key: cord-0725508-7xm2voxq
authors: Abou-Arab, Osama; Bennis, Youssef; Gauthier, Pierre; Boudot, Cedric; Bourdenet, Gwladys; Gubler, Brigitte; Beyls, Christophe; Dupont, Hervé; Kamel, Said; Mahjoub, Yazine
title: Association between inflammation, angiopoietins and disease severity in critically ill COVID-19 patients: a prospective study
date: 2020-12-23
journal: Br J Anaesth
DOI: 10.1016/j.bja.2020.12.017
sha: 2eaf0a43ae4ea8b2d5f27d56f93a83a82a1ead19
doc_id: 725508
cord_uid: 7xm2voxq

nan

inflammation and endothelial injury biomarkers with regard to disease severity of critically ill COVID-19 patients in order to investigate the potential mechanism of disease progression.

We conducted a single centre prospective study at Amiens Hospital University (France) as an ancillary study of a prospective COVID-19 critically patient database (registered on ClinicalTrials.gov NCT04354558 and CNIL number PI2020_843_0026). The population study was adult patients admitted to our intensive care unit (ICU) with a confirmed diagnosis of COVID-19 (RT-PCR diagnosed from nasopharyngeal swab). Severity was defined according to the World Health Organization case definition 3 . The Severe group included patients with respiratory distress syndrome (rate ≥ 30 breaths min -1 ) or oxygen saturation ≤ 93% at rest or ratio of arterial partial pressure of oxygen to fractional concentration of oxygen in inspired air < 300 mm Hg (X kPa), or > 50% lesion progression over 24-48 h by pulmonary imaging. The Critical group included patients with respiratory failure and requiring mechanical ventilation or with shock or organ failure that requires ICU care.

Peripheral blood samples were collected on ICU admission in EDTA-containing tubes and centrifuged within 30 min of sampling for 10 min at 1000 × g. Plasma samples were collected and stored at -80°C until use. Cytokine levels were analysed in 2-fold diluted plasma samples using the ProteinSimple® (San Jose, CA, USA) microfluidic ELISA technology, according to the manufacturer's instructions. The inter-assay and intra-assay coefficients of variation for tumour necrosis factor-α (TNF-α) and IL-6 were all <9%. Levels of angiopoetin-1 (Ang-1) and Ang-2 were determined in 5-fold diluted plasma samples using commercially available ELISA kits according to the manufacturer's instructions (ELH-Angiopoietin1 and ELH-Angiopoietin2, RayBio-tech, Norcross, GA, USA). Assay sensitivity was 30 pg mL -1 for Ang-J o u r n a l P r e -p r o o f 1 and Ang-2 were all <12%.

From March to May, 2020, 65 patients were included in the study: 17 patients in the severe group (26%) and 48 in the critical group (74%). Patient data are reported in the Table. At ICU admission, TNF-α and IL-6 were significantly higher in the critical group in comparison with the severe group (p=0.006 and p=0.038, respectively). Ang-1 did not significantly differ between groups (p=0.221) whereas Ang-2 and Ang-2/Ang-1 were significantly higher in the critical group (p=0.025 and p=0.028, respectively). Ang-2 was positively correlated to IL-6 and TNF-α (p=0.029 and p<0.0001); Ang-2/Ang-1 was correlated to TNF-α (p<0.0001) (supplementary Figure) .

In univariate logistic regression analysis, TNF-α, IL-6, Ang-2 and Ang-2/Ang-1 were associated with vasopressor use (p<0.0001, p=0.038, p=0.013 and p=0.041, respectively).

TNF-α and Ang-2 levels were associated with renal replacement therapy (RRTY) requirement (p=0.001, p=0.046 respectively) but not IL-6 nor Ang2/Ang1. TNF-α was the only plasma marker associated with in-ICU death (p=0.025) (supplementary Table) . After adjustment for clinical severity, only TNF-α was an independent factor associated with vasopressor use (OR=54.03, CI 95%: 1.12-2611), RRT requirement (OR=142.7 CI 95%: 4.3-4716.1) and in-ICU death (OR=22.5, CI 95%: 1.1 -440.6).

This study has three main findings. First, IL-6, TNF-α and Ang-2 were increased in the critical group. Second, TNF-α was associated with organ failure and mortality. Third, TNF-α levels correlated with Ang-2. The first finding supports the so-called cytokine storm pathogenesis. Nevertheless, our findings on IL-6 are not in accordance with previous reports.

It was suggested that IL-6 is a key factor in COVID-19 associated cytokine response, allowing therapies with IL-6 antagonists such as tocilizumab. However, the first clinical trials with IL-6 antagonists were disappointing with negative results regarding mortality or disease progression 4 . Moreover, a recent meta-analysis confirmed that IL-6 elevation in COVID-19 is actually lower than in other respiratory virus-related disease 5 .

In our report, TNF-α was a better correlate of initial severity and progression of ICU stay.

Observations from large registries of COVID-19 patients with chronic rheumatism or inflammatory bowel diseases suggested that anti-TNF-α therapies may have prevented COVID-19 progression during the outbreak 6 . These results confirm the pro-inflammatory state associated with COVID-19 and emphasize the confirmed benefice of corticosteroid therapy on disease progression and mortality in critically ill patients 7 .

We also assessed endothelial injury with Ang-2 elevation. This result reinforces our previous hypothesis that COVID-19 is, at least partially, a vascular disease with endothelial damage, angiogenesis and thrombosis 8 . Ang-2 expression can be promoted by different stimuli including inflammation and hypoxaemia. We confirmed a positive correlation between Ang-2 and TNF-α levels suggesting endothelial activation induced by inflammation, although an additive effect of tissue hypoxia on Ang-2 release was not excluded 9 . Critical patients had higher Ang-2 elevation than less severe patients. Ang-2, by promoting endothelial permeability, may increase pulmonary oedema and hence disease severity. In accordance with our results, Smadja and colleagues 10 showed that Ang-2 level at hospital admission predicted COVID-19 severity.

The lack of a non-COVID-19 comparative group is a major limitation of this study. For example, Ang-2 levels in COVID-19 patients might be similar to those of patients with other causes of ARDS. Nevertheless, we compared Ang-2 according to COVID-19 severity showing that the most severe patients had higher markers of endothelial injury. Another major limitation is the single timepoint measurement.

Our main mechanistic hypothesis is that after the initial phase of viral infection, a proinflammatory state occurs aggravated by local hypoxaemia leading to progression of pulmonary vascular injuries.

Study conception, statistical analysis and manuscript drafting and final approval: OAA, SK, YM and YB. Clinical data collection: PG, CB and OAA. Vascular and cytokine measurement: BG, CB, GB, SK and YB. Manuscript revision: OAA, YM, HD, SK and YB.

The authors declare that they have no conflicts of interest.

Supported by Amiens Hospital University. (82) 52 (80) 17 (100) 17 (100) 36 (75) 35 (73) <0.0001

ICU mortality (%) 12 (18) 1 (6) 11 (23) 0.088 J o u r n a l P r e -p r o o f

Is a "Cytokine Storm