Generation of DYT1 dystonia-specific iPS cells with isogenic controls

具有等基因对照的 DYT1 肌张力障碍特异性 iPS 细胞的生成

基本信息

批准号：
8445111
负责人：
David Cristopher Bragg
金额：
$ 8.7万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-01 至 2014-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8445111
关键词：
Address Adrenergic Agents Adrenergic Receptor Affect Biological Models Brain Brain region Cell Line Cells Central Nervous System Diseases Childhood Chronic Collaborations Collection Communities Cultured Cells DYT1 gene Data Databases Defect Disease Disease model Dopamine D2 Receptor Dystonia Endoplasmic Reticulum Engineering Epigenetic Process Equipment and supply inventories Excision Exhibits Experimental Models Fibroblasts Functional disorder Future G-Protein-Coupled Receptors Gene Expression Gene Expression Profile Gene Mutation Gene Targeting Generations Genome Glutamic Acid Goals Hereditary Dystonia Human Individual Institutes Ligands Maps Mediating Membrane Proteins Modeling Modification Molecular Molecular Chaperones Molecular Profiling Movement Movement Disorders Muscle Contraction Mutant Strains Mice Mutation Neuroglia Neurologic Neurons Outcome Pathogenesis Pathway interactions Patients Pattern Peripheral Phenotype Plasmids Positron-Emission Tomography Posture Process Proteins Qualifying RNA Research Resources Signal Transduction Smooth Muscle Somatic Cell Southern Blotting Stem cells Study models Symptoms Synapses TOR1A gene Technology Teratoma Therapeutic Time TorsinA Transcription Coactivator Transgenes Work adrenergic base cell type cohort disease mechanisms study early onset homologous recombination induced pluripotent stem cell lymphoblast mouse model mutant mutation carrier nuclease repaired stem cell differentiation synaptic function transcription factor

项目摘要

DESCRIPTION (provided by applicant): A major challenge in understanding and treating dystonia is the limitation of current models for studying the disease. The dystonias are generally believed to represent chronic signaling dysfunctions in neurons within brain regions controlling movement, although characterizing subtle, dystonia-related disturbances in these cells at the molecular level has been difficult. Traditional approaches to model the disorder have relied on (1) peripheral patient cells (fibroblasts or lymphoblasts); (2) mutant mice; and/or (3) cultured cells bearing manipulations in proteins implicated in the disease process. While these strategies have value, there are limits to what they can reveal about effects of dystonia-related gene mutations on human neurons. Reprogramming patient somatic cells to induced pluripotent stem cells (iPSCs) which can be differentiated into neurons has become an increasingly common approach to modeling CNS disorders. Yet over time it has become clear that iPSCs can display substantial inter-individual variability which may confound identification of disease-specific phenotypes. Here we propose a strategy to develop a new human neuronal culture model of DYT1 dystonia that combines iPSCs reprogramming with gene targeting using transcription activator-like effector nucleases (TALENs). By using engineered TALENs to correct the DYT1 gene mutation in patient iPSCs, we will generate matched isogenic control lines from the same epigenetic background. As a first application of this model, we will perform transcriptional profiling with pathway analyses to identify functional networks that are perturbed in DYT1 iPSC-derived neurons compared to matched control cells. The outcome of this work will be a powerful new resource for probing neuron-specific deficits in DYT1 dystonia, with detailed expression profiling that will inform future studies of this model system. PUBLIC HEALTH RELEVANCE: Early onset (DYT1) dystonia is a crippling neurologic movement disorder. A major challenge in studying DYT1 is that the cells believed to be most affected by the disease mutation are neurons, which may be functionally distinct from the cell lines frequently used as experimental models. Here we propose to establish a new model system consisting of DYT1 patient-specific, induced pluripotent stem cells which can be differentiated into neurons, thereby facilitating new analyses of neuron-specific functional defects that may underlie this disease.

描述（由申请人提供）：理解和治疗肌张力障碍的主要挑战是研究该疾病的当前模型的局限性。通常认为肌张力蛋白代表了控制运动的脑区域内神经元中的慢性信号传导功能障碍，尽管表征了这些细胞在分子水平上的微妙，与肌张力障碍相关的干扰。传统的建模该疾病的方法依赖于（1）外周患者细胞（成纤维细胞或淋巴细胞）；（2）突变小鼠；和/或（3）与疾病过程有关的蛋白质中有操纵的培养细胞。尽管这些策略具有价值，但它们可以揭示与肌张力障碍相关基因突变对人神经元的影响的局限性。将患者的体细胞重新编程为可以区分神经元的多能干细胞（IPSC）已成为建模中枢神经系统疾病的一种越来越普遍的方法。然而随着时间的流逝，很明显，IPSC可以显示出实质性的个体变异性，这可能会混淆疾病特异性表型的识别。在这里，我们提出了一种策略，以开发一种新的人类神经元培养模型的Dyt1肌张力障碍，该模型将IPSC重编程与使用转录激活剂样效应核酸酶（Talens）相结合的基因靶向。通过使用工程的TALEN纠正患者IPSC中的DYT1基因突变，我们将从相同的表观遗传背景中生成匹配的同基因控制线。作为该模型的第一个应用，我们将通过途径分析进行转录分析，以识别与匹配的对照单元相比，在DYT1 IPSC衍生的神经元中受到干扰的功能网络。这项工作的结果将是探测Dyt1肌张力障碍神经元特异性缺陷的有力新资源，并具有详细的表达分析，将为该模型系统的未来研究提供依据。公共卫生相关性：早期发作（DYT1）肌张力障碍是一种残酷的神经运动障碍。研究DYT1的一个主要挑战是，据信被认为受疾病突变影响最大的细胞是神经元，其在功能上可能与经常用作实验模型的细胞系不同。在这里，我们建议建立一个新的模型系统，该模型系统由DYT1患者特异性，诱导的多能干细胞组成，可以分化为神经元，从而促进了可能是这种疾病的神经元特异性功能缺陷的新分析。