臺灣博碩士論文加值系統

在亞洲地區，蜆被廣泛認為具有保護肝臟避免損傷的功效，然而
詳細的機轉並不清楚。肝星狀細胞在肝纖維化的發展中扮演著重要角
色，當肝星狀細胞受到刺激轉變成活化態時，細胞會增生並造成細胞
外基質的累積。活化態肝星狀細胞會大量表現α-型平滑肌蛋白
(α-smooth muscle actin; α-SMA)及膠原蛋白，細胞所分泌之腫瘤壞死
因子-α (tumor necrosis factor-α, TNF-α) 及轉型生長因子-β
(transforming growth factor-β, TGF-β)也會增加。此研究的目的是探討
蜆萃取物對於活化態肝星狀細胞的影響。先以蜆萃取物培養活化態肝
星狀細胞五天，結果發現蜆萃取物可以抑制活化態肝星狀細胞的增生，
並促使細胞週期停滯於G2/M期。而蜆萃取物亦會抑制活化態細胞中
α-SMA及膠原蛋白的表現量，並降低TNF-α及TGF-β等生長因子的表
現。此結果顯示蜆萃取物可能具有抑制肝星狀細胞活化的功效，達到
保護肝臟的效果。

The freshwater clam is a widely-consumed shellfish and is used as a
remedy for liver injury and anti-alcoholic toxicity in Asia. The hepatic
stellate cells (HSCs) are central for the development of fibrosis during
liver injury. The HSCs transit from quiescent type to activated type which
is characterized by upregulation of alpha-smooth muscle actin (α-SMA)
expression and extracellular matrix secretion in hepatic fibrosis. The
tumor necrosis factor-α (TNF-α) and transforming growth factor-β
(TGF-β) were increased in activated HSCs. The aim of this study was to
investigate the effect of clam extracts on the HSCs. Activated HSCs were
incubated with clam extracts. After 5 days incubation, the clam extracts
significantly inhibited activated HSCs proliferation and cause cell cycle
arrest in G2/M phase. Reverse transcriptase polymerasechain reaction
(RT-PCR) or western blotting revealed that the expression of α-SMA and
type I collagen in activated HSC were decreased. Clam extracts could
also inhibit TNF-α and TGF-β mRNA expression in activated HSCs.
These results suggest that clam extracts could inhibit activation of HSCs.

摘要 I
Abstract II
致謝 III
圖目錄 VII
縮寫表 VIII
第一章 緒論 1
第一節 前言 1
第二節 肝臟纖維化 1
一、肝損傷 1
二、肝星狀細胞 2
第三節 蜆 5
一、概述 5
二、蜆的化學組成 6
三、蜆的生理活性 7
第四節 研究動機與設計 8
第二章 材料與方法 9
第一節 實驗材料 9
一、化學藥品及試劑 9
二、實驗儀器 11
三、細胞來源 12
四、蜆萃取物製備 12
五、其他藥品配製 12
第二節 實驗方法 14
一、肝星狀細胞分離及培養 14
二、肝星狀細胞自發性螢光測定 14
三、肝星狀細胞定性免疫螢光染色 15
四、細胞增生與存活性試驗 15
五、細胞增生及死亡試驗 16
六、細胞週期測定 17
七、西方墨點法 17
八、萃取RNA 18
九、RNA反轉錄 18
十、聚合酶連鎖反應 19
十一、二維膠體電泳 19
十二、統計分析 20
第三章 實驗結果 21
第一節 肝星狀細胞之型態分析 21
一、肝星狀細胞活化之型態轉變 21
二、靜止態肝星狀細胞的辨識 21
三、活化態肝星狀細胞的辨識 21
第二節 蜆萃取物對活化態肝星狀細胞的影響 22
一、蜆萃取物減緩活化態肝星狀細胞的增生 22
二、蜆萃取物對活化態肝星狀細胞標記蛋白質表現的影響 23
三、蜆萃取物對活化態肝星狀細胞其細胞外基質累積的影響 24
四、蜆萃取物對活化態肝星狀細胞中其他生長因子的影響 24
第三節 以二維電泳分析蜆萃取物對肝星狀細胞蛋白質的變化 24
第四章 討論 26
第五章 結論 30
第六章 參考文獻 31
第七章 圖表 38

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