本文選題：脊髓損傷 + 載脂蛋白E類　； 參考：《昆明醫(yī)科大學(xué)》2017年碩士論文

【摘要】：[目的]本研究通過建立C57/BL6野生型小鼠和ApoE基因敲除小鼠脊髓損傷模型,從動物整體水平探討ApoE對小鼠脊髓損傷后運動功能活動和星形膠質(zhì)細胞影響,以期為ApoE對中樞神經(jīng)損傷后的作用機制的研究提供理論依據(jù)。[方法]本研究采用腦立體定位儀自制脊髓打擊器制作C57/BL6小鼠脊髓損傷動物模型,探究ApoE在脊髓損傷恢復(fù)進程的影響及其可能的作用機制。所有動物分為三組:假手術(shù)組、野生型小鼠組以及ApoE-/-組。對野生型小鼠運用免疫印跡分析和RT-PCR,定量測定ApoE蛋白和mRNA在脊髓損傷恢復(fù)過程中的表達變化。建模后每天行BBB功能評分直至實驗結(jié)束,表示脊髓損傷后小鼠運動功能恢復(fù)狀態(tài)。對脊髓損傷處組織行蘇木精-伊紅染色,觀察損傷節(jié)段細胞水腫及炎癥反應(yīng)情況。通過Western-Blot和免疫熒光染色觀察損傷節(jié)段星形膠質(zhì)細胞特異性標記物GFAP的表達變化及神經(jīng)纖維的生長情況。[結(jié)果]1.C57/BL6野生型小鼠SCI后,ApoE最早從第3天開始升高,第7d和14d時,ApoE的表達量顯著增加(P0.01)。其中第7d時,ApoE mRNA含量達到最高,第14d時ApoE蛋白含量達到最高,28天時含量有所下降,但是仍然高于損傷后第1天和假手術(shù)組(P0.05)。2.BBB評分結(jié)果顯示,SCI后第7d開始,C57/BL6野生型小鼠得分優(yōu)于ApoE-/-小鼠,第14d時差異最明顯;第28d時,兩組評分差異減小,但野生型小鼠BBB評分仍高于ApoE-/-(P0.05),說明ApoE存在的狀態(tài)下,有利于小鼠脊髓損傷后的運動功能恢復(fù)。3.小鼠脊髓損傷節(jié)段HE染色結(jié)果顯示,SCI后第14d,野生型小鼠脊髓組織結(jié)構(gòu)較ApoE-/-小鼠完整,脊髓灰、白質(zhì)組織結(jié)構(gòu)可辨認,炎性細胞較少,細胞核固縮較輕,可見清晰胞質(zhì)胞核,形態(tài)較正常。ApoE-/-小鼠組織結(jié)構(gòu)紊亂,廣泛水腫,有較多炎性細胞浸潤,細胞核呈游散狀態(tài),固縮嚴重。4.Western blot結(jié)果顯示各組脊髓損傷后均有GFAP蛋白表達。且在第7d,14d,28d,ApoE-/-小鼠的GFAP表達量高于野生型小鼠。5.與野生型小鼠相比,ApoE-/-組小鼠損傷節(jié)段的GFAP陽性染色細胞較多,軸突數(shù)目相對較多,這提示星形膠質(zhì)細胞活化程度相對較高。[結(jié)論]1.小鼠脊髓損傷后,ApoE表達量隨著病程進展呈進行性增加;2.ApoE能加快脊髓損傷小鼠后肢運動功能恢復(fù)進程,對脊髓損傷后的神經(jīng)恢復(fù)有正向作用;3.ApoE對脊髓損傷的修復(fù)作用可能與下調(diào)GFAP的含量,抑制損傷周圍星形膠質(zhì)瘢痕的形成有關(guān)。
[Abstract]:[objective] to investigate the effects of ApoE on motor function and astrocytes after spinal cord injury in C57 / BL6 wild-type mice and ApoE knockout mice. To provide a theoretical basis for the study of the mechanism of ApoE after central nerve injury. [methods] the spinal cord injury model of C57 / BL6 mice was made by using the brain stereotactic locator to investigate the effect of ApoE on the recovery process of spinal cord injury and its possible mechanism. All animals were divided into three groups: sham operation group, wild type mice group and ApoE-P-group. The expression of ApoE protein and mRNA during the recovery of spinal cord injury in wild type mice was determined by Western blot and RT-PCR. After modeling, BBB scores were performed every day until the end of the experiment, indicating the recovery of motor function after spinal cord injury. Hematoxylin-eosin staining was used to observe the edema and inflammatory reaction in the injured spinal cord tissue. The expression of GFAP and the growth of nerve fibers were observed by Western-Blot and immunofluorescence staining. [results] 1. The expression of ApoE in C57 / BL6 wild-type mice increased at the earliest from the 3rd day after sci, and the expression of ApoE increased significantly on the 7th and 14th days (P0.01). On the 7th day, the ApoE mRNA content reached the highest level, and the ApoE protein content reached the highest on the 14th day, and decreased at 28 days. But it was still higher than that of the first day after injury and the sham-operation group (P0.05). 2. The results of BBB score showed that the score of C57 / BL6 wild-type mice was better than that of ApoE-r-mice on the 7th day after sci, and the difference was the most obvious at the 14th day, and the difference between the two groups decreased on the 28th day. But the BBB score of wild type mice was still higher than that of ApoE-r- (P0.05), which indicated that the presence of ApoE was beneficial to the recovery of motor function after spinal cord injury. 3. The results of HE staining showed that the structure of spinal cord of wild type mice was more intact than that of ApoE-r-mice on the 14th day after sci, the structure of gray and white matter tissue was recognizable, the inflammatory cells were less, the nucleus was less pyknosis, and the cytoplasmic nucleus was clear. Compared with the normal. ApoE-r- mice, the tissue structure was disordered, extensive edema, more inflammatory cells infiltrated, and the nucleus was swimming away. 4. The results of Western blot showed that GFAP protein was expressed in all groups after spinal cord injury. The expression of GFAP in ApoE-r-mice was higher than that in wild-type mice. Compared with the wild type mice, the GFAP positive staining cells and the number of axons in the injured segments of the ApoE-r- group were more than those in the wild type mice, which suggested that the astrocytes were more activated than the wild type mice. [conclusion] 1. The expression of ApoE in mice after spinal cord injury increased progressively with the progression of the disease. 2. ApoE could accelerate the recovery of motor function in the hind limbs of mice with spinal cord injury. The effect of ApoE on the repair of spinal cord injury may be related to the down-regulation of GFAP content and the inhibition of astrocyte scar formation around the injury.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R651.2

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