本文選題：遠(yuǎn)隔預(yù)處理 + 血管生成��； 參考：《復(fù)旦大學(xué)》2014年碩士論文

【摘要】：背景與目的：遠(yuǎn)隔預(yù)處理(Remote ischemic preconditioning, RPC)是繼缺血預(yù)處理(IPC)后發(fā)現(xiàn)的一種在不同器官之間誘導(dǎo)缺血耐受的方法。它通過預(yù)先給予遠(yuǎn)隔部位的器官或組織亞致死性的缺血刺激,來誘導(dǎo)另一器官或組織對致死性缺血損傷的耐受力。遠(yuǎn)隔預(yù)處理的器官保護作用自發(fā)現(xiàn)以來,備受神經(jīng)學(xué)界的關(guān)注。目前,動物實驗以及臨床實驗均已證實遠(yuǎn)隔預(yù)處理是一種有效且可行的腦保護誘導(dǎo)方式。但是遠(yuǎn)隔預(yù)處理的分子生物學(xué)機制仍不明確。血管生成是從先存血管產(chǎn)生新生血管的過程,它受血管生成因子和血管生成抑制因了的精細(xì)調(diào)節(jié),是機體一項重要的修復(fù)再生程序,也是腦缺血后神經(jīng)功能修復(fù)的主要途徑之一。本實驗的目的在于研究遠(yuǎn)隔預(yù)處理對局部腦缺血后缺血半暗帶區(qū)血管生成過程以及相關(guān)因子表達模式的影響,進一步揭示遠(yuǎn)隔預(yù)處理介導(dǎo)腦保護的分子機制。方法：雄性SD大鼠(鼠齡11-12周,體重270g-330g),隨機分為遠(yuǎn)隔預(yù)處理(RPC)組、缺血對照(ISC)組和假手術(shù)(SHO)組。腦缺血模型采用遠(yuǎn)端大腦中動脈閉塞模型。遠(yuǎn)隔預(yù)處理以同側(cè)股動脈夾閉15分鐘、復(fù)流15分鐘,循環(huán)三次的方式誘導(dǎo)。采用免疫組化CD31染色法計數(shù)RPC組、ISC組、SHO組缺血后1天、2天、3天、7天、14天缺血半暗帶區(qū)的微血管密度。免疫熒光CD31/BrdU雙標(biāo)計數(shù)各小組缺血后2天、7天缺血半暗帶區(qū)新生血管數(shù)目。免疫組化法半定量計算各實驗小組缺血半暗帶區(qū)膜受體VEGFR-2、Tie-2的表達水平,Western Blotting法半定量計算血管生成因子VEGF-A、Ang-1、Ang-2、HIF-lalpha的表達水平。圖片分析采用Image Proplus、Image J軟件,統(tǒng)計分析采用SPSS19.0軟件,p0.05顯示差異有統(tǒng)計學(xué)意義。結(jié)果：RPC組缺血半暗帶區(qū)微血管密度在缺血后1天、2天、7天、14天均高于ISC對照組,兩組差異有統(tǒng)計學(xué)意義,RPC組與ISC組半暗帶區(qū)微血管密度均于缺血后7天達峰。RPC組半暗帶區(qū)VEGFR-2蛋白表達水平亦高于ISC組,兩組差異在缺血后1天、2天、3天、7天有統(tǒng)計學(xué)意義,且RPC與ISC組VEGFR-2表達量也在缺血后7天達峰,其時間演變規(guī)律與微血管密度相仿。缺血后半暗帶區(qū)Tie-2的上調(diào)在RPC組更為明顯,兩組蛋白水平差異在缺血后2天、3天、7天、14天有統(tǒng)計學(xué)意義,且RPC組蛋白表達上調(diào)較ISC組出現(xiàn)更早且持續(xù)時間更長。在缺血后2天、7天時RPC組VEGF-A、Ang-2蛋白表達量均高于ISC組,與之相反,Ang-1的表達量在缺血2天時RPC組低于ISC組,7天時兩組無差異。RPC組與ISC組缺血后2天、7天半暗帶區(qū)HIF-lalpha蛋白的表達量無差異,但在缺血早期(缺血后6小時內(nèi)),RPC組HIF-lalpha表達的上升較ISC組略滯后但明顯增強。結(jié)論：遠(yuǎn)隔預(yù)處理顯著增強了缺血半暗帶區(qū)血管生成過程,并改變了半暗帶區(qū)血管生成因子的表達模式。這很可能是遠(yuǎn)隔預(yù)處理介導(dǎo)腦保護的主要的分子生物學(xué)機制
[Abstract]:Background & AIM: remote ischemic preconditioning, RPC) is a method to induce ischemic tolerance between different organs after ischemic preconditioning. It induces the tolerance of another organ or tissue to fatal ischemic injury by giving sublethal ischemic stimulation to the distant organ or tissue in advance. The role of remote preconditioning in organ protection has been concerned by neuroscientists since it was discovered. At present, animal experiments and clinical trials have proved that remote preconditioning is an effective and feasible way to induce brain protection. However, the molecular biological mechanism of remote preconditioning is still unclear. Angiogenesis is a process of angiogenesis from pre-existing blood vessels, which is carefully regulated by angiogenic factors and angiogenesis inhibition, and is an important procedure for the body to repair and regenerate. It is also one of the main ways to repair nerve function after cerebral ischemia. The purpose of this study was to investigate the effects of remote preconditioning on angiogenesis and expression patterns of related factors in ischemic penumbra after local cerebral ischemia, and to further reveal the molecular mechanism of remote preconditioning mediated brain protection. Methods: male Sprague-Dawley rats (11-12 weeks old, weight 270g-330g) were randomly divided into remote preconditioning (RPC) group, ischemic control group (ISCC) and sham operation group. The middle cerebral artery occlusion model was used in cerebral ischemia model. Distal preconditioning was induced by ipsilateral femoral artery occlusion for 15 minutes, reflow for 15 minutes and circulation for three times. The microvessel density of ischemic penumbra in RPC group was counted by immunohistochemical CD31 staining method. The number of neovascularization in ischemic penumbra was counted by double labeling immunofluorescence CD31/BrdU 2 days after ischemia and 7 days after ischemia. The expression level of VEGFR-2Tie-2 in ischemic penumbra was semi-quantitatively calculated by immunohistochemical method and the expression level of angiogenic factor VEGFR-2HIF-lalpha was semi-quantitatively calculated by Western Blotting method. Image Proplusus Image J software was used for image analysis, and SPSS19.0 software for statistical analysis showed significant difference. Results the microvessel density of the ischemic penumbra in the control group was significantly higher than that in the control group on the 1st day, 2nd day, 7th day and 14th day after ischemia. There was significant difference between the two groups. The microvessel density of penumbra in RPC group and ISC group was significantly higher than that in ISC group 7 days after ischemia. The difference between the two groups was statistically significant at 1 day, 2 days, 3 days and 7 days after ischemia, and the expression level of VEGFR-2 protein in the penumbra region of the two groups was significantly higher than that in the ISC group on the 7th day after ischemia, and the difference between the two groups was statistically significant. The expression of VEGFR-2 in RPC and ISC groups reached the peak 7 days after ischemia, and its temporal evolution was similar to that of microvessel density. The upregulation of Tie-2 in the posterior penumbra of ischemia was more obvious in the RPC group. The difference of protein level between the two groups was statistically significant 2 days after ischemia and 7 days and 14 days after ischemia, and the up-regulation of RPC protein expression appeared earlier and lasted longer than that in the ISC group. The expression of VEGF-AfAng-2 protein in RPC group was higher than that in ISC group at 7 days after ischemia. On the contrary, the expression of Ang-1 in the RPC group was lower than that in the ISC group on the 7th day after ischemia. There was no difference in the expression of HIF-lalpha protein in the penumbra between the RPC group and the ISC group on the 7th day after ischemia. However, in the early stage of ischemia (within 6 hours after ischemia), the increase of HIF-lalpha expression in ISC group was slightly slower than that in ISC group. Conclusion: the process of angiogenesis in ischemic penumbra was significantly enhanced by remote preconditioning and the expression pattern of angiogenic factors in penumbra was changed. This may be the main molecular biological mechanism of remote preconditioning mediating brain protection.
【學(xué)位授予單位】：復(fù)旦大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R743.3

【共引文獻】

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