AGING BRAIN--IMMUNOHISTOLOGY AND BIOCHEMISTRY

大脑老化——免疫组织学和生物化学

• 批准号: 2442201
• 负责人: SHU-HUI C YEN
• 金额: $ 4.44万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-09-15 至 1997-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2442201
• 关键词: Alzheimer's disease; Downs syndrome; adult human (21+); aging; antigens; complementary DNA; electron microscopy; fibrous protein; gel electrophoresis; gene expression; histochemistry /cytochemistry; human tissue; immunochemistry; laboratory mouse; laboratory rabbit; microfilaments; microtubules; monoclonal antibody; neurofibrillary tangles; neurofilament proteins; nonhistone nucleoprotein; nucleic acid sequence; paired helical filament; protein sequence; protein structure

This project is aimed at understanding the biology and molecular pathogenesis of aging in the nervous system. Alzheimer's disease (AD), the most common organic dementia seen in old age, is characterized histopathologically by the presence of neurofibrillary tangles which consist primarily of paired helical filaments. AD type of neuropathology is found in subjects with Down syndrome who lived to middle age. Alzheimer's neurofibrillary tangles (ANT) and numerous fine processes in the disease affected brain contain epitopes unique to ANT and epitopes shared with normal proteins such as neurofilament proteins, microtubule associated-proteins, and ubiquitin. It is unknown how and when these different components are incorporated into ANT. Using brain tissues with Down syndrome from various age groups and immunocytochemical methods we will determine if there are differences in the sequence of acquiring various epitopes into the abnormal structures. In a previous study, we used monoclonal anti- ANT antibodies to screen a human brain cDNA expression library and have isolated and characterized a MAP2 cDNA that encodes ANT epitopes. In continuing studies, we will produce antibodies to human MAP2 fusion protein in which the known ANT epitopes are deleted. These antibodies will be used to identify and localize additional ANT epitopes in MAP2. The position of ANT epitopes in intact MAP2 molecule will be determined by immunoblotting of the MAP2 peptide fragments (generated by limited proteolysis and chemical cleavage) with anti-ANT antibodies. We hope to find out if ANT-related epitopes are clustered in a specific region of the MAP2 molecule. Using peptide fragments generated from MAP2 fusion protein (by lambda gt 11 containing the MAP2 cDNA insert), we will obtain the partial amino acid sequencing data of small fragments. This data will allow us to confirm the results obtained from the sequencing of MAP2 cDNA. Recent studies showed the cross- reactivity between ubiquitin and neurofibrillary inclusions found in various neuropathological conditions, suggests that the ubiquitin system plays a significant role in neurofibrillary degeneration. In continuing studies, we hope to find out if neurocytoskeletal proteins are acceptors for ubiqutination, to compare the distribution of ubiquitin in different neuronal cytoplasmic compartments, and to determine if cytoskeletal proteins, under pathological conditions, are ubiquitinated to a different extent.

该项目旨在了解生物学和分子 神经系统衰老的发病机理。 阿尔茨海默氏病 （AD）是老年最常见的有机痴呆症，是 通过存在的组织病理学来表征 神经原纤维缠结，主要由配对的螺旋形成 细丝。 广告类型的神经病理学类型在患有的受试者中发现 唐氏综合症的生活到中年。 阿尔茨海默氏症的神经原纤维 疾病影响的缠结（ANT）和许多细微过程 大脑包含蚂蚁独有的表位，并与之共享的表位 正常蛋白质，例如神经丝蛋白，微管 相关蛋白和泛素。 未知如何以及何时 这些不同的组件被纳入蚂蚁。 使用大脑 来自各个年龄组的唐氏综合症的组织和 免疫细胞化学方法我们将确定是否存在 将各种表位获取到该序列的差异 异常结构。 在先前的研究中，我们使用了单克隆抗 筛选人脑cDNA表达文库的蚂蚁抗体和 已经孤立并表征了编码蚂蚁的MAP2 cDNA 表位。 在持续研究中，我们将产生抗体 人Map2融合蛋白，其中已知的蚂蚁表位是 删除。 这些抗体将用于识别和本地化 MAP2中的其他蚂蚁表位。 蚂蚁表位的位置 完整的MAP2分子将通过免疫印迹确定 MAP2肽片段（由有限的蛋白水解和 化学裂解）抗抗体。 我们希望找出答案 如果将与蚂蚁相关的表位聚集在特定区域 MAP2分子。 使用MAP2融合生成的肽片段 蛋白质（由Lambda GT 11包含MAP2 cDNA插入物），我们将 获得小片段的部分氨基酸测序数据。 这些数据将使我们能够确认从 MAP2 cDNA的测序。 最近的研究表明跨 发现泛素和神经原纤维夹杂物之间的反应性 在各种神经病理学条件下，表明 泛素系统在神经原纤维中起重要作用 退化。 在继续学习中，我们希望找出是否 神经细胞骨骼蛋白是ubiqutination的受体， 比较不同神经元中泛素的分布 细胞质隔室，并确定是否细胞骨架 蛋白质在病理条件下，被泛素化为 不同的程度。