Background Traumatic injury can lead to dysregulation of the normal clotting system, resulting in hemorrhagic and thrombotic complications. Platelet activation is robust following traumatic injury and one process of platelet activation is to release of extracellular vesicles (PEV) that carry heterogenous cargo loads and surface ligands. Objectives We sought to investigate and characterize the release and function of PEVs generated following traumatic injury. Methods PEV content and quantity in circulation following trauma in humans and mice was measured using flow cytometry, size exclusion chromatography, and nanoparticle tracking analysis. PEVs were isolated from circulation and the effects on thrombin generation, bleeding time, hemorrhage control, and thrombus formation were determined. Finally, the effect of hydroxychloroquine (HCQ) on PEV release and thrombosis were examined. Results Human and murine trauma results in a significant release of PEVs into circulation compared with healthy controls. These PEVs result in abundant thrombin generation, increased platelet aggregation, decreased bleeding times, and decreased hemorrhage in uncontrolled bleeding. Conversely, PEVs contributed to enhanced venous thrombus formation and were recruited to the developing thrombus site. Interestingly, HCQ treatment resulted in decreased platelet aggregation, decreased PEV release, and reduced deep vein thrombosis burden in mice. Conclusions These data demonstrate that trauma results in significant release of PEVs which are both pro‐hemostatic and pro‐thrombotic. The effects of PEVs can be mitigated by treatment with HCQ, suggesting the potential use as a form of deep vein thrombosis prophylaxis.

Treatment of bleeding disorders using transfusion of donor-derived platelets faces logistical challenges due to their limited availability, high risk of contamination, and short (5 to 7 days) shelf life. These challenges could be potentially addressed by designing platelet mimetics that emulate the adhesion, aggregation, and procoagulant functions of platelets. To this end, we created liposome-based platelet-mimicking procoagulant nanoparticles (PPNs) that can expose the phospholipid phosphatidylserine on their surface in response to plasmin. First, we tested PPNs in vitro using human plasma and demonstrated plasmin-triggered exposure of phosphatidylserine and the resultant assembly of coagulation factors on the PPN surface. We also showed that this phosphatidylserine exposed on the PPN surface could restore and enhance thrombin generation and fibrin formation in human plasma depleted of platelets. In human plasma and whole blood in vitro, PPNs improved fibrin stability and clot robustness in a fibrinolytic environment. We then tested PPNs in vivo in a mouse model of thrombocytopenia where treatment with PPNs reduced blood loss in a manner comparable to treatment with syngeneic platelets. Furthermore, in rat and mouse models of traumatic hemorrhage, treatment with PPNs substantially reduced bleeding and improved survival. No sign of systemic or off-target thrombotic risks was observed in the animal studies. These findings demonstrate the potential of PPNs as a platelet surrogate that should be further investigated for the management of bleeding.

Introduction-Recent evidence demonstrated that prehospital plasma in patients at risk of hemorrhagic shock was safe for ground transport and resulted in a 28-day survival benefit for air