Mechanisms of age-dependent nigral neuron loss in PINK1 knockout rats

描述（由申请人提供）：帕金森病（PD）是老年人神经变性、残疾和过早死亡的常见原因。黑质多巴胺能神经元的丧失是帕金森病的主要神经病理学标志。目前还没有证实的治疗方法可以减缓PD患者的进行性神经细胞丢失，这导致临床症状的严重程度增加。最近将Parkin、DJ-1和PINK1等基因突变与隐性遗传形式的PD联系起来，为发现致病机制以及基于与人类帕金森病相关的生理机制开发和测试神经保护疗法的动物模型提供了新的机会。十多年来，我们一直在使用基因敲除（KO）小鼠来追求这一策略。由于未知的原因，Parkin KO， DJ-1 KO和PINK1 KO小鼠不会复制在这些基因中发生功能丧失突变的人类中发生的黑细胞损失。我们和其他人在KO小鼠、果蝇和培养细胞中发现了线粒体功能障碍（呼吸链缺陷），这表明这是这些基因功能缺失突变导致黑质细胞损失的常见和可能的早期机制事件。然而，在Parkin KO、DJ-1 KO或PINK1 KO小鼠中，缺乏黑质细胞损失排除了将黑质细胞损失作为直接测试这种和其他潜在关键致病机制的结果测量。最近发展的PINK1 KO大鼠表现出年龄依赖性的神经细胞损失，从6个月开始，到7个月时达到50%。PINK1 KO大鼠中年龄依赖性的神经神经元丢失使得首次在哺乳动物大脑中直接测试主要的候选致病机制，如线粒体损伤和帕金森介导的线粒体自噬减少，从而再现PD的中枢神经病理特征。我们建议使用PINK1 KO大鼠来实现以下相关的特定目标，这些目标之所以被选择，是因为它们对治疗发展的重要性：1)确定PINK1缺乏导致大鼠黑质细胞损失的机制；2)表征帕金蛋白的抑制作用

DESCRIPTION (provided by applicant): Parkinson's disease (PD) is a frequent cause of neurodegeneration, disability and premature mortality in older adults. Loss of dopaminergic neurons in the substantia nigra is the primary neuropathological hallmark of PD. There are currently no treatments proven to slow down the progressive nigral cell loss in PD, which causes increasing severity of the clinical symptoms. The recent linking of human mutations in genes such as Parkin, DJ-1, and PINK1 to recessively inherited forms of PD provides new opportunities to discover pathogenic mechanisms and to develop and test neuroprotective therapies in animal models with nigral cell loss based on mechanisms physiologically relevant to human parkinsonism. We have pursued this strategy for over a decade using knockout (KO) mice. For unknown reasons, Parkin KO, DJ-1 KO and PINK1 KO mice do not reproduce the nigral cell loss that occurs in humans bearing loss-of-function mutations in these genes. We and others identified mitochondrial dysfunction (respiratory chain defects) in KO mice, Drosophila and cultured cells, suggesting that this is a common and perhaps early event in the mechanism by which loss-of-function mutations in these genes cause nigral cell loss. However, the lack of nigral cell loss precludes using nigral cell loss as an outcome measure to directly test this and other potential key pathogenic mechanisms in Parkin KO, DJ-1 KO or PINK1 KO mice. Recently developed PINK1 KO rats show age-dependent nigral cell loss beginning at age 6 months and reaching 50% by age 7 months. The age-dependent nigral neuron loss in PINK1 KO rats makes it possible for the first time to test the leading candidate pathogenic mechanisms such as mitochondrial impairment and diminished Parkin-mediated mitochondrial autophagy directly in a mammalian brain that reproduces this central neuropathological feature of PD. We propose to use PINK1 KO rats to achieve the following interrelated specific aims which have been chosen because of their significance for therapeutic development: 1) To identify mechanisms by which PINK1 deficiency leads to nigral cell loss in the rat, 2) To characterize Parkin as an inhibitor of age-dependent nigral cell loss in PINK1 KO rats and 3) To stimulate the mitochondrial respiratory chain as a potential therapy for PD. We expect to contribute a systematic characterization of PINK1 KO rats as an apt test bed for therapeutic development by testing the principal hypothesis that PINK1 deficiency diminishes Parkin-mediated degradation of impaired mitochondria and the second main hypothesis that respiratory chain stimulation via dietary intake of methylene blue is neuroprotective in PINK1 KO rats. We have previously shown that methylene blue prevents neurodegeneration in the rotenone rat PD model. The use of this novel rat model of nigral cell loss is innovative and the proposal will significantly impact the understanding of PD by shifting emphasis to disease mechanisms present in PD brain tissue selected for their strong therapeutic potential. The knowledge gained from our study will be broadly applicable to diseases associated with mitochondrial dysfunction and neurodegeneration.