Stimulatory effect of platelet releasates for induction of α_IIbβ₃ activation in platelets tested for recruitment.

Role of erythrocytes and aspirin ex vivo. PRP (2 × 10⁸platelets/mL), PRP plus RBCs, or WB were stimulated with collagen (1 μg/mL) and centrifuged to obtain a cell-free releasate within 1 minute. An aliquot of releasate was used as agonist for aspirin-free autologous PRP. Releasate-induced α_IIbβ₃ activation was monitored by flow cytometry with FITC–PAC-1 on recruiting platelets (“Materials and methods”). Releasates were obtained in duplicate for each data point, and duplicate flow cytometric measurements of each sample of releasate were performed. The 4 values so obtained from each donor were averaged; n represents the number of different volunteers studied. Data are mean ± SEM of percentage of platelets with bound PAC-1. (A) Effects of releasates from collagen-stimulated PRP, WB, and solvent controls (n = 22; *P < .001, PRP versus WB). (B) Relationship to hematocrit (n = 4; *P < .001). (C) Normal versus glutaraldehyde-fixed RBCs, compared with PRP (Hct 40%; n = 4;P < .001, normal RBCs versus PRP or fixed RBCs). (D) Effect of aspirin (n = 6; Hct 40%; before versus after ASA ingestion: *P < .005 for PRP, and *P < .001 for PRP plus RBCs). Significance was evaluated by Studentt test.