| The objective of this study was to acquire a better understanding of the physical and biochemical responses of human platelets to fluid shear stress in the presence or absence of platelet antagonists. Abnormal fluid shear stresses are present, for instance, in partially occluded coronary arteries. These forces are known to contribute to platelet-mediated thrombus formation in these areas through the initiation of platelet activation, aggregation, secretion, and microvesicle release. However, the mechanism of action of these antagonists, as well as the basic process underlying thrombus formation, has not been clearly delineated. Moreover, these antagonists have not been compared to each other with regard to their efficacy under shear stress. The studies described herein focused on the structural, functional, and biochemical responses of blood platelets to physiologic and pathophysiologic levels of fluid shear stress and specifically examined if: (1) the conformation of integrin α IIbβ3 is differentially affected by the receptor antagonists eptifibatide (Integrilin™), abciximab (ReoPro™), and tirofiban (Aggrastat®); (2) these antagonists cause intracellular signaling upon binding to the αIIbβ3 receptor under static conditions; (3) shear stress-mediated platelet activation occurs via intracellular signaling pathways that are distinct from those utilized by traditional platelet agonists; and (4) shear stress-induced platelet activation, aggregation, and procoagulant activity are differentially inhibited by αIIbβ3 receptor antagonists.; Overall, this study has provided evidence that the antagonists were not dissociated from the receptor under shear stress, nor was their efficacy compromised. Results from the immunoblot studies indicated that the tyrosine phosphorylation signals elicited by shear stress were a result of engagement of the α IIbβ3 receptor under shear stress rather than any direct impact of shear stress on the intracellular signaling pathways. Importantly, although aggregation was inhibited, the procoagulant activity of platelets was not and, in the case of tirofiban, there was an enhancement of expression of PS residues on the platelet surface. A complete characterization of the interactions of future αIIbβ3 antagonists with platelets under physiologic and pathophysiologic shear stresses will serve to improve αIIbβ3 antagonist design and help to elucidate the general role of fluid forces in platelet thrombus formation and hemostasis. (Abstract shortened by UMI.)... |
