Circulating Von Willebrand factor and high molecular weight multimers as markers of endothelial injury predict COVID-19 in-hospital mortality

Abstract

Background: Coronavirus disease 2019 (COVID-19) is a respiratory disease associated with endotheliitis and microthrombosis.

Objectives: To correlate endothelial dysfunction to in-hospital mortality in a bi-centric cohort of COVID-19 adult patients.

Methods: Consecutive ambulatory and hospitalized patients with laboratory-confirmed COVID-19 were enrolled. A panel of endothelial biomarkers and von Willebrand factor (VWF) multimers were measured in each patient ≤ 48 h following admission.

Results: Study enrolled 208 COVID-19 patients of whom 23 were mild outpatients and 189 patients hospitalized after admission. Most of endothelial biomarkers tested were found increased in the 89 critical patients transferred to intensive care unit. However, only von Willebrand factor antigen (VWF:Ag) scaled according to clinical severity, with levels significantly higher in critical patients (median 507%, IQR 428-596) compared to non-critical patients (288%, 230-350, p < 0.0001) or COVID-19 outpatients (144%, 133-198, p = 0.007). Moreover, VWF high molecular weight multimers (HMWM) were significantly higher in critical patients (median ratio 1.18, IQR 0.86-1.09) compared to non-critical patients (0.96, 1.04-1.39, p < 0.001). Among all endothelial biomarkers measured, ROC curve analysis identified a VWF:Ag cut-off of 423% as the best predictor for in-hospital mortality. The accuracy of VWF:Ag was further confirmed in a Kaplan-Meier estimator analysis and a Cox proportional Hazard model adjusted on age, BMI, C-reactive protein and D-dimer levels.

Conclusion: VWF:Ag is a relevant predictive factor for in-hospital mortality in COVID-19 patients. More than a biomarker, we hypothesize that VWF, including excess of HMWM forms, drives microthrombosis in COVID-19.

Keywords: COVID-19; Endothelial activation; Microthrombosis; Mortality; Multimers; Von Willebrand factor.

Fig. 1

Levels of endothelial activation biomarkers according to critical patients, non-critical patients, outpatients, and non-COVID-19 individuals. Datapoints indicate individual measurements, whereas horizontal bars represent the means with standard deviations. Green shaded areas indicate the normal ranges of values. p-value comes from the Mann–Whitney for comparison between two groups and the Kruskal Wallis test for comparison between the four groups. a Measurement of circulating endothelial cells (CECs) in 37 critical and 40 non-critical COVID-19 patients at admission. Plasma levels of angiopoietin-1 (b), soluble endoglin (c), soluble endothelial protein C receptor (sEPCR) (d), soluble vascular cell adhesion molecule (VCAM-1) (e), soluble E-selectin (f), sTM (soluble thrombomodulin) (g), angiopoietin-2 (h), von Willebrand factor antigen (VWF:Ag) (i) in 89 critical and 96 non-critical COVID-19 patients, 23 COVID-19 outpatients and 29 non-COVID-19 individuals at admission

Fig. 2

Comparisons of von Willebrand factor multimers pattern in 40 critical and 37 non-critical COVID-19 patients. Datapoints indicate individual measurements, whereas horizontal bars show mean with standard deviation. P-value comes from the Mann–Whitney test for comparison between the two groups. a Low molecular weight multimers (LMWM) expressed as a percentage of total multimer (left panel) or a ratio compared to healthy individuals’ pool of plasma (right panel). b Intermediate molecular weight multimers (IMWM) expressed as a percentage of total multimers (left panel) or a ratio compared to healthy individuals’ pool of plasma (right panel). c High molecular weight multimers (HMWM) expressed as a percentage of total multimers (left panel) or a ratio compared with healthy individuals’ pool of plasma (right panel). d Curves showing Von Willebrand factor multimers pattern analysed in densitometric analysis of one critical COVID-19 patient, one non-critical COVID-19 patient and one non-COVID-19 individual. HMWM are located at the right part of each curves. (Refer to materials and methods for detailed multimers classification)

Fig. 3

Receiver operating curves evaluating unadjusted von Willebrand factor antigen’s ability to predict in-hospital-mortality. The diagonal black dotted segment is the reference line. AUC area under the curve, Se sensitivity, NPV negative predictive value, CI confidence interval

Fig. 4

Survival curves according to von Willebrand factor antigen concentration using a Kaplan–Meier estimator. Data are shown for patients with low von Willebrand factor antigen (< 423%) and high von Willebrand factor antigen (> 423%). Survival curves are compared using the log-rank test

Fig. 5

Forest plot showing the Cox proportional hazards model for von Willebrand factor antigen adjusted for age, body mass index, d-dimer and C-reactive protein. Values are hazard ratios with 95% confidence intervals. VWF:Ag von Willebrand factor antigen, BMI body mass index, CRP C-reactive protein, aHR adjusted hazard ratio, CI confidence interval