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研究生:郭佳惠
研究生(外文):Chia-Hui Kuo
論文名稱:以醣皮質類固醇治療大鼠腦出血對血腦屏障的影響
論文名稱(外文):The effects of dexamethasone treatment on blood-brain barrier of brain in intracerebral hemorrhage rat
指導教授:沈國愉楊仁宗
學位類別:碩士
校院名稱:嘉南藥理科技大學
系所名稱:生物科技系暨研究所
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2007
畢業學年度:95
語文別:中文
論文頁數:55
中文關鍵詞:腦內出血、基質金屬蛋白酶
外文關鍵詞:Matrix metalloproteinasesIntracerebral hemorrhage
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腦內出血是一個快速變化且不易診斷的疾病,往往在幾個小時或幾天
之內有急速的變化。但目前腦內出血後的腦部變化並無清楚的機制。基質
金屬蛋白酶是一個內含鋅的蛋白酶家族,可分解基底層及細胞外間質。目
前有些研究指出腦內出血後第二型和第九型基質金屬蛋白酶的產生是有害
的,但其活性的多寡與對腦部的二次傷害是否有關尚未釐清。而內皮細胞
黏附因子在受傷時會產生,可吸附白血球並且活化使血腦屏障通透性變高
白血球易進入腦部中,為發炎反應的重要指標。在本實驗中我們採用以膠
原蛋白分解酶誘導大鼠腦出血的動物模式來探討第二型和第九型基質金屬
蛋白酶及內皮細胞黏附因子的表現,並且以免疫組織化學染色法和西方墨
點分析來印證其表現量。在實驗中採用廣泛使用的抗發炎藥物-類固醇 (15
mg/kg) 來治療,看是否能抑制第二型和第九型基質金屬蛋白酶及內皮細胞
黏附因子的表現量及減少腦水腫的程度。最後,結果發現第二型和第九型
基質金屬蛋白酶及內皮細胞黏附因子可能在腦出血後扮演了有害的角色,
使用類固醇可以抑制此兩者基質金屬蛋白酶及內皮細胞黏附因子的表現及
腦水腫,或野i以減少腦出血後的二次傷害。
Intracerebral hemorrhage (ICH), an acute neurological disorder, has a
rapidly evolving process that may progress over hours and days. The
mechanisms of brain insult after ICH remain to be clarified. Matrix
metalloproteinases (MMPs) is a family of zinc-dependent proteases which are
involved in the degradation of basal lamina and extracellular matrix components.
Although a few studies suggested a detrimental role of MMP-2 and MMP-9 in
ICH, the relationships between it’s activity and the secondary brain changes
after ICH are not determined. The intercellular adhesion molecule-1 (ICAM-1)
binds to its leukocyte ligands and allows activated leukocytes entry into the CNS.
In this study, we examine the expression of MMP-2, MMP-9 and ICAM-1 in
vivo using a collagenase-induced rat model of ICH. Immunohistochemistry and
Western blot revealed that MMP-2, MMP-9 and ICAM-1 was upregulated after
ICH. The treatment with a broad-spectrum inflammation inhibitor
dexamethasone (DEX) (15 mg/kg) could cause a decrease of MMP-2, MMP-9
and ICAM-1 expression and local neutrophil infiltration, then prevented the
neuron death. The block of local brain edema around ICH was also observed. In
conclusion, MMP-2, MMP-9 and ICAM-1 play a deleterious role in acute brain
injury after ICH. The inhibition of MMP-2, MMP-9 and ICAM-1 expression
with DEX during this critical period may be useful in treatment of secondary
brain edema, which could be a therapeutic strategy for ICH.
目錄 (contents)
中文摘要....................................................................................................... I
Abstract......................................................................................................... II
縮寫表............................................................................................................Ⅳ
第一章 緒論....................................................................................................1
第二章 材料與方法..................................................................................... 10
2-1 實驗動物………………………………………………………...10
2-2 腦出血動物模式………………………………………………..10
2-3 給予醣皮質類固醇 (Dexamethasone, DEX)………………….11
2-4 核磁共振檢查(MRI)…………………………………………...11
2-5 擷取腦組織及冷凍切片……………………………………….12
2-6 H&E 染色…………………………………………………….12
2-7 抽取蛋白質…………………………………………………….13
2-8 蛋白質定量…………………………………………………….14
2-9 組織免疫化學染色法(immunohistochemistry staining,
IHC)………………………………………………………….14
2-10 西方墨點法 (Western blotting)………………………………15
2-11 統計分析……………………………………………………..17
第三章 結果…………………………………………………………………18
第四章 討論…………………………………………………………………22
第五章 結論…………………………………………………………………26
參考文獻………………………………………………………………….......27
附圖…………………………………………………………………………32
圖目錄
圖1. 大鼠經立體定位儀定位後注射 collagenase 之情形……………..32
圖2. 腦出血動物模式定位圖…………………………………………….33
圖3. 腦切片後之檢體經 H&E 染色後結果…………………………….34
圖4. 大鼠腦出血後經 MRI 檢查後的出血情形………………………..35
圖5. MRI 影像計算水腫面積示意圖………………………………….....36
圖6. 大鼠腦出血後經 MRI 檢查後換算水腫之比率…………………..37
圖7. 腦切片檢體經 IHC (MMP 2) 染色後之結果……………………...38
圖8. 腦切片檢體經 IHC (MMP 9) 染色後之結果………………………39
圖9. 腦切片檢體經 IHC (ICAM-1) 染色後之結果……………………..40
圖10. 腦組織抽出之蛋白質經 SDS-PAGE 電泳後,進行 western blotting
分析 (MMP 2) 之結果…………………………………………..41
圖11. 腦組織抽出之蛋白質經 SDS-PAGE 電泳後,進行 western blotting
分析 (MMP 9) 之結果…………………………………………...42
圖12. 腦組織抽出之蛋白質經 SDS-PAGE 電泳後,進行 western blotting
分析 (ICAM-1) 之結果…………………………………………..43
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