Thrombin stimulation of platelets induces plasminogen activation mediated by endogenous urokinase-type plasminogen...

Gene knockout mice studies indicate that urokinase-type plasminogen activator (u-PA) is importantly involved in fibrinolysis, but its physiologic mechanism of action remains poorly understood. We postulated that platelets may be involved in this mechanism, as they carry a novel receptor for u-PA and a portion of the single-chain u-PA (scu-PA) intrinsic to blood is tightly associated with platelets. Therefore, plasminogen activation by platelet-associated u-PA was studied. When washed platelets were incubated with plasminogen, no plasmin was generated as detected by plasmin synthetic substrate (S2403) hydrolysis; however, after the addition of thrombin, but not other agonists, platelet-dependent plasminogen activation occurred. Plasminogen activation was surface-related, being inhibited by blocking platelet fibrinogen receptors or by preventing plasminogen binding to the thrombin-activated platelet surface. U-PA was identified as the only plasminogen activator responsible and enrichment of platelets with exogenous scu-PA significantly augmented plasminogen activation. These findings appeared paradoxical because thrombin inactivates scu-PA. Indeed, zymograms showed inactivation of scu-PA during the first hour of incubation with even the lowest dose of thrombin used (1 u/mL). However, this was followed by a thrombin dose-dependent (1 to 10 u/mL) partial return of u-PA activity. Reactivation of u-PA was not due to the direct action of thrombin, but required platelets and was found to be related to a platelet lysosomal thiol protease, consistent with cathepsin C. In conclusion, a new pathway of plasminogen activation by platelet-associated endogenous or exogenous scu-PA was demonstrated, which is specifically triggered by thrombin activation of platelets. These findings may help explain u-PA-mediated physiological fibrinolysis and have implications for therapeutic thrombolysis with scu-PA.