本文選題：間歇性缺氧 + HIF-1α　； 參考：《廣州中醫(yī)藥大學(xué)》2017年碩士論文

【摘要】：目的:本實驗通過建立常壓慢性間歇性缺氧和慢性持續(xù)性缺氧的幼齡大鼠模型,探討不同缺氧方式對幼齡Wistar大鼠血清HIF-1 α、BDNF的水平表達(dá)和海馬形態(tài)學(xué)和認(rèn)知功能的影響及其可能的相關(guān)機(jī)制。方法:選取(20±2)日齡雄性Wistar大鼠24只,隨機(jī)數(shù)字表法分為3組:NC組,CIH組和CSH組。NC組正常飼養(yǎng),CIH組間歇缺氧8h每天,CSH組持續(xù)缺氧8h每天,兩組大鼠連續(xù)造模30天。造模完成后記錄三組大鼠體重,觀察其生長發(fā)育情況,Morris水迷宮檢測大鼠學(xué)習(xí)和記憶能力,ELISA檢測血清HIF-1 α、BDNF的水平,海馬組織行HE染色、尼氏染色觀察海馬各亞區(qū)神經(jīng)元細(xì)胞形態(tài)學(xué)變化,電鏡觀察海馬CA1區(qū)神經(jīng)元超微結(jié)構(gòu)。結(jié)果:1.大鼠生長發(fā)育NC組大鼠精神狀況良好,毛發(fā)光亮,行動自如,反應(yīng)敏捷。CSH組和CIH組大鼠消瘦,精神萎靡,嗜睡,毛發(fā)無光澤或稀疏,行動遲緩,反應(yīng)遲鈍。NC組體重[(218.63± 15.287)g]明顯高于 CIH 組[(195.75±6.497)g]和 CSH 組[(180.88 ± 12.017)g]體重,P0.05。2.Morris水迷宮實驗2.1定位航行實驗4天訓(xùn)練中,三組間不同時間點逃避潛伏期有顯著差異(F=64.18,P0.05),CIH組和CSH組不同時間點逃避潛伏期有顯著差異(P0.05)。NC組、CIH組、CSH組逃避潛伏期隨著訓(xùn)練時間和次數(shù)的增加逐漸縮短(F=54.355,P0.05;F=109.809,P0,05;F=272.514,P0.05)。訓(xùn)練第1、2天,三組間逃避潛伏期有顯著差異(F=19.248,P0.05;F=5.021,P0.05);第3天、4天,三組間逃避潛伏期差距逐漸縮短,無顯著差異(P=0.706,P0.05;F=0.219,P0.05)。2.2空間探索實驗三組間穿越平臺的次數(shù)有顯著差異(F=8.913,P0.05),組間兩兩對比,CIH組和CSH組之間差異無統(tǒng)計學(xué)意義(P0.05)。3.血清中HIF-1α、BDNF水平表達(dá)三組間HIF-1α、BDNF表達(dá)有顯著差異(F=19.524,P0.05;F=8.552,P0.05),CIH組和CSH組HIF-1 α、BDNF表達(dá)顯著高于NC組(P0.05),且CIH組HIF-1 α、BDNF表達(dá)較CSH組升高(P0.05)。三組間HIF-1 α、BDNF分別做相關(guān)性分析,均無明顯相關(guān)性(P0.05)。4.海馬神經(jīng)細(xì)胞形態(tài)學(xué)變化4.1 HE染色NC組海馬CA1區(qū)細(xì)胞帶完整,細(xì)胞排列整齊緊密,形態(tài)規(guī)則,細(xì)胞核清晰;CSH組和CIH組細(xì)胞排列分散、紊亂,少量胞核固縮、胞質(zhì)深染的早期壞死樣改變。CIH組細(xì)胞帶部分脫失,活細(xì)胞數(shù)量下降。4.2尼氏染色CA1區(qū)NC組海馬細(xì)胞排列緊密,細(xì)胞核清晰,有豐富的細(xì)顆�；虬邏K樣尼氏體;CIH組錐體細(xì)胞排列散亂、數(shù)目減少,在Ⅰ和Ⅳ層均有細(xì)胞壞死。CSH組錐體細(xì)胞排列紊亂,細(xì)胞腫脹、空泡樣變,少量壞死細(xì)胞。CIH組和CSH組均有部分尼氏體溶解、減少或者消失。細(xì)胞計數(shù)示,CIH組和CSH組陽性細(xì)胞數(shù)少于NC組(F=321.25,P0.05),組間對比,CIH組陽性細(xì)胞數(shù)少于CSH組(P0.05)。CA3區(qū)NC組海馬錐體細(xì)胞排列緊密,結(jié)構(gòu)清晰,尼氏體豐富;CIH組和CSH組均錐體細(xì)胞帶中斷,細(xì)胞排列紊亂,Ⅱ?qū)映霈F(xiàn)細(xì)胞空泡樣改變和少量壞死細(xì)胞。CIH組和CSH組尼氏體有不同程度溶解、減少。DG區(qū)NC組海馬顆粒細(xì)胞排列緊密,結(jié)構(gòu)清晰,尼氏體豐富;CIH組活細(xì)胞數(shù)量廣泛下降,壞死細(xì)胞集中出現(xiàn)在Ⅱ和Ⅲ層。CSH組活細(xì)胞數(shù)量減少,在齒狀回尾部,Ⅱ?qū)映霈F(xiàn)大量細(xì)胞質(zhì)深染,核消失的壞死細(xì)胞。4.3透射電鏡檢查NC組海馬CA1區(qū)神經(jīng)元胞核呈圓形或橢圓形,核膜清晰完整,常染色質(zhì)分布均勻,細(xì)胞器豐富,結(jié)構(gòu)形態(tài)清晰。CIH組海馬神經(jīng)元細(xì)胞核圓形,染色質(zhì)邊集,內(nèi)質(zhì)網(wǎng)不清晰,線粒體腫脹、嵴斷裂,細(xì)胞器減少。CSH組海馬神經(jīng)元細(xì)胞變形,線粒體和內(nèi)質(zhì)網(wǎng)部分破壞,細(xì)胞器顯著減少。結(jié)論:慢性缺氧抑制大鼠生長發(fā)育,使幼齡大鼠學(xué)習(xí)記憶認(rèn)知功能下降;HIF-1 α參與神經(jīng)系統(tǒng)缺氧應(yīng)答,BDNF水平代償性增加可能參與神經(jīng)元細(xì)胞損傷調(diào)節(jié),兩者可能獨立參與維持神經(jīng)認(rèn)知功能;而CIH高水平的HIF-1 α和BDNF提示機(jī)體可能處于持續(xù)缺氧應(yīng)激狀態(tài)。CIH獨有的缺氧模式對海馬神經(jīng)細(xì)胞損傷程度更重,出現(xiàn)更嚴(yán)重認(rèn)知功能障礙,海馬不同亞區(qū)(CA1、CA3、DG)對缺氧損傷敏感度不同,神經(jīng)元細(xì)胞損傷程度不同;CIH海馬CA1區(qū)對缺氧損傷更敏感,出現(xiàn)顯著的認(rèn)知障礙。
[Abstract]:Objective: To explore the effects of different hypoxia methods on the expression of HIF-1 alpha, BDNF and hippocampal morphology and cognitive function of young Wistar rats, and to explore the possible mechanism of related mechanisms by establishing a young rat model of chronic intermittent hypoxia and chronic persistent hypoxia. Methods: 24 male Wistar rats of (20 + 2) days of age were selected. The random digital table method was divided into 3 groups: group NC, group CIH and group.NC in group CSH, CIH group intermittent hypoxia 8h every day, CSH group continuous hypoxia 8h every day, two groups of rats for 30 days. After completing the model, the body weight of the rats was recorded, the growth and development of the rats were recorded, the learning and memory ability of the rats was detected by the Morris water maze, ELISA tested HIF-1 alpha in serum ELISA. BD The level of NF, hippocampal tissue HE staining, Nishi staining observed the morphological changes of neurons in the hippocampus and the ultrastructure of hippocampal neurons in the hippocampus CA1 area. Results: the 1. rats grew and developed NC rats with good mental condition, bright hair and action, reaction to agile.CSH group and CIH group of rats, lethargy, lethargy, hairs and hair. Lustre or sparsity, slow action, and slow reaction.NC group weight [(218.63 + 15.287) g] significantly higher than group CIH [(195.75 + 6.497) g] and CSH Group [(180.88 + 12.017) g] weight, P0.05.2.Morris water maze experiment 2.1 navigation Experiment 4 days training, three groups at different time points escape latency has significant difference (F=64.18, P0.05), CIH and CSH groups There were significant differences in the escape latency at different time points (P0.05), group CIH, group CIH, and the escape latency of group CSH gradually shortened with the increase of training time and number of times (F=54.355, P0.05; F=109.809, P0,05; F=272.514, P0.05). Training for 1,2 days, there were significant differences between the three groups of escape latency (F=19.248, 4 days, three escape). There was no significant difference (P=0.706, P0.05; F=0.219, P0.05).2.2 space exploration experiment between three groups, there were significant differences (F=8.913, P0.05), and between group 22, CIH group and CSH group had no statistical significance (P0.05).3. serum HIF-1 alpha in three groups. The differences (F=19.524, P0.05, F=8.552, P0.05), HIF-1 alpha and BDNF in group CIH and CSH group were significantly higher than that in group NC (P0.05), and CIH group HIF-1 alpha. The cells arranged neatly closely, the morphology was regular, the nucleus was clear, the cells in group CSH and CIH were arranged and dispersed, the cells were in disorder, a small number of nuclei were fixed, the early necrotic samples of the cytoplasm were changed in.CIH group, the number of living cells decreased and the number of living cells decreased by.4.2 Nissl staining CA1 region NC group, the hippocampus cells were closely arranged, the nuclei were clear, and there were rich fine particles or spots. The pyramidal cells in the CIH group were arranged and scattered, the number of pyramidal cells was reduced. The pyramidal cells in group.CSH of group I and IV were disorganized, the cells were swollen, and the vacuoles were changed. Some of the.CIH and CSH groups of necrotic cells were dissolved, reduced or disappeared. The number of cells in the CIH and CSH group was less than that of the NC group (F=321.25,) P0.05), the number of positive cells in group CIH was less than that of group CSH (P0.05) in group.CA3 (P0.05), the pyramidal cells in the hippocampal pyramidal cells were closely arranged, the structure was clear and the Nissl body was rich; the pyramidal cell bands of the CIH group and the CSH group were interrupted, the cell arrangement was disorganized, the cell vacuoles like changes in the second layer and the small number of necrotic cells and the CSH group Nissl bodies were dissolved in varying degrees and decreased. The hippocampal granulosa cells in group DG NC were arranged closely, the structure was clear and the Nissl body was rich. The number of living cells in group CIH decreased widely. The necrotic cells concentrated in the number of living cells in group II and III.CSH group, at the tail of dentate gyrus, and there was a lot of cytoplasmic deep staining in the dentate gyrus, and the bad dead cells of the nucleus disappeared in the NC group CA1 region neurons by.4.3 transmission electron microscope. The nucleus is round or oval, the nuclear membrane is clear and complete, the chromatin is evenly distributed, the organelle is rich and the structure shape is clear. The nucleus of the hippocampal neurons is round, the chromatin is set, the endoplasmic reticulum is not clear, the mitochondria swollen, the crista breaks, the organelle reduces the deformation of the hippocampal deity in the.CSH group, the mitochondria and endoplasmic reticulum are partially destroyed and the organelles are destroyed. Conclusion: chronic hypoxia inhibits the growth and development of rats and reduces the cognitive function of learning and memory in young rats; HIF-1 alpha participates in the hypoxia response of the nervous system, and the compensatory increase of BDNF may participate in the regulation of neuronal cell damage. Both of them may participate in the maintenance of neuro recognition function independently; and CIH high level HIF-1 alpha and BDNF prompting machine The hypoxic mode of.CIH, which may be in the state of persistent hypoxia stress, has more severe impairment of hippocampal neurons, more serious cognitive impairment, different hippocampal subregions (CA1, CA3, DG) are sensitive to hypoxia injury, and the damage degree of neuron cells is different. CIH sea horse CA1 area is more sensitive to hypoxia injury, and a significant cognitive impairment appears. Hindering.

【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R766

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