Calpain functions in a caspase-independent manner to promote apoptosis-like events during platelet activation
Calpain functions in a caspase-independent manner to promote apoptosis-like events during platelet activation
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DOI：
10.1182/blood.v94.5.1683.417k37_1683_1692
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发表时间：
1999-09-01
期刊：
影响因子：
20.3
通讯作者：
Green, DR
中科院分区：
文献类型：
--
作者：
Wolf, BB;Goldstein, JC;Green, DR
细胞凋亡和血小板活化具有共同的形态学和生化特征。由于半胱天冬酶是细胞凋亡的重要介质，我们研究了血小板是否含有这些蛋白酶，并在血小板活化过程中使用它们。十二烷基硫酸钠-聚丙烯酰胺凝胶电泳和Western印迹显示，人血小板含有caspase-9、caspase-3、caspase激活剂APAF-1和细胞色素c。细胞色素c和dATP处理后，血小板胞质提取物重演凋亡事件，包括半胱天冬酶原-9和半胱天冬酶原-3的顺序激活和随后的半胱天冬酶底物的蛋白水解。钙离子载体刺激的血小板也重演了凋亡事件，包括细胞收缩、质膜微泡形成、磷脂酰丝氨酸外化和蛋白酶原-9、蛋白酶原-3、凝溶胶蛋白和蛋白激酶C-δ的蛋白水解。然而，引人注目的是，这些事件的发生没有半胱天冬酶激活或释放线粒体细胞色素c，这表明一个noncaspase蛋白酶的作用。支持这一点，抑制钙依赖性蛋白酶，钙蛋白酶，防止半胱天冬酶蛋白水解，'凋亡'底物裂解，和血小板微泡形成。在体外，纯化的钙蛋白酶裂解重组半胱天冬酶原-9和半胱天冬酶原-3，而不激活半胱天冬酶，确认抑制剂的研究。这些数据暗示钙蛋白酶作为半胱天冬酶的潜在调节剂，并表明钙蛋白酶，而不是半胱天冬酶，促进血小板活化过程中的血小板减少样事件。(C)1999年，美国血液学会。
Apoptosis and platelet activation share common morphological and biochemical features. Because caspases are essential mediators of apoptosis, we examined whether platelets contain these proteinases and use them during platelet activation. Human platelets contained caspase-9, caspase-3, and the caspase activators APAF-1 and cytochrome c as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting. Upon treatment with cytochrome c and dATP, platelet cytoplasmic extracts recapitulated apoptotic events, including sequential activation of procaspase-9 and procaspase-3 and subsequent proteolysis of caspase substrates. Calcium ionophore-stimulated platelets also recapitulated apoptotic events, including cell shrinkage, plasma membrane microvesiculation, phosphatidyl serine externalization, and proteolysis of procaspase-9, procaspase-3, gelsolin, and protein kinase C-delta. Strikingly, however, these events occurred without caspase activation or release of mitochondrial cytochrome c, suggesting a role for a noncaspase proteinase. Supporting this, inhibition of the calcium-dependent proteinase, calpain, prevented caspase proteolysis, 'apoptotic' substrate cleavage, and platelet microvesiculation. In vitro, purified calpain cleaved recombinant procaspase-9 and procaspase-3 without activating either caspase, confirming the inhibitor studies. These data implicate calpain as a potential regulator of caspases and suggest that calpain, not caspases, promotes apoptosis-like events during platelet activation. (C) 1999 by The American Society of Hematology.