Exploring Disease-Toxicant Interactions in a Human Huntington's Disease Model

探索人类亨廷顿病模型中的疾病与毒性相互作用

• 批准号: 8396851
• 负责人: Andrew M Tidball
• 金额: $ 2.69万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-01 至 2014-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8396851
• 关键词: Affect; Age; Basic Science; Biological Assay; Biological Models; Brain; Brain region; CAG repeat; Calcium; Cell Culture Techniques; Cell Death; Cell Line; Cell Survival; Cells; Cellular Stress; Cellular Stress Response; Codon Nucleotides; Communities; Complex; Corpus striatum structure; Data; Development; Disease; Disease Progression; Disease model; Environment; Environmental Health; Environmental Medicine; Environmental Risk Factor; Etiology; Exhibits; Exposure to; Freedom; Functional disorder; Funding; Generations; Genes; Genetic; Genetic Risk; Genotype; Goals; Heat shock proteins; Human; Huntington Disease; In Vitro; Individual; Inheritance Patterns; Juvenile-Onset Huntington Disease; Knowledge; Laboratories; Left; Length; Measures; Membrane Potentials; Messenger RNA; Metabolic; Metals; Mitochondria; Modeling; Modification; Multivariate Analysis; Mutate; Nerve Degeneration; Neurodegenerative Disorders; Neurologic; Neuronal Dysfunction; Neurons; Neurotoxins; Other Genetics; Ownership; Oxidative Stress; Pathway interactions; Patients; Phenotype; Population; Predisposition; Principal Investigator; Process; Production; Promoter Regions; Proteins; Protocols documentation; Reactive Oxygen Species; Regression Analysis; Research; Research Personnel; Research Project Grants; Risk; Staging; Statistical Models; Stress; Study models; Techniques; Testing; Toxic Environmental Substances; Toxic effect; Toxicant exposure; Training; Variant; base; cell type; design; disorder control; environmental agent; environmental stressor; excitotoxicity; genetic risk factor; human Huntingtin protein; human subject; induced pluripotent stem cell; inhibitor/antagonist; mitochondrial dysfunction; mitochondrial membrane; mutant; nerve stem cell; neuropathology; regional difference; research study; response; stress protein; stressor; therapy development; toxicant; toxicant interaction

DESCRIPTION (provided by applicant): Environmental agents are thought to interact with an individual's genetic makeup to influence or cause neurodegenerative disease. Huntington's disease (HD) is a debilitating neurological condition with a dominant monogenic inheritance pattern. Still, environmental factors are thought to strongly affect disease age at onset and progression. We postulate that the environment causes cellular stress that neurons in the HD brain are susceptible to such as oxidative stress, mitochondrial dysfunction, and calcium dysregulation. We hypothesize that mutant Huntingtin impinges upon the cellular stress response to produce this disease-specific vulnerability. We will test this hypothesis by exposing cells to toxicants known to elicit implicated cellular stress pathways and measure toxicological phenotypes. Here we propose using induced pluripotent stem cells differentiated into striatal neural progenitors from patients with Huntington's disease and controls as our model system. These cell lines contain the mutated Huntingtin gene with the complete promoter region as well as patient- specific genetic background in a neural cell type. In Specific Aim 1, we will optimize the technique for generating DLX2-expressing neural progenitors. We will then expose DLX2-expressing cultures from hiPSCs from two juvenile HD subjects and two control subjects to cell stress model toxicants and evaluate for differential viability. In Specific Aim 2, these same cells will be used to assay the effects toxicant exposure on reactive oxygen species production, mitochondrial membrane potential, ATP content, and cytosolic calcium concentration. The data generated in Aim 1 and Aim 2 will be analyzed by multivariate ANCOVA statistical modeling for disease-toxicant and/or patient-toxicant interactions. For toxicants eliciting a statistically significant disease-toxicant interaction, we will then perform protein and mRNA arrays for important cellular response factors. In Aim 3, any disease-toxicant interactions identified in Aim 1 and Aim 2 will be further tested in hiPSC-derived cells from HD patients with a range of different pathological repeat lengths. Results will also be analyzed by multivariate ANCOVA to determine if any of the phenotypes are repeat length dependent. This proposal will identify types of cellular stress and potential toxicants that could alter the progression of Huntington's disease and investigate the manner by which mutant Huntingtin causes this susceptibility. The proposed research will seek to identify patient-specific risk for these toxicants potentially providing individualized environmental health advice for patients (i.e. personalized environmental medicine). This funding will also contribute to my individual development as an independent researcher by allowing me to pursue these research aims and present the resulting data to the broader scientific community. As the principal investigator, I will have increased ownership of this research project and the freedom to take full advantage of my graduate training by reducing potential financial constraints. PUBLIC HEALTH RELEVANCE: This proposal will use human induced pluripotent stem cells (hiPSCs) from Huntington's disease and control subjects to evaluate the effects of toxicants (e.g. metals, mitochondrial complex inhibitors, and calcium dysregualtors) producing types of cell stress implicated in Huntington's disease progression. We will test cell viability, reactive oxygen species, mitochondrial membrane potential, ATP content, and cytosolic calcium concentration in patient-specific neural progenitors to identify differential toxicity and assess te mechanisms behind these differences. This proposal uses basic science to define potential environmental toxicant modification of Huntington's disease.

描述(由申请人提供)：环境因子被认为与个体的基因构成相互作用，从而影响或导致神经退行性疾病。亨廷顿病(HD)是一种具有显性单基因遗传模式的衰弱神经系统疾病。尽管如此，环境因素被认为对发病和进展的年龄有很大影响。我们推测，环境导致HD大脑中的神经元容易受到细胞应激的影响，如氧化应激、线粒体功能障碍和钙调节失调。我们假设，突变的Huntingtin会影响细胞的应激反应，从而产生这种疾病特异性的脆弱性。我们将通过将细胞暴露在已知可引发牵连的细胞应激途径的毒物中并测量毒物表型来检验这一假说。在这里，我们建议使用从亨廷顿病患者和对照组分化为纹状体神经前体细胞的诱导多能干细胞作为我们的模型系统。这些细胞系含有突变的亨廷顿基因，具有完整的启动子区域以及特定于患者的神经细胞类型的遗传背景。在具体目标1中，我们将优化表达DLX2的神经前体细胞的生成技术。然后，我们将来自两名青少年HD受试者和两名对照受试者的HiPSCs中表达DLX2的培养物暴露于细胞应激模型毒物中，并评估不同的活性。在特定目标2中，这些相同的细胞 将被用来分析毒物暴露对活性氧产生、线粒体膜电位、ATP含量和胞浆钙浓度的影响。AIM 1和AIM 2中产生的数据将通过疾病-毒物和/或患者-毒物相互作用的多变量ANCOVA统计模型进行分析。对于引起统计学上显著的疾病-毒物相互作用的毒物，我们将对重要的细胞反应因子进行蛋白质和信使核糖核酸阵列。在目标3中，目标1和目标2中确定的任何疾病-毒物相互作用都将在来自HD患者的HiPSC来源的细胞中进行进一步测试，这些细胞具有一系列不同的病理重复长度。结果还将用多变量ANCOVA进行分析，以确定是否有任何表型与重复长度有关。这项提案将确定可能改变亨廷顿病进展的细胞应激类型和潜在毒物 并调查突变的亨廷顿是如何导致这种易感性的。拟议的研究将寻求确定这些毒物可能为患者提供个性化环境健康建议的患者特定风险(即个性化环境医学)。这笔资金还将有助于我作为一名独立研究人员的个人发展，使我能够追求这些研究目标，并将结果数据呈现给更广泛的科学界。作为首席研究员，我将增加对这个研究项目的所有权，并通过减少潜在的财务限制来充分利用我的研究生培训的自由。 公共卫生相关性：这项建议将使用亨廷顿病患者和对照受试者的人类诱导多能干细胞(HiPSCs)来评估毒物(如金属、线粒体复合体抑制剂和钙调节障碍因子)产生的与亨廷顿病进展有关的细胞应激类型的影响。我们将测试患者特定神经前体细胞的细胞存活率、活性氧物种、线粒体膜电位、ATP含量和胞浆钙浓度，以确定不同的毒性并评估这些差异背后的TE机制。这项建议使用基础科学来定义亨廷顿氏病潜在的环境毒物修饰。