呼吸链电子漏在电离辐射诱导肺成纤维细胞分化中的促进作用及OMT的抑制效应研究

Myofibroblasts，mainly differentiated from lung fibroblasts, play a pivotal role in pulmonary fibrosis pathogenesis. Recent experiments have indicated that radiation-induced fibroblast differentiation into myofibroblasts was mediated by reactive oxygen species (ROS). It has been shown that electron leakage from mitochondrial respiratory chain is the primary source of ROS generation in nonphagocytic cells. However, the precise roles of electron leakage in radiation-related lung fibroblasts differentiation have yet to be determined. . Our previous studies have found that oxymatrine (OMT), an alkaloid extracted the dried roots of Sophora flavescens Ait., Sophora subprostrata, could markedly improve the radiation-induced lung fibrosis in mice. It displayed inhibitory effects on lung fibroblast differentiation, which may be correlated with attenuating electron leakage. .In this project, the relations of electron leakage with radiation-induced fibroblast differentiation will be studied in vivo and in vitro using polarography, BN-PAGE, RNAi and other methods. Furthermore, focus on complex III , the changes of radiation-induced mitochondrial function state will be explored. Also, the targets and characteristics of OMT bonding with complexes located in mitochondrial membrane will be assayed.. Based on the results of the study, the role that electron leakage from complex III of respiratory chain may contribute to driving fibroblasts differentiation will be elucidated. And the molecular mechanisms responsible for OMT's inhibitory effects on fibroblast differentiation in response to radiation will be clarified. Thus new strategies for the prevention and treatment of radiation-induced lung fibrosis will be provided from the experiments.

主要由肺成纤维细胞（LF）分化而来的肌成纤维细胞（MFB）在肺纤维化发展过程中发挥着关键作用。新近发现，电离辐射诱导LF分化依赖于细胞活性氧（ROS）水平，而线粒体呼吸链复合体III电子漏程度是决定细胞ROS水平的关键因素。以电子漏为切入点，探讨电离辐射诱导LF分化机制的相关研究，目前尚未见文献报道。我们前期实验提示，氧化苦参碱（OMT）对放射性肺纤维化动物具有肯定的保护作用，该药显著抑制电离辐射诱导LF分化的效应可能与改善电子漏程度有关。本项目拟应用极谱法、蓝绿非变性胶电泳、RNAi等先进技术，结合动物和细胞模型，以复合体III为中心，研究辐射后线粒体功能状态改变，OMT在呼吸链的作用位点等科学问题，旨在阐明复合体III底物端电子漏在电离辐射致细胞转化中的地位及OMT抑制LF分化的分子机制，为探寻放射性肺纤维化的防治措施提供新思路。

主要由肺成纤维细胞（LF）分化而来的肌成纤维细胞（MFB）在肺纤维化发展过程中发挥着关键作用。新近发现，电离辐射诱导LF分化依赖于细胞活性氧（ROS）水平，而线粒体呼吸链电子漏程度是决定细胞ROS水平的关键因素。以电子漏为切入点，探讨电离辐射诱导LF分化机制的相关研究，目前尚未见文献报道。.本课题通过研究证实，电离辐射后肺成纤维细胞发生分化，表达α-SMA，成为肌成纤维细胞。辐射后细胞内ROS显著升高，ATP含量下降、线粒体O2-•含量显著增加。辐射后细胞呼吸功能明显受抑，表现为细胞基础呼吸、最大呼吸、ATP偶联反应、质子漏和电子漏均显著增加。氧化苦参碱和线粒体电子漏抑制剂MitoQ均抑制电离辐射所诱导的α-SMA表达。但MitoQ作用更显著。MitoQ显著抑制辐射后细胞内ROS升高和细胞ATP下降，降低线粒体O2-•含量，改善辐射后细胞呼吸功能，降低线粒体质子漏和电子漏。综上通过本课题证明，电离辐射通过影响呼吸链的电子漏产生过量自由基，引起肺成纤维细胞分化，而改善呼吸链的电子漏，降低线粒体O2-•和细胞内ROS含量，可抑制电离辐射引起的肺成纤维细胞分化。