α-突触核蛋白上调细胞色素C氧化酶亚基2引起多巴胺能神经元损伤的机制

The misfolded α-Synuclein(α-Syn) transferring from cells to cells as prion protein is an important pathogenesis of Parkinson’s disease(PD). However, the mechanism of how α-Syn serves as “seeds” to injure the dopaminergic neuron is still unclear. The results of gene microarray and western blot showed that the expression of cytochrome c oxidase subunit 2(MT-CO2) had increased significantly in cells stimulated by α-Syn for 24h. Furthermore, the decreased mitochondrial membrane potential (MMP) and enhanced ROS in cells treated by α-Syn had been reversed by inhibiting MT-CO2 gene expression with siRNA. Accumulating researches have suggested that the aberrant expression and posttranslational modification of MT-CO2 contribute to the abnormal vitality of mitochondrial complex IV and dysfunction of mitochondria. Thus we supposed that α-Syn led to abnormal vitality of mitochondrial complex IV, mitochondrial dysfunction and apoptosis of dopaminergic neurons by regulating the expression and posttranslational modification of MT-CO2. Based on the PD cellular and animal models induced by α-Syn oligomer, we will detect the expression and phosphorylated modification of MT-CO2 by western blot and uncover the conformation and vitality of mitochondrial complex IV by CD spectroscopy and spectrophotometry, respectively. The mitochondrial function will be determined by flow cytometry. We hope to reveal the initial step of the process by which α-Syn injures the dopaminergic neurons and provide new therapeutic targets of PD.

异常折叠的α-突触核蛋白（α-Syn）在细胞间如朊蛋白样传播是帕金森病（PD）的重要发病假说，但其如何作为“种子”触发多巴胺能神经元损伤的机制尚未阐明。预实验基因芯片和WB检测发现α-Syn刺激细胞24h后细胞色素C氧化酶亚基2（MT-CO2）表达显著增高，用siRNA下调MT-CO2可逆转α-Syn导致的线粒体膜电位下降和ROS含量增高。MT-CO2异常表达及翻译后修饰可使线粒体呼吸链复合体IV活性改变、线粒体功能障碍。故我们假设：α-Syn通过调控MT-CO2表达及翻译后修饰引起复合体IV活力异常，导致线粒体功能紊乱，加速多巴胺能神经元死亡。本课题拟采用α-Syn诱导的PD细胞模型和大鼠模型，WB法检测MT-CO2表达和翻译后修饰，圆二色光谱法和分光光度法检测细胞色素C氧化酶构象及活性，流式细胞仪检测线粒体功能，从而明确α-Syn引起多巴胺神经元损伤的早期机制，为PD提供新的防治靶点。

α-突触核蛋白（α-Syn）的异常聚集是帕金森病（PD）的核心发病机制，异常折叠的α-Syn在细胞间如朊蛋白样传播是PD的重要发病假说，但其如何作为“种子”触发多巴胺能神经元损伤的机制尚未阐明。我们前期通过基因芯片和WB检测发现α-Syn刺激细胞24h后细胞色素C氧化酶亚基2（MT-CO2）表达显著.增高，用siRNA下调MT-CO2可逆转α-Syn导致的线粒体膜电位下降和ROS含量增高。MT-CO2异常表达及翻译后修饰可使线粒体呼吸链复合体IV活性改变、线粒体功能障碍。故我们假设：α-Syn通过调控MT-CO2表达及翻译后修饰引起复合体IV活力异常，导致线粒体功能紊乱，加速多巴胺能神经元死亡。本项目采用α-Syn诱导的PD细胞模型和大鼠模型，WB法检测MT-CO2表达和翻译后修饰，圆二色光谱法和分光光度法检测细胞色素C氧化酶构象及活性，流式细胞仪检测线粒体功能。我们的结果证实，外源性的α-Syn通过调控MT-CO2上调，引起细胞内线粒体功能障碍，从而影响细胞色素c合成及其代谢循环障碍，进一步加剧.线粒体损伤，介导凋亡过程。同时外源性α-Syn能通过调控MT-CO2介导线粒体内和胞浆内线粒体相关的前凋亡因子和抗凋亡因子的表达异常，导致细胞凋亡的调节过程紊乱。本研究为α-Syn相关的神经退行性疾病提供了新的治疗靶点。