Effect of physiologic shear stresses and calcium on agonist-induced platelet aggregation, secretion, and thromboxane A2 formation

IntroductionPlatelets in vivo react under low-shear venous-flow as well as high-shear arterial-flow conditions. Because most studies were carried out at low shear stresses, platelet granule secretion at high shear has not been examined thoroughly. We investigated the secretion of all three types of platelet granules and thromboxane A2 formation at high shear after stimulation with ADP or thrombin.Materials and methodsWashed human platelets were reacted rapidly (< 5 s) in a quenched-flow system simulating high-shear arterial-flow conditions (30 dyn/cm2). For comparison, we employed a low-shear stirring system (1–5 dyn/cm2). Serotonin release and membrane exposure of P-selectin (alpha), CD63 (dense), and CD107a (lysosomes) were used to assess granule secretion. Aggregation was evaluated by resistive-particle counting of the remaining platelet singlets.Results and conclusionsADP and thrombin induced similar strong levels of aggregation (∼ 70%) at high shear by 5 s. Thrombin also caused release of about 40% of all alpha and dense granules within 5 s. However, by 5 s at high shear, ADP failed to induce significant granule secretion or thromboxane A2 formation (< 5%, p > 0.05). By 10 min at low shear, ADP caused secretion and thromboxane A2 formation only at non-physiological, micromolar extracellular Ca2+ concentrations. These results emphasize the ability of thrombin to initiate multiple aspects of platelet function within seconds, while ADP was only able to induce rapid aggregation.