整合素β1在百草枯诱导肺纤维化去细胞支架促成纤维细胞活化中的作用与机制研究

Irreversible pulmonary fibrosis is the main cause of death in the late stage of paraquat poisoning, however, the mechanism remains unclear. The research shows that fibroblasts is one of the main sources of myofibroblasts in the fibrous lesions. Further study on the regulation and mechanism of phenotypic transformation of lung fibroblasts is of great significance to delay or even reverse the pulmonary fibrosis induced by paraquat poisoning. The research shows that the change of microenvironment of extracellular matrix (ECM) can lead to the change of the stress factors, which are the important inducing factors of the differentiation of fibroblasts into myofibroblasts. In this study, pulmonary fibrosis will be induced by paraquat in rats, and lung acellular biological scaffold will be prepared, then the fibroblasts will be cultured with 3D lung acellular scaffold. To observe fibroblast phenotype transformation, the markers of mesenchy (a-SMA、vimentin)and the expression and phosphorylation levels of mechanochemical pathway related protein mediated by integrin beta 1 will be detected by immunofluorescence staining and WB. Moreover, the mRNA will be detected by RT-PCR. To elucidate the effect of ECM of pulmonary fibrosis induced by paraquat on fibroblast activation and to clarify its possible mechanism，will lay the experimental foundation for clinical prevention and treatment of pulmonary fibrosis.

百草枯中毒患者后期出现不可逆性肺纤维化，是主要致死原因，机制不明。研究表明成纤维细胞是纤维病灶肌成纤维细胞产生的主要来源之一，深入研究肺成纤维细胞表型转化调控及机制对延缓乃至逆转百草枯中毒诱导的肺纤维化来说意义重大。研究发现细胞外基质（ECM）微环境结构变化可致细胞受力因素改变，后者是成纤维细胞向肌成纤维细胞分化的重要诱导因素。本研究通过百草枯诱导大鼠肺纤维化，制备纤维化肺去细胞支架，将成纤维细胞与3D纤维化肺去细胞支架共同培养，观察成纤维细胞表型转化情况。免疫荧光染色及WB检测间质细胞标志物（a-SMA、vimentin）及整合素β1介导的机械化学通路相关蛋白表达及其磷酸化水平；RT-PCR检测间质细胞标志物及该机械化学通路相关mRNA表达。阐明百草枯诱导的肺纤维化ECM对成纤维细胞活化作用与可能机制，为后续从整合素β1介导的机械化学通路寻找新的肺纤维化临床治疗靶点奠定实验基础。

百草枯中毒患者后期出现不可逆性肺纤维化，是主要致死原因，机制不明。研究表明成纤维细胞是纤维病灶肌成纤维细胞产生的主要来源之一，深入研究肺成纤维细胞表型转化调控及机制对延缓乃至逆转百草枯中毒诱导的肺纤维化来说意义重大。本研究通过百草枯诱导大鼠肺纤维化，制备纤维化肺去细胞支架，将成纤维细胞与3D纤维化肺去细胞支架共同培养，观察成纤维细胞表型转化情况。免疫荧光染色及WB检测间质细胞标志物（a-SMA、vimentin）及整合素β1介导的机械化学通路相关蛋白表达及其磷酸化水平；RT-PCR检测间质细胞标志物及该机械化学通路相关mRNA表达。研究发现纤维化细胞外基质（ECM）微环境结构变化可促进成纤维细胞向肌成纤维细胞分化，其机制可能是通过激活整合素β1介导的FAK-Scr-p130Cas机械化学信号传导通路及其下游FAK- Ras-MAPK通路。通过阐明百草枯诱导的肺纤维化ECM对成纤维细胞活化作用与可能机制，为后续从整合素β1介导的机械化学通路寻找新的肺纤维化临床治疗靶点奠定实验基础。