Lenich C, Liu J N, Gurewich V
Department of Medicine, Vascular Research Laboratory, Institute for the Prevention of Cardiovascular Disease, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA 02115, USA.
Blood. 1997 Nov 1;90(9):3579-86.
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.
基因敲除小鼠研究表明，尿激酶型纤溶酶原激活剂（u-PA）在纤维蛋白溶解过程中起重要作用，但其生理作用机制仍不清楚。我们推测血小板可能参与了这一机制，因为血小板携带一种新型的u-PA受体，且血液中一部分单链u-PA（scu-PA）与血小板紧密相关。因此，我们研究了血小板相关的u-PA对纤溶酶原的激活作用。当用纤溶酶原孵育洗涤后的血小板时，通过纤溶酶合成底物（S2403）水解检测未产生纤溶酶；然而，加入凝血酶后（而非其他激动剂），发生了血小板依赖性纤溶酶原激活。纤溶酶原激活与表面相关，可通过阻断血小板纤维蛋白原受体或阻止纤溶酶原与凝血酶激活的血小板表面结合来抑制。已确定u-PA是唯一负责的纤溶酶原激活剂，用外源性scu-PA富集血小板可显著增强纤溶酶原激活。这些发现似乎自相矛盾，因为凝血酶会使scu-PA失活。实际上，酶谱分析显示，即使使用最低剂量的凝血酶（1 u/mL）孵育一小时内，scu-PA也会失活。然而，随后凝血酶剂量依赖性（1至10 u/mL）地部分恢复了u-PA活性。u-PA的重新激活并非由于凝血酶的直接作用，而是需要血小板，并且发现与血小板溶酶体硫醇蛋白酶有关，与组织蛋白酶C一致。总之，我们证明了血小板相关的内源性或外源性scu-PA激活纤溶酶原的新途径，该途径由血小板的凝血酶激活特异性触发。这些发现可能有助于解释u-PA介导的生理性纤维蛋白溶解，并对scu-PA治疗性溶栓有启示意义。
