Flow cytometry analysis of the importance of Rac1 in shear-induced PS exposure and microvesiculation in activated platelets. (A-C) Washed human platelets (n = 3) preincubated with 0.1% DMSO or 100 µM NSC23766 were treated with or without 0.1 U/mL thrombin and 5 µg/mL collagen, in the absence or presence of shear. The samples were analyzed for Annexin V binding and MVs using flow cytometry. (A) Histograms and quantification of (B) Annexin V binding and (C) MV release are shown (mean ± SEM, n = 3). (D-F) Washed WT and Rac1^−/− platelets were treated with or without 0.03 or 0.1 U/mL thrombin and with or without shear. (D) Histograms of Annexin V binding. (E) Quantification of PS exposure. (F) Quantification of MV release. (E-F, mean ± SEM, n = 4). (G-I) Washed WT and Rac1^−/− platelets were treated with or without 2 μg/mL collagen and 50 nM A23187, in the absence or presence of shear (n = 8 and 4, respectively). (G) Histograms of Annexin V binding. (H) Quantification of Annexin V binding. (I) Quantification of MV release (H-I, mean ± SEM). For all data, *** represents statistical significance of P < .001.