Ishii-Watabe A, Uchida E, Mizuguchi H, Hayakawa T
Division of Biological Chemistry and Biologicals, National Institute of Health Sciences, Tokyo, Japan.
Life Sci. 2001 Jul 13;69(8):945-60. doi: 10.1016/s0024-3205(01)01189-4.
Plasmin-induced platelet activation is considered to be a cause of reocclusion after thrombolytic therapy with plasminogen activators. However, little is known regarding its mechanism and regulation, particularly with respect to the initial step shape change. We here demonstrate that a Ca2+-independent pathway is involved in plasmin-induced human platelet shape change, and that Rho-kinase plays an important role in this pathway. When the increase in cytosolic Ca2+ was prevented by an intracellular Ca2+ chelator, 5,5'-dimethyl-BAPTA, plasmin-induced platelet shape change was partially inhibited but still occurred. In the presence of 5,5'-dimethyl-BAPTA, a specific Rho-kinase inhibitor, Y-27632, completely inhibited the shape change. Phosphorylation of myosin light chain, a key regulator of platelet shape change, was completely inhibited by Y-27632 in 5,5'-dimethyl-BAPTA-treated platelets. Although plasmin caused tyrosine phosphorylation of the 80 kDa protein during the shape change, it did not seem to have a critical role. cAMP-elevating agents inhibited plasmin-induced shape change in 5,5'-dimethyl-BAPTA- or Y-27632-treated platelets with similar efficiency. These results indicated that plasmin causes platelet shape change by activating Ca2+-dependent and Ca2+-independent-Rho-kinase-dependent pathways, both of which are sensitive to cAMP.
纤溶酶诱导的血小板活化被认为是纤溶酶原激活剂溶栓治疗后再闭塞的一个原因。然而，关于其机制和调节，尤其是初始步骤形状变化方面，人们了解甚少。我们在此证明，一条不依赖Ca2+的途径参与了纤溶酶诱导的人血小板形状变化，并且Rho激酶在该途径中起重要作用。当细胞内Ca2+螯合剂5,5'-二甲基-1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸（5,5'-dimethyl-BAPTA）阻止胞质Ca2+增加时，纤溶酶诱导的血小板形状变化被部分抑制但仍会发生。在存在5,5'-二甲基-1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸的情况下，一种特异性Rho激酶抑制剂Y-27632完全抑制了形状变化。肌球蛋白轻链的磷酸化是血小板形状变化的关键调节因子，在5,5'-二甲基-1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸处理的血小板中，Y-27632完全抑制了其磷酸化。尽管纤溶酶在形状变化过程中导致80 kDa蛋白的酪氨酸磷酸化，但它似乎没有关键作用。cAMP升高剂以相似的效率抑制了5,5'-二甲基-1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸或Y-27632处理的血小板中纤溶酶诱导 的形状变化。这些结果表明，纤溶酶通过激活依赖Ca2+和不依赖Ca2+-Rho激酶的途径导致血小板形状变化，这两条途径均对cAMP敏感。
