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系統識別號 U0026-2601201012535100
論文名稱(中文) 硫辛酸在硫代乙醯胺誘發肝纖維化的保護作用及可能機制探討
論文名稱(英文) Protective effects of α-lipoic acid in thioacetamide-induced liver fibrosis and its related mechanisms
校院名稱 成功大學
系所名稱(中) 環境醫學研究所
系所名稱(英) Institute of Environmental and Occupational Health
學年度 98
學期 1
出版年 99
研究生(中文) 林舒慧
研究生(英文) Shu-Huei Lin
學號 S7696406
學位類別 碩士
語文別 中文
論文頁數 69頁
口試委員 指導教授-王應然
口試委員-何元順
口試委員-潘敏雄
口試委員-郭靜娟
口試委員-羅至佑
中文關鍵字 硫辛酸  硫代乙醯胺  肝纖維化  肝星狀細胞 
英文關鍵字 α-lipoic acid  thioacetamide  liver fibrosis  hepatic stellate cell 
學科別分類
中文摘要 肝纖維化是在肝臟受到損傷時,因反覆修復癒合作用而形成疤痕組織的病理現象,而肝星狀細胞的活化為造成肝纖維化的關鍵。硫辛酸(α-lipoic acid, α-LA)是一種在體內可自行生合成、作用類似脂溶性維生素、具有雙硫化合物的強力抗氧化劑。目前對於α-LA在硫代乙醯胺(thioacetamide, TAA)誘導大鼠肝纖維化中的角色尚不明確。因此,本研究的主要目的在於探討α-LA在TAA誘導大鼠肝纖維化模式中預防肝纖維化的能力;以及藉由轉化生長因子(transforming growth factor β, TGF-β)及血小板衍生生長因子(Platelet-derived growth factor, PDGF)誘導肝星狀細胞株(HSC-T6)活化與增生的模式探討α-LA抑制纖維化的活性與分子機制。動物實驗以雄性Wistar大鼠進行體內試驗,每週給予α-LA及TAA腹腔注射三次連續八週。實驗結果指出,單獨處理TAA八週後於組織型態學上觀察到大鼠肝臟中肝細胞變性及膠原沉積,且在血清肝功能指標AST及ALT數值相對於控制組皆有明顯上升;另在合併處理TAA及α-LA的組別中則發現α-LA可以抑制TAA造成之肝細胞變性、膠原沉積以及血清肝功能指標數值下降。此外在細胞試驗中發現TGF-β與PDGF分別可誘導肝星狀細胞株表現α-平滑肌肌動蛋白(α-SMA)與促進細胞增生;而當細胞合併處理TGF-β與α-LA之還原態DHLA後,實驗結果發現DHLA能抑制TGF-β所造成之α-SMA表現上升,並抑制TGF-β誘導之活性氧物種生成與降低ERK及JNK蛋白質之磷酸化;另外在PDGF誘導星狀細胞增生的實驗中亦發現DHLA可明顯的抑制PDGF造成之增生,實驗結果顯示其抑制增生的機制可能為藉由降低PDGF生成之活性氧物種與PI3K-Akt-p70S6K傳遞路徑中Akt及p70S6K的活化。此研究證明α-LA能改善TAA所誘發之肝纖維化。
英文摘要 Hepatic fibrosis is the result of the wound-healing response of the liver to repeated injury. α-Lipoic acid (α-LA) was postulated to be an effective antioxidant with dithiol groups, its functions like fat-soluble vitamin. However, it is unknown about the precise role of α-LA on liver fibrosis induced by thioacetamide (TAA). The purpose of this study is to assess (1) the preventive efficacy of α-LA on liver fibrosis induced by TAA; (2) the related protective mechanism of α-LA on transforming growth factor-β (TGF-β) and platelet-derived growth factor (PDGF) induced-activation and proliferation in hepatic stellate cells (HSC). In this study, male Wistar rats were treated with TAA and α-LA three times per week for eight weeks. The results indicated that treated with TAA induced degeneration of hepatocyte and collagen deposition. We also found that the levels of AST and ALT, in TAA-treated group, was significant increased with normal group (p<0.05), whereas both were decreased in combine-treated group (TAA and α-LA). In the in vitro study, the results show that TGF-β-induce transformation and PDGF-induce cell proliferation of HST-T6 cells both in dose-dependent manner. In addition, treated with TGF-β combine DHLA could reduce α-smooth muscle actin protein expression, phosphorylation of ERK and JNK, as well as the reactive oxygen species (ROS) production. Indeed, DHLA treatment also reduces PDGF-induced cell proliferation. The results show the possible mechanism may through inhibit ROS production and down-regulate PI3K-Akt-p70S6K pathway. This study demonstrated that α-LA could ameliorate TAA-induced liver fibrosis.
論文目次 第一章、緒論 1
第二章、文獻回顧 2
第一節、肝臟組織的功能及其細胞組成 2
第二節、肝臟纖維化之形成 3
第三節、肝臟星狀細胞(Hepatic stellate cells)於肝纖維化中的角色 3
第四節、細胞激素(cytokines)於肝纖維化中的角色 5
第五節、硫辛酸(α-Lipoic acid) 6
第六節、硫代乙醯胺(Thioacetamide)誘發肝纖維化之動物模式 8
第三章、研究目的 10
第四章、研究架構 11
第五章、研究材料與方法 13
第一節、研究材料 13
細胞株: 13
儀器: 13
試劑: 14
溶液: 15
第二節、研究方法 16
體外試驗(In vitro) 16
體內試驗(In vivo) 21
第六章、結果 32
第一節、 實驗中雄性Wistar大鼠體重與器官重量變化情形 32
第二節、 血清生化值比較 32
第三節、 組織學檢查 32
第四節、 肝臟組織中Col 1α1、Col 1α2、TGF-β及α-SMA基因表現分析 33
第五節、 肝組織中相關蛋白質表現量測定 34
第六節、 DHLA對HSC-T6是否具有細胞毒性 34
第七節、 PDGF-BB對HSC-T6增生的影響 34
第八節、 DHLA對抑制PDGF-BB造成HSC-T6增生的效果 35
第九節、 DHLA對抑制PDGF造成HSC-T6生成ROS的效果 35
第十節、 DHLA對抑制PDGF造成HSC-T6增生相關蛋白分析 35
第十一節、 TGF-β對HSC-T6造成促纖維化之影響 36
第七章、討論 38
第八章、結論及建議 42
第九章、參考文獻 43
圖表 51
附錄 68
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