OXIDATIVE STRESS IN PARKINSON'S DISEASE

特发性帕金森病（PD）是一种影响至少100万美国人的主要神经退行性疾病，PD的细胞病因尚不确定。本研究将进一步探讨线粒体电子传递链（ETC）功能缺陷是帕金森病细胞过早死亡的主要原因这一中心假设，并将解决以下四个具体目标：1)定义线粒体过渡孔功能的病理生理学，以及帕金森病中膜电位和细胞内钙信号的调节如何改变；2)确定Bcl蛋白在PD杂交体中的调节机制，以及转染Bcl过表达载体是否会改变线粒体功能，提高生存率；3)进一步明确PD中MAPKinase信号通路与NFkappaBeta转录因子的相互作用；4)表征从人类死后PD脑中分离的线粒体过渡孔复合物，并将其与对照脑分离的线粒体过渡孔复合物进行功能比较。该项目将利用最先进的细胞内离子成像技术，RT-PCR技术，基因转染策略，并将开发无细胞系统来检查几个相互关联的假设。在所有这些实验室实验的背后是一种治疗的必要性，这将在细胞和无细胞模型中进行探索。由于本应用中提出的新数据支持PD患者系统性氧化应激增加的假设，因此在已建立的细胞模型中探索这些事件更具吸引力。该提案还将比较PD杂交与暴露于慢性鱼藤酮治疗的SY5Y细胞的发现，这是一种基于细胞的复合物I丢失的药理学模型。最终，本研究的结果将确定遗传获得性线粒体ETC功能障碍作为散发性PD的病因因素的核心重要性。评估神经保护疗法的范例也将被开发出来，以允许有针对性的方法来纠正细胞中氧化应激增加的后果。

Idiopathic Parkinson's Disease (PD) is a major neurodegenerative disease affecting at least 1 million Americans, and the cellular cause of PD is not yet known with certainty. This proposal will explore further the central hypothesis that defects in mitochondrial electron transport chain (ETC) function are a major contributor to premature cell death in PD and will address four Specific Aims 1) define the pathophysiology of mitochondrial transition pore function, and how regulation of membrane potential and intracellular calcium signaling are altered in PD; 2) determine mechanisms of Bcl protein regulation in PD cybrids, and whether transfection with Bcl-overexpression vectors alters mitochondrial function and improves survival; 3) further define the interactions among MAPKinase signaling pathways and NFkappaBeta transcription factor in PD; and 4) characterize mitochondrial transition pore complexes isolated from human postmortem PD brain and compare their function to those isolated from control brain. This project will make use of state-of-the- art intracellular ion imaging technology, RT-PCR techniques, gene transfection strategies, and will develop cell-free systems to examine several inter-related hypotheses. Behind all of these laboratory experiments is a therapeutic imperative, which will be explored in cell and cell-free models. Because new data presented in this application supports the hypothesis of systemically increased oxidative stress in PD patients, exploring these events in an established cell model is even more compelling. This proposal will also compare findings in PD cybrids with those in SY5Y cells exposed to chronic rotenone treatment, a pharmacological cell-based model of complex I loss. Ultimately, the results from this proposal will establish the central importance of genetically acquired mitochondrial ETC dysfunction as an etiologic factor in sporadic PD. Paradigms for evaluating neuroprotective therapies will also be developed to allow targeted approaches to correcting consequences of increased oxidative stress in cells.