Following vascular injury, platelet activation and thrombin generation occur in concert in a mutually amplifying scheme to promote the rapid and localized formation of a hemostatic plug. While the important role of platelets in coagulation is now established, interactions between platelets and the fibrinolytic system during clot dissolution have not been elucidated. Studies performed in this laboratory during the previous project period have shown that the fibrinolytic protease plasmin regulates platelet activation in a complex manner that is distinct from the actions of the coagulation protease thrombin. Furthermore, preliminary experiments have shown that plasmin also exerts different actions on vascular endothelial cells (EC).
The aim of this proposal is to investigate in detail the effects of plasmin and the fibrinolytic system on platelets and cultured vascular cells. Hypotheses to explain the complex actions of plasmin on platelets and EC will be pursued by examining the sequence and causal relationships of biochemical events in signal transduction induced by plasmin, and comparing and contrasting these events with those induced by the coagulation protease thrombin as well as other platelet and EC agonists. Studies will specifically focus on the control of phospholipid composition and turnover, protein kinase C activation and arachidonic acid metabolism. Biochemical mechanism of platelet and EC deactivation following stimulation by plasmin and other agonists by negative termination signals will be examined, an area of research in cellular signal- response coupling that has received little attention. Evidence for possibly selective actions of plasmin on the G proteins of adenylate cyclase will be followed with detailed studies of the control of the plasmin effect on platelets and EC by cyclic nucleotides. Preliminary evidence for synergistic inhibitory effects of plasmin and prostacyclin on platelet activation, a potentially important new platelet-EC interaction in controlling hemostasis and clot dissolution, will lead to studies of basic mechanisms of previously unknown synergistic inhibitory signaling pathways in platelets and EC. Studies of the regulation of EC function by plasmin will examine not only blood vessels as targets for actions of the fibrinolytic system but also elucidate basic biochemical mechanisms of signal transduction in vascular cells which are largely unknown. Understanding the modulation of platelet and EC actions and interactions by the fibrinolytic system will have pathophysiologic and pharmacologic importance in clarifying our concepts of thrombosis and hemostasis.
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