PKCs-p66Shc信号通路介导的气道上皮细胞线粒体损伤在COPD发病中的作用机制研究

我们前期研究发现，与线粒体相关的衔接蛋白p66Shc可能通过影响气道上皮细胞线粒体功能，参与慢性阻塞性肺疾病（COPD）发病过程。为进一步阐明p66Shc及其上游调节分子PKCs在COPD发病中的作用，本项目拟首先用香烟烟雾提取物刺激原代培养的大鼠气道上皮细胞，采用基因转染、RT-PCR、Wb、免疫共沉淀、GST Pull-down等技术，明确p66Shc对气道上皮细胞线粒体损伤的分子机制以及与线粒体膜蛋白FKBP38相互作用对线粒体损伤的影响；其次通过基因沉默和PKCs抑制剂干预，确定p66Shc对气道上皮细胞线粒体损伤的调控依赖PKCs信号分子；最后香烟烟雾刺激建立COPD大鼠模型，同步应用PKCs抑制剂下调PKCs活性，明确PKCs-p66Shc信号通路在COPD大鼠气道上皮细胞线粒体损伤中的作用。本课题的实施与完成，为从线粒体角度探索COPD的发病机制及靶向治疗提供重要科学依据。

Our previous study has demonstrated that p66Shc adapter protein related to mitochondria may be involved in the progression of COPD, through modulating mitochondrial function of airway epithelial cell. In order to further elucidate the role of PKCs-p66Shc signaling pathway in the pathogenesis of COPD, this project has been designed. First, primary cultured airway epithelial cell will be stimulated with cigarette smoke extract. The role of p66Shc and its interaction with FKBP38 in the course of mitochondrial damage of airway epithelial cell will be studied using gene transfection, RT-PCR, Wb, co-immunoprecipitation, GST Pull-down and so on. Second, the regulative effect of PKCs on p66Shc will be explored during the mitochondrial damage of airway epithelia cell by gene silencing and PKCs inhibitor intervention. Last, a rat model of COPD will be induced by cigarette smoke, and the rats will be treated with PKCs inhibitors. The effect of PKCs-p66Shc signaling pathway on mitochondrial damage of airway epithelium will be studied in those rats. The purpose of this project is to provide scientific basis for further studying the role of mitochondria in the pathogenesis of COPD and targeted therapy for COPD.