本文選題：TRIM32 + 放射膠質(zhì)細(xì)胞��； 參考：《南方醫(yī)科大學(xué)》2017年碩士論文

【摘要】：研究背景大腦皮層的發(fā)育(cortex development)是一個復(fù)雜而又精細(xì)的過程。在嚙齒類動物中,神經(jīng)元主要在胚胎發(fā)育的E12到E18之間產(chǎn)生,E15時達(dá)到高峰,出生后停止;神經(jīng)膠質(zhì)細(xì)胞在E18天左右開始產(chǎn)生,在新生期達(dá)到高峰。在腦室區(qū)域(ventricular zone,VZ)的背側(cè),分布著一群放射狀膠質(zhì)細(xì)胞(Radial glial cells,RGCs)的細(xì)胞,這群細(xì)胞通過增殖分化產(chǎn)生了皮層的興奮性椎體神經(jīng)元。RGCs在自我更新的同時,遷移分化為椎體神經(jīng)元,組成分為六層的腦皮層。TRIM32(Tripartite motif 32)是一種具有E3泛素鏈接酶活性的蛋白,其蛋白結(jié)構(gòu)中具有TRIM家族特征性的鋅指結(jié)構(gòu)域、B-boxes結(jié)構(gòu)域和卷曲結(jié)構(gòu)域,以及特征性的6個重復(fù)性的NHL結(jié)構(gòu)域。TRIM32參與了某些中樞神經(jīng)系統(tǒng)的疾病發(fā)生,例如:阿爾茨海默病(Alzhermer' s Disease,AD)、注意力缺陷多動癥(Attention Deficit Hyperacitivity Disorder,ADHD),我們前期研究發(fā)現(xiàn),TRIM32高表達(dá)于小鼠成熟的大腦皮層神經(jīng)元以及胎腦的MGE區(qū)中間神經(jīng)前體細(xì)胞中,TRIM32的缺失能夠?qū)е轮虚g神經(jīng)節(jié)隆起(Medial ganglionic eminence,MGE)區(qū)域的中間神經(jīng)前體細(xì)胞增殖分裂減少,進(jìn)而導(dǎo)致了 γ-氨基丁酸能(GABAergic)抑制性神經(jīng)元的減少,從而引起了小鼠自閉癥樣的行為學(xué)表現(xiàn)。本研究主要探討的是TRIM32在調(diào)控皮層興奮性神經(jīng)元的產(chǎn)生過程中的作用。研究目的1.TRIM32基因在小鼠大腦皮層細(xì)胞表達(dá)情況的研究;2.TRIM32的遺傳缺失對小鼠胚胎時期大腦皮層神經(jīng)元產(chǎn)生的影響研究;3.TRIM32的遺傳缺失對小鼠胚胎時期皮層VZ/SVZ區(qū)神經(jīng)前體細(xì)胞產(chǎn)生的影響;4.TRIM32的遺傳缺失影響小鼠胚胎時期皮層VZ/SVZ區(qū)神經(jīng)前體細(xì)胞產(chǎn)生的機(jī)制研究。研究方法1.TRIM32基因在小鼠大腦皮層細(xì)胞表達(dá)情況野生型小鼠交配后母鼠孕第E13.5、E16.5、E18.5時取大腦標(biāo)本,切片后進(jìn)行TRIM32熒光染色。2.TRIM32的遺傳缺失對小鼠皮層成熟神經(jīng)元產(chǎn)生的影響DAPI染色比較皮層CNx和CP的厚度,然后通過大腦各皮層神經(jīng)元的特異性marker來標(biāo)記相應(yīng)神經(jīng)元,計(jì)數(shù)統(tǒng)計(jì)后比較差異。3.TRIM32的遺傳缺失對小鼠胚胎時期皮層VZ/SVZ區(qū)神經(jīng)前體細(xì)胞產(chǎn)生的影響PAX6、TBR2熒光染色標(biāo)記皮層VZ/SVZ區(qū)神經(jīng)前體細(xì)胞,計(jì)數(shù)統(tǒng)計(jì)后比較差異。4.TRIM32的缺失對神經(jīng)前體細(xì)胞分裂、增殖的影響Edu和PH3熒光分別標(biāo)記增殖和分裂的細(xì)胞,計(jì)數(shù)統(tǒng)計(jì)后比較差異。5.TRIM32的缺失對皮層神經(jīng)細(xì)胞凋亡的影響Caspase3熒光染色標(biāo)記凋亡狀態(tài)細(xì)胞,計(jì)數(shù)統(tǒng)計(jì)后比較差異。6.統(tǒng)計(jì)學(xué)分析統(tǒng)計(jì)結(jié)果采用均數(shù)±標(biāo)準(zhǔn)差(x± s)表示,采用SPSS20.0軟件結(jié)果,以P0.05認(rèn)為有統(tǒng)計(jì)學(xué)差異。研究結(jié)果1.TRIM32基因在小鼠大腦皮層細(xì)胞表達(dá)情況E13.5時,TRIM32主要表達(dá)在VZ與SVZ區(qū)細(xì)胞的胞質(zhì)中,E16.5時TRIM32高表達(dá)于IZ區(qū)細(xì)胞,而到E18.5時,TRIM32高表達(dá)于TBR1陽性成熟神經(jīng)元胞核。2.TRIM32的遺傳缺失導(dǎo)致了皮層厚度縮小以及小鼠皮層成熟神經(jīng)元數(shù)量減少E18.5 時,TRIM32-/-胎鼠皮層 NCx 和 CP 的均縮小(NCx:t=-4.709,P=0.026;CP:t=-7.634,P=0.011);E14.5,E16.5 和 E18.5 三個時期,TRIM32-/-胎鼠 TBR1陽性神經(jīng)元數(shù)量均減少,有統(tǒng)計(jì)學(xué)差異(E14.5,t=-4.433,P=0.032;E16.5,t=-5.549,P=0.022;E18.5,t=-7.793,P=0.021);E16.5和 E18.5 兩個時期,TRIM32-/-胎鼠BCL11b陽性神經(jīng)元數(shù)量均減少,有統(tǒng)計(jì)學(xué)差異(E16.5,t=-6.691,P=0.037;E18.5,t=-12.047,P=0.036);E18.5 和 P30 兩個時期,TRIM32-/-胎鼠 CUX1 陽性神經(jīng)元數(shù)量明顯減少,有統(tǒng)計(jì)學(xué)差異(E18.5,t=-7.622,P=0.047;P30,t=33.956,P=0.014)。3.TRIM32的遺傳缺失導(dǎo)致神經(jīng)前體細(xì)胞的數(shù)量減少E14.5和E16.5兩個時期,TRIM32-/-胎鼠VZ/SVZ區(qū)PAX6和TBR2陽性的神經(jīng)前體細(xì)胞數(shù)量減少,有統(tǒng)計(jì)學(xué)差異(PAX6,E14.5,t=9.125,P=0.018;E16.5,t=13.230,P=0.018;TBR2,E14.5,t=7.143,P=0.028,E16.5,t=12.548,P=0.024)。4.TRIM32的遺傳缺失導(dǎo)致神經(jīng)前體細(xì)胞增殖、分裂能力減弱E14.5,E16.5時,TRIM32-/-胎鼠Edu陽性細(xì)胞數(shù)目減少,有統(tǒng)計(jì)學(xué)差異(E14.5,t=14.061,P=0.000;E16.5,t=8.715,P=0.038)。E14.5時,TRIM32-/-胎鼠basal區(qū)和apical區(qū)PH3陽性細(xì)胞數(shù)減少,有統(tǒng)計(jì)學(xué)差異(basal 區(qū):t=16.234,P=0.000;apical 區(qū):t=26.415,P=0.000)。E16.5時,TRIM32-/-胎鼠basal區(qū)PH3陽性細(xì)胞數(shù)減少,但差異不顯著(t=1.784,P=0.231),apical區(qū)細(xì)胞減少差異仍然有顯著性(t=4.051,P=0.022)。5.TRIM32的遺傳缺失對大腦皮層神經(jīng)細(xì)胞的凋亡無影響E14.5,E18.5時,TRIM32-/-與TRIM32+/+胎鼠之間Caspase3陽性細(xì)胞數(shù)目無學(xué)統(tǒng)計(jì)差異(E14.5,t=0.083,P=0.938;E18.5,t=-1.350,P=0.283)。
[Abstract]:Cortex development is a complex and fine process. In rodents, neurons are produced mainly between E12 and E18 from embryonic development. E15 reaches the peak and stops after birth; glial cells begin to produce at about E18 days and peak in the newborn period. In the ventricle region (ventricula) The back side of R zone, VZ, distributed in a group of cells of Radial glial cells, RGCs, which proliferate and differentiate to produce cortical excitatory vertebral neurons.RGCs, while self renewal, migration and differentiation into vertebral neurons, and the composition of the six layers of cerebral cortex.TRIM32 (Tripartite motif 32) is a kind of E3. Ubiquitin linked enzyme activity protein with TRIM family characteristic domain of zinc finger, B-boxes domain and curl domain, and 6 repetitive NHL domain.TRIM32, characterized by the disease in some central nervous systems, such as Alzheimer's disease (Alzhermer's Disease, AD), and attention deficit Attention Deficit Hyperacitivity Disorder (ADHD), our previous study found that TRIM32 was highly expressed in the mature cerebral cortex neurons in mice and the intermediate nerve precursor cells in the MGE region of the fetal brain. The deletion of TRIM32 could lead to the increase of the intermediate ganglion (Medial ganglionic eminence, MGE) region of the intermediate nerve precursor cells. The decrease of the colonization resulted in the reduction of the inhibitory neurons of gamma aminobutyric acid (GABAergic), which resulted in the behavioral performance of the autistic mice. This study mainly discussed the role of TRIM32 in the regulation of the production of cortical excitatory neurons. The purpose of the study was to express the expression of 1.TRIM32 gene in the cerebral cortex cells of mice. A study of the effects of the genetic deletion of 2.TRIM32 on the production of cortical neurons in the embryonic stage of the mouse; the effect of the genetic deletion of 3.TRIM32 on the production of neural precursor cells in the VZ/SVZ region of the mouse embryo; the genetic deletion of 4.TRIM32 affects the mechanism of the production of neural precursor cells in the VZ/ SVZ region of the mouse embryo. The expression of 1.TRIM32 gene in the mouse cerebral cortex cells was expressed in the cerebral cortex cells of mice. The brain specimens were taken at E13.5, E16.5, and E18.5 after mating in the wild type mice. The effects of the genetic deletion of TRIM32 fluorescent staining.2.TRIM32 on the production of mature neurons in the cortex of mice were compared with the thickness of CNx and CP compared with the cortical CNx and CP, and then through the brain. The specific marker of cortical neurons was used to mark the corresponding neurons. After counting the statistical difference, the effects of the genetic deletion of.3.TRIM32 on the production of neural precursor cells in the VZ/SVZ region of the mouse embryonic period were PAX6, and the TBR2 fluorescence staining was used to mark the neuronal precursor cells in the cortical VZ/SVZ region, and the difference of.4.TRIM32 loss was compared to the neural precursor after the counting system. Cell division and proliferation effects of Edu and PH3 fluorescence labeling of cells with proliferation and division respectively. After counting statistics, the effects of.5.TRIM32 loss on apoptosis of cortical neurons were compared and Caspase3 fluorescent staining was used to mark apoptotic cells. After counting the statistical difference, the statistical results of.6. statistical difference (x + s) were indicated by the statistical difference (x + s). The results of SPSS20.0 software showed that there were statistical differences between P0.05 and 1.TRIM32. When the expression of 1.TRIM32 gene in the cerebral cortex cells of mice was E13.5, TRIM32 was mainly expressed in the cytoplasm of VZ and SVZ cells, and TRIM32 was highly expressed in IZ region cells when E16.5, and TRIM32 was highly expressed in the positive mature neuron nucleus when E18.5. When the cortical thickness was reduced and the number of mature neurons in the cortex was reduced by E18.5, the NCx and CP in the cortex of TRIM32-/- mice were reduced (NCx:t=-4.709, P=0.026; CP:t=-7.634, P=0.011), and the number of positive neurons in E14.5, E16.5 and E18.5 decreased in the three periods. 4.433, P=0.032, E16.5, t=-5.549, P=0.022; E18.5, t=-7.793, P=0.021); E16.5 and E18.5 two periods, the number of BCL11b positive neurons in TRIM32-/- fetal mice decreased. The genetic deletion (E18.5, t=-7.622, P=0.047; P30, t=33.956, P=0.014).3.TRIM32 causes the decrease of the number of neural precursor cells in the two periods of E14.5 and E16.5. The genetic deletion of =7.143, P=0.028, E16.5, t=12.548, P=0.024).4.TRIM32 leads to the proliferation of neural precursor cells, and the division ability weakens the E14.5, E16.5, the number of Edu positive cells in TRIM32-/- fetal mice decreases. Fewer, there were statistical differences (basal region: t=16.234, P=0.000; apical region: t=26.415, P=0.000).E16.5, the number of PH3 positive cells in basal region of TRIM32-/- fetal mice decreased, but the difference was not significant (t=1.784, P=0.231). When E14.5 and E18.5 were affected, there was no statistical difference in the number of Caspase3 positive cells between TRIM32-/- and TRIM32+/+ fetal rats (E14.5, t=0.083, P=0.938, E18.5, t=-1.350, P=0.283).
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R741

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 陳罡;金明;王開宇;薛曉靜;鄭望;丁玉強(qiáng);袁小兵;;大腦皮層神經(jīng)元放射狀遷移導(dǎo)向機(jī)制的新發(fā)現(xiàn)[J];中國基礎(chǔ)科學(xué);2008年02期

2 李小剛,董為偉,范生堯,張志,楊艷,曾曉榮;缺氧對大鼠大腦皮層神經(jīng)元鈣激活性鉀通道的影響[J];卒中與神經(jīng)疾病;2002年04期

3 徐靜;伏·格·依利謝耶夫;;大腦皮層神經(jīng)元的有絲分裂[J];解剖學(xué)報;1958年02期

4 ;大腦皮層神經(jīng)元放射狀遷移的導(dǎo)向機(jī)制新發(fā)現(xiàn)[J];科學(xué)通報;2008年04期

5 應(yīng)水旺;張家駒;郝蘇陽;張建瑜;;微電泳給予褪黑激素對大鼠大腦皮層神經(jīng)元自發(fā)和谷氨酸誘發(fā)放電的影響[J];福建醫(yī)學(xué)院學(xué)報;1988年02期

6 彭博;吳哲;張朝東;;鉛對小鼠大腦皮層神經(jīng)元磷脂酰肌醇3激酶影響[J];中國公共衛(wèi)生;2010年02期

7 鄒哲華;陶陶;徐堅(jiān);劉智;羅開儉;;大鼠大腦皮層神經(jīng)元缺氧后細(xì)胞凋亡情況的動態(tài)觀察[J];第三軍醫(yī)大學(xué)學(xué)報;2012年24期

8 董文成;張燕豐;;納洛酮對針刺穴位抑制大腦皮層神經(jīng)元傷害性反應(yīng)的影響[J];中國針灸;1985年02期

9 王慧娟;郭建國;張金平;張惠欣;趙昱;李陳莉;;大腦皮層神經(jīng)元微管相關(guān)蛋白-2在人胚發(fā)育過程中的表達(dá)變化[J];山東醫(yī)藥;2008年11期

10 李紅;陳曉蓉;趙正梅;思盼盼;張金秋;湯俊峰;;胚鼠大腦皮層神經(jīng)元培養(yǎng)方法的改進(jìn)[J];四川解剖學(xué)雜志;2009年02期

相關(guān)會議論文 前5條

1 裘瑋;李惠明;田毓華;黃倩;;條件復(fù)制型腺病毒輔助的腺相關(guān)病毒對大腦皮層神經(jīng)元的轉(zhuǎn)導(dǎo)[A];第九屆全國腫瘤生物治療學(xué)術(shù)會議論文集[C];2006年

2 楊坤;陳廷福;金先慶;;丙泊酚對發(fā)育中大腦皮層神經(jīng)元細(xì)胞活性及凋亡的影響[A];中華醫(yī)學(xué)會第八次全國小兒外科學(xué)術(shù)會論文集[C];2010年

3 董麗萍;韓明;袁芳;;一組mGluRs反義寡核苷酸抗谷氨酸引起的小鼠大腦皮層神經(jīng)元的興奮毒作用[A];中國神經(jīng)科學(xué)學(xué)會第六屆學(xué)術(shù)會議暨學(xué)會成立十周年慶祝大會論文摘要匯編[C];2005年

4 李妙齡;楊艷;曾曉榮;劉智飛;裴杰;;新生大白鼠大腦皮層神經(jīng)元的培養(yǎng)及基本電生理學(xué)特性研究[A];中國生理學(xué)會第21屆全國代表大會暨學(xué)術(shù)會議論文摘要匯編[C];2002年

5 鮑嵐;;微管蛋白乙�；揎椗c大腦皮層神經(jīng)元發(fā)育[A];細(xì)胞—生命的基礎(chǔ)——中國細(xì)胞生物學(xué)學(xué)會2013年全國學(xué)術(shù)大會·武漢論文摘要集[C];2013年

相關(guān)博士學(xué)位論文 前3條

1 巨興達(dá);肌球蛋白10調(diào)控大腦皮層神經(jīng)元輻射遷移的研究[D];東北師范大學(xué);2013年

2 崔繼紅;漢坦病毒感染體外培養(yǎng)的乳鼠大腦皮層神經(jīng)元誘導(dǎo)凋亡發(fā)生的初步研究[D];第四軍醫(yī)大學(xué);2009年

3 李瑋琦;Dok6在原代培養(yǎng)的小鼠大腦皮層神經(jīng)元神經(jīng)突起生長過程中的功能研究[D];北京協(xié)和醫(yī)學(xué)院;2010年

相關(guān)碩士學(xué)位論文 前4條

1 陳文錦;TRIM32的缺失導(dǎo)致小鼠大腦皮層神經(jīng)元發(fā)生障礙的研究[D];南方醫(yī)科大學(xué);2017年

2 張會春;八肽膽囊收縮素對體外培養(yǎng)大鼠大腦皮層神經(jīng)元的保護(hù)作用研究[D];重慶醫(yī)科大學(xué);2009年

3 楊坤;丙泊酚對發(fā)育中大腦皮層神經(jīng)元細(xì)胞活性及凋亡的影響[D];重慶醫(yī)科大學(xué);2010年

4 唐華非;羥基喜樹堿腦植入緩釋片的神經(jīng)毒性研究[D];第三軍醫(yī)大學(xué);2010年

，

本文編號：1904446