AGING BRAIN--IMMUNOHISTOLOGY AND BIOCHEMISTRY

该项目旨在了解生物学和分子生物学 神经系统衰老的发病机制。 阿尔茨海默病 (AD)老年人最常见的器质性痴呆是 组织病理学特征为存在 神经纤维缠结，主要由成对的螺旋 细丝。 AD型神经病理学在患有 唐氏综合症患者活到中年。 阿尔茨海默氏神经系统 缠结（ANT）和许多受影响的疾病的细过程 脑含有ANT特有的表位和与ANT共有的表位。 正常蛋白质如神经丝蛋白、微管蛋白 相关蛋白和泛素。 目前还不清楚如何以及何时 这些不同的组件被合并到ANT中。 使用大脑 唐氏综合症患者的组织， 免疫细胞化学方法，我们将确定是否有 在获得各种表位的序列中的差异， 异常结构 在以前的研究中，我们使用单克隆抗- ANT抗体筛选人脑cDNA表达文库， 分离并鉴定了编码ANT的MAP 2 cDNA 表位 在接下来的研究中，我们将生产抗体， 人MAP 2融合蛋白，其中已知的ANT表位是 的页面不存在或 这些抗体将用于识别和定位 MAP 2中的额外ANT表位。 ANT表位在 完整的MAP 2分子将通过免疫印迹来确定。 MAP 2肽片段（通过有限的蛋白水解产生， 化学切割）与抗ANT抗体。 我们希望找出 如果ANT相关表位聚集在免疫球蛋白的特定区域， MAP 2分子。 使用由MAP 2融合产生的肽片段 蛋白质（由λ gt 11含有MAP 2 cDNA插入），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 MAP 2 cDNA序列测定。 最近的研究表明，交叉- 发现泛素和神经细胞内含物之间的反应性 在各种神经病理学条件下，表明 泛素系统在神经细胞凋亡中起着重要的作用 退化 在接下来的研究中，我们希望发现， 神经细胞骨架蛋白是泛素化的受体， 比较泛素在不同神经元中的分布 细胞质区室，并确定是否细胞骨架 在病理条件下，蛋白质被泛素化为 不同程度。

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.