noggin基因修饰神经干细胞移植治疗衰老模型小鼠的实验研究

发布时间：2018-05-26 13:32

本文选题：神经干细胞 + 基因转染　；参考：《第三军医大学》2006年博士论文

【摘要】： 阿尔茨海默病(Alzheimer disease,AD)是一种中老年人常见的中枢神经退行性疾病,主要表现为进行性痴呆、记忆和认知功能减退,其患病率随年龄增长而上升。随着世界人口日趋老龄化,AD已经成为继心血管疾病、肿瘤和脑卒中之后的第4位死亡原因,当前AD已成为老年医学的重要研究课题。在神经退行性疾病中,神经元的变性、坏死是其功能障碍的主要原因,因此采取有效措施来补充和取代已死亡的神经元,并恢复其功能,将是治疗神经退行性疾病的重要途径。已有研究表明,神经干细胞(Neural stem cells,NSCs)具有自我更新能力,能够分化成神经元和胶质细胞。利用NSCs植入受损的神经组织,替代和修复受损的神经元,以恢复大脑的正常功能,是未来治疗AD的一个重要方向。目前,利用NSCs治疗AD的研究尚处于初步阶段。已经证实NSCs不仅存在于发育中的神经系统,也持续存在于成年中枢神经系统多个脑区。近年来对于NSCs在神经系统疾病应用方面作了大量的研究,然而真正有效的策略是直接移植或作为基因治疗的载体细胞。但移植的NSCs成活率低,分化为神经元的比例非常低,因此治疗作用非常有限。目前需要解决体外培养NSCs保持其强大的增殖能力和诱导其定向分化的问题。已有研究表明,noggin基因在体内外均具有明显的诱导作用,能够促进NSCs的定向分化,为了提高移植NSCs的治疗效果,我们用noggin基因转染NSCs后移植入海马内,利用NSCs自身的修复和noggin基因定向诱导的双重作用,给予衰老小鼠的海马补充有一定功能的神经元,以达到更好的治疗效果。在本实验中,我们通过基因重组技术构建了表达noggin基因的重组腺病毒,通过病毒上清感染培养的NSCs,将表达noggin的NSCs移植入衰老模型小鼠的海马内,观察noggin修饰的NSCs是否对衰老小鼠具有一定的治疗作用,并与仅含GFP的对照病毒pAdEasy-1-GFP感染的NSCs移植的疗效进行比较,实验结果如下: 1.成功构建表达noggin基因的重组腺病毒载体pAdEasy-1-noggin和对照病毒载体pAdEasy-1-GFP,并鉴定其在靶细胞中表达; 2.分离、培养并鉴定新生昆明小鼠NSCs;
[Abstract]:Alzheimer disease (AD) is a common degenerative disease of central nervous system in middle and old people. The main manifestations of AD are progressive dementia, memory and cognitive impairment, and the prevalence rate increases with age. With the aging of the world population, AD has become the fourth leading cause of death after cardiovascular disease, tumor and stroke. At present, AD has become an important research topic in geriatrics. In neurodegenerative diseases, degeneration and necrosis of neurons are the main causes of dysfunction. Therefore, it is an important way to treat neurodegenerative diseases to take effective measures to supplement and replace the dead neurons and to restore their functions. It has been shown that neural stem cells have the ability of self-renewal and can differentiate into neurons and glial cells. The use of NSCs to implant damaged nerve tissue to replace and repair damaged neurons in order to restore normal brain function is an important direction in the treatment of AD in the future. At present, the treatment of AD with NSCs is still in the preliminary stage. It has been proved that NSCs exists not only in the developing nervous system, but also in multiple brain regions of the adult central nervous system. In recent years, a lot of research has been done on the application of NSCs in nervous system diseases. However, the effective strategy is to transplant directly or as vector cells for gene therapy. However, the survival rate of transplanted NSCs is low and the proportion of differentiated neurons is very low, so the therapeutic effect is very limited. At present, it is necessary to solve the problem that NSCs can maintain its strong proliferative ability and induce its directional differentiation in vitro. Previous studies have shown that the gene can induce the directional differentiation of NSCs in vitro and in vivo. In order to improve the therapeutic effect of NSCs transplantation, noggin gene was transfected into NSCs and implanted into hippocampus. By using the dual effects of NSCs repair and directed induction of noggin gene, the hippocampal neurons of aging mice were supplemented with some functional neurons, so as to achieve a better therapeutic effect. In this experiment, we constructed recombinant adenovirus expressing noggin gene by gene recombination technique. NSCs expressing noggin was transplanted into hippocampus of aging model mice by infection of NSCs with virus supernatant. To observe whether noggin modified NSCs has a certain therapeutic effect on aging mice, and to compare the effect of NSCs transplantation with GFP only virus pAdEasy-1-GFP infection. The results are as follows: 1. The recombinant adenovirus vector pAdEasy-1-noggin expressing noggin gene and the control vector pAdEasy-1-GFPwere successfully constructed and expressed in target cells. 2. The NSCs of newborn Kunming mice were isolated, cultured and identified.
【学位授予单位】：第三军医大学
【学位级别】：博士
【学位授予年份】：2006
【分类号】：R-332

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