本文選題：轉(zhuǎn)錄激活因子3　切入點(diǎn)：苔蘚纖維出芽　出處：《中南大學(xué)》2014年碩士論文

【摘要】：目的通過觀察戊四氮(pentylenetetrazole; PTZ)大鼠點(diǎn)燃模型海馬CA3區(qū)及齒狀回區(qū)轉(zhuǎn)錄激活因子3(activating transcription factor3; ATF3)的動(dòng)態(tài)表達(dá)變化,結(jié)合海馬苔蘚纖維出芽(mossy fiber sprouting; MFS)的程度,探討其與MFS的關(guān)系及在癲癇過程中的作用。 方法健康SD雄性成年大鼠150只,隨機(jī)分為生理鹽水對(duì)照組(60只)和PTZ實(shí)驗(yàn)組(90只)。實(shí)驗(yàn)組每日腹腔注射PTZ (30mg/kg)；同時(shí)生理鹽水組注射等量生理鹽水。按照首次腹腔注射后第3天,1周,2周,4周和6周的時(shí)間點(diǎn)分配,將PTZ組及生理鹽水組隨機(jī)分為5個(gè)亞組。根據(jù)實(shí)驗(yàn)方法的不同再將每個(gè)亞組分為三個(gè)小組：第一組用于免疫組化及免疫熒光,檢測各時(shí)間點(diǎn)ATF3蛋白在海馬各區(qū)的動(dòng)態(tài)表達(dá)及在細(xì)胞內(nèi)表達(dá)定位；第二組用于Western Blot,將海馬各區(qū)ATF3表達(dá)變化進(jìn)行半定量分析；第三組用于Timm染色,檢測苔蘚纖維出芽情況,其中實(shí)驗(yàn)組每小組各6只,對(duì)照組每小組各4只。 結(jié)果 1.動(dòng)物行為學(xué)表現(xiàn) 大鼠多在連續(xù)注射PTZ21-28天(平均23.7±1.9天)后被點(diǎn)燃。模型點(diǎn)燃率為94.4%,死亡率為5.33%。除了4周和6周組各有一只大鼠未出現(xiàn)癇性發(fā)作外,點(diǎn)燃成功后大鼠出現(xiàn)誘發(fā)發(fā)作及自發(fā)發(fā)作,對(duì)照組未出現(xiàn)癇性發(fā)作。 2.苔蘚纖維出芽評(píng)分 實(shí)驗(yàn)組CA3區(qū)與對(duì)照組相對(duì)應(yīng)時(shí)間點(diǎn)相比MFS評(píng)分均有顯著性差異(P0.05)。從3天起評(píng)分逐漸增加,4周時(shí)達(dá)高峰,至6周仍維持在較高水平。齒狀回區(qū)MFS評(píng)分在實(shí)驗(yàn)組與對(duì)照組之間以及各組內(nèi)不同時(shí)間點(diǎn)之間比較均無顯著性差異(P0.05)。 3.ATF3蛋白的動(dòng)態(tài)表達(dá) 免疫組化示在癲癇點(diǎn)燃過程中,實(shí)驗(yàn)組：海馬CA3區(qū)ATF3蛋白表達(dá)逐漸上升,在4周組達(dá)最高峰,6周組稍有下降(P0.05)；齒狀回區(qū)ATF3蛋白在點(diǎn)燃過程中無明顯變化(P0.05)。對(duì)照組：海馬CA3區(qū)及齒狀回區(qū)在3天到6周組均無明顯變化(P0.05)。Western Blot結(jié)果示海馬ATF3的表達(dá)自3天組逐漸上升,至4周組達(dá)高峰,6周組較前明顯下降；除了3天組之外,其他各時(shí)間點(diǎn)ATF3表達(dá)在實(shí)驗(yàn)組均較對(duì)照組高(P0.05)。免疫熒光結(jié)果示ATF3蛋白定位于大鼠海馬神經(jīng)元細(xì)胞核及細(xì)胞漿中,且主要集中在細(xì)胞漿。 結(jié)論 ATF3蛋白在海馬CA3區(qū)的表達(dá)上調(diào)可能促進(jìn)苔蘚纖維出芽,進(jìn)而導(dǎo)致顳葉癲癇的發(fā)生及發(fā)展。
[Abstract]:Objective to observe the dynamic expression of 3(activating transcription factor 3 (ATF3) in the hippocampal CA3 and dentate gyrus of pentylenetetrazole (PTZ) kindling rat model, and to investigate the extent of mossy fiber sprouting in hippocampal mossy.To explore the relationship between MFS and Epilepsy and its role in epilepsy.Methods 150 healthy male SD rats were randomly divided into normal saline control group (n = 60) and PTZ experimental group (n = 90).The experimental group was intraperitoneally injected with 30 mg 路kg ~ (-1) of PTZ and the same amount of saline was injected into the saline group.According to the time points of 1 week, 2 weeks, 4 weeks and 6 weeks after the first intraperitoneal injection, the PTZ group and the saline group were randomly divided into 5 subgroups.Each subgroup was divided into three groups according to the different experimental methods. The first group was used for immunohistochemistry and immunofluorescence to detect the dynamic expression and localization of ATF3 protein in hippocampal regions at different time points.The second group was used for Western blot.The third group was used for Timm staining to detect the budding of moss fiber, 6 rats in each group in experimental group and 4 in control group.Result1.Animal behavior manifestationMost of the rats were kindled after continuous PTZ21-28 injection (mean 23.7 鹵1.9 days).The model ignition rate was 94. 4% and the mortality was 5. 33%.Except for 4 weeks and 6 weeks group, there was no epileptic seizure in each group, but there was no epileptic attack in the control group after the success of kindling.2.Mossy fiber budding scoreCompared with the control group, the MFS scores in the CA3 area of the experimental group were significantly different from those in the control group (P 0.05).The score gradually increased from 3 days to reach the peak at 4 weeks and remained at a relatively high level at 6 weeks.There was no significant difference in MFS score of dentate gyrus between experimental group and control group and between groups at different time points (P 0.05).Dynamic expression of 3.ATF3 proteinImmunohistochemistry showed that the expression of ATF3 protein in the hippocampal CA3 region increased gradually during epileptic kindling, and decreased slightly in the 4th week group and the dentate gyrus region ATF3 protein in the 6th week group, while the ATF3 protein in the dentate gyrus area did not change significantly during the kindling process.Control group: there was no significant change in CA3 and dentate gyrus in hippocampus from 3 days to 6 weeks. The results of Western Blot showed that the expression of ATF3 in hippocampus increased gradually from 3 days to 6 weeks, and decreased significantly in the group of 4 weeks and 6 weeks, except for the group of 3 days.At other time points, the expression of ATF3 in the experimental group was higher than that in the control group (P 0.05).The results of immunofluorescence showed that the ATF3 protein was located in the nucleus and cytoplasm of hippocampal neurons of rats and mainly concentrated in the cytoplasm.ConclusionThe upregulation of ATF3 protein in hippocampal CA3 may promote mossy fiber sprouting, which may lead to the occurrence and development of temporal lobe epilepsy.
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R742.1

【共引文獻(xiàn)】

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