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研究生:林書瑋
研究生(外文):Shu-wei Lin
論文名稱:多酚類化合物對蛋白質降解體作用之探討
論文名稱(外文):Studies on 26S Proteasome Effected by Polyphenols
指導教授:張翠玲張翠玲引用關係
指導教授(外文):Tsui-ling Chang
學位類別:碩士
校院名稱:國立臺南大學
系所名稱:生物科技學系碩士班
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:2012
畢業學年度:100
語文別:中文
論文頁數:53
中文關鍵詞:多酚類化合物蛋白質降解體細胞凋亡
外文關鍵詞:Poly ADP ribose polymeraseMethyl gallateGallic acidEllagic acidPro-caspase-3
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根據行政院衛生署對國人所做的十大死亡原因報告中,近幾年惡性腫瘤一直是國人死因的第一名。蛋白質降解體在真核生物的細胞質中普遍存在,是真核生物中對於蛋白質降解作用的主要方法之一。1980年有學者發現在惡性腫瘤病患中都有蛋白質降解體活性異常增高現象;近年來有眾多學者以抑制蛋白質降解體活性為主要目的,提供作為癌症以及其他疾病的一種潛在性治療方式,以及癌症發生的預防。多酚類的化合物具有抗發炎以及預防細胞癌化作用;本研究目的為運用多酚類化合物來對蛋白質降解體進行活性抑制,首先選用了methyl gallate、gallic acid、ellagic acid、tannic acid等四種多酚類化合物及將前三項混合在一起的複方M+G+E,探討對於不同種類的癌症細胞內之蛋白質降解體的活性影響;運用對蛋白質降解活性分析、細胞存活、西方墨點法以及細胞週期,來觀察四種多酚類化合物對於肺癌細胞Hep-2和A549以及肝癌細胞HepG-2和Huh7癌細胞的影響。從目前的研究中顯示:(1)在四種的癌細胞中,這些多酚類化合物都有效的抑制蛋白質降解體的活性。(2)對於這四種癌細胞,各類的多酚類化合不僅有效的抑制蛋白質降解體並且進一步的促進細胞凋亡。(3)多酚類化合物在肺癌細胞A549中對於S5a和20S core具有調控作用,以及造成凋亡蛋白pro- caspase-3和Poly ADP ribose polymerase表現量的減少。(4)Tannic acid和M+G+E可以促進細胞週期走向細胞凋亡。綜合以上研究,多酚類化合物對於肝癌和肺癌細胞有效的抑制26S proteasome活性;複方M+G+E的效果也比單一methyl gallate、gallic acid、ellagic acid好。
According to the top ten causes of death report of the Department of Health, malignant tumors have been the first cause of death in recent year. 26S proteasome, which is widespread in the cytoplasm of eukaryotes, is one of the roles of protein degradation in eukaryotes. In 1980, researchers have found extraordinarily high activity of 26S proteasome in patients with cancer. In recent years, inhibiting the activity of 26S proteasome can provide potential treatments for cancer and other diseases, even for the cancer prevention. The compounds of polyphenols, have anti-inflammatory effects and can prevent cancer. At the first part of the study, four kinds of polyphenols are chosen: methyl gallate, gallic acid, ellagic acid, tannic acid and combinations M+G+E. The purpose of this study is to inhibit the activity of 26S proteasome by using polyphenol. These compounds are used to test the effect on 26S proteasome activity in different cancer cells. Hep-2 and A549 cells from lung cancer cell, HepG-2 and Huh7 cells from liver cancer cell are chosen 26S proteasome activity, cell viability, Western blotting and cell cycle are used to observe the effect of the four polyphenols on in cancer cells. In conclusions, the present study showed: (1) Among these four cancer cell lines, polyphenols inhibit the activity of 26S proteasome. (2) Polyphenols not only effectively inhibit the activity of 26S proteasome but also promote cell apoptosis. (3) The levels of S5a and 20S core were regulated by polyphenols in A549 cells. Reduced pro-caspase-3 and Poly ADP ribose polymerase were shown in this study. (4) Tannic acid and M+E+G promoter cell cycle toward apoptosis. In our study, we find polyphenols can inhibit the 26S proteasome activity in lung cancer and liver cancer. The effect of combinations M+G+E is better than that of methyl gallte, gallic acid, or ellagic acid.
目錄
中文摘要 i
Abstract ii
誌謝 iv
目錄 v
圖目錄 vii
表目錄 viii
縮寫對照表 1
第一章 緒論 2
1-1 研究背景 2
1-2 蛋白質降解體與癌症的關係 3
1-3 蛋白質降解體抑制劑 4
1-4 蛋白質降解 8
1-5 蛋白質降解體 9
1-6 泛素 11
1-7 細胞凋亡 13
1-8 多酚類化合物 14
1-9 細胞株簡介 16
1-10 研究目的 18
1-11 研究架構 19
第二章 材料與方法 20
2-1 材料 20
2-1-1 化學藥品及試劑 20
2-1-2 儀器與設備 21
2-1-3 多酚類化合物 22
2-1-4 細胞株 23
2-2 實驗方法 24
2-2-1 細胞培養 24
2-2-2 繼代細胞培養 24
2-2-3 細胞冷凍保存 24
2-2-4 細胞計數 25
2-2-5 多酚類化合物對細胞存活測試 26
2-2-6 多酚類化合物對26S proteasome的活性分析 26
2-2-7 蛋白質含量測定 27
2-2-8 蛋白質電泳 (SDS-PAGE) 27
2-2-9 西方墨點法 (Western Blotting) 28
2-2-10 細胞週期分析 (Cell cycle analysis) 29
第三章 結果 30
3-1 多酚類化合物對癌症細胞內26S proteasome 之活性分析 30
3-2 多酚類化合物對癌症細胞存活性測試 31
3-3 細胞凋亡特定蛋白表現現象在A549肺癌細胞中 32
3-4 多酚類化合物對於細胞週期影響現象在A549肺癌細胞中 34
第四章 討論 36
第五章 參考資料 50
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