Braat E A, Los P, Rijken D C
Gaubius Laboratory, TNO Prevention and Health, Leiden, The Netherlands.
Blood Coagul Fibrinolysis. 1998 Jul;9(5):419-27. doi: 10.1097/00001721-199807000-00004.
Thrombin cleaves single-chain urokinase-type plasminogen activator (scu-PA) into a virtually inactive two-chain form (tcu-PA/T), a process which may contribute to the maintenance of a fresh blood clot. We have examined the inactivation of scu-PA by thrombin in a plasma milieu to get more insight in the physiological relevance of this phenomenon. Citrated pooled normal plasma was treated with thrombin in the absence and presence of thrombomodulin. After an incubation period of 30 min the concentrations of scu-PA and tcu-PA/T were measured using specific bioimmunoassays. The inactivation of scu-PA in citrated plasma was found to be stimulated fourfold by thrombomodulin. Kinetic experiments showed that the inactivation of scu-PA by thrombin in the absence and presence of thrombomodulin occurred rapidly and declined within 1 min as a result of rapid inhibition by antithrombin III (ATIII) and other possible inhibitors. Calcium had no direct effect on the inactivation of scu-PA by exogenously added thrombin in the absence and presence of thrombomodulin. However, recalcification of plasma induced significant inactivation of scu-PA in plasma as a result of endogenous thrombin generation through the contact activation system. This calcium-induced inactivation of scu-PA was completely abolished in the presence of thrombomodulin, most likely as a result of activation of protein C by the complex formed between thrombomodulin and endogenously generated thrombin. Thrombomodulin thus appeared to play a dual role both by stimulating the inactivation of scu-PA by thrombin, and by inhibiting calcium-induced inactivation of scu-PA in plasma. In the plasma from a patient heterozygous for protein C deficiency, thrombomodulin could not prevent calcium-induced generation of tcu-PA/T, whereas the stimulating effect of thrombomodulin predominated instead. This result implied that disturbance of the protein C pathway may lead to the inactivation of substantial amounts of scu-PA in plasma under (patho)physiological circumstances and may provide an additional explanation for the association found between thromboembolism and deficiencies in the protein C pathway. This study shows that the amount of scu-PA that is inactivated in plasma depends mainly on the generation of thrombin and on thrombomodulin. We conclude that the inhibition of scu-PA-induced fibrinolysis appears to be regulated by activation of the coagulation system, providing a link between coagulation and fibrinolysis.
凝血酶将单链尿激酶型纤溶酶原激活剂（scu-PA）裂解为几乎无活性的双链形式（tcu-PA/T），这一过程可能有助于维持新鲜血凝块。我们研究了在血浆环境中凝血酶对scu-PA的灭活作用，以更深入了解这一现象的生理相关性。用凝血酶处理枸橼酸化的混合正常血浆，分别在有无血栓调节蛋白的情况下进行。孵育30分钟后，使用特异性生物免疫测定法测量scu-PA和tcu-PA/T的浓度。发现血栓调节蛋白可使枸橼酸血浆中scu-PA的灭活增强四倍。动力学实验表明，在有无血栓调节蛋白的情况下，凝血酶对scu-PA的灭活均迅速发生，并且由于抗凝血酶III（ATIII）和其他可能的抑制剂的快速抑制作用，在1分钟内就下降了。在有无血栓调节蛋白的情况下，钙对外源性添加的凝血酶对scu-PA的灭活没有直接影响。然而，血浆重新钙化会导致血浆中scu-PA的显著灭活，这是由于通过接触激活系统产生内源性凝血酶所致。在血栓调节蛋白存在的情况下，这种钙诱导的scu-PA灭活被完全消除，这很可能是由于血栓调节蛋白与内源性产生的凝血酶形成的复合物激活了蛋白C。因此，血栓调节蛋白似乎发挥了双重作用，既刺激凝血酶对scu-PA的灭活，又抑制血浆中钙诱导的scu-PA灭活。在一名蛋白C缺乏杂合子患者的血浆中，血栓调节蛋白无法阻止钙诱导的tcu-PA/T生成，而血栓调节蛋白的刺激作用却占主导地位。这一结果表明，在（病理）生理情况下，蛋白C途径的紊乱可能导致血浆中大量scu-PA的灭活，这可能为血栓栓塞与蛋白C途径缺陷之间的关联提供了另一种解释。这项研究表明，血浆中被灭活的scu-PA量主要取决于凝血酶的生成和血栓调节蛋白。我们得出结论，scu-PA诱导的纤维蛋白溶解的抑制似乎受凝血系统激活的调节，这为凝血和纤维蛋白溶解之间提供了联系。
