MOLECULAR BIOLOGY OF ALZHEIMER DISEASE NEURODEGENERATION

阿尔茨海默病神经退行性变的分子生物学

• 批准号: 3415681
• 负责人: RACHAEL Lee NEVE
• 金额: $ 16.72万
• 依托单位: MCLEAN HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-01 至 1994-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3415681
• 关键词: Alzheimer's disease; PC12 cells; amyloid proteins; disease /disorder model; endopeptidases; gene expression; genetically modified animals; glia; hamsters; hippocampus; human tissue; immunocytochemistry; in situ hybridization; laboratory mouse; laboratory rat; membrane proteins; model design /development; molecular cloning; molecular pathology; nervous system transplantation; neural degeneration; neurogenesis; neurons; neuropharmacology; neurotoxins; neurotrophic factors; radioassay; tau proteins; tissue /cell culture; transfection

The isolation by our laboratory and others of the gene for the precursor of the Alzheimer's disease amyloid polypeptide, has made it possible to begin dissecting at a molecular level the processes whereby this normal protein may become altered in Alzheimer's disease. The sequence of events which leads to the deposition of the small self-aggregating amyloid peptide in Alzheimer's disease is not known. A key question concerns the causal relationship of amyloid to the pathology of Alzheimer's disease. Projects are proposed to address these questions. In the original application, we proposed to do a detailed analysis of amyloid protein precursor (APP) transcription in defined regions of Alzheimer's disease and normal brains, to test the hypothesis that the development of pathology in Alzheimer's disease is related to changes in the expression of the APP mRNAs. This specific aim has been completed: we were able to show that the mRNA encoding APRP-563, an APP-related mRNA lacking the amyloid-encoding sequence, is specifically elevated in pathologically affected regions of Alzheimer's disease brain. We have shown, by the construction and characterization of recombinant transfected cell lines expressing particular domains of APP, that the carboxyterminal 105 amino acids of the human amyloid protein precursor is neurotoxic. Studies are proposed to examine the relationship of the neurotoxicity of this fragment (termed AB1) to Alzheimer's disease pathology, by analyzing the pattern of segregation of the microtubule associated protein tau in neurons treated with AB1, and by assaying for differential vulnerability of CA1 hippocampal neurons to AB1 neurotoxicity. Characterization of AB1 neurotoxicity will include definition of the mechanism of glial protection of neurons against AB1 toxicity, determination of the vulnerability of hippocampal neurons from rats of various ages to AB1 neurotoxicity, and assessment of the minimum exposure time of neurons to the AB1 conditioned medium that is necessary to cause their subsequent degeneration. We recently discovered that the neurotoxic AB1 fragment can, under the appropriate conditions, be trophic: experiments to quantify and characterize this trophic effect, and to relate it to the neurotoxicity, are described. Both genetic and pharmacologic methods will be utilized in an attempt to neutralize or block the neurotoxicity of the carboxyterminal 105 amino acids of AB1. The development of alternate methods of synthesis of this AB1 fragment will facilitate the performance of the proposed studies. Studies to follow up on our recent finding that the AB1 fragment binds specifically to the surface of NGF-differentiated PC12 cells but not to undifferentiated PC12 cells, are proposed. Finally, we show that mouse brains transplanted with AB1-transfected PC12 cells demonstrate progressive neurodegeneration in the region of the transplant, and show immunoreactivity with the Alz50 antibody. The possibility of developing these transplanted mice as an animal model for Alzheimer's disease is proposed.

我们的实验室和其他实验室分离出前体基因 阿尔茨海默病淀粉样蛋白多肽，使开始成为可能 在分子水平上剖析这种正常蛋白质的过程 阿尔茨海默病可能会发生改变。 事件发生的顺序 导致小的自聚集淀粉样肽沉积在 阿尔茨海默病尚不清楚。 一个关键问题涉及因果关系 淀粉样蛋白与阿尔茨海默病病理学的关系。 项目 建议解决这些问题。 在最初的应用程序中，我们 建议对淀粉样蛋白前体（APP）进行详细分析 阿尔茨海默病和正常大脑特定区域的转录， 检验阿尔茨海默病病理发展的假设 疾病与 APP mRNA 表达的变化有关。 这 具体目标已经完成：我们能够证明 mRNA 编码 APRP-563，一种缺乏淀粉样蛋白编码的 APP 相关 mRNA 序列，在受病理影响的区域中特别升高 阿尔茨海默病大脑。 我们已经表明，通过重组体的构建和表征 表达 APP 特定结构域的转染细胞系， 人类淀粉样蛋白前体的羧基端105个氨基酸是 神经毒性。 建议进行研究以检验两者之间的关系 该片段（称为 AB1）对阿尔茨海默病的神经毒性 病理学，通过分析微管的分离模式 经 AB1 处理的神经元中的相关蛋白 tau，并通过测定 CA1 海马神经元对 AB1 神经毒性的不同脆弱性。 AB1 神经毒性的表征将包括以下定义： 神经胶质细胞保护神经元免受AB1毒性的机制， 大鼠海马神经元脆弱性测定 不同年龄对 AB1 神经毒性的影响，以及最低暴露量的评估 神经元接触 AB1 条件培养基的时间是引起 他们随后的退化。 我们最近发现，神经毒性 AB1 片段在适当的条件下可以是营养性的： 量化和表征这种营养效应的实验，并将 对它的神经毒性，进行了描述。 无论是遗传还是药理学 方法将被用来试图中和或阻止 AB1 羧基末端 105 个氨基酸的神经毒性。 这 开发该 AB1 片段的替代合成方法将 促进拟议研究的进行。 后续研究 我们最近发现 AB1 片段特异性结合 NGF 分化的 PC12 细胞表面，但不包括未分化的 PC12 细胞，提出了。 最后，我们证明小鼠大脑移植了 AB1转染的PC12细胞表现出进行性神经变性 移植区域，并显示与 Alz50 的免疫反应性 抗体。 将这些移植小鼠开发为 提出了阿尔茨海默病的动物模型。