Pluripotent stem cell models of sporadic Alzheimer's Disease

描述(由申请人提供)：散发性阿尔茨海默病(SAD)患者的基因组变异如何促进阿尔茨海默病(AD)的发展？为了开始解决这个问题，人类诱导多能干细胞(HiPSC)技术将被用来开始测试这一假设，即单个SAD患者的基因组包含在人类神经元中产生生化可检测表型的遗传变异。人类多能干细胞(HiPSC)允许在多能干细胞系中捕获患有SAD的人类个体的基因组。然后，这些细胞可以在体外分化为人类神经元或神经胶质细胞，以评估捕获的基因组是否以类似于携带FAD突变的细胞的方式改变神经元或神经胶质表型，或如ASS假说等SAD的机械模型所预测的那样。HiPSC模型也可能有助于解决人类特有的效应，并避免动物模型众所周知的某些方面的限制，如转基因模型中的高拷贝数和缺乏人类遗传背景。我们的基本假设是，SAD患者的基因组包含一组易感变异，这些变异可能会以可检测的方式改变神经元、神经胶质细胞或其他相关细胞表型。总的来说，SAD患者的基因组变异可能通过多种方式导致疾病，从影响炎症通路、Ass的胶质周转或由神经元和其他细胞产生的其他潜在有毒分子，或通过神经元对APP处理途径的影响、Ass中毒的易感性、tau过度磷酸化、氧化损伤等。在这项R21探索性提案中，我们建议迈出第一步，并使用hiPSC技术来检验这样一种假设，即至少一些SAD基因组导致APP表达、APP加工或tau磷酸化表型在携带SAD患者基因组的人类神经元中。这些基因组的鉴定将为进一步研究提供原材料，使用高分辨率的分子遗传分析来确定不同的基因组变异如何影响神经元表型，并可能识别新的药物靶点和途径。还可以生成可能的预测性措施和诊断。在这个试点项目中，我们将通过完成两个特定的目标来探讨这些问题：1)我们将从5名SAD患者和5名年龄匹配的正常对照(NC)各建立3个HiPSC系。2)我们将从每个HiPSC系中提取纯化的神经元，并测试SAD神经元是否表现出典型的FAD的APP表达、APP处理或tau磷酸化变化。 与公共卫生相关：散发性阿尔茨海默病是一种主要的公共卫生问题，在65岁以上的人口中有10%的人会发病，在85岁以上的人口中有50%会发病。目前还没有疾病改变治疗策略，尽管一些药物提供了暂时的症状缓解。我们建议使用干细胞技术来发现遗传变异是如何导致阿尔茨海默病的，希望这项工作将允许开发新的疗法和预测性诊断。

DESCRIPTION (provided by applicant): How do the genomic variants in individuals with sporadic Alzheimer's Disease (SAD) contribute to the development of Alzheimer's Disease (AD)? To begin addressing this question human induced pluripotent stem cell (hIPSC) technology will be used to begin testing the hypothesis that the genomes of individual SAD patients contain genetic variants that generate biochemically detectable phenotypes in human neurons. Human pluripotent stem cells (hIPSC) allow the genomes of human individuals afflicted with SAD to be captured in a pluripotent stem cell line. Such cells can then be differentiated to human neurons or glia in vitro for evaluating whether the captured genome alters neuronal or glial phenotype in a manner similar to that seen in cells carrying FAD mutations or as predicted by mechanistic models of SAD such as the Ass hypothesis. An hIPSC model may also be useful for addressing human specific effects and avoiding some aspects of the well known limitations of animal models such as high copy number in transgenic models and the absence of a human genetic background. Our underlying hypothesis is that the genome of a patient with SAD contains a set of susceptibility variants that may alter neuronal, glial, or other relevant cellular phenotypes in a detectable manner. Broadly speaking, genomic variants in SAD patients could contribute to disease in a variety of ways ranging from effects on inflammatory pathways, glial turnover of Ass or other potentially toxic molecules generated by neurons and other cells, or by effects in neurons on pathways of APP processing, susceptibility to Ass poisoning, tau hyperphosphorylation, oxidative damage, etc. In this R21 exploratory proposal, we propose to take the first step and use hIPSC technology to test the hypothesis that at least some SAD genomes cause APP expression, APP processing, or tau phosphorylation phenotypes in human neurons that carry the genomes of patients who developed SAD. Identification of such genomes would provide the raw material for further studies using high resolution molecular genetic analyses to define how different genomic variants contribute to neuronal phenotypes and might allow new drug targets and pathways to be identified. Possible predictive measures and diagnostics might also be generated. In this pilot project, we will probe these issues by completing two specific aims: 1) We will generate 3 hIPSC lines each from 5 SAD patients and 5 age-matched normal controls (NC). 2) We will generate purified neurons from each hIPSC line and test whether SAD neurons exhibit APP expression, APP processing, or tau phosphorylation changes typical of FAD. PUBLIC HEALTH RELEVANCE: Sporadic Alzheimer's Disease is a major public health problem that develops in as much as 10% of the population older than 65 and 50% of the population older than 85. There are currently no disease altering therapeutic strategies although some agents offer temporary symptomatic relief. We propose to use stem cell technology to discover how genetic variation predisposes to Alzheimer's Disease with the hope that this work will allow new therapeutics and predictive diagnostics to be developed.