Epigenetic Analysis of iPS Cell Models of Aging and Alzheimer's Disease

描述(由申请人提供)：本申请涉及广泛的挑战领域(14)干细胞，以及特定的挑战主题14-AG-101：用于衰老和神经变性研究的诱导多能干细胞(IPS)。在很大一部分人口中，衰老伴随着认知能力的下降，是阿尔茨海默病(AD)的主要危险因素。尽管这在疾病的发病机制和发病率中发挥了核心作用，但在分子水平上对大脑的衰老还没有很好的了解。这项应用的总体目标是通过探索不同年龄和AD患者皮肤成纤维细胞产生的iPS细胞和iPS细胞衍生神经元的生理学和表观遗传学，获得对人类衰老和神经退行性疾病的新见解。与年龄匹配的对照组相比，AD成纤维细胞的转录图谱显示细胞死亡基因、突触基因和参与染色质表观遗传修饰的编码蛋白的基因表达发生了变化。此外，其中一些变化也出现在AD患者前额叶皮质的表达谱中。我们的工作假设是，突触和细胞死亡调控基因的表观遗传学变化有助于大脑老化，并在AD的发病机制中发挥作用。这一观点得到了我们已发表的研究的支持，这些研究表明，抑制神经元基因的表达，特别是突触基因，是人类大脑老化的一个显著特征，这种衰老已经在长寿的灵长类动物中进化(1，2)。我们未发表的结果进一步表明，这些与年龄相关的基因表达变化可能反映了染色质的表观遗传变化，并在AD中表现出导致基因表达变化的变化模式。该项目将汇集来自哈佛医学院、Dana Farber癌症研究所和布罗德研究所的一个在神经生物学、疾病生物学和基因组学方面拥有广泛专业知识的多学科团队，以鉴定iPS细胞系和从成人(24-90岁)和散发性AD患者的皮肤成纤维细胞中提取的神经元。免疫细胞化学、电生理学和基因组学鉴定iPS细胞在体外向神经元分化的特征，并与原代人类皮质神经元和神经前体细胞进行比较。尤其令人感兴趣的是iPS神经元形成功能性突触的能力，以及在与衰老和AD相关的应激相关条件下存活的能力，例如氧化应激和暴露于淀粉样蛋白(A？)，以及这种变化是否发生在AD患者的细胞中。供体来源的iPS细胞和iPS神经元的转录和表观基因组图谱将使用微阵列和应用最新的深度测序染色质免疫沉淀(CHIP-SEQ)技术进行。这一多学科的方法将评估iPS细胞作为衰老和神经退化的模型，并探索基因调控和表观遗传学的作用。由于我们已经建立了来自正常老年和AD捐赠者的iPS样细胞系，我们预计这些目标可以在挑战申请的两年时间框架内实现。 公共卫生相关性：目前约有400万美国人患有阿尔茨海默病(AD)，给社会带来了巨大的经济、情感和生产力负担。我们的研究表明，基因组的表观遗传修饰可能在人脑老化中发挥重要作用，并可能导致老化的大脑对AD的脆弱性。这项研究将确定从诱导的祖细胞干细胞获得的神经元是否可以作为研究表观遗传学在衰老和AD中的作用的模型，为分析人类大脑在健康和疾病中的作用提供一个新的水平。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (14) Stem Cells, and specific Challenge Topic, 14-AG-101: Induced Pluripotent Stem (iPS) Cells for Aging and Neurodegeneration Research. Aging is accompanied by cognitive decline in a major segment of the population and is the primary risk factor for Alzheimer's disease (AD). Despite this central role in disease pathogenesis and morbidity, the aging of the brain is not well understood at a molecular level. The overall goal of this application is to gain new insights into human aging and neurodegenerative disease by exploring the physiology and epigenetics of iPS cells and iPS cell-derived neurons generated from skin fibroblasts of individuals at different ages and with AD. Transcriptional profiling of AD fibroblasts compared with age-matched controls shows changes in the expression of cell death genes, synaptic genes and genes encoding proteins involved in epigenetic modification of chromatin. Moreover, some of these changes also appear in the expression profile of the prefrontal cortex in AD. Our working hypothesis is that epigenetic changes in synaptic and cell death regulatory genes contribute to the aging of the brain and play a role in the pathogenesis of AD. This idea is supported by our published studies which show that repression of neuronal gene expression, particularly synaptic genes, is a prominent feature of human brain aging that has evolved in long-lived primates (1, 2). Our unpublished results further suggest that these age-related gene expression changes may reflect epigenetic changes in chromatin, and exhibit an altered pattern in AD leading to altered gene expression. This project will bring together a multidisciplinary team with broad expertise in neurobiology, disease biology and genomics from Harvard Medical School, the Dana Farber Cancer Institute and the Broad Institute to characterize iPS cell lines and neurons derived from dermal fibroblasts of individuals across the adult human lifespan (age 24-90 years) and patients with sporadic AD. Differentiation of iPS cells to neurons in vitro will be characterized by immunocytochemical, electrophysiologic and genomic criteria and compared with primary human cortical neurons and neural progenitors. Of particular interest will be the ability of iPS neurons to form functional synapses and survive under stress-related conditions associated with aging and AD, such as oxidative stress and exposure to amyloid ¿-protein (A¿), and whether this changes in cells derived from AD patients. Transcriptional and epigenomic profiling of donor-derived iPS cells and iPS neurons will be performed using microarrays and by applying recent innovations in deep sequencing chromatin immunoprecipitation (ChIP- seq). This multidisciplinary approach will evaluate iPS cells as models of aging and neurodegeneration, and explore the roles of gene regulation and epigenetics. Since we have already established iPS-like cell lines from normal aged and AD donors, we anticipate that these goals can be accomplished in the two year time frame of a challenge application. PUBLIC HEALTH RELEVANCE: Presently about 4 million Americans suffer from Alzheimer's disease (AD) constituting an enormous financial, emotional and productivity burden on society. Our studies suggest that epigenetic modifications of the genome may play a major role in the aging of the human brain, and may contribute to the vulnerability of the aging brain to AD. This study will determine if neurons derived from induced progenitor stem cells can serve as a model to study the role of epigenetics in aging and AD, providing a new level of analysis of the human brain in health and disease.