A2 blocked activated VWF in TBI mice. (A) HUVECs were incubated with BDEVs for 3 hours at 37°C with increasing doses of A2. The supernatants were collected and analyzed for VWF using ELISA (n = 25, 1-way ANOVA, *P < .01 vs untreated). (B) The culture medium (EBM-2), sham plasma (sham-PPP), plasma from TBI mice receiving TBST (TBI-PPP) or A2 (TBI-PPP+A2), purified EVs (TBI-EVs and TBI-EVs+A2), EV-free plasma (EVFP) from the TBI mice, and purified VWF were incubated with confluent HUVECs for 3 hours at 37°C. The HUVECs were washed and incubated with FITC-dextran (100 μg/mL) for 1 hour at 37°C to quantify FITC intensity in the bottom chamber (n = 56, 1-way ANOVA). (C,D) VWF:Ag (C) and VWF:CB (D) in plasma samples collected at 3 and 6 hours post-TBI (n = 36, 1-way ANOVA). (E) VWF:CB measured in mice receiving increasing doses of A2 (n = 12, 1-way ANOVA). (F) The VWF:CB-to-VWF:Ag ratio calculated for the 3 groups of mice (n = 21, 1-way ANOVA). (G) The expression of CD62p on platelets from sham mice and TBI mice receiving TBST or A2 (n = 21, 1-way ANOVA). (H) Plasma levels of platelet EVs with surface-bound VWF (CD41a⁺/VWF⁺, n = 21, 1-way ANOVA).