Gene Environment Interactions in Parkinson's Disease

帕金森病的基因环境相互作用

• 批准号: 7894952
• 负责人: KIM TIEU
• 金额: $ 19.13万
• 依托单位: UNIVERSITY OF ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-16 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7894952
• 关键词: 1-Methyl-4-phenylpyridinium; Attenuated; Biological Assay; Cell Death; Cell Survival; Cells; Cessation of life; Chemicals; Chimeric Proteins; Data; Dominant-Negative Mutation; Dopamine; Dose; Electron Transport; Experimental Models; Exposure to; Gene Mutation; Genetic; Goals; Herbicides; High Pressure Liquid Chromatography; Human; Immunoblotting; Induced Mutation; Intervention; Lead; Measures; Mediating; Mitochondria; Molecular; Mutation; Nerve Degeneration; Neurodegenerative Disorders; Neurotoxins; PTEN-induced putative kinase; Paraquat; Parkinson Disease; Pathogenesis; Pathway interactions; Patients; Phosphotransferases; Play; Polarography; Predisposition; Process; Production; Proteins; Reporting; Research; Role; Shapes; Small Interfering RNA; System; Testing; Time; Toxic Environmental Substances; Toxic effect; Translating; base; design; functional outcomes; gene environment interaction; genetic manipulation; inhibitor/antagonist; insight; mitochondrial dysfunction; mutant; neuroprotection; neurotoxicity; novel; overexpression; prevent; public health relevance; research study; response; small molecule; stable cell line; toxicant; vector; vector control

DESCRIPTION (provided by applicant): Our long term goal is to study the mechanism of neurodegeneration induced by environmental neurotoxicants. This proposal is submitted to investigate gene-environment interactions in the pathogenesis of Parkinson's disease mediated through the mitochondrial pathway. Mitochondrial dysfunction has been proposed as a major mechanism of neurodegeneration in PD for years, but direct evidence in humans was inadequate until the recent discoveries of mutations in PTEN-induced putative kinase 1 (PINK1), which encodes a mitochondrial kinase. Although PINK1 mutations are associated with autosomal recessive PD, interestingly, an increasing number of PD patients carrying single heterozygous mutations have been reported. These observations suggest that a heterozygous PINK1 mutation may act as a susceptibility factor that interplays with environmental insults. To determine the interactions between PINK1 mutations and environmental toxicants, we have created stable cell lines with inducible over-expression of various PINK1 mutants. Based on our preliminary results, we hypothesize that PINK1 mutations increase cell susceptibility to environmental toxicants such as paraquat (PQ) through a novel mechanism: mitochondrial fragmentation via the mitochondrial fission / fusion machinery. In the first specific aim, we will characterize neurotoxicity induced by PQ and MPP+ (two toxicants representing different mechanisms of toxicity) through the mitochondrial fission and fusion pathway in N27 cells with a PINK1 mutation (L347P) and empty vector control, as well as in N27 cells with PINK1 knockdown mediated by siRNA. We will assess cell viability, functional outcomes (ATP production, dopamine release and electron transport chain activity), as well as mitochondrial fragmentation (size/shape) and alterations in mitochondrial fission and fusion proteins. In specific aim 2, we will perform neuroprotective experiments against PQ and MPP+ toxicity in mutant PINK1 cells by targeting the mitochondrial fission and fusion pathway, through genetic manipulations and a small molecule. We will transfect cells with relevant constructs and use a chemical inhibitor to attenuate mitochondrial fragmentation. The neuroprotective effects of these two strategies against MPP+ and PQ toxicity will be determined using cell viability and functional assays as described in specific aim 1. PUBLIC HEALTH RELEVANCE This research may provide insights into the complexity of gene-environment interactions in the cause of cell death as seen in Parkinson's disease by unraveling a still unrecognized molecular pathway. Furthermore, this novel mechanism may offer an additional avenue to develop neuroprotective therapy for patients with Parkinson's disease.

描述（由申请人提供）：我们的长期目标是研究环境神经毒物诱导的神经变性机制。本研究旨在探讨线粒体途径介导的帕金森病发病机制中基因与环境的相互作用。多年来，线粒体功能障碍一直被认为是帕金森病神经退行性变的主要机制，但在人类身上的直接证据不足，直到最近发现pten诱导的推定激酶1 （PINK1）突变，PINK1编码线粒体激酶。尽管PINK1突变与常染色体隐性PD相关，但有趣的是，越来越多的PD患者携带单杂合突变。这些观察结果表明，杂合子PINK1突变可能是与环境损害相互作用的易感因子。为了确定PINK1突变与环境毒物之间的相互作用，我们创建了稳定的细胞系，诱导各种PINK1突变体过表达。基于我们的初步结果，我们假设PINK1突变通过一种新机制增加了细胞对环境毒物如百草枯（PQ）的易感性：通过线粒体裂变/融合机制的线粒体断裂。在第一个特定目标中，我们将在PINK1突变（L347P）和空载体对照的N27细胞中，以及由siRNA介导的PINK1敲低的N27细胞中，通过线粒体裂变和融合途径，表征PQ和MPP+（两种代表不同毒性机制的毒物）诱导的神经毒性。我们将评估细胞活力，功能结果（ATP产生，多巴胺释放和电子传递链活性），以及线粒体碎片（大小/形状）和线粒体裂变和融合蛋白的变化。在特定目标2中，我们将通过遗传操作和小分子，针对线粒体裂变和融合途径，对突变PINK1细胞进行PQ和MPP+毒性的神经保护实验。我们将用相关构建体转染细胞，并使用化学抑制剂来减弱线粒体断裂。这两种策略对MPP+和PQ毒性的神经保护作用将通过细胞活力和功能测定来确定，如具体目标1所述。这项研究可能通过揭示一个尚未被识别的分子途径，为帕金森病中细胞死亡原因中基因-环境相互作用的复杂性提供见解。此外，这一新机制可能为开发帕金森病患者的神经保护疗法提供额外的途径。