PATHOGENIC MECHANISMS OF NEURONAL CELL DEATH IN PD

这是四个项目小组中的第二个项目，该小组正在研究与帕金森氏病(PD)相关的线粒体功能和线粒体缺陷的多个方面。帕金森氏症是一种主要的神经退行性疾病，困扰着至少100万美国人。项目2侧重于导致帕金森病患者神经元进行性神经细胞死亡的致病机制。这项研究将使用Cybrid模型神经元和原代培养的神经元进行。Cybrid模型神经元是由缺乏线粒体DNA(MtDNA)和患者来源的线粒体的克隆人神经细胞系构建的。该项目将检验一个单一的中心假设：线粒体参与氧化应激、钙稳态和调节信号与帕金森病衍生胞体(PD胞体)脆弱性增加有关。线粒体在钙信号网络的维持中起着重要的作用，在钙信号网络的维持和作用中发挥着重要作用，尤其是在神经元中。这反映在线粒体和包括内质网在内的网络中的其他细胞元素之间特定的形态和功能关系上。钙信号网络在协调胞吐分泌、细胞代谢、基因表达和细胞器间通讯等方面起着至关重要的作用。线粒体还起到抵御兴奋性毒性事件的细胞毒性的作用，并被认为是包括细胞色素c释放在内的凋亡信号的关键来源。实验将通过以下几个目标来研究帕金森氏病患者胞质对细胞死亡的易感性变化与钙信号网络中线粒体功能的变化之间的关系：1)ETC复合体I活性、钙稳态缺陷和胞体对细胞死亡的易感性之间存在什么定量关系？神经保护是否包括恢复正常的钙稳态？2)长期抑制ETC复合体I活性的影响与急性抑制的影响是否不同？关于钙的动态平衡？细胞死亡？氧自由基的产生？线粒体的形态和细胞器间的相互作用？3)PD囊体中线粒体与其他细胞器的形态和功能关系是如何受到影响的？4)慢性鱼藤酮治疗后PD囊体和原代神经元中线粒体的运动是如何改变的？5)PD囊体中的线粒体缺陷如何影响溶酶体的活性、蛋白泛素化、降解和类似路易小体的包涵体的形成？所获得的结果将在精心控制的模型系统中揭示这种重要疾病的可能发病机制的新方面，并可能揭示治疗靶点。

This is the second Project in a four Project group which is examining multiple aspects of mitochondrial functioning and mitochondrial defects associated with Parkinson's Disease (PD), a major neurodegenerative disease afflicting at least 1 million Americans. Project 2 is focused upon the pathogenetic mechanisms leading to the progressive neuronal cell death of the neurons affected in PD. The study will be conducted using cybrid model neurons and primary neurons in culture. Cybrid model neurons are constructed from a clonal human neural cell line lacking mitochondrial DNA (mtDNA) and patient-derived mitochondria. The project will examine a single central hypotheses: Mitochondrial involvement in oxidative stress, Ca2+ homeostasis and regulatory signaling is linked to increased vulnerability in Parkinson's disease- derived cybrids (PD cybrids). Mitochondria play an important role in the maintenance play an important role in the maintenance and efficacy of the calcium signaling network, especially in neurons. This is reflected in specific morphological and functional relationships between mitochondria and other cellular elements in the network, including the endoplasmic reticulum. The calcium signaling network plays a critical role coordinating exocytotic secretion, cellular metabolism, gene expression and inter-organelle communication. Mitochondria also function as a defense against cytotoxic of excitotoxic events and are implicated as a critical source of apoptotic signals including release of cytochrome c. Experiments will investigate the relationship between altered vulnerability to cell death in the cybrids derived from patients with Parkinson's disease and altered mitochondrial functioning in the calcium signaling network by addressing several aims: 1) What quantitative relationship exists between defects in ETC complex I activity, Ca2+ homeostasis and vulnerability to cell death in cybrids? Does neuroprotection involve restoration in normal Ca2+ homeostasis? 2) Does the impact of long-term inhibition of ETC complex I activity differ from that of acute inhibition? With respect to Ca2+ homeostasis? Cell death? Generation of oxyradicals? Mitochondrial morphology and inter- organelle interactions? 3) How are morphological and functional relationships of mitochondrial of mitochondria to other organelles affected in the PD cybrids? 4) How is mitochondrial movement altered in PD cybrids and in primary neurons after chronic rotenone treatment? 5) How do the mitochondrial defects in PD cybrids affect lysosomal activity, protein ubiquitination, degradation and the formation of inclusions that resemble Lewy bodies? The results gained will reveal novel aspects of the probable pathogenesis of this important disease in a carefully controlled model systems and may reveal therapeutic targets.