Fig. 3

The activation NLRP3 inflammasome and the release of IL-1β increase coagulation factors mediated by S protein. Human umbilical vein endothelial cells (HUVEC) were treated with viral S protein at 7, 35 and 70 nM or IL-1β at 2.5 ng/mL for 18 h. The levels of von Willebrand Factor (vWF) were determined A by Western blot in total cell lysates (n = 6), and B visualized in HUVEC stimulated with S (35 nM) or IL-1β (2.5 ng/mL) by indirect immunoflorescence (red). Representative images are shown in which vWF can be seen both as granules within the HUVEC cytoplasm and multimeric filaments in the extracellular space. Nuclei were counter-stained with DAPI (blue). Scale bar represents 50 μm. C vWF was also measured by ELISA in cell supernatants (n = 4). In the same experimental conditions, the protein levels of D ADAMTS-13, E FVIII, and F TF were quantified in total cell lysates by Western blot. Representative gels are shown on top of the corresponding graphs, with β-actin used as a loading control. Bar graphs represent mean ± SEM. Statistical differences were analyzed by t-test. *p < 0.05