Fig 1: Schematic Illustration of Structural and Functional Changes of VWF in an LVAD-Driven Blood Flow(A) High shear stress (HSS) activates VWF (gain of function) and facilitates VWF cleavage by ADAMTS-13 (loss of function), with the former being the predominant pathway in patients on LVAD supports. (B) The 2 vicinal cysteines in the A2 domain can be oxidized to disrupt the A1-A2 complex so that the A1 exposed (activated) VWF can form a complex with isolated A2. (C) Cysteine thiols of VWF can be oxidized to form intermultimeric disulfide bonds under HSS and thus cannot be precipitated by the active thiol beads, which form mix disulfide bonds with surface exposed free thiols on VWF.68 Abbreviations as in Figures 1 and 3.
Fig 2: Shear-Induced VWF Activation and Cleavage(A) ADAMTS-13 antigen (ADAMTS-13:Ag) and (B) its ratio to VWF:Ag in samples from HS and patients sampled at BL, at Post, and at B or T events (sample sizes at the bottom of the panel figures, *P < 0.05 and ***P < 0.001, HS vs other groups). (C) A2 precipitated VWF from the plasma of LVAD patients who developed bleeding or stroke post-LVAD and the precipitation was blocked by A1 (representative from 26 patients analyzed). A2 precipitated VWF purified from cryoprecipitate only after ristocetin (Risto.) treatment. (D) A2 also blocked SIPA of HS (n = 8/group). (E) Thiol-containing VWF precipitated from HS (n = 13) and patients at BL and Post (n = 26, ***P < 0.001). (F) Cleaved VWF (cVWF) and uncleaved VWF (ucVWF) from HS exposed to 5 minutes of high shear stress (lanes 1-5) or in the static condition with 1.5 M urea and 1 mM of BaCl2 (lane 6). (G) A2 cleavage in the static condition. (H) (Top) cVWF and ucVWF from HS exposed to high shear stress for 60 minutes (lanes 1-8), under static incubation of 16 hours (lane 9), or exposed to a constant vortex for 60 minutes (lane 10). (Bottom) Densitometry of 8 independent experiments. All quantitative data were analyzed using 1-way analysis of variance. cA2 = cleaved A2; MM = molecular mark in kDa; RBC = red blood cells; ucA2 = uncleaved A2; ucVWF = uncleaved VWF; other abbreviations as in Figures 1 and 3.
Fig 3: Immune-thrombosis in COVID-19: endothelial and plasma coagulation: (1) Viral infection leading to endothelial damage and inflammation. (2) this leads to increased expression of cytokines, and procoagulant factors like VWF, activation of platelets and neutrophil extracellular traps (NETs) release; VWF is cleaved by ADAMTS13. (3) increased thrombin and plasmin generation potential in presence of thrombomodulin leading to increase in fibrin degradation products and D-dimer. (4) thrombus formation and (5) subsequent fibrin degradation that results in increased D-dimer in Covid-19. TM, thrombomodulin; tPA, tissue plasminogen activator; TF, tissue factor; VWF, von Willebrand factor; NETs, neutrophil extracellular traps (Figure created with Biorender.com).
Fig 4: VWF/ADAMTS13 axis changes and coagulation in acutely ill COVID-19 (-) and (+) patients: (A) VWF antigen, 1.868 (IQR, 1.257–2.770) (-), 2.736 (IQR, 1.822–4.060) (+), p < 0.0001; (B) VWF collagen binding activity (9) 2.989 (IQR, 1.958–4.252) (-), 3.745 (IQR, 2.506–5.262) (+), p < 0.0001; (C) FVIII antigen 1.79 (IQR, 0.898–3.283) (-), 1.769 (IQR, 1.031–3.366) (+), p = 0.4154; (D) ADAMTS13 antigen 0.806 (IQR, 0.592–1.023) (-), 0.813 (IQR, 0.614–1.069) (+), p = 0.539; (E) ADAMTS13 activity 0.540 (IQR, 0.420–0.689) (-), 0.597 (IQR, 0.427–0.767) (+), p = 0.027; (F) VWF:AG/ADAMTS13 activity 5.567 (IQR, 3.352–8.245) (-), 6.051 (IQR, 3.824–10.17) (+), p < 0.0044. Datapoints indicate individual measurements, and p-values are from the Mann-Whitney analysis for comparison within groups. Values are presented as median and interquartile range (IQR, 25th-75th percentile) for continuous variables. ns, P > 0.05; *P = 0.05; **P = 0.01; ***P = 0.001; ****P = 0.0001.
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