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研究生:吳靜芬
研究生(外文):Jin - Fen Wu
論文名稱:桑椹花青素誘發程序性死亡及細胞週期停止之機轉
論文名稱(外文):Induction apoptosis and cell cycle arrest by Mulberry Anthocyanins
指導教授:王朝鐘王朝鐘引用關係
指導教授(外文):Chau-Jong Wang
學位類別:碩士
校院名稱:中山醫學大學
系所名稱:生物化學研究所
學門:生命科學學門
學類:生物化學學類
論文種類:學術論文
論文出版年:2003
畢業學年度:91
語文別:中文
論文頁數:82
中文關鍵詞:桑椹花青素人類胃癌細胞細胞凋亡細胞週期抗氧化作用
外文關鍵詞:MulberryAnthocyaninsAGSapoptosiscell cycleantioxidant
相關次數:
  • 被引用被引用:4
  • 點閱點閱:295
  • 評分評分:
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:2
近年來,花青素已廣泛被證實其具有抗氧化以及抗腫瘤促進作用之功效。本研究室先從桑椹果實中萃取出花青素,並加以作定性及定量。在定量方面,計算萃取桑椹花青素中含有85-95%的花青素。而定性方面,利用HPLC分析出桑椹花青素中含有cyanidin的成份。當確定桑椹花青素之後,就可以進行下列實驗:篩選八種細胞株( Chang liver, Hep G2, MCF-7, KB, Caco-2, Hep 3B,HL-60, and AGS )的存活測試。結果顯示:人類胃癌細胞( Human gastric carcinoma, AGS )對於桑椹花青素抑制細胞存活的敏感度最高,而且呈現dose-dpendent。往後以 AGS 為實驗模式,處理桑椹花青素在不同劑量( 1.0, 2.0, 3.0, 5.0 mg/ml )下,觀察細胞凋亡( apoptosis )的現象。我們發現隨著桑椹花青素濃度增加對於造成細胞凋亡的現象也愈顯著:包括細胞型態的改變,以及 DNA fragmentation 、造成細胞凋亡的蛋白質( Cytochrome c, Caspase 3, p53, bax )有明顯表現。進一步,流式細胞儀分析中,處理桑椹花青素 5.0mg/ml 的AGS 在hypodiploid phase中 apoptotic cell 高達 33.86%;在Western Blots中,也發現p21有被活化,導致細胞週期停滯在G0/G1 期。另外,在處理AGS細胞後,在MAPK pathway中,JNK的蛋白質有明顯有活化的表現,因此,推斷桑椹花青素會活化p53路徑而導致AGS細胞有cell cycle arrest;並透過誘發Fas產生而進一步在活化JNK而使細胞走向凋亡。
As shown in previous studies Mulberry Anthocyanins (MACs) had demonstrated widely antioxidant and antitumor promotion effects previously. In our study, first we used Mulberry is a nature fruit containing a lot of pigments that was found to possess antioxidative activity. In this study, Mulberry anthocyanins (MACs) were isolated from the dried fruit of Mulberry to know the effect of MACs on eight kinds of cells (Chang liver, Hep G2, MCF-7, KB, Caco-2, Hep 3B, HL-60, and AGS ). These results showed that human gastric carcinoma AGS cells was the most susceptible to MACs inhibit the survival of cells in a concentration-dependent manner. And then the study revealed that AGS cells underwent DNA fragmentation and morphological changes characteristic of apoptosis after a 24-hrs treatment with MACs ( 0, 1, 2, 3, 5 mg/ml ). Flow cytometric analysis of cells treated with MACs for 24 hrs showed that the cells were distributed mainly in the hypodiploid phase ( apoptotic peak, 33.86% ). Moreover, MACs treatment caused an increase in the level of apoptotic proteins ( p-P53, Bax, Cytochrome c, and Caspase 3 ) and the expression of p21 was induced significantly. Furtherrmore, we found that the cell cycle arrest. On the other hand, we also detected the MAPK pathway including the expression of phosphor-JNK affected cell apoptosis. Thus, our results suggest that MACs is an apoptosis inducer in human gastric carcinoma cells. Conclusively, we suggest that MACs can arrest the cell cycle through activating p53 pathway; and MACs can induce cell apoptosis via activing JNK pathway.
壹、中文摘要..............................................1
貳、英文摘要..............................................2
參、緒論..................................................3
一、花青素簡介………………………................................................................3
二、細胞凋亡及停滯細胞週期的機制................................................................8
肆、研究動機.............................................12
伍、實驗方法.............................................13
陸、實驗藥品與器材........................................28
柒、結果..................................................32
捌、討論..................................................38
玖、實驗結果圖表..........................................43
拾、參考文獻..............................................76
拾壹、參考圖表............................................82
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