Mitochondrial mechanisms and vulnerability to alpha-synuclein toxicity

线粒体机制和α-突触核蛋白毒性的脆弱性

基本信息

批准号：
9385532
负责人：
DAVID K. SIMON
金额：
$ 21.63万
依托单位：
BETH ISRAEL DEACONESS MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-07-01 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9385532
关键词：
Accounting Age Behavioral Bioenergetics Biogenesis Brain Cell Line Cells Cessation of life Complex Corpus striatum structure DNA-Directed DNA Polymerase Data Defect Dopamine Functional disorder Future Gene Dosage Gene Expression Goals Gray unit of radiation dose Hair Human Impairment Incidence Inherited Injection of therapeutic agent Lead Mediating Mitochondria Mitochondrial DNA Mus Mutation Nerve Degeneration Neurodegenerative Disorders Neurons Parkinson Disease Parkinsonian Disorders Pathologic Pathology Patients Phenotype Premature aging syndrome Proteins Role Substantia nigra structure Testing Therapeutic Toxic effect Tyrosine 3-Monooxygenase Viral Work age related age related neurodegeneration alpha synuclein alpha synuclein gene base brain cell dopaminergic neuron mitochondrial DNA mutation mitochondrial dysfunction mouse model mouse synuclein alpha mutant neuron loss normal aging novel novel therapeutics overexpression vector control

项目摘要

Acquired (somatic) mitochondrial DNA (mtDNA) mutations accumulate with age and reach high levels in dopaminergic neurons at early pathological stages in Parkinson's disease (PD). POLG “mutator” mice have an accelerated accumulation of somatic mtDNA mutations and develop a premature aging phenotype at levels of mutations comparable to levels found in neurons in PD patients, demonstrating that somatic mtDNA mutations can be functionally significant. Furthermore, some patients with POLG mutations develop parkinsonism associated with α-synuclein (αSyn) pathology and loss of dopaminergic neurons in the substantia nigra (SN). Based on these and other data, we hypothesize a “two-hit” hypothesis whereby somatic mtDNA mutations contribute to mitochondrial dysfunction, and thereby exacerbate vulnerability to αSyn. This may help to explain the dramatic rise with age in the incidence of PD. The overall goal of this proposal is to assess the relationship between somatic mtDNA mutations and vulnerability to αSyn toxicity. We will do this using 2 strategies. First, we we will assess the impact of increased levels of somatic mtDNA mutations on αSyn toxicity by assessing heterozygous and homozygous POLG mutator mice that overexpress double-mutant (A30P and A53T) αSyn in dopaminergic neurons. Double-mutant αSyn mice (dMutαSyn) develop a slowly progressive phenotype including striatal dopamine deficiency with behavioral deficits and loss of dopaminergic SN neurons. We predict earlier and more severe behavioral deficits, mitochondrial dysfunction, impaired mitophagy, more severe loss of striatal dopamine and dopaminergic terminals, and increased SN dopaminergic neuronal loss compared to αSyn overexpression in the SN of WT mice. Second, we will perform stereotaxic SN injections of AAV-αSyn (wild-type or A53T) or a control vector (GFP-degron) in wild-type and heterozygous and homozygous POLG mutator mice. We again predict that somatic mtDNA mutations will exacerbate the phenotype associated with increased αSyn expression, including the progressive death of neurons normally seen following AAV-αSyn injections, resulting in earlier and more severe deficits. These studies may yield the first experimental evidence that somatic mtDNA mutations, at levels relevant to early stages in PD, influence vulnerability to αSyn toxicity, a result that may in part explain the dramatic rise in the incidence of PD with age. These studies also will have characterized a novel mouse model of dopaminergic neuronal degeneration that combines two mechanisms of pathophysiological relevance to PD.

获得的（体细胞）线粒体DNA（mtDNA）突变随着年龄的增长而积累，达到高水平帕金森氏病（PD）早期病理阶段的多巴胺能神经元。 POLG“突变器”小鼠有一个体细胞mtDNA突变的加速积累，并在水平的水平上发展过早的衰老表型与PD患者神经元中发现的水平相当的突变，表明体细胞mtDNA突变可能在功能上很重要。此外，一些患有POLG突变的患者会发展出帕金森氏症与黑质Nigra（SN）中α-突触核蛋白（αSyn）病理学和多巴胺能神经元的丧失有关。基于这些数据和其他数据，我们假设一个“两次命中”假设，从而使体细胞mtDNA突变有助于线粒体功能障碍，从而加剧了对αSyn的脆弱性。这可能有助于解释 PD事件中随着年龄的增长而急剧上升。该提案的总体目标是评估关系在体细胞mtDNA突变与αSyn毒性的脆弱性之间。我们将使用两种策略来做到这一点。第一的，我们将通过评估评估体细胞mtDNA突变水平增加对αSyn毒性的影响杂合子和纯合的POLG突变器小鼠过表达双重突变剂（A30p和A53T）αSyn 多巴胺能神经元。双突变αSyn小鼠（DMUTαSyn）发展了缓慢进行的表型包括纹状体多巴胺缺乏症和行为缺乏和多巴胺能SN神经元的丧失。我们预测较早和更严重的行为定义，线粒体功能障碍，线粒体受损，更多纹状体多巴胺和多巴胺能末端的严重丧失，并增加了SN多巴胺能神经元丧失与WT小鼠SN中的αSyn过表达相比。其次，我们将执行立体定位SN注射 AAV-αSyn（野生型或A53T）或野生型和杂合中的对照载体（GFP-Degron）以及纯合POLG突变器小鼠。我们再次预测，体细胞mtDNA突变会加剧与αSyn表达增加相关的表型，包括正常神经元的进行性死亡请参阅以下AAV-αSyn注射，导致早期和更严重的缺陷。这些研究可能会产生第一个实验证据表明，在与PD早期相关的水平上的体细胞mtDNA突变会影响对αSyn毒性的脆弱性，这可能部分解释了PD随年龄的急剧上升。这些研究还将表征一种新型的多巴胺能神经元变性的小鼠模型结合了两种与PD的病理生理相关性的机制。