年龄相关的BubR1水平下降对成体海马神经发生受损的研究

发布时间：2018-05-17 10:19

本文选题：BubR1 + 衰老　；参考：《吉林大学》2017年博士论文

【摘要】：目的:衰老可以引起成年海马神经发生明显下降,导致认知功能障碍。年龄是伴有认知功能受损的神经退行性障碍的主要危险因素。无论人类还是小鼠,海马都是体内两处神经起源的小生境之一,在整个生命过程中神经干细胞在这里持续生成新生神经元,这个过程叫做成体海马神经发生。实验发现衰老过程中成体海马神经发生存在缺陷,这种缺陷被认为是年龄相关认知功能障碍的基础。然而,是什么样的分子机制支配着这种损害我们并没有完全知晓。新近的证据表明自然的衰老过程中伴随着有丝分裂检验点激酶BubR1水平的下降,BubR1水平的下降导致嵌合体杂色非整倍性综合征的小鼠模型或患儿出现早衰的特点。作为有丝分裂检查点激酶,BubR1已经成为一种与年龄相关疾病和整个生命过程中关键的遗传因素。BubR1是否会造成年龄相关的成体海马神经发生的缺陷还需要进一步确认。方法:本课题拟设三个部分进行实验研究。第一部分通过体内免疫组化实验明确小鼠不同年龄段齿状突中的BubR1的表达,从而明确其具体表达位置;通过其在各类型细胞中的表达明确其年龄依赖性的变化;此外还通过体外免疫组化实验明确BubR1有丝分裂后期的角色变化,并明确其在新生神经元的表达位置。在实验第二部分中通过免疫组化染色和腹腔注射Ed U的方法明确BubR1对于神经元前体细胞增殖过程的影响,同时明确BubR1H/H小鼠成熟新生神经元的数量的变化是否单独由神经元前体细胞增殖数量下降导致。在实验的第三部分中,基于前两部分结果,使用逆转录病毒注射、腹腔注射Ed U及免疫组化染色的方法研究BubR1是否损伤了神经元的成熟以及BubR1对于神经元的成熟和树突形态发生的影响。结果:在第一部分实验中:BubR1在出生后的小鼠齿状突中表达,其在颗粒下层的表达水平要稍高于齿状颗粒层。此外,BubR1还在放射状胶质样神经干细胞表达,同样其水平会以年龄依赖的方式降低。体外免疫组化染色实验显示BubR1集中表达在细胞质和树突。在第二部分实验中:我们发现在BubR1H/H小鼠中活跃期的放射状胶质样神经干细胞、中间祖细胞和成神经细胞的数量是明显减少的。然而,虽然在BubR1H/H小鼠中静止期的放射状胶质样神经干细胞轻度减少的。还发现BubR1H/H小鼠新生神经元数量以年龄依赖方式急剧减少。显而易见的是,在胚胎14天至生后7天之间细胞增殖没有受到BubR1的表达水平下降的影响。此外,BubR1H/H小鼠齿状突内成熟新生神经元细胞密度明显下降。然而,幸存的新生神经元数量没有受到BubR1缺陷的影响。在第三部分的试验中,我们发现BubR1H/H小鼠非成熟神经元数量有显著性上升,而成熟神经元数量同期是明显下降的。与此同时,还发现sh RNA-BubR1表达组与其对照组相比GFP+新生神经元起始树突长度、树突总长度缩短,树突分支点数量减少。此外,这些在BubR1低表达鼠树突形态上的变化(不包括树突总长度)在BubR1过度表达鼠上明显改善挽救。结论:大量数据表明海马神经发生的年龄相关性缺陷造成了认知水平下降。BubR1缺陷对活跃期的放射状胶质样神经干细胞、中间祖细胞和成神经细胞的损伤是高度易损的,而静止期的放射状胶质样神经干细胞是轻度受损的。其对在体小鼠调控神经元前体增殖过程起到了必要的和成年特有角色的作用。BubR1H/H成年小鼠海马神经发生的减少主要是由于神经元前体细胞增殖数量的减少而并非是其影响了细胞的存活。BubR1对于神经元的成熟和树突形态发生是必要的,而且这些结果也支持BubR1在体内有丝分裂后期的功能及其关键作用。由此我们认定BubR1可以作为一种新发现的关键性的调控因子控制成年海马神经发生的顺序过程,它可能是调控年龄相关海马病理状态的关键介质。
[Abstract]:Objective: aging can cause a significant decline in adult hippocampal neurogenesis, leading to cognitive impairment. Age is a major risk factor for neurodegenerative disorders associated with impaired cognitive function. Both in humans and in mice, the hippocampus is one of the two niches in the body of the nerve origin in the body, and the neural stem cells are held here throughout the life process. This process is called adult hippocampal neurogenesis. Experiments have found defects in adult hippocampal neurogenesis during aging. This defect is considered to be the basis of age related cognitive impairment. However, what kind of molecular mechanism dominates this damage is not fully known. The new evidence table is not fully known. The decline in the level of mitosis checkpoint kinase BubR1 in the natural aging process, the decrease of BubR1 level leads to the characteristics of premature failure in mice model of chimerism aneuploidy syndrome or in children. As mitosis checkpoint kinase, BubR1 has become a kind of age related disease and the whole life process. The genetic factor.BubR1 of the key will further confirm the defects of the age-related adult hippocampal neurogenesis. Methods: three parts of the subject were set up to carry out the experimental study. The first part is to clarify the expression of BubR1 in the odontoid process of different age segments of the mice by the body immunization test, so as to clarify the specific expression position. Through the expression of its expression in various types of cells, the changes in age dependence are clearly defined. In addition, the role changes in the late mitosis of BubR1 are determined by in vitro immunohistochemical test, and its expression in the newborn neurons is clearly defined. In the second part of the experiment, the method of immunohistochemistry and intraperitoneal injection of Ed U is used to clarify BubR1 for the God. In the third part of the experiment, a retrovirus injection, an intraperitoneal injection of Ed U and immunohistochemical staining were used in the third part of the experiment. Whether BubR1 damage the maturation of neurons and the effect of BubR1 on the maturation of neurons and the morphogenesis of dendrites. Results: in the first part of the experiment, BubR1 was expressed in the odontoid process of the mice after birth, and the expression level in the subgranular layer was slightly higher than that of the dentate granular layer. In addition, the expression of BubR1 was also expressed in the radiate glial neural stem cells. The same level would be reduced in age dependent manner. In vitro immunohistochemical staining showed that BubR1 was concentrated in cytoplasm and dendrites. In the second part of the experiment, we found that the number of radial glial neural stem cells in the active phase of BubR1H/H mice, the number of intermediate progenitor cells and neurocytes, was significantly reduced. However, there was a slight decrease in the radiative glial neural stem cells of the stationary phase in the BubR1H/H mice. It was also found that the number of new neurons in the BubR1H/H mice decreased rapidly in the age dependent manner. It was obvious that the proliferation of cells between 14 days and 7 days after birth was not affected by the decrease of BubR1 expression level. In addition, the BubR1H/H mice teeth. However, the number of surviving new neurons was not affected by BubR1 defects. In the third part of the experiment, we found that the number of immature neurons in BubR1H/H mice increased significantly, while the number of mature neurons decreased significantly in the same period. At the same time, the number of SH RNA was also found. Compared with the control group, the length of the initial dendrites of the new GFP+ neurons, the total length of the dendrites and the number of branches of the dendrites decreased. In addition, these changes in the dendrite morphology of the BubR1 low expression rat (excluding the total dendrite length) were obviously improved and saved on the BubR1 overexpressed rats. Conclusion: a large number of data indicate the occurrence of hippocampal neurogenesis. Age related defects cause a decline in cognitive levels of.BubR1 defects in the active stage of radiative glial neural stem cells, intermediate progenitor cells and neurocytes, which are highly vulnerable to damage, while static glia like neural stem cells are slightly damaged. The decrease in the hippocampal neurogenesis of.BubR1H/H adult mice is mainly due to the decrease in the number of neuronal precursors, not the effect of the survival of the cells on the survival of the cells, which is necessary for the maturation and dendrite morphogenesis of neurons, and the results also support BubR1 in the body. The function and key role of the anaphase, thus we have identified that BubR1 can be a key regulatory factor in the control of the sequential process of adult hippocampal neurogenesis, which may be the key medium for regulating the pathological state of the age related hippocampus.
【学位授予单位】：吉林大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R749.1

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