PATHOGENIC MECHANISMS OF PRESENILIN MUTATION

描述(申请人摘要)：编码早老素的基因突变 1和2(PS-1和PS-2)是家族性早发的主要原因 阿尔茨海默病(AD)。操纵早老素表达和表达的研究 将突变形式引入转基因小鼠和培养细胞 证实PS-1在早期脑发育和 暗示了突变的潜在致病机制，但有技术上的 限制其实用性的限制。拟议的研究将延长 通过评估PS-1的正常和致病作用，这些努力 使用新的小鼠模型和培养的神经元来源的大脑成熟和老化 从他们那里。我们使用基因打靶技术来创建小鼠实验模型 忠实地模仿家族性阿尔茨海默病(突变体)的遗传学 “敲入”)或在成年期过低表达PS-1(“低畸形症”)。五个老鼠品系 将研究：(1)PS-1和APP野生型；(2)PS-1P264L敲入；(3)APPswe (4)PS-1/APP双重敲入；(5)PS-1低构型。具体目标1将 确定PS-1在小鼠脑成熟和衰老中的功能并进行测试 FAD相关的PS-1突变或PS-1功能部分丧失是否导致 阿尔茨海默型退行性神经病理学。比较组织学分析 将评估FAD连锁的PS-1P264L突变或PS-1的敲入效应 神经元死亡率与神经元大小、拓扑和局部的亚同态 成熟和老化的小鼠大脑的体系结构，并将解决细胞 以及PS-1相关神经病理的生化基础。特定目标2将 验证与AD相关的突变体PS-1在正常表达时的假设 水平，危及体外和体内脑神经元的退化。神经元 对萎缩、细胞凋亡和坏死的易感性将被评估为 PS-1基因对不同来源原代培养神经元的作用 对于成熟的状态和受伤的成人大脑。《特定目标3》将进行测试 阿尔茨海默病相关突变体PS-1增加脑组织产量的假说 通过促进淀粉样蛋白Abeta1-42片段的募集而获得 β-淀粉样前体蛋白。阿贝塔1-42地层的限速措施 将由培养的原代细胞的分子和药理学分析确定 神经元，以及“招募假说”所做的预测将会被评估 在培养的神经元和小鼠大脑中至关重要。这些研究将取得进展 我们对PS-1在大脑中的正常和致病功能的理解，以及 从而为制定针对性治疗策略提供了重要的基础 在减缓阿尔茨海默病的进行性恶化方面。

DESCRIPTION (Applicant's Abstract): Mutations in the genes encoding presenilins 1 and 2 (PS-1 and PS-2) are a leading cause of familial, early-onset Alzheimer's disease (AD). Studies manipulating presenilin expression and introducing mutant forms into transgenic mice and cultured cells have demonstrated an important role for PS-1 in early brain development and suggested potential pathogenic mechanisms for the mutations, but have technical limitations which constrain their utility. The proposed research will extend these efforts by evaluation of the normal and pathogenic roles of PS-1 in the maturing and aging brain, using novel mouse models and cultured neurons derived from them. We use gene targeting to create mouse experimental models that emulate faithfully the genetics of familial Alzheimer's disease (mutant "knock-in") or underexpress PS-1 into adulthood ("hypomorph"). Five mouse lines will be studied: (1) PS-1 and APP wild type; (2) PS-1P264L knock-in; (3) APPswe knock-in; (4) PS-1/APP double knock-in; (5) PS-1 hypomorph. Specific Aim 1 will define functional roles of PS-1 in mouse brain maturation and aging and test whether an FAD-linked PS-1 mutation or partial loss of PS-1 function cause Alzheimer-type degenerative neuropathology. Comparative histological analyses will evaluate effects of knock-in of the FAD-linked PS-1P264L mutation or PS-1 hypomorphism on neuronal death rates and the size, topology and regional architecture of the maturing and aging mouse brain, and will address cellular and biochemical bases for PS-1-related neuropathologies. Specific Aim 2 will test the hypothesis that an AD-linked mutant PS-1, when expressed at normal levels, endangers brain neurons in vitro and in vivo to degeneration. Neuronal vulnerability to atrophy, apoptosis and necrosis will be evaluated as a function of PS-1 genotype for cultured primary neurons of different maturational states and for the injured adult brain. Specific Aim 3 will test the hypothesis that an AD-linked mutant PS-1 increases production in brain of the amyloid Abeta1-42 peptide by enhancing recruitment of a fragment of the beta-amyloid precursor protein. The rate-limiting step in Abeta1-42 formation will be determined by molecular and pharmacologic analyses of cultured primary neurons, and predictions made by the "recruitment hypothesis" will be evaluated critically in cultured neurons and the mouse brain. These studies will advance our understanding of normal and pathogenic functions of PS-1 in the brain, and so provide an important foundation for developing therapeutic strategies aimed at slowing the progressive deterioration of Alzheimer's disease.