The selective uptake of high molecular weight urokinase-type plasminogen activator by human platelets

Abstract

Summary In a previous study it was shown that about 20% of the endogenous pro-urokinase (pro-UK) in whole blood was associated with platelets and that there was uptake of pro-UK by platelets from the ambient plasma or buffer. In the present study, the association of urokinase-type plasminogen activator (u-PA) with platelets was further characterized. Isolated platelet suspensions were exposed to high and low molecular weight u-PA or tissue plasminogen activator (t-PA), then washed and analyzed by SDS gel electrophoresis and zymography. Only the high molecular weight u-PA was found associated with platelets and this association resisted acid washing, aggregation by thrombin, or osmotic swelling of platelets to obliterate the open canalicular system. Ultrasonic disruption of platelets and isolation of the membrane portion indicated that u-PA was associated with the membrane. The catalytic site of u-PA remained available on the platelet surface as evidenced by its inhibition by the specific inhibitor glutamyl-glycyl-arginyl chloromethylketone (GGAck). Uptake by platelets, but not red cells, of pro-UK from whole blood was demonstrated, and this was detectable at physiological concentrations of pro-UK suggesting that endogenous u-PA in platelets may originate from the ambient plasma. A Western blot of platelet membrane proteins with antibody to u-PA receptor (u-PAR) showed no evidence of u-PAR. It was postulated that u-PA binds to a platelet membrane protein distinct from u-PAR, via the A-chain of u-PA, and that this interaction becomes stabilized resisting dissociation.