Synergistic activities of shear stress and RBC-derived ADP. (A) Levels of CD62p⁺ platelets in response to 10 Pa shear stress exposure after reconstitution with fRBCs or sRBCs; n = 3 per condition; t test). (B) ADP released from washed RBCs subjected to increasing levels of shear stress (n = 5-7 per condition; 1-way ANOVA). The white bar indicates the amount of ADP from lysed RBCs at 40% hematocrit (n = 5). (C) ADP-induced aggregation of resting platelets and those primed with shear stress (10 Pa for 5 minutes at 37°C; n = 7-8 per condition; 2-way ANOVA); a representative aggregation curve in response to 2 μM ADP (left). (D) SIPA in response to 8 Pa when PRP was reconstituted with unwashed, washed, and apyrase-treated RBCs exposed to high shear stress (n = 8-12 per condition; 1-way ANOVA). (E) Collagen-induced aggregation of resting platelets and those primed with shear stress (10 Pa for 5 minutes at 37°C; n = 7-8 per condition; 2-way ANOVA) with representative curve in response to 2 μg/mL collagen on the left. (F) Plasma levels of RBC-derived EVs (CD235⁺ EVs) of patients at baseline and after LVAD implantation (n = 12; paired t test). BL, baseline; fRBC, fresh RBC; sRBC, cold-stored RBC.