摘要
蛋白激酶C(Protein Kinase C,PKC)在血小板信号转导和粘附、聚集、释放等多种生理功能的发挥中均起到重要的作用。目前报道,PKC通过激活去整合素金属蛋白酶(ADAM)10或ADAM17诱导血小板膜表面糖蛋白受体发生酶切,对血小板活化后产生负调控作用。那么PKC活化是否可诱导血小板凋亡进而负调控血小板功能,目前还不清楚。本研究目的是探讨血小板内PKC活化对血小板凋亡的影响。取健康志愿者外周静脉血并分离血小板。选择PKC特异性激活剂和抑制剂,并设置不同的时间梯度和浓度梯度,与洗涤血小板共同孵育,应用流式细胞术和Western blot等技术,检测PKC激活剂和抑制剂对血小板线粒体膜电位(△ψm)、半胱氨酸的天冬氨酸蛋白水解酶(Caspase)和磷脂酰丝氨酸(PS)的影响。实验结果表明,PKC激活剂呈时间依赖性诱导血小板发生△ψm去极化;PKC激活剂呈浓度依赖性地诱导血小板内caspase-3活化裂解,而PKC抑制剂则不能诱导血小板发生△ψm去极化和PS暴露的变化。结论:血小板内PKC活化后,可以通过影响线粒体功能和激活Caspase-3而诱导血小板凋亡,提示PKC可能参与调控活化血小板的凋亡过程,本研究结果对探明血小板活化的负调控功能及PKC活化相关疾病的发病机理具有意义。
Platelet apoptosis elucidated by either physical or chemical compound or platelet storage occurs wildly, which might play important roles in controlling the numbers and functions of circulated platelets, or in the development of some platelet-related diseases. However, up to now, a little is known about the regulatory mechanisms of platelet apoptosis. Protein kinase C (PKC) is highly expressed in platelets and plays central roles in regulating platelet functions. Although there is evidence indicating that PKC is involved in the regulation of apoptosis of nucleated cells, it is still unclear whether PKC plays a role in platelet apoptosis. The aim of this study was to investigate the role of PKC in platelet apoptosis. The effects of PKC on mitochondrial membrane potential (AtOm), phosphatidylserine (PS) exposure, and caspase-3 activation of platelets were analyzed by flow cytometry and Western blot. The results showed that the A,m depolarization in platelets was induced by PKC activator in time-dependent manner, and the caspase-3 activation in platelets was induced by PKC in concentration-dependent manne. However, the platelets incubated with PKC inhibitor did not results in △ψm depolarization and PS exposure. It is concluded that the PKC activation induces platelet apoptosis through influencing the mitochomdrial functions and activiating caspase 3. The finds suggest a novel mechanism for PKC in regulating platelet numbers and functions, which has important pathophysiological implications for thrombosis and hemostasis.
出处
《中国实验血液学杂志》
CAS
CSCD
北大核心
2013年第5期1207-1210,共4页
Journal of Experimental Hematology
基金
国家自然科学基金(编号81130008、30971067、30770795)
江苏省临床医学中心(编号ZX201102)
江苏省科技厅生命健康科技专项资金(编号BL2012005)资助