本文關(guān)鍵詞： 腦低灌注 白質(zhì) CX3CR1 認(rèn)知 p38MAPK　出處：《吉林大學(xué)》2014年博士論文　論文類(lèi)型：學(xué)位論文

【摘要】：缺血性白質(zhì)損傷可以引起認(rèn)知功能障礙，嚴(yán)重?fù)p害人類(lèi)健康，其主要的發(fā)生機(jī)制有免疫炎性損害、氧化應(yīng)激損傷、突觸結(jié)構(gòu)及功能異常等。目前研究的較多的為免疫炎性損傷機(jī)制。小膠質(zhì)細(xì)胞作為中樞神經(jīng)系統(tǒng)免疫吞噬細(xì)胞發(fā)揮著雙重作用，少量被激活時(shí)，可以釋放神經(jīng)營(yíng)養(yǎng)因子、吞噬廢棄的組織碎片發(fā)揮保護(hù)性作用；大量被激活則產(chǎn)生炎性損害作用。CX3CR1作為小膠質(zhì)細(xì)胞表面的一種受體分子，在生理?xiàng)l件下，使小膠質(zhì)細(xì)胞保持靜息狀態(tài)、維持內(nèi)環(huán)境的穩(wěn)定狀態(tài)；在急性缺血、變性疾病、神經(jīng)創(chuàng)傷時(shí)發(fā)揮著損害性作用。在缺血性白質(zhì)損傷時(shí)，如何變化、起什么樣的作用尚未有研究。p38MAPK是MAPK家族中重要成員，主要參與炎性應(yīng)激反應(yīng)。p38MAPK是否參與CX3CR1介導(dǎo)的缺血性白質(zhì)損傷的發(fā)病過(guò)程目前還不清楚。 目的：觀(guān)察CX3CR1是否參與了慢性缺血性白質(zhì)損傷發(fā)病過(guò)程；CX3CR1對(duì)慢性缺血性白質(zhì)損傷是否通過(guò)p38MAPK途經(jīng)實(shí)現(xiàn)。方法：雄性Wistar大鼠320只，隨機(jī)分為正常組、假手術(shù)組、缺血組、陰性對(duì)照組、干預(yù)組，Morris水迷宮觀(guān)察造模后28d學(xué)習(xí)記憶功能。造模后不同時(shí)間點(diǎn)（1d、3d、7d、14d、28d），HE染色及Luxol Fast Blue（LFB）染色法評(píng)價(jià)胼胝體、內(nèi)囊、視神經(jīng)纖維變化；Western Blot觀(guān)察CX3CR1、P-p38/p38蛋白表達(dá)；免疫熒光雙標(biāo)技術(shù)觀(guān)察CX3CR1與小膠質(zhì)細(xì)胞關(guān)系；免疫組織化學(xué)染色評(píng)價(jià)小膠質(zhì)細(xì)胞變化規(guī)律；側(cè)腦室內(nèi)注射不同濃度CX3CR1中和抗體、p38抑制劑進(jìn)一步觀(guān)察缺血后上述指標(biāo)的變化。結(jié)果：（1）造模后28d，缺血組逃避潛伏期、空間探索路徑較正常組延長(zhǎng)，跨越平臺(tái)次數(shù)顯著減少（P0.01）。（2）隨著缺血時(shí)間的延長(zhǎng)，HE染色觀(guān)察到胼胝體、內(nèi)囊及視神經(jīng)區(qū)域白質(zhì)纖維疏松，，排列紊亂且空泡樣改變；LFB染色可見(jiàn)上述區(qū)域髓鞘崩解范圍擴(kuò)大，分層明顯，部分髓鞘出現(xiàn)空泡狀；CX3CR1表達(dá)量持續(xù)增加，與CD11b標(biāo)記的小膠質(zhì)細(xì)胞數(shù)目持續(xù)增多相一致且高于假手術(shù)組和正常組（P0.01）。（3）隨著缺血時(shí)間的延長(zhǎng)，P-p38/p38比值在缺血3d時(shí)出現(xiàn)明顯變化，與CX3CR1變化具有相關(guān)性。（4）不同濃度CX3CR1中和抗體對(duì)缺血性白質(zhì)損傷的認(rèn)知功能、纖維損傷程度、CD11b小膠質(zhì)細(xì)胞數(shù)目及P-p38/p38比值均有改善作用，且呈現(xiàn)劑量依賴(lài)性。應(yīng)用p38抑制劑后，缺血性白質(zhì)的纖維損傷程度及小膠質(zhì)細(xì)胞數(shù)目均有不同程度改善。結(jié)論：（1）CX3CR1過(guò)表達(dá)通過(guò)介導(dǎo)小膠質(zhì)細(xì)胞的變化對(duì)缺血性白質(zhì)產(chǎn)生損傷作用，進(jìn)而影響空間學(xué)習(xí)記憶功能。（2）CX3CR1/p38MAPK信號(hào)通路是缺血性白質(zhì)損傷發(fā)生機(jī)制之一。
[Abstract]:Ischemic white matter injury can cause cognitive impairment and serious damage to human health. Its main mechanisms are immune inflammatory damage and oxidative stress injury. Abnormal synaptic structure and function. The mechanism of immune inflammatory injury is studied. Microglia, as immune phagocytes of the central nervous system, play a dual role, when a small amount of activated, can release neurotrophic factors, Phagocytosis of abandoned tissue fragments plays a protective role; a large number of activated inflammatory damage. CX3CR1, as a receptor molecule on the surface of microglia, keeps the microglia in a resting state under physiological conditions. Maintain a stable state of the internal environment; play a damaging role in acute ischemia, degenerative diseases, and neurotrauma. In ischemic white matter injury, how and what role to play has not been studied. P38 MAPK is an important member of the MAPK family. It is unclear whether p38 MAPK is involved in the pathogenesis of ischemic white matter injury mediated by CX3CR1. Objective: to observe whether CX3CR1 was involved in the pathogenesis of chronic ischemic white matter injury (CX3CR1) and whether it was achieved by p38 MAPK. Methods: 320 male Wistar rats were randomly divided into normal group, sham operation group and ischemic group. In the negative control group, the learning and memory function was observed by Morris water maze on the 28th day after modeling, and the expression of CX3CR1P-p38 / p38 protein was evaluated by Luxol Fast BlueLFB staining and Luxol Fast BlueLFB staining at different time points. The expression of CX3CR1P-p38 / p38 protein was evaluated by Western Blot in corpus callosum, internal capsule and optic nerve fibers. The relationship between CX3CR1 and microglia was observed by immunofluorescence double labeling technique, and the changes of microglia were evaluated by immunohistochemical staining. The changes of above indexes after ischemia were further observed by intraventricular injection of different concentrations of CX3CR1 neutralizing antibody p38 inhibitor. Results the escape latency and spatial exploration pathway of ischemia group were longer than that of normal group 28 days after modeling. With the prolongation of ischemia time, the white matter fibers of corpus callosum, internal capsule and optic nerve region were loose, and the disordered arrangement and vacuolar change LFB staining showed that the area of myelin disintegrating was enlarged. The expression of CX3CR1 was increased in some myelin sheath. The number of microglia labeled with CD11b was increased and higher than that of sham operation group and normal group (P 0.01). The ratio of P-p38 / p38 was significantly changed with the prolongation of ischemic time on the 3rd day after ischemia. The cognitive function of different concentrations of CX3CR1 neutralizing antibody to ischemic white matter injury, the number of CD11b microglia and the ratio of P-p38 / p38 were improved in a dose-dependent manner. The degree of fibrous injury and the number of microglia in ischemic white matter were improved in different degrees. Conclusion the overexpression of CX3CR1 can induce damage to ischemic white matter by mediating the change of microglia. Furthermore, affecting the spatial learning and memory function of CX3CR1 / p38 MAPK signaling pathway is one of the mechanisms of ischemic white matter injury.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2014
【分類(lèi)號(hào)】：R742

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