Characterization of von Willebrand Factor-Enriched Extracellular Vesicles and Their Association with Platelet Adhesion in COVID-19

Microvascular thrombosis is a hallmark of severe COVID-19 and a major contributor to adverse clinical outcomes. Extracellular vesicles have emerged as key mediators of thrombo-inflammation, yet their functional and molecular characteristics remain incompletely understood. Here, we investigated the prothrombotic activity, cellular origin, and von Willebrand factor (VWF) composition of plasma-derived small extracellular vesicles (SEVs) from hospitalized COVID-19 patients. We demonstrate that SEVs from critically ill COVID-19 patients markedly enhance platelet adhesion and thrombus formation under arterial shear conditions and promote endothelial activation, as evidenced by increased release of ICAM-1 and VCAM-1. These SEVs were highly enriched in ultra-large VWF multimers, with significantly elevated high-to-low molecular-weight ratios compared with controls. Flow cytometric analysis showed increased circulating SEVs derived from leukocytes, platelets, and endothelial cells in severe disease, with VWF-positive SEVs predominantly originating from endothelial cells. The enrichment of VWF-SEVs was independent of gender and diabetes status, suggesting a primary association with disease severity. Collectively, these findings indicate that SEVs, particularly those rich in endothelial-derived VWF, are associated with platelet activation and may contribute to the prothrombotic milieu in COVID-19. These findings underscore the importance of VWF-positive SEVs as biomarkers of disease severity and warrant further investigation into their role in COVID-19 related vascular pathology.