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研究生:蔡德誠
研究生(外文):De-Chang Tsai
論文名稱:3-amino-1,2,4-triazole影響人類肝癌細胞中鎘所誘導之金屬硫蛋白表現的研究
論文名稱(外文):Study on the effect of 3-amino-1,2,4-triazole on the cadmium-induces metallothioneins in HepG2 cells
指導教授:林立元林立元引用關係
指導教授(外文):Lih-Yuan Lin
學位類別:碩士
校院名稱:國立清華大學
系所名稱:生命科學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:65
中文關鍵詞:金屬硫蛋白人類肝癌細胞
外文關鍵詞:metallothioneinHepG2 cell linescadmium
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Metallothionein (MTs)是一種低分子量、含高量cystein並且容易被重金屬誘導的蛋白質。除了重金屬以外,氧化逆境也是誘導MT基因表現的一個因子。在本篇研究當中,我使用catalase的抑制劑3-amino-1,2,4-triazole (3-AT)來研究鎘在人類肝癌細胞株(HepG2)內對MT基因表現的影響。首先我以北方墨漬法,發現隨著3-AT劑量的提高能夠有效的抑制鎘或鋅所引起的MT基因的表現。 此外,另一種catalase抑制劑,salicylic acid也能夠隨著劑量的提高有效的抑制鎘所引起的MT基因的表現。其次我分析了3-AT 與salicylic acid對鎘進入細胞內的影響,結果顯示隨著salicylic acid劑量的提高能夠有效的減少細胞內鎘的含量,低劑量(10 mM)的3-AT也能夠有效的減少細胞內鎘的含量,然而高劑量(80 mM)的3-AT只能夠稍微減少細胞內鎘的含量。另一方面,我發現高劑量(80 mM)的3-AT在鎘所處理的細胞中,能夠有效的降低細胞增殖以及存活度,而低劑量的3-AT或單獨以鎘做處理,則在細胞增殖以及存活度上沒有明顯的抑制效果。 此外,以過氧化氫與鎘共同處理的細胞並不能夠增強MT的表現。這些結果均顯示3-AT抑制鎘所誘導的MT基因表現,有可能不是借由氧化逆境的產生,而是因為3-AT減少鎘進入細胞所造成的結果。

Metallothioneins (MTs) are a group of low molecular weight proteins which rich in cystein residues, and induced markedly by metal ions. In addition, oxidative stress was also a factor to induce the MT gene expression. Previous reports have been demonstrated that cadmium was able to induce the gene expression of metallothionein (MT), but the underlying mechanisms were not well understood. In this study, we used the catalase inhibitor, 3-amino-1, 2, 4-triazole (3-AT), to investigate the effects of cadmium on the gene expression of MT in human hepatoma cell lines (HepG2). First, we found that 3-AT significantly suppressed the gene expression of MT induced by cadmium or zinc using the Northern blot. 3-AT inhibited the gene expression of MT was a dose-dependent manner. Another catalase inhibitor, salicylic acid, also suppressed the gene expression of MT and shown a dose-dependent manner. Second, we tested the effects of 3-AT and salicylic acid on the intracellular level of cadmium in cadmium treatment cells. The results shown that salicylic acid significantly decreased the intracellular level of cadimium in dose-dependent manner. Low dose of 3-AT (10 mM) significantly decreased the intracellular level of cadmium, however, high dose of 3-AT (80 mM) slightly decreased the intracellular level of cadmium. Third, we found that high concentration of 3-AT (80 mM) significantly decreased the proliferation rate and viability in cadmium treatment cells. Low concentration of 3-AT or cadmium alone exhibited no significant effect in the inhibition of cell proliferation and viability. Furthermore, cotreatment of hydrogen peroxide and cadmium was not able to enhance the gene expression of MT compare with cadmium alone. These results suggested that induction of MT gene expression by cadmium might not mediate the generation of oxidative stress. It is possible that 3-AT decreased the cadmium into the cells and resulted in low intracellular level of cadmium and MT expression.

中文摘要................................ 1
英文摘要................................ 3
緒言.................................... 5
材料與方法.............................. 14
結果.................................... 24
討論.................................... 30
參考資料................................ 36
附圖.................................... 50

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