【作者】 董素珍；

【导师】 胡应和；

【作者基本信息】 华东师范大学 ， 生理学， 2006， 博士

【摘要】 老年痴呆(Alzheimer’s disease，AD)是老年人中最普遍的一种以脑内Aβ水平显著升高为特征的中枢神经系统退行性疾病，早老素基因(Presenilin，PS)在AD病理学中起重要作用，揭示其生理功能对阐明AD的具体分子机制具有重要意义。早老素是跨膜的天冬氨酸蛋白酶，有PS1和PS2两种形式，基因敲除研究表明，条件性PS1和PS2双敲除(PS conditional knock out，cDKO)小鼠表现出类似AD的进行性的神经退行性病理变化，包括认知能力衰退和前脑退化等，但是这些病理症状不依赖于脑内Aβ水平，其确切的分子机理还未阐明。本研究采用基因芯片和定量PCR技术分别考察3月龄和10月龄cDKO小鼠皮层和海马内发生的基因表达变化，分析差异表达基因和相关的信号通路，初步探讨cDKO小鼠发生退行性病变的分子机理。 复杂环境处理(Environmental enrichment，EE)是把动物以更大的群体放在一个更大的、放有多种玩具的笼子或其他饲养环境里，使动物有更大的机会进行复杂的社会性交流和探索的一种处理方式。EE已被证实能改善多种正常和认知能力缺陷动物的学习记忆能力，诱导动物脑部的形态学变化，还对许多神经退行性疾病及其他病理过程具有良好的延缓和逆转作用。为了考察EE是否能减轻cDKO小鼠的记忆缺陷和前脑退化，本实验对1月龄的cDKO小鼠进行5个月的每天3小时的复杂环境暴露，然后进行行为学和组织学检测，以考察EE诱导的变化。接着运用高密度基因芯片技术、荧光实时定量PCR技术、免疫组化以及Western blotting等分子生物学技术考察EE引起的基因表达变化，初步探讨EE对cDKO小鼠的生理效果的分子机理，同时为开发AD的治疗方案提供一些理论基础。 1．PS1／PS2条件性双敲除小鼠神经退行性早期基因表达谱分析 运用基因芯片技术考察了cDKO小鼠3月龄时皮层和海马基因表达的变化，分别鉴别到56和46个基因的表达发生了改变。用定量PCR技术检测了一些重要基因的表达，其结果与芯片结果相符。功能分析揭示，这些差异表达基因主要可以归入免疫和炎症、代谢、细胞黏附、信号转导、结构蛋白和肿瘤相关基因等几类。其中，免疫炎症相关的基因包括补体成员和Cathepsin家族蛋白酶的表达显著上调。此外，突触后受体AMPA受体亚基GluR1和GABA受体α亚基Gabra2及蛋白激酶Mapk8等都与AD神经病理学有关，它们的基因在cDKO小鼠海马内表更多还原

【Abstract】 Alzheimer’s disease is an age-related neurodegenerative disease characterized by significant elevation of Aβ level in patients’ brain. Presenilins (PS) play an important role in AD pathology, so it is of great importance to identify the function of PS to elucidate the accurate molecular mechanism of AD. PS is a transmembrane aspartic protease. There are two PS genes, PS1 and PS2. Mice lacking both PS1 and PS2 (cDKO) in the forebrain showed AD-like progressive neurodegenerative disease phenotypes, including age-dependent memory decline and forebrain degeneration, which were independent of Aβ level. However, the molecular mechanism of the neurodegenerative phenotype in PS cDKO mice is to be elucidated. The gene expression profiles of the cortex and hippocampus of 3- and 10-month cDKO mice were examined by oligonucletide microarray and Real-time PCR techniques to identify the related genes and signaling pathways.The classical paradigm of the environmental enrichment (EE) involves placing animals in large cages that contain running wheels, colorful tunnels, and assorted toys. It has been well established that EE can enhance the memory of normal and memory-deficient animals, induce brain structural changes and delay onset and progression of several neurodegenerative animal models. In this study, we exposed one-month-old cDKO mice to EE 3 hr daily for 5 months, then tested the effects using behavioral and histological approaches, and analyzed the EE-induced gene expression changes in the cDKO brain using the high-intensity microarray, fluorescence real-time quantitative PCR, immunohistology and western blotting technologies to explore the underlying mechanism of the effects of EE, which might provide some clues for AD therapeutics.1. Gene expression changes in the forebrains of PS cDKO mice at early peroid of neurodegeneration.Fifty-six and forty-six differently expressed genes were identifitied in the cortex and hippocampus of cDKO mice using microarray techniques. Expression changes of更多还原