Circulating Human Platelet Membrane Microparticles

Abstract

“Platelet dust” providing coagulant activity has been postulated to exist in human plasma. We studied human subjects to identify platelet membrane microparticles (MP) by immunologic methods in cell-free plasma. Blood was drawn into a syringe containing ACD-PGE1 and centrifuged at 30,000 g⋅min to obtain plasma with no identifiable platelets. The plasma concentration of platelet factor 4 was normal (9.4 ng/ml, n = 14), demonstrating no in vitro platelet activation. MP were isolated from diluted plasma by centrifugation at 7 × 105 g⋅min, the pellet was washed once, and solubilized in Triton X-100. Antibody from rabbits immunized with whole platelets did not cross-react with red cells or white cells. Using tandem crossed immunoelectrophoresis with MP and whole platelets, platelet antigens were identified in the MP. A specific antibody to platelet membrane glycoprotein II-III was prepared by adsorption of the anti-whole platelet antibody with cryoprecipitate and throm- basthenic platelets and used to quantify MP by rocket immunoelectrophoresis. Normal plasma MP concentration was 4.37 ± .71 (SE) μg/ml (n = 20). MP concentration was greater in serum (65.2 ± 13.2 μg/ml, p < .001) demonstrating platelet microparticle formation during coagulation. In a patient with thrombocytosis (1.76 × 106 platelets/μl) plasma MP were normal (7.5 μg/ml) but serum MP were increased (185 μg/ml). Plasma MP were assayed in 11 normal subjects before and after aspirin (640 mg/day × 7 days) and MP decreased in 9 (mean values: 4.45 μg/ml pre- and 1.92 μg/ml post-aspirin, paired t test: p <.02). Therefore platelet membrane fragmentation occurs during normal circulation and is inhibited by aspirin.