呼吸链电子漏在PM2.5诱导肥大细胞脱颗粒中的关键作用与机制研究

The recent increase in the prevalence of allergic diseases attributed to increased exposure to air pollutants , especially particulate matter with a diameter of 2.5μm or less (PM2.5). It was suggested that PM2.5-induced mast cell degranulation was mediated by reactive oxygen species (ROS). It has been shown that the electron leak from mitochondrial respiratory chain is the primary source of ROS generation in cells. Our previous studies have shown that the increasing of electron leak from the respiratory chain may play an important role in the mast cell degranulation induced by PM2.5. Morever, the results of our study indicated that the electron leak may come from complex III of the respiratory chain. However, the precise mechanisms of the electron leak increase in PM2.5 exposed mast cell have yet to be determined. In this project, the mitochondrial dysfunction, such as basal oxygen consumption, glycolysis rates, ATP production, and respiratory capacity induced by PM2.5 will be explored by using cellular bioenergetics analysis, blue native-PAGE, RNAi and other technological methods. And the the changes of the components in complex Ⅲ and the construuction of supermolecular complex will be stuided. Based on the results of the study, the key role that the electron leak from complex Ⅲ of the respiratory chain may contribute to mast cell degranulation will be elucidated. And the molecular mechanisms responsible for the electron leak in response to PM2.5 will be clarified. Thus new strategies for the prevention and treatment of PM2.5-induced allergic diseases could be provided.

PM2.5是过敏疾病发病率迅速增加的重要原因。研究提示PM2.5通过增加细胞活性氧（ROS）刺激肥大细胞（MC）脱颗粒。本课题组进一步发现呼吸链电子漏是PM2.5引起ROS增加的关键因素，且电子漏可能主要发生于呼吸链复合体Ⅲ。为深入探讨PM2.5引起MC脱颗粒的分子机制，本项目拟应用细胞能量代谢实时测定、Blue native-page、RNAi等先进技术手段，系统研究PM2.5作用后MC线粒体功能状态、复合体Ⅲ结构功能及超分子复合物稳定性、构成比的变化等科学问题，以明确呼吸链复合体Ⅲ底物端电子漏在PM2.5刺激MC脱颗粒中的重要地位，阐明复合体Ⅲ电子漏发生的生化及分子机制，为探寻PM2.5诱发过敏疾病的防治新措施提供新思路。

大气污染可加重变态反应性疾病的发生，具体机制尚不明确。本项研究发现低浓度的PM2.5对肥大细胞LAD2可能具有刺激作用，PM2.5浓度增高可对肥大细胞产生毒性作用；研究发现了PM2.5对肥大细胞的作用可能是通过氧化应激使LAD2细胞产生ROS，致使肥大细胞产生炎症因子和细胞因子（β-氨基己糖苷酶、组胺及IL-4）。PM2.5刺激P815肥大细胞后，ROS生成增多，氧化应激水平提高，线粒体功能发生障碍，ATP含量、正常MMP细胞占比及呼吸链复合物III活性均降低，复合物I活性无显著变化。实验结果显示，加入抗氧化剂N-乙酰-L-半胱氨酸(NAC)后，可抵抗PM2.5对肥大细胞的损伤，降低IL-4释放水平，并抑制肥大细胞脱颗粒反应的发生，说明PM2.5影响肥大细胞生物学功能，引发脱颗粒可能与细胞内氧化应激水平相关。研究发现PM2.5诱导角质形成细胞内NF-KB信号通路激活及细胞因子TSLP, IL-33, IL-1β, CCL-20的表达上调，进而可能加剧慢性皮肤炎症反应。以上结果提示呼吸链复合物III是PM2.5诱导肥大细胞产生ROS的重要靶点，线粒体损伤可能是PM2.5引起肥大细胞生物功能改变的重要机制。