本文選題：高血脂　切入點：腦缺血　出處：《河北聯合大學》2014年碩士論文

【摘要】：目的探討跑臺運動對大鼠高血脂腦缺血再灌注后海馬區(qū)Ku蛋白和細胞周期蛋白的作用。 方法本實驗將SPF級雄性高血脂誘導的Wistar大鼠120只隨機分為：對照組（n=40），模型組（n=40）、實驗組（n=40）。高血脂模型采用高血脂飼料喂養(yǎng)6周形成高血脂大鼠后。實驗組于建模前15天開始進行跑臺運動20m/min，30min/day，應用四血管阻斷法（Pulsinelli4VO法）制備全腦缺血模型，分別于缺血15min再灌注后3h、6h、24h和48h四個時間點處死大鼠，缺血再灌注后24h和48h大鼠在處死前采用水迷宮檢測各組大鼠視空間記憶。各時間點的標本采用蘇木素-伊紅染色法（hematoxylin-eosin staining，HE）觀察各組大鼠腦組織神經元細胞形態(tài)的變化情況；采用免疫組織化學染色和免疫印跡蛋白（western-blotting）方法檢測各組大鼠腦組織中Ku70、Ku80、CycinA和CyclinE蛋白的表達情況。應用SPSS17.0統(tǒng)計分析軟件對所得實驗數據進行重復設計的單因素方差分析，以P0.05，表示差異具有統(tǒng)計學意義。 結果1水迷宮檢測結果：與對照組比較，模型組大鼠在各時間點（24h、48h）的潛伏期時間顯著延長，穿臺次數顯著減少，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（24h、48h）的潛伏期時間顯著縮短而穿臺次數顯著增加，差異有統(tǒng)計學意義（P0.05）。2形態(tài)觀察結果（HE染色）對照組大鼠海馬區(qū)神經元細胞排列整齊，胞漿淡染；模型組與對照組比較3h結構形態(tài)即出現形態(tài)改變，表現為細胞失去正常結構，部分細胞形態(tài)異常，6h海馬區(qū)可見較多腫脹的神經元，間質水腫，結構疏松，神經元和膠質細胞出現核固縮、深染，也有部分神經元胞核完全消失，形成空泡狀結構；24h腦組織神經細胞破壞，可見較多變性壞死的神經細胞，神經細胞呈空泡狀或海綿狀，神經元細胞出現核固縮、碎裂和核仁消失現象，間質水腫明顯；48h可見壞死神經細胞，腦組織結構破壞，結構疏松，核膜碎裂、核仁消失，各時間點中以48h損傷最嚴重；實驗組大鼠海馬區(qū)神經元損傷較模型組明顯減輕，可見壞死的神經元減少，間質水腫較輕，核固縮和空泡較少。3Ku70免疫組化和Western blot分析結果：免疫組化結果：Ku70免疫組化染色呈棕黃色，定位于細胞胞漿中，主要表達在神經元細胞。與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）Ku70蛋白的表達增加，且24h達高峰，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）Ku70蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）；Westernblot分析結果：與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）Ku70蛋白的表達增加，且24h達高峰，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）Ku70蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）。4Ku80免疫組化和Western blot分析結果：免疫組化結果：Ku80免疫組化染色呈棕黃色，定位于細胞胞核中，主要表達在海馬區(qū)神經細胞。與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）Ku80蛋白的表達明顯增加，且6h達高峰，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）Ku80蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）；Western blot分析結果：與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）Ku80蛋白的表達明顯增加，且6h達高峰，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）Ku80蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）。5CyclinA免疫組化和Western blot分析結果：免疫組化結果：CyclinA免疫組化染色呈棕黃色，，定位于細胞胞核中，主要表達在海馬區(qū)神經細胞。與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）CyclinA蛋白的表達量逐漸增加，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）CyclinA蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）；Western blot分析結果：與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）CyclinA蛋白的表達量逐漸增加，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）CyclinA蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）。6CyclinE免疫組化和Western blot分析結果：免疫組化結果：CyclinE免疫組化染色呈棕黃色，定位于細胞胞核中，主要表達在海馬區(qū)神經細胞。與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）CyclinE蛋白的表達逐漸降低，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）CyclinE蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）；Western blot分析結果：與對照組比較，模型組大鼠在各時間點（3h、6h、24h、48h）CyclinE蛋白的表達降低，差異有統(tǒng)計學意義（P0.05）；與模型組比較，實驗組大鼠在各時間點（3h、6h、24h、48h）CyclinE蛋白的表達明顯增加，差異有統(tǒng)計學意義（P0.05）。 結論1跑臺運動可改善全腦缺血再灌注大鼠空間記憶功能。2跑臺運動可增加Ku蛋白的表達，同時調控CyclinA和CyclinE的表達，對腦缺血有保護作用。
[Abstract]:Objective to investigate the effect of treadmill exercise on Ku protein and cyclin protein in hippocampus of rats after hyperlipidemia and cerebral ischemia reperfusion.
Methods the Wistar rats induced by SPF male hyperlipidemia 120 rats were randomly divided into control group (n=40), model group (n=40), experimental group (n=40). The model of hyperlipidemia with high lipid diet for 6 weeks after the formation of hyperlipemia rats. The experimental group began running 20m/min. In the 15 days before modeling 30min/day, using four vessel occlusion method (Pulsinelli4VO method) to prepare the model of whole brain ischemia in ischemic reperfusion after 15min of 3H, 6h, 24h and 48h four time points and the rats were killed after ischemia reperfusion 24h and 48h rats were sacrificed before using in water maze test in rats visuospatial memory. At different time points were detected by hematoxylin eosin staining (hematoxylin-eosin, staining, HE) to observe the morphological changes of brain tissue in rats of each group of neurons; using chemical staining and Western blot immunohistochemistry (Western-blotting) method to detect the rat brain The expression of Ku70, Ku80, CycinA and CyclinE protein in tissues was analyzed. SPSS17.0 analysis software was used to analyze the experimental data by one-way ANOVA, and the difference was statistically significant with P0.05.
The 1 water maze test results: compared with the control group, the rats in the model group at each time point (24h, 48h) the incubation time was prolonged and the frequency of crossing the platform was significantly reduced, the difference was statistically significant (P0.05); compared with the model group, the rats in the experimental group at each time point (24h, 48h). The latency time was significantly shortened and the times across the platform significantly increased, the difference was statistically significant (P0.05.2) the results of morphology (HE staining) in control neurons in hippocampus of rats in order, cytoplasmic staining; model group compared with the control group 3H morphology appeared morphological changes showed the cells lose their normal structure, abnormal part of the cell morphology, 6h in hippocampus neurons showed more swelling, interstitial edema, loose structure, nuclear pyknosis, appearance of neurons and glial cells stained, some neurons disappeared completely, the formation of air bubbles; 24h brain nerve cells There were lots of damage, degeneration and necrosis of nerve cells, nerve cells were vacuolated or cavernous, pyknosis neurons cells, fragmentation and nucleoli disappeared, interstitial edema obviously; 48h necrosis of nerve cells, brain structure damage, loose structure, nuclear fragmentation, nucleolus disappeared, at each time point in the most serious 48h damage; neurons in hippocampus of rats in experimental group injury significantly reduced compared with the model group, reduce the necrosis of neurons, interstitial edema, pyknosis and vacuolation less.3Ku70 immunohistochemistry and Western blot analysis results: immunohistochemical results: Ku70 immunohistochemical staining was brown, located in the cytoplasm that is mainly expressed in neuronal cells. Compared with the control group, the rats in the model group at each time point (3H, 6h, 24h, 48h) the expression of Ku70 protein increased, and reached the peak at 24h, the difference was statistically significant (P0.05); compared with the model group, 瀹為獙緇勫ぇ榧犲湪鍚勬椂闂寸偣(3h,6h,24h,48h)Ku70铔嬬櫧鐨勮〃杈炬槑鏄懼鍔

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