CPI17、ILK、ZIPK及PKCα、ε在低氧大鼠血管平滑肌中相互作.doc

CPI17、ILK、ZIPK及PKCα、ε在低氧大鼠血管平滑肌中相互作 　 作者：徐竞，明佳，李涛，杨光明，陈玮，刘良明 【摘要】 目的 本实验室前期研究发现蛋白激酶Cα（protein kinase Cα,PKCα）、蛋白激酶Cε（protein kinase Cε,PKCε）可能通过改变CPI17（PKCpotentiated inhibitory protein for protein phosphatase 1 of 17kDa）、ILK（integrinlinked kinase）、ZIPK（zipperinteracting protein kinase）的蛋白表达和活性，来调节失血性休克后血管钙敏感性，但这些分子是否存在直接相互作用，尚不清楚。本实验观察CPI17、ILK、ZIPK与PKCα、ε在低氧血管平滑肌细胞（vascualr smooth muscle cell，VSMC）中的相互作用。方法 采用低氧培养的大鼠VSMC，运用免疫共沉淀技术，观察PKCα、ε与下游分子CPI17、ILK、ZIPK，以及CPI17、ILK、ZIPK之间的相互作用。结果 （1）在PKCα抗体捕获的免疫沉淀中，ILK、ZIPK抗体，尤其是ILK抗体可杂交出明显蛋白条带，CPI17抗体不能杂交出蛋白条带；（2）在PKCε抗体捕获的免疫沉淀中，ILK、ZIPK抗体均可杂交出明显蛋白条带，CPI17抗体不能杂交出蛋白条带；（3）在CPI17抗体捕获的免疫沉淀中，ILK、ZIPK抗体可杂交出明显蛋白条带；在ILK抗体捕获的免疫沉淀中，ZIPK抗体可杂交出明显蛋白条带。结论 PKCα可直接作用于ILK，PKCε可直接作用于ILK和ZIPK，ILK、ZIPK可直接作用于CPI17，ILK可直接作用于ZIPK。上述分子相互作用，形成网络，共同调节大鼠失血性休克后的血管钙敏感性。 【关键词】 失血性休克；钙敏感性；蛋白激酶Cα；蛋白激酶Cε；CPI17；ILK；ZIPK Abstract： Objective Our previous research showed that protein kinase Cα(PKCα), protein kinase Cε(PKCε) regulated vascular calcium sensitivity through PKCpotentiated inhibitory protein for protein phosphatase 1 of 17kDa(CPI17),integrinlinked kinase(ILK) and zipperinteracting protein kinase(ZIPK),while their interaction is not clear,so this research is to observe the interaction between CPI17,ILK,ZIPK and PKCα,PKCε in vascular smooth muscle cell(VSMC) after hypoxia.Methods The hypoxic VSMC of rats were adopted to observe the interaction between PKCα,PKCε and the downstream molecular CPI17,ILK,ZIPK,and the interaction among CPI17,ILK,ZIPK through coimmunoprecipitation.Results (1)ILK and ZIPK,especially ILK,were present in immunoprecipitates of PKCα,while CPI17 was not observed.(2)ILK and ZIPK were both obviously present in immunoprecipitates of PKCε,while CPI17 was not either.(3)ILK and ZIPK were both obviously present in immunoprecipitates of CPI17,and ZIPK was present in immunoprecipitates of ILK.Conclusion PKCα interacts with ILK.PKC ε interacts with ILK and ZIPK.ILK and ZIPK interacts with CPI17,ILK interacts with