發(fā)布時間：2018-03-20 07:19

  本文選題：堿性螺旋-環(huán)-螺旋　切入點：神經(jīng)源性分化因子　出處：《福建醫(yī)科大學》2008年碩士論文　論文類型：學位論文

【摘要】： 目的 探討神經(jīng)源性分化因子(Neurogenic Differentiation,NeuroD)在大鼠神經(jīng)系統(tǒng)發(fā)育過程中,表達量及部位的變化情況,進而研究其對神經(jīng)分化的影響。 方法 利用神經(jīng)細胞標志分子NSE和星形膠質(zhì)細胞標記分子GFAP,對發(fā)育不同時期的大鼠腦組織進行NeuroD與NSE、GFAP的共標記免疫熒光染色,激光共聚焦顯微鏡檢測蛋白在鼠腦中的分布情況;利用蛋白質(zhì)免疫印跡(Western Blot)對NeuroD蛋白在各腦組分和不同發(fā)育階段的表達進行半定量分析,利用RT-PCR方法檢測NeuroD mRNA在大鼠腦不同發(fā)育時段的表達變化,進而確定NeuroD在鼠腦發(fā)育過程中表達的時空特異性。 結(jié)果 免疫熒光染色結(jié)果顯示:NeuroD蛋白(綠)分布區(qū)域與神經(jīng)元(NSE)(紅)相重合,與星形膠質(zhì)細胞(GFAP)(紅)部分相符。NeuroD主要分布于大腦皮質(zhì),其次在小腦也有部分表達,在間腦內(nèi),NeuroD在側(cè)腦室附近見有多量表達。蛋白質(zhì)免疫印跡結(jié)果顯示,NeuroD在腦組織各組分中均有表達,其中以大腦皮質(zhì)表達最為豐富,其次為間腦和小腦。NeuroD蛋白在大鼠腦發(fā)育的不同時期都有表達,從神經(jīng)管發(fā)生的E9.5天到腦形態(tài)初步建立的E12.5天,NeuroD的表達呈現(xiàn)明顯升高,E12.5天達到最高峰,在胚胎發(fā)育的晚期,腦組織形態(tài)得到進一步的完善,在E21.5天腦組織各部分進一步成熟,此時NeuroD達到第二次高峰,而出生后NeuroD表達無明顯的差異。NeuroD mRNA在E7.5天時即開始有表達,目的基因和β-actin在E10.5天至E12.5天之間先后達到高峰,出生前(E21.5天)形成第二次表達高峰,RT-PCR結(jié)果與Western-blotting結(jié)果基本一致。 結(jié)論 在不同發(fā)育時期的大鼠腦組織中,NeuroD的表達在時間和空間上呈現(xiàn)出明顯的差異性:在神經(jīng)系統(tǒng)發(fā)育早、中期,即從E9.5天神經(jīng)管閉合至E12.5天形成前腦、中腦和后腦的發(fā)育原基,和胚胎發(fā)育的后期,即E21.5天腦組織各部分進一步成熟,腦組織形態(tài)得到進一步的完善,NeuroD的表達均有明顯升高,而出生后NeuroD表達無明顯的差異,推測NeuroD的主要作用有:1、對神經(jīng)細胞的早期形成、分化起作用;2、在神經(jīng)系統(tǒng)成熟階段對神經(jīng)細胞間的突觸聯(lián)系的進一步完善也有一定作用;3、NeuroD在側(cè)腦室附近也見有明顯表達,推測其可能與室管膜上皮下的神經(jīng)干細胞遷移分化有關(guān),還有待進一步實驗證實。
[Abstract]:Purpose. To investigate the changes of neurogenic differentiation factor NeuroDduring the development of rat nervous system, and to study the effect of neurogenic differentiation on neurogenic differentiation. Method. NSE and GFAP were used to detect the distribution of NeuroD and NSE GFAP in the brain of rats at different stages of development. The distribution of protein in the brain was detected by confocal laser microscopy. The expression of NeuroD protein in different brain components and different developmental stages was semi-quantitatively analyzed by Western blotting. The expression of NeuroD mRNA in different developmental stages of rat brain was detected by RT-PCR method. Furthermore, the spatiotemporal specificity of NeuroD expression during brain development was determined. Results. The results of immunofluorescence staining showed that the distribution of neuroD protein (green) coincided with that of NSEP (red), and that the neuroD was mainly distributed in the cerebral cortex, and was also partially expressed in the cerebellum. In diencephalon, neuroD was expressed in the vicinity of lateral ventricle. Western blot showed that NeuroD was expressed in all components of brain, especially in cerebral cortex. Secondly, diencephalon and cerebellar neuroD protein were expressed at different stages of brain development in rats. The expression of NeuroD increased significantly from the day of neurotubule development to the day 12. 5, and reached the peak at day 12. 5, and reached the peak at the late stage of embryonic development. The morphology of brain tissue was further improved, and the parts of brain tissue matured further on the day of E21.5, at which NeuroD reached the second peak, but there was no significant difference in the expression of NeuroD after birth. NeuroD mRNA began to express at E7.5. Objective the gene and 尾 -actin reached the peak between E10.5 and E12.5 days, and the second peak expression of 尾 -actin was observed on E21.5 days before birth. The results of RT-PCR were consistent with the results of Western-blotting. Conclusion. The expression of NeuroD in brain tissues of rats at different developmental stages showed significant difference in time and space: the developmental primordia of forebrain, midbrain and hindbrain were formed in the early stage of nervous system development, in middle stage, i.e. from day 9.5 to day 12.5. The expression of neuroD was significantly increased in the later stage of embryonic development, i.e. the maturation of all parts of brain tissue at E21.5 day, but there was no significant difference in the expression of NeuroD after birth. It is speculated that NeuroD plays a major role in the early formation and differentiation of nerve cells, and in the further improvement of synaptic connections between nerve cells in the mature stage of the nervous system. It may be related to the migration and differentiation of neural stem cells under ependymal subcutaneous.
【學位授予單位】：福建醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2008
【分類號】：R329.21

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1 郭瑋;NeuroD在大鼠腦發(fā)育過程中的差異性表達[D];福建醫(yī)科大學;2008年

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本文編號：1638040