HCMV先天性潜伏感染老年小鼠脑AD样病理改变的形态学观察

发布时间：2018-06-01 18:03

本文选题：人巨细胞病毒 + 先天性潜伏感染　；参考：《安徽医科大学》2005年硕士论文

【摘要】：目的本研究在建立人巨细胞病毒(human cytomegalovirus, HCMV)先天性潜伏感染老年小鼠模型的基础上,重点观察了HCMV先天性潜伏感染引起小鼠大脑皮质AD样病理改变,探求HCMV先天性潜伏感染在AD发生中的可能作用,为AD病因学研究及发生机制提供新的思路和合适的动物模型。方法从确定为HCMV先天性潜伏感染的小鼠及DMEM组小鼠中随机各取6只,分别按照实验设计处死小鼠,无菌取小鼠大脑组织。一部分用多聚甲醛固定,常规制作石蜡切片。一部分用戊二醛固定,制作电镜标本。其余脑组织做体外共培养激活实验。取石蜡切片用于以下试验:(1)HE染色检测小鼠脑组织构筑、病理改变和生物学特性;(2)刚果红染色检测脑组织中的淀粉样斑块;(3)镀银染色检查脑组织中的神经原纤维缠结和淀粉样斑块;(4)间接免疫荧光检测脑组织中的磷酸化Tau蛋白;(5)透射电镜观察小鼠大脑神经元超微结构改变;(6)体外共培养激活实验观察:①HCMV特征性CPE ②PCR和RT-PCR分别检测HCMV IE DNA和UL83 DNA及其相应mRNA表达③间接免疫荧光检测细胞爬片中的HCMV IE和pp65蛋白 ④透射电镜观察细胞爬片中的疱疹样病毒颗粒:(7)对大脑皮层的神经元数目、神经原纤维缠结数和突触数密度进行计数。结果 HE染色发现病毒感染组小鼠大脑皮质神经元着色较浅,数目明显减少,神经元发生退行性病变;尤其是海马区神经细胞脱失,数目减少,胞质疏松,细胞层数减少。刚果红染色发现病毒感染组小鼠大脑皮质中有淀粉样斑块。镀银染色发现小鼠大脑皮质中有神经原纤维缠结和淀粉样斑块。间接免疫荧光检测发现病毒感染组小鼠大脑皮质中有磷酸化Tau蛋白沉积,呈团块状分布。透射电镜发现在小鼠大脑皮质神经元超微结构出现明显病理改变,虽未找到疱疹样病毒颗粒,但在核内发现双股螺旋细丝、膜性环状物和长线形丝状物。体外共培养激活后,
[Abstract]:Objective to establish an old mouse model of congenital latent infection of human cytomegalovirus (HCMV), and to observe the AD like pathological changes in cerebral cortex of mice caused by congenital latent infection of human cytomegalovirus (HCMV). To explore the possible role of congenital latent infection of HCMV in the pathogenesis of AD and to provide a new idea and a suitable animal model for the study of the etiology and pathogenesis of AD. Methods six mice in HCMV congenital latent infection group and six mice in DMEM group were randomly selected. According to the experimental design, the mice were killed and the cerebrum tissues of the mice were removed aseptically. Some of them were fixed with paraformaldehyde, and paraffin sections were made routinely. Part of the specimens were fixed with glutaraldehyde. The other brain tissues were cocultured and activated in vitro. The paraffin sections were used to detect the brain tissue architecture of mice by using the following test: 1. Pathological changes and biological characteristics) Detection of amyloid plaques in brain tissue by Congo red staining (Ag-plated staining) Detection of neurofibrillary tangles and amyloid plaques in brain tissue) indirect immunofluorescence detection of phosphorylation in brain tissue Observation of ultrastructural changes of mouse brain neurons by transmission electron microscope (Tau protein 5) in vitro co-culture activation experiment observation on the expression of HCMV IE DNA and UL83 DNA and their corresponding mRNA expression 3 indirect immunofluorescence in CPE 2PCR and RT-PCR, respectively The transmission electron microscopy of HCMV IE and pp65 protein 4 in the cell climbing tablets was used to observe the number of neurons in the cerebral cortex induced by Herpesvirus particles: 7). The number of neurofibrillary tangles and synaptic number density were counted. Results HE staining showed that the number of neurons in cerebral cortex of mice infected with virus was light, the number of neurons decreased and the number of neurons decreased, especially in hippocampal area, the number of neurons was decreased, the cytoplasm was loose and the number of cell layers was decreased. Congo red staining revealed amyloid plaques in the cerebral cortex of mice infected with virus. Silver plated staining revealed neurofibrillary tangles and amyloid plaques in the cerebral cortex of mice. Indirect immunofluorescence assay showed that phosphorylated Tau protein was deposited in the cerebral cortex of mice infected with virus and distributed as a mass. The ultrastructure of neurons in the cerebral cortex of mice was observed by transmission electron microscope. Although no herpes like virus particles were found in the nucleus, double helical filaments, membranous rings and long linear filaments were found in the nucleus. After in vitro co-culture activation,
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2005
【分类号】：R361

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