becomes tolerant to this individual. Thus, a recipient of a successful donor bone marrow (BM) transplant (BMT) becomes tolerant toward the donor. Triggered by this observation experimental studies have been systematically addressing why and how donor BM transplantation achieves tolerance. The concept of mixed chimerism -in which a substantial donor hematopoietic cell proportion co-exists with the recipient hematopoietic repertoire following donor BMT -has emerged as a promising BMT-based tolerance strategy.

Mixed chimerism leads to a particularly robust and durable type of tolerance. Central clonal deletion of immature donorreactive T cells was identified as a key mechanism of tolerance in mixed chimeras and seems to be in large part responsible for the robust nature of tolerance achieved. Peripheral clonal deletion of mature donor-reactive T cells and non-deletional (i.e. regulatory) mechanisms also contribute to tolerance induction in some models. Overall, donor BMT emulates many of the physiologic mechanisms controlling self-tolerance, extending them to donor antigens.

While clinical translation of the mixed chimerism approach has been achieved in pilot trials, large-scale application in routine clinical practice awaits minimally toxic BMT regimens. Myelosuppressive and cytotoxic pre-treatments commonly employed in 'conventional' BMT recipients would be unacceptable in organ transplant recipients. Therefore, efforts to develop advanced, non-toxic murine BMT protocols are ongoing. The use of costimulation blockers and, more recently, adjunctive T regulatory cell therapy have yielded dramatically less toxic regimens which hold promise for clinical translation. Aspergillus und seltene Schimmelpilzinfektionen. Die Inzidenz der Aspergillose bei der allogenen Stammzelltransplantation beträgt zwischen 8-25 %. Aspergillussporen kommen ubiquitär in der Umwelt vor; ungefilterte Luft aus Belü ftungssystemen und Staub bilden die Hauptinfektionsquellen. Aspergillussporen werden exogen durch Inhalation erworben. Die Reduktion der Sporenlast in der Umgebung von Patienten "at risk" ist die wichtigste prophylaktische Maßnahme. Der klinisch wichtigste Vertreter ist Aspergillus fumigatus. Die Letalität invasiver Aspergillosen liegt bei 70-90 %. Zu den anderen Schimmelpilzinfektionen zählen die Zygomykosen (Absidia-, Mucor-und Rhizopus-Spezies), die insbesondere bei Patienten mit Diabetes mellitus oder zunehmend bei immunsupprimierten Patienten unter Voriconazol Therapie auftreten. Die Letalität beim Immungeschwächten ist annähernd 100 %. Bei Empfängern solider Organtransplantate konnte durch Neuerungen in der Chirurgie und Intensivierung der Immunsuppression das Transplantatü berleben verbessert werden. Mit bis zu 15 % ist die Inzidenz bei Herzund Lungentransplantation am höchsten. Nach Lungentransplantation treten neben der typischen pulmonalen Aspergillose gehäuft ulzerierende Tracheobronchitis und Aspergillose der Bronchusanastomose auf. Das geringste Risiko einer invasiven Aspergillose besteht bei Nierentransplantierten mit 1 % Inzidenz, die Mortalität ist generell hoch.

Institut für Hygiene, Mikrobiologie und Umweltmedizin, Medizinische Universität Graz, Graz, Österreich Pilzinfektionen spielen bei Immunsupprimierten, vor allem bei Patientinnen und Patienten nach Organ-und Stammzelltransplantationen, eine steigende Rolle. Dabei reicht das Spektrum der verursachenden Pilze von Hefen der Gattung Candida zu Schimmelpilzen der Gattung Aspergillus, wobei auch eine Zunahme von Infektionen mit anderen Schimmelpilzen wie den Zygomyzeten, Fusarien, Scedosporium/Pseudallescheria und anderen zu beobachten ist. Um eine zielgerichtete Therapie einleiten zu können, ist eine schnelle und genaue Diagnostik unumgänglich, da die einzelnen Pilzarten teils unterschiedliche Antimykotika-Empfindlichkeitsspektren aufweisen.

Neben den klassischen Kulturmethoden, die trotz ihrer relativen "Langsamkeit" nach wie vor unverzichtbar sind, wurden in den letzten Jahren Techniken entwickelt, die teilweise innerhalb weniger Stunden Ergebnisse bringen können -vorausgesetzt, sie werden richtig angewandt und interpretiert. Dazu zählen insbesondere bildgebende Verfahren, molekularbiologische Methoden und Antigen-Nachweise.

Idealerweise beruht die Diagnose einer Pilzinfektion auf der Kombination mehrerer Techniken, man spricht auch von einer Puzzle-Diagnostik.

Die einzelnen Techniken zur Diagnose von Pilzinfektionen in der Organ-und Stammzelltransplantation sollen vorgestellt und deren Vor-und Nachteile diskutiert werden.

Therapie von Pilzinfektionen in der Organ-und Stammzelltransplantation

Universitätsklinik für Innere Medizin, Medizinische Universität Graz, Graz, Österreich Pilze verursachen bei Immunsupprimierten infektiöse Komplikationen, die von oberflächlichen Infektionen der Haut/ Schleimhäute bis zu systemischen, lebensbedrohlichen Organinfektionen oder Fungämien reichen. Auslöser dieser Infektionen sind meist Candida-Arten, Aspergillus-Arten, andere Schimmelpilze wie Zygomyceten, und seltene andere Pilze. Zur Therapie der systemischen Infektionen stehen Polyene (z. B. Amphotericin B, lipid-assoziierte Amphotericin B Präparate), Azole (z. B. Fuconazol, Itraconazol, Voriconazol, Posaconazol) und Echinocandine (z. B. Caspofungin, Anidulafungin, Micafungin) zur Verfügung. Fluconazol ist ausschließlich gegen Candida Arten wirksam. Die anderen oben genannnten Antimykotika sind in vitro sowohl gegen Candida-als auch Aspergillus-Arten wirksam. Für die Therapie von Candida-Infektionen eignen sich abhängig von der lokalen Epidemiologie Fluconazol, Echinocandine oder Amphotericin B-Präparate. Für die Therapie von Aspergillus-Infektionen eignen sich Voriconazol, Posaconazol, Amphotericin B-Präparate oder Echinocandine. Für Zygomkosen stehen Amphotericin B-Präparate und Posaconazol zu Verfügung. Für seltene Pilzinfektionen sind adjuvante Therapeutika (wie z. B. Flucytosin) vorhanden. Für den Behandlungserfolg der schweren lebensbedrohlichen Infektionen ist der Zeitpunkt des Therapiebeginns und das Spektrum der antifungalen Therapie relevant. Es werden empirische, kalkuliert-präemptive und zielgerichtete Therapien unterschieden. Bei machne Pilzinfektionen sind chirurgische Sanierungen zusätzlich zur antimykotischen Therapie notwendig.

Die einzelnen Therapioptionen zur Behandlung von Pilzinfektionen in der Organ-und Stammzelltransplantation sollen vorgestellt und deren Vor-und Nachteile diskutiert werden.

Erlebenüberlebenund dann? Lebensqualität nach Herztransplantation

Klinische Abteilung für Herzchirurgie, Universitätsklinik für Chirurgie, Medizinische Universität Wien, Wien, Österreich Aus wissenschaftlicher Perspektive ist der Begriff "Lebensqualität" ein nicht direkt beobachtbares Konstrukt, das die Bewertung des physischen, psychischen und sozialen Zustandes einer Person in seiner Zusammenschau bezeichnet. Eine gute Annäherung kann auch durch den Begriff "Lebenszufriedenheit" erfolgen. Im Vortrag soll vor allem auf Langzeit-Lebensqualitätsstudien und die psychische Verarbeitung der Herztransplantation Bezug genommen werden: wie ändert sich das Leben der PatientInnen nach einer oft schicksalhaften schweren Erkrankung und lebenserhaltenden Transplantation? In einer prospektiven Studie unserer Abteilung über Lebensqualität ein, fünf und 10 Jahre nach Herztransplantation zeigte sich z. B. eine Schere zwischen konstant durchaus guter Lebensqualität in physischer und zunehmend abnehmender Lebensqualität in physischer Hinsicht (v. a. Depression, Labilität und Dysphorie). Es werden Gründe für diese Differenz diskutiert, Lösungsmöglichkeiten besprochen. Ferner ist die Frage interessant, ob Persönlichkeitsfaktoren Einfluß haben auf Langzeitüberleben und Lebensqualität bzw. -zufriedenheit: wer lebt länger, wer lebt besser? Es werden klinisch-psychologische Studien vorgestellt, die belegen, dass ein bestimmtes Persönlichkeitsmuster (Typ D, Denollet 1996) Für die drei am häufigsten durchgeführten pädiatrischen Organtransplantationen (Niere, Leber, Herz) wird der jeweils aktuelle Forschungsstand referiert. Darüber hinaus werden zwei spezifische Problembereiche in der pädiatrischen Organtransplantation dargestellt, das Problem der Compliance/Adherence und der Transition von der Kinder-und Jugendmedizin in die Erwachsenenmedizin. Die Studien unterscheiden sich hinsichtlich der Operationalisierung der LQ, des Studiendesigns, der Dauer des Follow-ups und des Altersbereichs der Kinder. Prospektive Längsschnittstudien mit größeren Stichproben in multizentrischer Zusammenarbeit unter Einbeziehung möglichst spezifischer Erhebungsmethoden sind erforderlich, um die postoperative Entwicklung der Kinder besser beurteilen zu können. Vor dem Hintergrund einer neuen Ä ra der immunsuppressiven Therapie (Steroidminimierung, individualisierte Therapie) kann hier zukünftig eine weitere Steigerung der LQ erwartet werden. Background. Autologous stem cell transplantation (ASCT) is a curative procedure for a variety of haemato-oncological malignancies and depends on the transplantation of sufficient CD34þ cells (threshold of >2Â10 6 /kg) to ensure sustained engraftment. In the last decade, peripheral blood stem cells (PBSC) have completely replaced bone marrow (BM) as preferred stem cell graft in ASCT. However, a number of patients fail to mobilize the threshold of >2Â10 6 /kg CD34þ cells. In these poor mobilizers, BM harvest was the only possibility to collect sufficient numbers of stem cells so far. Recently, the chemokine-receptor antagonist plerixafor has shown to enhance stem cell mobilization in patients who demonstrated with previous mobilization failure with an overall success rate of about 70%. We here report 4 patients with mobilization failure in whom G-CSF primed BM as well as plerixafor primed PBSC were collected.

Methods. Four heavily pretreated patients (median of 10 chemotherapy cycles, range 8-17) with non-Hodgkin lymphoma (NHL, n ¼ 3.) and germ cell cancer (n ¼ 1) presented with mobilization failure after 1-7 mobilization attempts. In the 3 NHL patients BM was harvested after unsuccessful PBSC collection with plerixafor. In the patient with germ cell tumor before plerixafor primed PBSC collection was done after insufficient BM harvest.

Results. In the 3 NHL patients plerixafor enhanced PBSC revealed an insufficient number of CD34þ cells (i.e. 0.64-1.15Â10 6 /kg CD34þ cells in 2-3 leukaphereses procedures). Therefore BM was harvested after G-CSF stimulation. However, again we failed to obtain enough CD34þ cells in all of them (0.16 to 0.62Â10 6 /kg CD34þ cells). In the patient with germ cell cancer a collection of CD34þ cells from the BM was carried out as first salvage procedure after the failure of conventional mobilization (0.9Â10 6 /kg CD34þ cells). Thus, in a next attempt PBSCs were mobilized with G-CSF and plerixafor resulting in a successful harvest (3.36Â10 6 /kg CD34þ cells in one leukapheresis).

Conclusions. Reviewing our cases one might speculate that a BM harvest after failure of stem cell mobilization with G-CSF and plerixafor does not yield in sufficient CD34þ cell numbers.

Transient Elastography (FibroScan) for the prediction of liver toxicity following autologous or allogeneic HSCT J. Auberger 1,2 , I. Graziadei 3 , W. Vogel 3 , J. Clausen 1,2 , D. Grundlagen. Lebertoxizität oder das Sinusoidale Obstruktions-Syndrom (SOS) gehören zu den häufigsten Organtoxizitäten nach zytoreduktiver Hochdosis-Chemo-/Radiotherapie bei autologer oder allogener HSCT. Die transiente Elastographie (Fibroscan, FS) ist eine neue, schnell zugängliche und nicht-invasive Methode eine Leberfibrose ü ber die Lebersteifigkeit (LS) zu bestimmen und wird bereits erfolgreich bei Patienten mit Hepatitis C und alkoholischer Lebererkrankung eingesetzt.

Methodik. Zwischen April 2009 und Oktober 2010 wurden 67 Patienten FS-Messungen während der Routine-Abdomensonographie im Rahmen der Transplantvoruntersuchungen vor autologer (n ¼ 37) oder allogener (n ¼ 30) Transplantation zugeführt. Die Konditionierung bestand aus HD MEL (140-200 mg/m 2 ), BEAM oder anderen krankheitsspezifischen Hochdosisschemata im Rahmen autologer Transplantationen. Allogene Transplantationen wurden nach myeloablativer Konditionierung (MAC, n ¼ 18) oder in reduzierter Intensität, hauptsächlich BUFLU basiert (RIC, n ¼ 12) durchgeführt. Die Lebertoxizität wurde anhand des maximalen Serumbilirubins, der Anzahl der Tage mit einem Bilirubin >2 mg/dL und der prozentualen, maximalen Gewichtszunahme bis zum Tag þ20 definiert.

Ergebnisse. Das maximale Serum Bilirubin vor Tag þ20 betrug 1,18 (range, 0,44-11,72) mg/dL, der mediane Zeitpunkt mit einem Serumbilirubin !2 mg/dL war 10,5 (range, 1-20) Tage und die mediane prozentualen Gewichtszunahme war 5,2 % (range, 0,1-17,2). Lebertoxizität wurde häufiger nach MAC vs. RIC Konditionierung beobachtet, wenngleich nicht statistisch signifikant (50 % vs. 33 %, p ¼ 0,25). Patienten, die eine Immunsuppression (IS) mit CsA/MTX erhielten, zeigten häufiger Anzeichen einer Lebertoxizität verglichen mit Pat., die CsA/MMF oder keine IS erhielten (83 % vs. 18 %). Die mediane LS vor Transplantation war 5,4 kP (range, 2,3-62,7). Die LS korrelierte signifikant mit der GOT und GPT vor Transplantation in der gesamten Kohorte und mit GPT, gGT und AP bei Pat. mit mehr als 10d Bilirubin >2 mg/dl bis zum Tag þ 20. Zwischen der LS vor HSCT und Zeichen der Lebertoxizität (definiert als max. Serumbilirubin, Anzahl der Tage mit Bilirubin >2 mg/dL und prozentualer Gewichtszunahme) nach HSCT konnte keine signifikante Korrelation hergestellt werden.

Schlussfolgerungen. Die nicht-invasive transiente Elastographie (Fibroscan) ist nicht wegweisend, um Patienten zu identifizieren, die ein hohes Risiko für die Entwicklung einer Lebertoxizität nach autologer oder allogener Transplantation aufweisen.

Norovirus associated hemophagocytic lymphohistiocytosis (HLH) after unrelated bone marrow transplantation (BMT) in a boy with refractory AML G. Kropshofer, C. Salvador, R. Crazzolara, B. Meister

Background. HLH is a potentially life threatening macrophage activating syndrome which leads to massive hyperinflammation through an exaggerated but ineffective immune response. Diagnosis after BMT is complicated by other inflammatory syndroms like engraftment syndrome, capillary leak syndrome, venoocclusive disease and systemic infections.

Methods and Results. We report on a meanwhile 24month-old boy with AML (M4) relapse. Already in phase of maintenance chemotherapy he acquired a gastrointestinal infection with norovirus. Reinduction chemotherapy was performed with FLAG/Daunoxome and FLAMSA -he never reached a complete remission. So we decided to perform an allogeneic BMT (MUD) in aplasia. Conditioning regimen consisted of iv busulfan, cyclophosphamide and melphalan. There were no early complications despite his ongoing norovirus infection. Leukocyte engraftment was obtained on day þ20. Bone marrow at day þ28 showed a nearly normocellular bone marrow and full donor chimerism. At day þ40 the patient impressed with persistent fever despite antimicrobial therapy, his general status deteriorated. He developed CNS symptoms with vertical nystagmus, increasing hepatosplenomegaly and respiratory insufficiency. Laboratory findings showed hyperferritinemia (45000 mg/l), a subsequent pancytopenia, hypofibrinogenemia, elevated D-Dimer (34000 mg/l) and hypertriglyceridemia (723 mg/dl). Intensive care was necessary including high doses of catecholamines. Subsequent bone marrow aspiration revealed an aplastic bone marrow and hemophagocytosis. The patient underwent ventilation support for 17 days. Treatment included DXM 10 mg/m 2 /d for 14 days, then DXM was tapered to 5 mg/kg/d. Etoposide (150 mg/kg) was given three times until now. Considering the long aplastic phase and the ongoing hemophagocytosis we decided to give him stem cell support (remaining frozen bone marrow from the same donor) without reconditioning. Four days later we went on with a monoclonal anti-CD25 antibody (Simulect + ). The boy improved but therapy is still ongoing. The outcome at the moment is unclear.

Conclusions. HLH still is a rarely seen complication after BMT, therefore early diagnosis is necessary to start therapy promptly. The role of monoclonal anti-CD25 antibody and anti TNF-antibody as part of therapy is unclear yet. Up to now there is no report of norovirus triggering HLH.

Therapeutic effect of allogeneic bone marrow mesenchymal stem cell transplantation on liver chronic damage in rats Methods. 75 male rats divided in 3 groups in which chronic hepatitis was modeled by means of CCL 4 . 1st (n ¼ 25) received single transplantation of 2.5Â10 6 MSC on 3rd day after modelling. In 2nd (n ¼ 25) MSC were injected twice in a dose of 2.5Â10 6 cells on days 3 and 10 after modelling. 3rd group was control group (CG, n ¼ 25). Biochemical indices: ALT, AST, ALP; cytokines (IFN-, TNF-, IL-4, IL-10) were studied during 5 weeks. Liver histology were studied after 7, 14, 21, 28, 60, 90 days. Immunohistochemical methods: -SMA, caspase-3, AFP and PCNA were used.

Results. At the 1-3 weeks the AST, ALT and ALP were increased in all groups, but in the CG level of indices above was higher. Analysis of morphologic changes in liver resulted in following findings: on 7th and 14th day liver fatty degeneration was in all groups, but its appearance was stronger in the 1st and especially in CG. In 60 and 90 days the fibrosis became more severe in CG, but after MSC application a resorption of fibrosis occurred (especially in 2nd). It is important to note that connective tissue forming in liver after MSC application had two-phase dynamics: at first an area of connective tissue sharply increased and then gradually decreased. It was found out that the -SMA and caspase 3 expression after MSC application had also two-phase dynamics: at 1-2 weeks these indices were increased and then (3-4 weeks) began to decrease, becoming lower than in CG. At first weeks cytokines disbalance was identified in all groups, but in MSC groups cytokines disbalance was less expressed on 5th week. AFP and PCNA expression rates were higher in MSC groups, than in CG up to end of examination.

Conclusions. MSCs transplantation for therapy of liver damage can reduce liver fibrosis, but this process has two-phase dynamics (firstly increasing and then decreasing of fibrosis) which can explain the contradictory information on the cell therapy of chronic hepatitis. Background. Treatment of chronic liver failure in pretransplant period is an actual problem of medicine. This investigation was undertaken for working out of biounit containing viable liver cell (LC) and multipotent mesenchymal stromal cells (MMSC), which could support the functioning and recovered regeneration of damaged liver for a long time.

Methods. Chronic liver damage was modeled on Wistar rats by means of 0.3 ml CCl 4 on 100 g rat weights within 6 weeks. Adult Wistar rats were also used as donor cells. Isolated cells were cultivated in 10 days: at first MMSC during 7 days then the same MMSC together with LC in 3 days on Cytodex. Suspension of LC 2.5-4.0Â10 6 cells/cm 3 and MMSC 0.5-0.8Â10 6 cells/ cm 3 was applied on biodegradable matrixes "Sphero + Gel". Formed biounits were transplantated into rat liver. Survival of transplanted cells and liver morphology were investigated in 365 days after transplantation by using hematoxylin and eosin staining, Masson's trichrome staining. Cell viability and cellular phenotype were investigated by trypan blue staining and immunohistochemically (cytokeratine 18 and mitochondrial antigen).

Results. Before biounit transplantation liver damage was characterized by: fatty, lymphoid-cellular infiltration and proliferation of histioblasts and macrophages; porto-portal sclerosis; hydropic dystrophia and focal necrosis of hepatocytes. In 365 days after biounit transplantation it was detected: viable hepatocytes, neogenic plethoric vessels, neogenic plethoric bile duct in biounit. Liver damage (dystrophia, safe structure of a liver, beam structure, fatty vacuoles inside hepatocytes and other indices) were significant less after biounit transplantation than in control group without biounit transplantation. Restoration of hepatic lobe structure was better in studied group.

Conclusions. Our preliminary studies demonstrate that in biounit after transplantation into damage liver there is formation of morphologic structures of liver (viable cell, vessels, bile ducts); other words the used method allows to carry out organotypic liver remodeling into biounit and to support the function of the damaged liver. Thereby the suggested method is a perspective one and can be used as technology of building intracorporeal biounit for the long-term auxiliary supporting of the damaged organs.

Liposomales Amphotericin B als Prophylaxe invasiver Pilzinfektionen nach hämatopoietischer Stammzelltransplantation bei pädiatrischen und adoleszenten Patienten photericin B (LAMB) sind ein breites antifungales Spektrum, eine lange Eliminations-Halbwertszeit, und im Vergleich zu den neuen Azolen weniger Medikamenten-Interaktionen.

Methodik. Im Zeitraum 2009-2010 wurden bei 20 konsekutiven Patienten (medianes Alter: 11,4 Jahre; m:w ¼ 8:12) insgesamt 23 HSZT durchgeführt: 12 autologe (davon 3 Tandem-Transplantationen) und 11 allogene SZT, wegen soliden Tumoren (n ¼ 7), rezidivierter schwerer aplastischer Anämie (n ¼ 5), malignen hämatologischen Erkrankungen (n ¼ 5), Mb. Hurler (n ¼ 2) bzw. Mb Crohn (n ¼ 1). Autolog transplantierte Patienten erhielten im Median 7,46Â10 6 /kg CD34þ Zellen/kg und erreichten am Median Tag þ 9,5 das leukozytäre Engraftment. Allogene Stammzellquellen waren periphere Stammzellen (CD34þ selektioniert und/oder CD3/19 depletiert) in 9 und Knochenmark in 2 Fällen; Spender waren HLA-idente Fremd-(n ¼ 9) bzw. Geschwisterspender (n ¼ 1) und ein haploidenter Elternteil. Die mediane Stammzellzahl lag bei 15,37Â10 6 CD34þZellen/kg; das leukozytäre Engraftment erfolgte im Median am Tag þ 11. Bei 5 Patienten trat eine Graft-versus-host-Erkrankung (GVH) auf, davon einmal eine GVH IV. Die LAMB-Prophylaxe wurde in einer Dosis von 5 mg/kg in 3-tägigen (n ¼ 15) oder 2-tägigen (n ¼ 8) Intervallen gestartet, bei 2 Patienten mit refraktärer AML am Tag À46 bzw. À42, bei allen anderen Patienten im Median am Tag À3. Die Patienten erhielten im Median 11, prophylaktische Dosen bis zu Median Tag þ20,5 (þ11 bis þ43).

Ergebnisse. Bei 8 Patienten wurden wegen Fieber oder CRP-Anstieg die Infusionsintervalle verkürzt. Eine intermittierende Candida-Kolonisation wurde durch wöchentliche Ü berwachungskulturen bei 9 Patienten detektiert. Wöchentliche Galaktomannan-Test blieben bei 17 Patienten durchgehend negativ, 2 Patienten hatten je ein einzelnes positives Testergebnis; keiner dieser 19 Patienten entwickelte während einer medianen Nachbeobachtungszeit von 10 Monaten eine invasive Pilzinfektion. Eine Patientin mit GVH IV unter multimodaler Immunsuppression und wiederholt positiven Galaktomannan-Tests entwickelte trotz kontinuierlicher präemptiver Therapie mit LAMB alternierend mit Caspofungin präfinal eine Aspergillus-Pneumonie und verstarb am Tag þ140. Bei keinem Patienten mußte LAMB wegen infusionsassoziierter Nebenwirkungen abgesetzt werden; die Hauptnebenwirkung war eine Hypokaliämie, die in 20/23 Anwendungen substitutionsbedürftig war.

Schlussfolgerungen. Die intermittierende antifungale Prophylaxe mit LAMB nach autologer bzw. allogener SZT war bei 19/20 Patienten erfolgreich. Die Nebenwirkungen beschränkten sich auf eine substitutionsbedürftige Hypokaliämie. Background. Late complications including diabetic vasculopathy, retinopathy and neuropathy are still serious problems in the treatment of diabetes mellitus. Whole pancreas transplantation or human islet cell transplantation are alternatives but limited due to the organ shortage and lifelong immunosuppression. The use of immune isolated porcine islet cells (PIC) could offer a feasible alternative. In this study, the safety and applicability of three possible implantation sites for Sodium Cellulose Sulfate (SCS) microencapsulated PIC were evaluated in five healthy beagle dogs.

Methods. Five healthy, male castrated Beagle dogs were used for this study. SCS microencapsulated PIC were implanted in the subcutaneous tissue, the omentum and the gastric submucosa. Surgery time and intraoperative complications were monitored. Implantation in the omentum and gastric submucosa was performed laparoscopic assisted. All animals were continuously monitored including clinical examinations and complete blood counts for possible occurring side effects. On day À3, 30 and 90 a glucagon stimulation test was performed to assess -cell function and glucose metabolism. On day À3, 1, 3, 12, 30, 60 and 90 fasting glucose and insulin were measured. Blood samples were taken to check a possible porcine endogenous retrovirus (PERV) transmission. Biopsies of all implantation sites were obtained for histopathology on day 90.

Results. No technical complications occurred at the subcutis and omentum site. At the gastric submucosa only a limited amount of cells could be implanted. Delayed wound healing at the subcutaneous implantation site was observed in three out of five dogs. No further side effects occured during the study period. Transmission of PERV was not detected. Glucagon stimulation tests showed a trend towards a faster response to a glucagon stimulus and an increased insulin peak. Histopathology results have not been completely evaluated yet.

Conclusions. First results show that SCS microencapsulated PIC do not cause any harmful side effects to the recipients concerning application. No transmission of PERV was detected. The outcome of histopathology in combination with the clinical results will display the efficacy of this treatment and the expected differences in transplantation sites.

Background. Heart failure following acute myocardial infarction (AMI) is a major cause of morbidity and mortality. Our previous observation that injection of apoptotic peripheral blood mononuclear cells (PBMC) was able to restore cardiac function in a rat acute ischaemia model prompted us to study the effect of soluble factors derived from apoptotic PBMC on ventricular remodelling after AMI.

Methods. Cell culture supernatants derived from irradiated apoptotic peripheral blood mononuclear cells (APOSEC) were collected and injected as a single dose intravenously after myocardial infarction in an experimental AMI rat model and in a porcine reperfused AMI model. MRI and echocardiography were used to quantitate cardiac function. Immunohistology and flowcytometry were used to analyse the cellular components. Analysis of soluble factors present in APOSEC was performed with proteome membrane arrays and activation of signalling cascades in human cardiomyocytes by APOSEC in vitro was studied by immunoblot-analysis.

Results. Intravenous administration of APOSEC resulted in a reduction of scar extension in both models. Hearts explanted from animals infused with APOSEC evidenced less myocardial necrosis after 24 hrs. Troponin-I release was less than in animals treated with medium as control. In the porcine AMI model APOSEC led to an improvement of ejection fraction (57.0% vs. 40.5%, p < 0.01), cardiac output (4.0 vs. 2.4 l/min., p < 0.001) and a reduced infarction size (12.6% vs. 6.9%, p < 0.02) as determined by MRI. Administration of APOSEC in the rat AMI model caused increased presence of CD68 þ macrophages and c-kitþ endothelial progenitor cells (EPC) in the infarcted myocardium within 72 hrs. Exposure of human cardiomyocytes with APOSEC in vitro triggered activation of pro-survival signalling-cascades (AKT, p38 MAPK, Erk1/2, CREB, c-Jun) and anti-apoptotic gene products (Bcl-2, BAG1).

Conclusions. Intravenous infusion of APOSEC attenuated myocardial remodelling in both models of experimental AMI. This effect seems to be due to the activation of pro-survival signalling cascades in the affected cardiomyocytes and to a higher presence of regenerative cells (EPCs, macrophages) within the ischaemic tissue. APOSEC represents a "biological" which prevents myocardial infarction by causing peri-infarct conditioning and stimulation of regenerative effects in the hypoxic myocardium.

Anti-thymocyte globulin (ATG) reduces damage caused by ischaemia and preserves cardiac function after experimental myocardial infarction Background. Myocardial infarction (MI) is one of the leading causes of death in the western world. Consequent inflammatory reactions initiate and sustain remodeling of the damaged myocardium, which can worsen the outcome additionally. Our previous findings suggest that apoptotic peripheral blood mononuclear cells (PBMCs), injected intravenously after MI in rats, can attenuate inflammation and subsequently improve cardiac function after MI. In this study we sought to investigate, if anti-thymocyte globulin (ATG) as a therapeutic agent inducing apoptosis in white blood cells, evidences similar effects after experimental MI in rats.

Methods. To study effects of ATG in vitro, we incubated whole blood and PBMC cell cultures with this polyclonal antibody for 24 hours. ELISA was utilized to assess changes in the secretion profile of various cytokines. Furthermore, we conducted an in vivo study in an experimental rat model of myocardial infarction. Rodents were injected with ATG (10 mg/rat) after ligation of the left anterior descending (LAD) artery. Untreated and sham operated animals served as controls. Short term effects were evaluated by immunohistology after three days. Echocardiography and assessment of infarction size by planimetry depicted the outcome after six weeks.

Results. Cytokines and chemokines held responsible for cardioprotective and neo-angiogenic effects (e.g. IL-1RA, IL-8, MCP-1) were significantly elevated in ATG groups over controls in vitro. Histology of in vivo experiments confirmed these data, as ATG treated animals had a reduced area of necrosis three days after AMI (10.7% vs. 20.6%), smaller infarct scars after six weeks and an increased infiltration of macrophages. Echocardiography revealed a treatment advantage of ATG, as rodents enrolled in this study group evidenced an ejection fraction of 52.35% AE 1.96% compared to 42.91% AE 2.22% in controls (n ¼ 13, p < 0.001).

Conclusions. Via the mechanism of apoptosis, ATG induces a plethora of pro-angiogenic and immune-modulatory factors. Secretion of this ensemble of cytokines and chemokines attracts macrophages/monocytes into the infarcted area, and attenuates inflammation after myocardial infarction. In conclusion, interaction of these factors can significantly reduce infarction size and improve left ventricular function after experimental myocardial infarction in rats.

Secretion of cytokines and chemokines by peripheral blood mononuclear cells is triggered by coagulation products these mediators systemic in immune activation/sepsis, wound healing, autoimmune-diseases, artherosclerosis and myocardial infarction. However, cytokines and chemokines are usually not considered to be very stable after blood collection, which might therefore alter test.

Results. Thus, the aim of the pilot study was to obtain better knowledge about stability of these mediators in blood samples for interpretation of test results.

Methods. Venous blood was taken from healthy probands (n ¼ 7) using different blood tubes (serum, heparin plasma and EDTA plasma). Blood tubes were either centrifuged initially within 20 minutes after venipuncture and kept frozen at À80

until further testing or were stored at 4 C, at room temperature (RT) or at 37 for up to 24 hours. Samples were evaluated for IL-1ß, IL-6, TNF-and for selected chemokines such as interleukin-8, epithelial neutrophil-activating protein 78 (ENA-78) and granulocyte chemotactic peptide-2 (GCP-2) using commercially available enzyme-linked immunosorbent assay (ELISA) kits.

Results. Interestingly all examined mediators rise when samples were stored above room temperature for more than 4 hours in serum tubes. The rise of serum chemokine and cytokine levels culminated in a 79-fold increase for IL-6 (p < 0.0081), a 22-fold increase for ENA-78 (p < 0.0006) and a 17-fold increase for GCP-2 (p < 0.0026) compared to basic values. Serum levels of IL-1 and TNF-where not detectable at basic samples but rise up to 1157 pg/ml (mean IL-1, p < 0.03) and 488 pg/ml (mean TNF-, p < 0.03).

Conclusions. These data indicate that the most chemokine and cytokine levels remain stable when analysed within a short interval after venipuncture. When tubes were exposed to temperatures higher than 24 (RT), levels of measured chemokines increased dramatically. 

Background. Rabbit antithymocyte globulin (rATG) is widely used as induction agent in solid organ transplantation. Beside depletion of circulating lymphocytes there is a growing body of evidence suggesting that rATG may also play a pivotal role in modulating the immune system. As blood circulating endothelial cells (CECs) and circulating hematopoietic progenitor cells (CPCs) represent two minute fractions (CECs: 0.1% to 6.0% and CPCs: 0.01-0.20%) of blood mononuclear cells that are thought to play important roles in tissue vascularisation, the study of both cell types is currently suggested as surrogate markers for numerous pathologies. Especially the noninvasive endothelial evaluation as an early index of vascular injury following kidney transplantation has been already demonstrated.

Methods. We used four surface markers to identify viable CECs as CD31bright, CD34dim, CD45-, CD133-and viable CPCs as CD34bright, CD133þ, CD45dim, CD31 þ cells in the peripheral blood of liver transplanted recipients (n ¼ 28) until day 20 post transplantation via FACS-analysis.

Results. An induction of CECs was exclusively observed for rATG-treated patients (n ¼ 17) increasing from 0.56% AE 0.98% pre transplantation to 1.83% AE 1.85% at day 1-2 post transplantation compared with control patients receiving standard immunosuppression (n ¼ 11) (p < 0.04). In addition, the induction of CPCs was even more pronounced illustrating an increase in rATG treated patients from 0.20% AE 0.26% pre transplantation to 1.55% AE 1.75% at day 1-2 post transplantation (p < 0.001). A significant elevation of blood CPCs is still detectable at day 5 (p ¼ 0.0379 compared with controls) and starts to decline at day 10 post transplantation.

Conclusions. In summary we illustrated that both CECs and CPCs were detectable in numbers that allows kinetic monitoring of these cell types post transplantation and that rATG treatment results in a transient induction. As clinical correlations between the concentration of these two populations and the effect of immunosuppressive regimens has been already proven, validation of these cell populations as biomarkers in the setting of solid organ transplantation remains to be determined. Background. Lipocalin-2 (Lcn-2) has been described as a marker and potential positive modulator of inflammation during ischemia/reperfusion injury (IRI) following solid organ transplantation. Data on Lcn-2 expression during allograft rejection have been missing so far.

Methods. Sera of 68 patients undergoing orthotopic liver transplantation were collected preoperatively and postoperatively from day 1 to 15. Lcn-2 was analyzed by ELISA and expression levels were correlated with parameters of allograft rejection. Lcn-2 expression was further correlated with preexistent malignancy, postoperative renal failure and various immunosuppressive regimens.

Results. Lcn-2 serum levels were elevated 3 to 7 fold immediately after liver transplantation due to IRI and also increased prior to clinically diagnosed acute rejection, however not statistically significant (p > 0.05) but closely related to an elevation of routinely used laboratory parameters.

Conclusions. Our data suggest Lcn-2 to be a chemoattractant stimulus for infiltrating immune competent cells into the allograft following solid organ transplantation. It is an inflammatory marker which is upregulated during acute graft rejection and its elevation prior to routinely used parameters of acute rejection might be an important tool for therapeutic intervention.

Effect of oxidative stress and endotoxin on albumin in brain death Background. Albumin binds and detoxifies endotoxin in healthy people. Oxidative stress leads to protein oxidation and thus to impaired binding properties of albumin. This, in combination with increased gut permeability leads to appearance of endotoxin in the systemic circulation and further to impaired organ function. We hypothesise that these processes occur in serum of brain-dead organ donors.

Methods. Endotoxin was determined with an adapted limulus amoebocyte lysate assay. Albumin fractions and binding capacity were determined by HPLC. FlowCytomixTM was used for determination of cytokine levels.

Results. Eighty-four brain-dead organ donors were enrolled and categorized by the length of intensive care unit (ICU) stay. Albumin binding capacity for dansylsarcosine was reduced in brain-dead patients compared to controls. Endotoxin positivity in 16.7% of donors was associated with decreased binding capacity in donors and worse survival of recipients. Lengths of ICU stay increased albumin oxidation. In addition, IL-6, IL-8, IL-10 and IL-1 levels were elevated in patients, whereas IFN-levels were within the normal range.

Conclusions. We conclude that oxidative stress and systemic endotoxemia is present in brain-dead organ donors what might affect recipient survival. High endotoxin levels might be due to increased gut permeability and decreased binding capacity of albumin influenced not only by higher albumin oxidation.

Evidence of lymphoid neogenesis in skin biopsies of human hand allografts during rejection Background. Excessive production of reactive oxygen species (ROS) is a major contributor to the development of ischemia-reperfusion injury (IRI) in the course of solid organ transplantation. In particular mitochondria-derived ROS are critical for the initiation and progression of IRI, which restricts the pool of donor organs and results in elaborate follow up treatments. In various in vivo (IR) and in vitro (hypoxia/reoxygenation, HR) models we observed a consistent pattern in the activation of key intracellular signaling pathways. Most strikingly the use of p38 specific inhibitors prevented mitochondrial ROS production and cell death. Here we further dissected the contribution of p38 to IR-and HR-induced damage and provide first evidence for a therapeutic benefit of p38 inhibition by BIRB-796 in vivo.

Methods. Kidney transplantation and kidney clamping in the rat were used for the induction of IRI. H/R was analyzed in HL-1 cardiomyocytes and primary MEFs. Intracellular signaling was monitored by using phosphorylation specific antibodies. Mitochondrial ROS levels were determined by imaging of Mitotracker Red CM-H2XROS. ROS/NOS-induced tissue damage was visualized by 3-nitrotyrosine specific antibodies. To assess acute kidney injury (AKI) HSP70 expression was monitored by immunoblotting, serum creatinine and urea were determined, and serum cystatin c and NGAL concentrations were measured by ELISA.

Results. The expression patterns for all p38 isoforms were established in HL-1 cells and siRNA-mediated knockdown of the predominant isoform (p38Alpha) reduced ROS production, confirming the critical role of p38. Preliminary data suggested the requirement of MAPKAP kinase 2 (MK2) rather than the transcription factor ATF-2 downstream of p38. As observed in other settings reperfusion following kidney clamping or transplantation was marked by a profound increase in the activity of p38, its upstream kinases MKK3/6 and the putative effector MK2. Application of BIRB-796 prevented deterioration of kidney function following IR based on reduced serum creatinine, urea, cystatin c and NGAL levels in animals treated with the inhibitor. P38 inhibition also protected from oxidative damage. Thus the inhibition of p38 prevents key processes, which are essential for the development of IRI.

Conclusions. Inhibiting p38 signaling during IR and HR may provide a potent strategy for limiting IRI.

The role of constitutively expressed nitric oxide synthases in ischemiareperfusion-injury Background. Single shot donor therapy with the essential nitric oxide synthase cofactor (NOS) tetrahydrobiopterin (H4B) was shown to attenuate ischemia-reperfusion-injury-related pancreatitis in a murine pancreas transplantation (PTX) model. Since underlying mechanisms of tetrahydrobiopterin-mediated protection are still controversially discussed, we aimed to investigate, whether the two constitutively expressed NOS-isoforms represent its major targets using endothelial (eNOS-/-) and neuronal NOS (nNOS-/-) knockout mice.

Methods. In a heterotopic PTX-model syngeneic C57BL6 mice (wild-type, eNOS-/-and nNOS-/-) were used as donorrecipient pairs. Non-transplanted animals served as controls. Following a reperfusion time of 4 h, graft microcirculation was analyzed by intravital fluorescence microscopy. Parenchymal damage as well as peroxynitrite-formation were assessed by H&E-staining and immunohistochemistry. H4B levels were determined by high-performance liquid chromatography (HPLC).

Finally, all groups where tested for recipient survival.

Results. Compared to non-transplanted controls, prolonged CIT significantly worsened microcirculation in untreated wildtype and eNOS-/-grafts (p < 0.05), whereas no deterioration was measured in untreated nNOS-/-. While H4B-pre-treatment significantly restored capillary blood flow in wild-types and eNOS-/-(p < 0.01), no further beneficial effect was observed in nNOS-/-(p > 0.05). In contrast to untreated wild-type and eNOS-/-graft, nNOS-/-grafts developed minor parenchymal damage following prolonged CIT, which could be slightly ameliorated by tetrahydrobiopterin pre-treatment. There weren't any significant differences between the analyzed groups regarding intragraft nitrotyrosine formation. While H4B pre-treatment extended survival of all recipients, significantly prolonged recipient survival was also achieved if donors were untreated nNOS-/-(p < 0.01).

Conclusions. These observations in pancreatic grafts lacking nNOS suggest, that instead of the endothelial isoform, it is the neuronal isoform to be crucially involved in the promotion of ischemia-reperfusion-injury in this model, representing therefore the major target of tetrahydrobiopterin mediated protection.

Nutrazeutika optimieren die Organfunktion in einem Rattennieren-Ischämie-Reperfusionsmode P. Gehwolf 1 , F. M. Struller 1 , A. Kostron 1 , M. Wolzt 2 , F. Bach 3 , L. Otterbein 3 , B. Wegiel 3 , J. Pratschke 1 , R. Öllinger 1 tion, and angiogenic sprouting in vitro, and angiogenesis in a chicken embryochorioallantoic membrane assay. 5 0 -methoxy leoligin was consequently analyzed in an in vivo rat MI model. The novel compound potently stimulated angiogenesis in the peri-infarction zone and led to a significant increase in the cardiac ejection fraction (plus 20% 28 days after MI) in animals treated with 5 0 -methoxy leoligin. Based on microarray analyses followed by knockdown and over-expression experiments in vitro Cyp26B1 was identified as the central players in 5 0 -methoxy leoligin-induced angiogenesis induction.

Conclusion. The data presented herein indicate that 5 0methoxy leoligin induces angiogenesis in the peri-infraction area after MI via upregulation of Cyp26B1. 5 0 -methoxy leoligininduced capillarisation is capable of partially restoring cardiac function after MI. Therefore this novel compound could be able to positively influence post MI myocardial remodeling and even induce myocardial recovery. This or other pharmacological strategies to induce neoangiogenesis could preserve organ function and avoid several patients ending on the waiting list for cardiac transplantation. Background. A major reason of vein graft failure after coronary artery bypass grafting is neointimal hyperplasia. Several clinical studies revealed that the patency rates of arterial conduits are better than those of venous conduits. In the current study, we aimed to improve the patency rate of experimental vein grafts by induction of graft arterialisation by transplantation of homologous skeletal myoblasts.

Methods. For the experiments we isolated skeletal myoblasts from C57/Bl6 mice. Syngenic C57/Bl6 mice underwent interposition of the vena cava inferior from donor mice into the common carotid artery of recipient mice. We used 2 experimental groups: one control group, where we applied only the control medium without any cells on the vein grafts and a treatment group where we applied skeletal myoblast periadventially on the grafts. After 4 weeks of follow up veins were harvested, embedded in paraffin and underwent histological evaluation.

Results. 4 weeks after surgery the neointimal thickness in the control group was 56 mm (15-60), whereas in the treatment group we found neointimal thickness levels of 183 mm (152-204) (p < 0.001). Apart from that we could show that cells remain on the outside of the vein wall even after 4 weeks.

Conclusions. Application of skeletal myoblasts does not induce vascular regeneration after vein graft failure and does not reduce neointimal hyperplasia. Moreover application of these cells leads to a severe induction of neointimal hyperplasia in a murine vein graft model. This might be due to secretion of growth factors by the cells. Background. Based on international recommendations combined immunosuppression is used after kidney transplantation in children. A common regimen is based on steroids, mycophenolat-mofetil and cyclosporin A with IL-2 antagonist induction. One third of the patients had to be switched from cyclosporin A to tacrolimus, due to dermatologic cosmetic side effects. Thus, TABIC was established as a novel immunosuppressive regimen. Aim of the present registry is to perform quality assurance and to evaluate a risk/benefit analysis of the TABIC-regimen compared to previous immunosuppressive regimens in children undergoing kidney transplantation at our center.

Methods. In this observational single-center study, 32 consecutive patients undergoing kidney transplantation between December 2005 and October 2011 at our center, were enrolled. Patients who were not treated according to the TABIC-scheme served as a historic control. The end-point was a combination of death and graft loss.

Results. 5 patients (16%) had to be switched from tacrolimus to rapamycin due to side effects. Furthermore, 5 patients (16%) suffered from acute rejection, but no graft loss was observed. Within follow-up, no patient reached the combined end-point (100% kidney transplant as well as 100% patients survival). A Kaplan-Meier-analysis revealed a significantly better outcome for patients treated with the TABIC-scheme compared to controls (5 years event free survival, TABIC vs. non-TABIC: 100% vs. 79%, p ¼ 0.048).

Conclusions. Our data show that the TABIC-regimen is efficient and safe compared to cyclosporin A based immunosuppression. Tacrolimus is associated with a significantly better graft and patient survival. Therefore, it should be considered as standard therapy after kidney transplantation in children.

Falsely elevated tacrolimus concentrations in two kidney allograft recipients using the affinity column-mediated immunoassay (ACMIA) method: identification of IgG isotype rheumatoid factor as causative endogenous antibodies Background. Therapeutic monitoring of tacrolimus is currently performed employing ACMIA in several transplant centers. Here we report on two renal transplant patients displaying excessive tacrolimus levels either not corresponding to oral drug dosage or without taking the drug at all. Various antibodies, such as anti ds-DNA antibodies or heterophilic antibodies, have been identified in previous cases of falsely elevated tacrolimus measurements and have been suggested as a possible cause for interaction with the tacrolimus assay.

Methods and Results. In our observations, both patients were on a stable, long-term tacrolimus-based immunosuppressive regimen. In the first patient the elevated tacrolimus trough levels were only observed after loss of graft function (6 years posttransplant) while the patient was not longer on tacrolimus, as the patient was on hemodialysis again. The second patient currently still has a functioning graft, although he is taking minute amounts of tacrolimus. All rheumatoid and immunologic tests (including anti ds-DNA antibodies) were negative, except IgG isotype rheumatoid factors, which proved to be highly positive in both patients.

Conclusions. Our results show that certain patients with positive IgG isotype rheumatoid factor can exhibit false-positive tacrolimus trough levels when using the ACMIA assay. It can therefore be concluded that abnormally high drug levels should be verified by mass spectrometry methods. Further studies are underway to determine whether non-transplanted patients with positive rheumatoid factors also display falsely elevated tacrolimus values and also if this can be suppressed after preincubation with immunoglobulin blocking reagents specific for the IgG isotype rheumatoid factor. Background. The impact of vesicoureteral reflux after renal transplantation has been a matter of debate in the past with small studies reporting controversial results. The aim of the present study was to evaluate the effect of early post-transplantation VUR on long-term renal allograft performance, incidences of urinary tract infections (UTI), rejection rates, graft and patient survival in a large cohort series.

Methods. Retrospective study of 646 consecutive renal transplant patients with routine post-transplantation vesicocystourethero-graphy (VCUG). Uretero-neocystostomy was performed in an antirefluxive fashion using extravesical submucosal tunneling. VCUG was performed prior to discharge according to the international grading system and reviewed by an independent radiologist and urologist.

Results. Overall, 263 of the 646 (40.7%) kidney transplant recipients were diagnosed with VUR by VCUG at discharge. Patients had VUR grade I, II, III and IV in 7.9%, 19.8%, 10.2%, and 2.8%, respectively. No grade V reflux was seen. VUR showed no significant impact on death-censored graft survival, patient survival, proteinuria or UTI. Patients with VUR had a significantly lower GFR at one year after transplantation than patients without VUR (60 vs. 52 ml/min/1,73 m 2 , p ¼ 0.021), this difference was no longer seen at 3 and 5 years.

Conclusions. Although VUR is a common finding in patients after renal transplantation, it has no impact on death-censored graft survival, patient survival, proteinuria or UTI and only a short term effect on renal graft function that was no longer seen in long term follow up.

Grundlagen. Der Einfluss von primär unentdeckten hepatozelluären Karzinomen (HCC) auf das Ü berleben nach Lebertransplantation wird in der Literatur kontroversiell diskutiert. Ziel dieser Studie war es, die Rate von okkulten Karzinomen in unserem Kollektiv an äthyltoxischer Zirrhose erkrankter und für diese Indikation transplantierter Patienten der letzten 10 Jahre zu erheben und den Einfluss auf das rezidivfreie uund Gesamtüberleben nach Lebertransplantation zu untersuchen.

Methodik. Retropektive Analyse von 285 wegen äthyltoxischer Zirrhose gelisteter und transplantierter Patienten im Zeitraum von 1998 bis 2008. Neben den Baseline Parametern wurden alle Explanthistologien ausgewertet.

Ergebnisse. Dreizehn Patienten (4,5 %) zeigten ein HCC in der Explanthistologie. Drei Patienten waren außerhalb der Mailand Kriterien und 19 von 27 Herden waren größer als 1 cm. Die Wartezeit der Patienten betrug im Median 2,7 Monate. Nach 2004 gab es keine okkulten HCCs in unserem Kollektiv. Ein Ü berlebensnachteil fü r Patienten mit okkultem HCC gegenü ber der Normalgruppe konnte in diesem Kollektiv nicht gezeigt werden.

Schlussfolgerungen. Die kurze Wartezeit der Patienten mit okkultem HCC sowie die Tumorgröße legt nahe, dass bei diesen Patienten bereits vor der Listung das HCC unentdeckt blieb, während es später mit Verbesserung der radiologischen Technik sowie längerer Wartezeit im Rahmen des Screenings entdeckt wurde. Rein deskriptiv zeigte sich in unserem Kollektiv kein Ü berlebensnachteil für Patienten mit okkultem HCC.

Ist die ungeplante Reoperation nach Nierentransplantation ein Indikator für Organverlust? K. Huber 1 , A. Krause 1 , O. Gangl 1 , W. Enkner 2 , R. Oberbauer 2 , R. Függer 1 Ö sophagusvarizen (Grad II; >5 mm unter Insufflation) während der oLT aufgetreten sind und mittels Ballontamponade behandelt wurden. Vergleicht man die beiden Patientenkollektive (mit und ohne Krampfadern), so ist das Ergebnis statistisch nicht signifikant: Chi-Quadrat-Test, p ¼ 0,61. Alle Patienten erhielten im Median 2 Einheiten Erythrozytenkonzentrate (Bereich 0 bis 70 Einheiten), 7 Einheiten Fresh Frozen Plasma (Bereich von 0 bis 60 Einheiten) und 0 Thrombozytenkonzentrate (Bereich 0 bis 4 Einheiten). Obwohl Patienten mit Krampfadern eine signifikant längere Prothrombinzeit und eine geringere Thrombozytenzahl aufwiesen, bestand gegenü ber Patienten ohne Varizen kein Unterschied in der Verwendung von Blutprodukten.

Schlussfolgerungen. Die TEE scheint bei oLT Patienten mit dokumentierten Ö sophagusvarizen eine allgemein sichere Methode zur Kontrolle der Herzfunktion zu sein.

Entwicklung des Anteils an Leberspendern mit erweiterten Spenderkriterien innerhalb der letzten Dekade an der Universitätsklinik für Transplantationschirurgie Wien Grundlagen. Die Anzahl der jährlich durchgeführten Lebertransplantationen ist von der Anzahl der gemeldeten Organspender abhängig. Allerdings eignet sich nicht jeder Organspender als Leberspender. Ziel dieser Studie ist es, die Evolution der Leberspender innerhalb der letzten 10 Jahre zu verfolgen.

Methodik Methodik. Im Zeitraum von Jänner 1994 bis Juli 2011 wurden an der Medizinischen Universität Wien, Abteilung für Transplantation 22 Patienten, davon 18 männlich und 4 weiblich, mit einem medianen Durchschnittsalter von 51 Jahren kombiniert Leber und Niere transplantiert. Inkludiert wurden bei dieser retrospektiven Analyse nur einzeitig transplantierte Patienten. Die Indikationen zur Lebertransplantation waren wie folgt verteilt: Virushepatitis (n ¼ 6), Malignom (n ¼ 4), metabolische Zirrhose (n ¼ 5), Oxalose (n ¼ 3), sklerosierende Cholangitis (n ¼ 1), Hämochromatose (n ¼ 1), Citrullinämie (n ¼ 1) und zystische Zirrhose (n ¼ 1). Die Nierentransplantation erfolgte aufgrund von: chronischer Niereninsuffizienz o. n. B. (n ¼ 7), Glomerulonephritis (n ¼ 4), metabolischer Nephropathie (n ¼ 4), Oxalose (n ¼ 3), Pyelonephritis (n ¼ 1), Hantavirus (n ¼ 1), IgA-Nephropathie (n ¼ 1) und Cyclosporin-induziert (n ¼ 1). 72 % (n ¼ 16) der Patienten waren zum Zeitpunkt der Transplantation dialysepflichtig, davon 13 an der Hämodialyse und 3 erhielten Peritonealdialyse.

Ergebnisse. Die Patienten waren durchschnittliche 121 Tage gleichzeitig für Leber und Niere gelistet und der mediane MELD (Model for End-Stage Liver Disease) Score betrug bei Transplantation 22. Die 1-und 5-Jahresüberlebensrate betrug bei unseren Patienten 81 % und 67 %. Unmittelbar postoperativ verstarben 3 Patienten, davon 2 an einer Sepsis und einer in Folge einer primären Nichtfunktion der Leber.

Schlussfolgerungen. Die KLNT stellt an unserem Zentrum eine gute Therapieoption bei terminaler Leber-und Nierenerkrankung dar. Laut ELTR beträgt die 1-und 5-Jahresüberlebensrate für alleinige orthotope Lebertransplantation 78 % bzw. 64 % und ist daher mit unseren Resultaten vergleichbar. Die hohe perioperative Mortalität der KLNT spricht für ein rezent diskutiertes zweizeitiges Vorgehen, um einen möglichen Organverlust zu vermeiden.

Orthotopic liver transplantation (OLT) under the use of a protective filter against phototoxicity in a patient suffering from erythropoietic protoporphyria (EPP) with liver cirrhosis Background. Erythropoietic protoporphyria (EPP) is a rare condition arising from a deficiency in the enzyme ferrochelatase, leading to abnormally high levels of protoporphyrin in the tissue. Apart from increased photosensitivity leading to acute and chronic skin changes, in a small number of cases protoporphyrin deposits and pigment loading of the hepatocytes cause liver cirrhosis. In these patients liver transplantation is the therapy of choice. Nevertheless intestinal ulceration, bleeding and ultimately multiorgan failure can occur if no protective measures are taken against phototoxic injury during surgery.

Methods and Results. We would like to present the case of a 36-year-old patient who was admitted to our department for liver transplantation. EPP had been diagnosed 32 years ago. The patient suffered from typical skin changes and cholestatic liver cirrhosis had been histologically confirmed. OLT was performed successfully using protective measures to prevent phototoxic injury to the abdominal organs. A flexible yellow filter omitting wavelengths below 470 nm was applied to operating room luminaires to avoid phototoxic injury while maintaining visual colour perception of the surgeons. The immediate postoperative period was without any complications related to the EPP. Seven months after transplantation the patient is in good general health and liver function tests show good results.

Conclusions. OLT is a suitable treatment for patients suffering from EPP with hepatic involvement and cholestatic cirrhosis if protective measures against phototoxic injuries are used during surgery.

Günstige Ergebnisse nach schweren immunologischen und infektiösen Komplikationen nach Dünndarm-Transplantation: ein Fallbericht C. Bösmüller 1 , A. Weißenbacher 1 , M. Biebl 1 , R. Öllinger 1 , S. Schneeberger 1 , F. Aigner 1 , S. Weiß 1 , R. Oberhuber 1 , C. Ensinger 2 , J. Pratschke 1 uneventful course after transplantation with only three mild acute rejection episodes within 11 years. Due to a fracture of the right allograft radius requiring surgery at 10.5 years posttransplant immunosuppression (IS) was switched from sirolimusmonotherapy to prednisone. One month thereafter the patient experienced metabolic deterioration (hypothyroidism, hyperlipidemia and hyperglycemia) and developed an exanthema and pruritus on the trunk and the upper extremities, including both the recipient's own skin and the allograft skin. BP was diagnosed upon direct immunofluorescence of skin biopsies (linear C3 and IgG deposits along the basement membrane zone) and detection of serum autoantibodies against the BP antigen 2 (BP180nc16a: 142 U/l). Histology showed a partial separation along the dermalepidermal junction and a mild perivascular infiltration consisting of lymphocytes, eosinophils and granulocytes. Cell counts were within normal range. No donor specific antibodies were detected. Skin lesions disappeared under intensified therapy with prednisone. While conversion of IS and metabolic deterioration might have contributed to development of BP after hand transplantation, the pathomechanism remains unclear.

Belatacept treatment for 2 years after liver transplantation is not associated with overt immunomodulation Background. Belatacept is a costimulation blocker with immunomodulatory properties in the experimental setting. Moreover, the liver graft itself has been claimed to have 'tolerogenic' properties. The belatacept multi-center phase II liver transplantation trial was terminated during the long-term extension period on the recommendation of the Data Monitoring Committee. This situation gave us the unique opportunity to evaluate the minimum immunosuppression required in liver transplant recipients after >2 years of treatment with belatacept.

Methods. In our center all belatacept patients (n ¼ 4) were switched to MMF monotherapy (2Â1 g/day) after discontinuation of belatacept. We prospectively assessed the occurrence of acute rejection and evaluated kidney function (calculated GFR; MDRD) over time in comparison to tacrolimus-treated patients enrolled in the control groups of the multi-center trial (n ¼ 4).

Results. The mean period from transplantation to withdrawal of belatacept was 30 months (range 25-35). GFR at the time of withdrawal was more than 60 ml/min/1.73 m 2 in the belatacept group (4/4; mean: 101 ml [range 89-114]), whereas 3 out of 4 patients in the control group had a GFR below 60 ml/min/ 1.73 m 2 with a mean of 57.92 ml/min/1.73 m 2 (range 36-98) (p ¼ 0.026). Five months after belatacept discontinuation kidney function declined on average by 19.22 ml (range À45 to þ3,5) (follow-up ongoing). After belatacept withdrawal all 4 patients developed 3-fold elevated liver enzymes (ASAT, ALAT) within 10.3 weeks after EOT (7-14) (Biopsies were not performed, mostly due to lack of consent.). Patients were therefore switched to triple therapy with corticosteroids, CNIs and MMF. Graft dysfunction resolved within one to three weeks after switch.

Conclusions. Consistent with results from the multi-center trial, patients treated with belatacept showed better kidney function compared to those treated with CNIs. MMF monotherapy following withdrawal of belatacept is associated with a high risk of rejection. Thus belatacept has no obvious immunomodulatory effect in liver transplant recipients that would be sufficient to allow a high success rate of minimization strategies.

Ausgezeichnete Blutdruckkontrolle nach Nierentransplantation durch antihypertensive Mehrfachtherapie -Entwicklungen der letzten 20 Jahre Grundlagen. Patienten mit terminalen Lebererkrankungen entwickeln gehäuft Ö sophagusvarizen auf Basis einer portalen Hypertension. Ziel dieser Studie ist es, die Inzidenz von Ö sophagusvarizen und deren Korrelation mit dem Grad der Lebererkrankung bei Patienten, die auf der Leberwarteliste stehen, zu errechnen.

Methodik. Hierfür wurden die Daten von 512 Patienten, die zwischen 2002 und 2010 an der Universitätsklinik für Chirurgie lebertransplantiert wurden, analysiert. Patienten, die die Lebertransplantation aufgrund eines akuten Leberausfalls benötigten und solche, bei denen die Daten unvollständig waren, wurden aus der Studie ausgeschlossen. Somit umfasst die Studie 396 Patienten (77 %). Die Ergebnisse werden median in der 25. und der 75. Perzentile angegeben.

Ergebnisse. Die Lebertransplantierten waren im Median 54 (48-60) Jahre alt. Im Median lagen 227 Tage (125-368) zwischen der letzten Ö sophagogastroduodenoskopie und der Lebertransplantation. 287 Patienten (72,5 %) zeigten Varizen: 130 (32,8 %) Varizen Grad I ( < 5 mm bei Insufflation) und 157 (39,6 %) Var-izen Grad II (>5 mm bei Insufflation). Red Spot Signs fand man bei 40 Patienten (10,1 %). 82,2 % der Varizen lokalisierten sich im Ö sophagus, 4,2 % im Magen und 13,6 % im Ö sophagus und Magen. Patienten mit ösophagealen Varizen zeigten signifikant niedrigere Serum-Natrium Werte und Thrombozytenzahlen und signifikant höhere Ammoniak-Spiegel, Prothrombinzeiten und MELD-Scores als solche ohne Varizen.

Schlussfolgerungen. Ö sophagusvarizen treten gehäuft bei Patienten mit terminalen Lebererkrankungen auf und werden von einer Verschlechterung der Labor-und MELD-Werte begleitet. Background. A project called Transplant Procurement Management (TPM) was created in Barcelona, Spain. The aim of this project is to provide profound knowledge and skills for health care professionals, working in the transplantation field with the final purpose of promoting and increasing donation rates. We evaluated the effects of this program on donor outcome and the organ procurement in a single center analysis.

Methods. Number of donor procurements over a 1-year period, from June 1st, 2010 to May 31st, 2011, evaluated at the transplant center of the Medical University Vienna, the sixmonth period before and after participation in the TPM-course (Nov 22nd to Nov 26th, 2010) .

Results. In the 1st evaluation period from June 1st to Nov 30th, 2010 74 potential donors were reported in our center out of which 31 became actual donors. Within the 43 missed donors 23 did not fulfill brain dead criteria. Moreover twelve donors could not be realized because of their co-morbidities and eight due to opposition against organ donation. In the comparable 2nd period from Dec 1st to May 31st, 2011 we registered an increase of 42 actual donors out of 69 potentials. Thirteen donors were missed because of unaccomplished brain dead criteria. Moreover eight donors were lost due to their co-morbidities and six because of opposition against organ donation. However during this evaluation period the donor rates increased for about 35 %. The procurement of transplantable kidneys increased from 60 to 66, transplantable lungs increased from 12 to 13 and in liver procurement, we could even register an increase from 13 to 22 transplantable organs. Only in heart procurement we registered a decrease from 16 to 13 transplantable organs. These results show a remarkable effect of this training program as well as the fluctuation of potential donors and the diverse potential of organ recovery during this year.

Conclusions. After all training programs in organ donation are an interesting field and seem to increase organ donation rates. Efforts have to be made to provide and spread profound knowledge and skills for health care professionals, working in the transplantation field. pneumonia and had to be admitted. The patient with BOS II died due to acute respiratory distress syndrome (ARDS). Both patients received no vaccination. The other two LTRs, one with BOS I the other with BOS II and active vaccination protection were treated with Oseltamivir at the time of the first clinical assessment, without any further respiratory complications. Therapy with Oseltamivir was only started with clinical assessment within the first 48 hours.

Conclusions. Our review shows a causal link between preexisting BOS, vaccination state and the severeness of respiratory symptoms after infection with community acquired respiratory viruses and the clinical effectiveness of seasonal influenza vaccination especially in BOS patients.

A. Scheed 1 , A. Aliabadi 2 , P. Jaksch 1 , S. Taghavi 1 , W. Klepetko 1 , A. Zuckermann 2

Bei 836 Patienten mit funktionierendem Transplantat ein Jahr nach der Transplantation, die in unterschiedlichen Perioden 1990/1991 (n ¼ 129)

sowie die im Rahmen von Patientenselbstmessungen erhobenen Blutdruckwerte analysiert

Bei Patienten, die auf der Leberwarteliste stehen, korrelieren ösophagogastrale Varizen mit dem Grad der Lebererkrankung R. Schwarzer 1 , M. Thum 1 , R. Karatosic 2 , U. Burger-Klepp

Mark 1 of recipient-, donor-and transplantcharacteristics were analyzed using uni-and multivariate analyses

Kinder mit hoch ungünstigem SVM hatten höhere 24h-RR Werte. Die RR-Differenz aus Klinik-RR und 24hRR war jedoch höher bei Kindern mit günstigem SVM. Es konnte kein Zusammenhang zwischen SVM und Klinik-RR gefunden werden. Schlussfolgerungen. Unsere Ergebnisse zeigen, dass Stressverarbeitungsmechanismen bei der Blutdruck-Regulation nach NTX eine Rolle spielen

Attitude towards xenotransplantation of patients prior and after human organ transplantation V

53 I Iberer, F. 30

43 Ö ttl, K. 41 P Pacini

Druck: Holzhausen Druck GmbH, 1140 Wien Ö sterreich. -Verlagsort: Wien. -Herstellungsort: Wien. Printed in Austria P. b. b.= =Erscheinungsort: Wien= =Verlagspostamt 1201 Wien

Ergebnisse. Unter konsequenter systemischer und topischer Therapie mit liposomalem Amphotericin B gefolgt von Voriconazol, und Reduktion der Immunosuppression (Tacrolimusspiegel um 10 ng/ml, Cortison 10 mg), war die Pneumonie rückläufig bis zum 9. po. Monat. Im 12. po. Monat ist die Patientin in gutem Allgemeinzustand mit klinisch-bioptisch bestätigter stabiler Transplantatfunktion, ausreichender oraler Ernährung, stabilem Körpergewicht, normaler Leukozytenzahl nach beherrschten Infektionen (Zystitis, Lidabszeß, Herpesstomatitis).Schlussfolgerungen. Unter sorgfältiger Anpassung der Immunosuppression und konsequenter antimikrobieller Therapie konnten eine stabile Transplantatfunktion und Lebensqualität erreicht werden nach schwerer akuter Abstoßungsreaktion, CMV-Transplantatenteritis und Aspergilluspneumonie.

Bullous pemphigoid 11 years after bilateral hand transplantation 

Background. Expression of peripheral-node-addressin (PNAd) on endothelial cells indicates presence of tertiary lymphoid organs (TLO) in chronic autoimmunity and allograft rejection. We herein investigated the expression of PNAd in skin biopsies of human hand allografts for evidence of TLO after composite tissue allotransplantation.Methods. 167 skin biopsies of 11 hand allografts were collected over 10 years and assessed by HE-histology and immunohistochemistry using antibodies for PNAd, CD3, CD4, CD8, CD20, C4d, CD68, LFA-1, ICAM-1, E-selectin, P-selectin, VE-cadherin, HLA-DR, Psoriasin, IDO and Foxp3. Levels of PNAd expression was assessed semiquantitatively (% of PNAdþ vessels: 0, 1, 2, 3 and PNAd staining intensity: 0, 1, 2, 3) and correlated with rejection grade, characterization of the infiltrate, expression of adhesion molecules and time after transplantation.Results. Rejection ranged from grade 0 to IV (mean score: 0.79 AE 1.05). Upon rejection, expression of PNAd was increased in endothelial cells (grade 0 : 0.24 AE 0.48 vs. all grades of rejection: 0.44 AE 0.62). Most often PNAd expression was only found in few vessels (1-10%). PNAd staining intensity was increased the higher the grade of rejection (grade 0: 0.38 AE 0.76; grade I: 0.41 AE 0.74; grade II: 0.67 AE 0.80; grade III: 0.73 AE 0.91; grade IV: 0.50 AE 0.58). Intense PNAd-staining was associated with more CD4þ and CD8 þ infiltrating T-cells, but less B-cells and macrophages, compared to mild PNAd staining intensity (CD4 þ cells 49.00% AE 29.89%; CD8 þ cells 31.00% AE 22.34%; CD20 þ B-cells 0.50% AE 1.54%; CD68 þ macrophages 0.57 AE 0.60 vs. CD4 þ cells 37.35% AE 40.82%; CD8þcells 27.35% AE 35.93%; CD20 þ B-cells 0.94% AE 2.02%; CD68þmacrophages 0.67 AE 0.66). PNAd expression correlated well with CD3þcells and CD20þBcells. Poor correlation was found for expression of adhesion molecules, IDO and Foxp3, except for LFA-1þ infiltrating cells. PNAd expression was observed at all time-points after transplantation; however, staining intensity was enhanced very early and late after transplantation.Conclusions. PNAd expression in endothelial cells is increased in skin biopsies of human hand allografts indicating presence of TLO. Further investigations are needed to enlighten the role of PNAd and TLOs in composite tissue allotransplantation. Grundlagen. Die zum Hämabbau notwendige induzierbare Hämoxygenase-1 (HO-1) wirkt stark antiinflammatorisch und schützt vor Ischämie-und Reperfusionsschäden (IRS). Eine klinische Anwendung im Sinne einer HO-1-Induktion ist aufgrund der Hepatotoxizität der experimentell verwendeten HO-1-Induktoren nicht möglich. Ziel der Studie war es, natürlich vorkommende Nutrazeutika auf Ihr Potential zur HO-1 Induktion zu untersuchen.Methodik. Verschiedene Nutrazeutika wurden in Hinblick auf die HO-1-Induktion mittels PCR getestet. In einem etablierten Ratten-Nierenarterien-Klemmmodell wurde ein IRS gesetzt, Nierenfunktionsparameter und HO-1 Expression wurden zu festgelegten Zeitpunkten bestimmt, histologische Untersuchungen wurde durchgefü hrt. Die Nutrazeutika wurden 24 h vor Ischämie und unmittelbar nach Reperfusion oral appliziert.Ergebnisse. Zwei der getesteten Nutrazeutika führten zu einer starken Hochregulation (Resveratrol: 11-fach, Ginseng: 17fach) der HO-1 Expression. Im Nierenarterien-Klemmmodell kam es nach 48 Stunden zu einem Anstieg des Serum-Kreatinin von 0,38 mg/dl AE 0,07 auf 3,06 mg/dl AE 0,86 in den Kontrolltieren. Die Applikation von beiden Nutrazeutika in einer Dosierung von 10 mg/kg (Resveratrol, 48h-Kreatinin 0,54 mg/dl AE 0,23) bzw. 30 mg/kg (Ginseng, 48h-Kreatinin 0,53 mg/dl AE 0,06) verhinderte dramatisch die Einschränkung der Nierenfunktion (p < 0,0001 für beide vs. Kontrolle). Die kompetitive Antagonisierung durch SnPP (5 mg/kg/KG) konnte diesen positiven Effekt vermindern. Histologische und immunhistochemische Auswertungen unterstützen die Ergebnisse.Schlussfolgerungen. Die Anwendung von fü r Menschen ungefährlichen Nutrazeutika stellt eine ausgezeichnete Möglichkeit dar, die HO-1 zu induzieren und den IRS zu minimieren.

The novel compound 5 0 -methoxy leoligin increases the ejection fraction of infarcted rat hearts by promoting CYP26B1dependent angiogenesis Background. In times of organ shortage strategies to recovery damaged organs seem to be valuable options to face this problem. Cellular based strategies to recover ischemic myocardium have shown promising experimental results but the clinical evidence is scarce. Especially the lack of angiogenesis in cardiac peri-infarction and infarction areas is one of the most important problems in functional cardiac recovery after myocardial infarction (MI). While searching for possible pharmacological strategies to stimulate angiogenesis after MI we conducted a screen for plant compounds with pro-neoangiogenetic properties.Methods and Results. 5 0 -methoxy leoligin, a compound isolated from the roots of Leontopodium alpinum (Edelweiss) potently stimulated endothelial cell migration, tube forma-Grundlagen. Ungeplante Reoperationen dienen als Indikator der Qualitätskontrolle in der Chirurgie. Daten für die Nierentransplantation liegen noch nicht vor.Methodik. Retrospektive Analyse von 320 Nierentransplantationen zwischen 9/2002 und 12/2010. Die Anzahl der ungeplanten Reoperationen und Ursache, Mortalität, Kreatininspiegel 1 Monat und 1 Jahr post TX und die Art der Immunsuppression in Bezug zum Organverlust wurden als mögliche Risikofaktoren evaluiert.Ergebnisse. In der Analyse unseres Patientenkollektives (320 Nierentransplantationen über 100 Monate) zeigt sich eine 14 % ige Reoperationsrate, dies entspricht 46 Reoperationen, wobei bei 16 % der Reoperierten mehr als eine Reoperation notwendig war. Ursächlich dafür waren vordergründig Gefäßkomplikationen (30 %) gefolgt von Hämatomausräumungen und Wundinfektionen, welche einer operativen Sanierung bedurften (28 %). Seltener kam es zu Reoperationen aufgrund urologischer Ursachen (20 %), Lymphozelen und abdomineller Komplikationen (je 6,5 %). Von insgesamt 40 Organverlusten waren 17 bei reoperierten Patienten (das entspricht 43 % der reoperierten Patienten) und 23 (9 %) bei nicht reoperierten Patienten zu verzeichnen (p < 0,001). In der Subanalyse der Komplikationen, welche zu Reoperationen führten, sind Gefäßkomplikationen die häufigste Ursache für vorzeitigen Organverlust (64 %, das entspricht 9 von 14 Reoperationen vs. 33 % bei Lymphozelen, 33 % bei abdominellen Komplikationen, 31 % bei Weichteilproblemen und 11 % bei urologischen Komplikationen). In Bezug zur Immunsuppression konnten keine statistisch signifikanten Unterschiede erhoben werden. Ein erhöhtes Serum-Kreatinin nach 1 Monat (Kreatinin >2) fand sich bei 42 % der reoperierten vs. 16,5 % der nicht reoperierten Patienten (p < 0,001). Nach einem Jahr war das Kreatinin bei 37 % der reoperierten Patienten >2, hingegen nur bei 16,5 % der nicht reoperierten Patienten (p ¼ 0,005 Ergebnisse. Varizen wurden mittels Ö sophago-Gastroskopie in 287 (72,5 %) von 396 untersuchten Patienten dokumentiert, wobei nur in einem Fall schwere Blutungen aus Case Report. 11 years after bilateral hand transplantation a patient developed a bullous pemphigoid (BP), which is an autoimmune blistering skin disease, characterized by autoantibodies targeting the type XVII collagen component of hemidesmosomes in the skin basement membrane zone. This is the first report on an autoimmune complication in a patient after vascularized composite allotransplantation (VCA). The patient showed a quite Background. The Mounier-Kuhn syndrome (MKS) is a rare disease characterized by a pathological dilation of the trachea and the bronchial system. The etiology of the disorder remains elusive but genetic alterations and degradation of elastic fibers are thought to be involved in the pathogenesis. No causative treatment is available although transplantation is an option for endstage disease. Here, we describe a patient suffering from MKS who received a lung transplant at our department.Methods. A 39-year-old male never-smoker presented at our department for evaluation for a lung transplantat. The patient suffered from Mounier-Kuhn syndrome (MKS) with greatly dilated trachea and bronchi. Since a familial clustering of Mounier-Kuhn syndrome is discussed in the literature, we performed a chromosomal analysis and an array-CGH to search for genetic abnormalities. At the time of transplantation we collected samples from the bronchi and performed hematoxylin and eosin (HE), Elastic van-Gieson's (EVG) and immunohistochemical stains. Specimens of main bronchi from the donor lung harvested for transplant served as control.Results. The chromosomal analysis and array-CGH revealed that the patient's genome was completely unremarkable, the karyogram as well as the genome hybridization showed no significant gain or loss in known encoding regions. Through the histological evaluation we found considerably lower amounts of elastic and collagen fibers in the submucosal layer in the patient compared to healthy controls. Furthermore inflammatory infiltrates were present in the connective tissue throughout the whole histological specimen. Since chronic inflammation is known to result in tissue remodeling we performed immunohistological staining of different matrix metalloproteinases. MMP1, MMP2, MMP3 and MMP9 were detected in MKS tissue, but were totally absent in control bronchi.Conclusions. Based on these findings we hypothesize that the pathophysiology of MKS is of a chronic inflammatory type leading to tissue destruction and a loss of elastic fibers, possibly due to upregulation of MMP. The triggering mechanism(s) for this inflammatory reaction remain elusive, but since MKS has been reported to be associated with autoimmune diseases, an autoimmune reaction against components of the major airways could be one possible explanation. Background. Lung transplant recipients (LTRs) are uniquely predisposed in developing severe complications associated with community acquired respiratory viral infections (CARV). We report the outcomes of influenza infections in a cohort of 82 screened lung transplant recipients at our center.Methods. Data were collected from December 2010 to March 2011 on using real-time polymerase chain reaction (PCR) from nasal secretion. During this period 245 patients frequented our out patient department for thoracic surgery. All LTRs (n ¼ 82) with respiratory symptoms were screened. There were 9 (10, 9%) confirmed cases. H1N1 infection was diagnosed in 5, influenza B in 4 lung transplant recipients, median age 36 (26-65) years, with a median of 6 (1,1-12) years post lung transplantation.Results. All patients with BOS grade 0 (bronchiolitis obliterans syndrome, n ¼ 5) were treated symptomatically alone, with no further impact on their lung function. Two patients of this group were vaccinated for seasonal influenza. Among the group of patients with pre-existing BOS (n ¼ 4), two lung transplant recipients, one with BOS I, the other with BOS II developed Grundlagen. Eine Vielzahl von Medikamenten mit potentiell toxischen Nebenwirkungen auf den Respirationstrakt sind bekannt. Zytotoxische Substanzen wie Methotrexat oder Bleomycin und andere Pharmaka wie Amiodaron oder diverse Antibiotika werden am häufigsten als Ursache fü r eine medikamentös induzierte interstitielle Lungenerkrankung genannt.Methodik und Ergebnisse. Wir berichten den Fall eines 64jährigen Patienten, der im Oktober 2010 wegen dilatativer Kardiomypathie mit terminaler Herzinsuffizienz eine orthotope Herztransplantation erhielt. Zu diesem Zeitpunkt bestand radiologisch bereits der Verdacht auf eine Amiodaron-induzierte Lungenfibrose mit funktionell reduzierten Werten: VC 3,49 l (73 %), FEV1 3,28 l (86 %), MEF50 6,52 l (135 %) und TLC 5,88 l (84 %). Der postoperative Verlauf gestaltete sich von kardialer Seite komplikationslos, allerdings war der Patient respiratorisch stark limitiert. Der Weaningprozess gestaltete sich aufgrund einer Lobärpneumonie protrahiert. Auch auf der Normalstation fiel bei geringsten Belastungen die Sauerstoffsättigung auf bis zu 87 %. Die Lungenfunktion 2 Monate nach HTX ergab eine restriktive Ventilationsstörung mit VC 1,33 l (37,9 %), FEV1 1,29 l (34,3 %), MEF50 3,3 l (71,2 %), TLC 2,76 (39,4 %), PaO2 52,8 % und PaCO2 38,7 %. Zur Histologiegewinnung wurde nach inkonklusiver Bronchoskopie eine offene Lungenbiopsie durchgeführt, welche das Vorliegen einer Lungenfibrose vom UIP-Typ ergab. Die Kontroll-Echokardiographien zeigten durchwegs eine gute rechtsund linksventrikuläre Funktion, allerdings verschlechterte sich der respiratorische Zustand des Patienten zunehmend, sodass er "high urgent" zur Lungentransplantation gelistet werden musste. Ein größen-und blutgruppenkompatibles Spenderorgan wurde verfügbar und eine bilaterale Lungentransplantation mit veno-arterieller ECMO Unterstützung durchgeführt. Die ECMO konnte am Ende der Operation bei hämodynamisch und respiratorisch stabilen Verhältnissen wieder explantiert werden. Die Ischämiezeit der rechten Lunge betrug 340 Minuten und links 460 Minuten. Am 19. postoperativen Tag konnte der Patient auf die Normalstation verlegt und nach weiteren 23 Tagen aus dem Krankenhaus entlassen werden.Schlussfolgerungen. Entgegen der allgemeinen Erwartung, dass eine Fibrose unter immunsuppressiver Therapie ein geringes Risiko fü r Progression hat, zeigte sich in diesem Fall eine dramatische Entwicklung nach der Herztransplantation. Deshalb sollte eine Lungenfibrose vor einer HTX genau abgeklärt und engmaschige postoperative Kontrollen durchgefü hrt werden.

Invasive mycoses after heart transplantation: outcome and long-term prognosis T. Haberl 1 , D. Hutschala 2 , C. Pelanek 1 , A. Aliabadi 1 , G. Laufer 1 , A. Zuckermann 1Background. Although orthotopic heart transplantation has become a routine procedure in treatment of end-stage cardiac failure, postoperative mycosal infections are still a serious risk for the patients. Objective of the current study is to show the influence of various parameters on the severity of infection and the outcome for patients.Methods. This is a single-center retrospective study including 338 patients who underwent heart transplantation from 2000-2008 at our center in Vienna. The analyzed data were collected during the post transplant course in the intensive care unit (ICU). Mycoses were diagnosed by cultures of blood, bronchoalveolar lavages (BAL) and smear tests of central arterial and venous catheters, wounds and drainages.Results. Mean age at time of transplantation was 49 years, 260 patients (76.9%) were male (age 5-71 years) and 78 patients (23.1%) were female (age 3 weeks-76 years). 90 patients (34%) acquired an invasive mycosal infection with candida species (94.3%), aspergillus (3.8%) and pneumocystis carinii (1.9%). Diagnosis was performed in 24.1% by cultures of blood, in 23.5% by BAL, 18.8% by smear tests of vena-cava catheters and in 11.1% from pulmonal artery catheters. In 77.6% treatment with a single antimycosal therapy was sufficient. 20.9% of patients were in need of a double-drug therapy and 1.5% needed a triple-drug-therapy. Patients with mycosal infection had an median stay in the ICU of 16 days (1st Quartile: 7.25, 3rd Quartile: 24.75) with a median of 4.5 (Q1: 0, Q3: 7) respirator-days compared with patient without infection who had median 9 (Q1: 4, Q3: 16) days in ICU with 1 (Q1: 0, Q3: 2) respirator-days. 100% of the patients with mycosis compared to 49% of the patients without mycosis developed or already had bacterial co-infection (p < 0.001). The 1-year survival of patients with mycosis was 63.3% compared to 248 patients without infection where 1-year survival was 83.3% (p < 0.001). Average time to first fungal infection was 7.6 AE 5.3 days.Conclusions. Invasive mycosal infection acquired by immune suppressed patients after heart transplantation is associated with prolonged time of stay on the ICU as well as higher rates of co-infections and a significant increase of mortality post transplant.Background. Xenotransplantation is a potential strategy to overcome the shortage of human donor organs. Since this technique has a major medical and psychological impact on patients and their family and friends, the attitude of patients currently waiting for organ transplantation is important.Medhods. Therefore we conducted a survey on the attitude towards xenotransplantation of patients on the waiting list and already transplanted patients. Patients received detailed information before being asked to fill in the questionnaire.Results. We found that 65% would accept xenotransplantation, irrespective of gender, education level or if the patients were on the waiting list or already transplanted. The most common concern was transmission of diseases or genetic material, followed by psychological concerns and ethical issues. More patients had a positive attitude towards accepting cell or tissue transplantation as compared to whole organs. Pig pancreas islet cell transplantation is generally well accepted, patients with diabetes mellitus show even higher acceptance rates than patients without diabetes.Conclusions. Xenotransplantation seems to be well accepted in patients who are potential future candidates for organ transplantation. Informing patients about the current status of research tended to decrease acceptance rates slightly.