METHOD FOR INVESTIGATING CHROMOSOMES IN HUMAN NEURONS

未分裂的人类神经元一直无法进行分析 依赖细胞周期的传统细胞遗传学技术 分裂成有丝分裂以便观察到染色体。早熟染色体 凝聚(PCC)方法，基于有丝分裂和细胞融合 非有丝分裂细胞群，不需要这样的周期。公众投诉委员会 将在人类神经组织上开发和标准化技术以 允许人们第一次看到未转化的染色体 人类神经元。 一旦标准化，PCC-神经元技术将提供一个有价值的工具 用于检查从大脑分离的神经元的染色体变化 阿尔茨海默氏型痴呆症的个体。其性质和范围 这些变化可能会产生关于病因机制的重要信息。 这种混乱的症状。 冰冻尸检、未冰冻尸检的纯神经元群体，以及 活检样本将被隔离。有丝分裂的CHO和HeLa种群 细胞将被准备好。神经元将与有丝分裂细胞融合 使用紫外线灭活的仙台病毒或聚乙二醇作为 扶桑。病毒技术是在悬浮液中进行的，聚乙烯 乙二醇法，单层。首先，这两种技术都将是 使用HeLa和CHO有丝分裂细胞。对融合和融合的评估 初步试验的PCC结果将决定方法和细胞 未来所有实验的选择类型。 过早浓缩的神经元染色体会被带状排列， 分析染色体数目、染色体断裂和染色体 形态学。这三种不同的组织来源将被比较 融合效率的差异，PCC的诱导，还是为了差异 在染色体分析中。 在PCC-神经元技术在正常组织上标准化后，样本 从10名家族性阿尔茨海默病患者和10名对照患者的颞叶中提取， 与年龄、性别和尸检时间匹配的人将被分析。仅限 年龄在65岁以下的个人将被包括在内。染色体分析 将包括对数字和结构变化的审查。

Nondividing human neurons have not been accessible to analysis by conventional cytogenetic techniques which rely upon the cycling of a cell into mitosis in order to visualize the chromosomes. Premature chromosome condensation (PCC) methods, based on cell fusion between mitotic and nonmitotic cell populations, do not require such cycling. The PCC technique will be developed and standardized on human neural tissue to permit one to see, for the first time, the chromosomes of nontransformed human neurons. Once standardized, the PCC-neuron technique will provide a valuable tool for the examination of chromosomal changes in neurons isolated from brains of individuals with dementia of the Alzheimer type. The nature and extent of such changes could yield important information on etiological mechanisms of this disorder. Pure populations of neurons from frozen autopsy, unfrozen autopsy, and biopsy specimens will be isolated. Populations of mitotic CHO and HeLa cells will be prepared. The neurons will be fused with the mitotic cells using either UV-inactivated Sendai virus or polyethylene glycol as a fusogen. The viral technique is done in suspension, the polyethylene glycol technique, in monolayer. At first, both techniques will be employed, using both HeLa and CHO mitotic cells. Evaluation of fusion and PCC outcomes from preliminary trials will determine the method and cell type of choice for all future experiments. The prematurely condensed chromosomes of the neurons will be banded and analyzed for chromosome number, chromosome breakage, and chromosome morphology. The three different sources of tissue will be compared for any differences in the efficiency of fusion, PCC induction, or for differences in chromosomal analyses. After the PCC-neuron technique is standardized on normal tissue, samples from temporal lobes from 10 familial Alzheimer cases and 10 control cases, matched for age, sex, and post-mortem time will be analyzed. Only individuals aged less than 65 years will be included. Chromosomal analysis will consist of the examination of numerical and structural changes.