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研究生:呂美津
研究生(外文):Mei-Chin Lu
論文名稱:牛樟菇子實體乙醇萃取物誘導HL60細胞凋亡之研究
論文名稱(外文):Effect of Ethanol Extract from Fruiting Bodies of Antrodis camphorata in Inducing Apoptosis of HL60 Cells
指導教授:賴宏亮賴宏亮引用關係
指導教授(外文):Horng-Liang Lay
學位類別:碩士
校院名稱:國立屏東科技大學
系所名稱:熱帶農業暨國際合作研究所
學門:農業科學學門
學類:一般農業學類
論文種類:學術論文
論文出版年:2003
畢業學年度:91
語文別:英文
論文頁數:64
中文關鍵詞:牛樟菇細胞凋亡DNA斷裂
外文關鍵詞:Antrodia camphorataApoptosisDNA fragmentationAnnexin-ⅤTUNEL
相關次數:
  • 被引用被引用:11
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牛樟菇 (Antrodia camphorata)又名樟菇、樟芝,屬擔子菌綱的藥用性真菌類,在台灣民間用於癌症治療,但是尚未有科學研究證實其抗癌作用。本研究提出牛樟菇抗癌的生物活性主要作用是誘導腫瘤細胞發生凋亡現象的假說,並由以下的實驗來證實。實驗設計中,以trypan blue exclusion、Liu’s stain及TUNEL (TdT-mediated dUTP-biotin nick end labeling)染色等檢查法,直接利用顯微鏡觀察牛樟菇子實體之乙醇萃取物誘導人類白血病前骨髓細胞(human promeylocytic leukemia cell) 發生細胞凋亡的現象。以DNA agarose electrophoresis來分析細胞凋亡後在膠體電泳呈現的DNA斷裂現象(DNA ladder),所產生清晰DNA ladder的膠體電泳結果證實細胞凋亡所得到DNA片段約為180bp倍數的特定長度。並以流式細胞儀 (flow cytometry)經propidium iodide (PI) 及 Annexin-Ⅴ-FITC雙重螢光染色後檢測凋亡細胞的比例。研究結果顯示,以200 μg/ml的牛樟菇萃取物處理6小時後,與對照組比較下,細胞凋亡的比例從少於5%增加至36%;牛樟菇乙醇萃取物處理24小時後,單染PI以流式細胞儀分析出凋亡細胞的比例從5%增加至42%。綜合以上的結果證實,以200 μg/ml的牛樟菇乙醇萃取物處理人類白血病前骨髓細胞,導致HL60細胞發生細胞凋亡的現象最為顯著。因此,本研究首度提出牛樟菇萃取物,確實可透過誘導人類白血病前骨髓細胞發生凋亡的現象而達到抗癌之作用。
Abstract
Antrodia camphorata (Niu-Chang- Ku; Zhan-Ku) is a medicinal fungus of the family Basidiomycetes that has been used as a traditional anti-tumor drug for a long time in Taiwan. Unfortunately, its anti-tumor effect has not yet been scientifically identified. In this study, we analyzed the effect of the crude extract from the fruiting bodies of Niu-Chang-Ku in inducing apoptosis of HL60 cells. After the Niu-Chang-Ku treatment, HL60 cells were observed directly under microscope by trypan blue exclusion, Liu’s stain and in situ TUNEL (TdT-mediated dUTP-biotin nick end labeling) staining for evidence of apoptosis of HL60 cells induced by ethanol extract from the fruiting bodies of Niu-Chang-Ku. Niu-Chang-Ku extract significantly increased apoptosis of HL 60 cells, and the best result was found in the treatment with 200 mg/ml Niu-Chang-Ku extract. The effect of Niu-Chang-Ku in inducing apoptosis was further quantified by flow cytometry with propidium iodide (PI) and Annexin-Ⅴ-FITC staining; increasing apoptotic cells from less than 5 % in the control groups to 36 % after 6 h treatment of 200 μg/ml Niu-Chang-Ku extracts. Furthermore, gel electrophoresis of the DNA extract of HL 60 cells showed DNA ladder phenomenon after the treatment with Niu-Chang-Ku extract. Evidence of DNA fragmentation was also shown by flow cytometry analysis of PI stained HL 60 cells. Hypodiploid population increased from less than 5 % in the control to 42 % in the 200 μg/ml Niu-Chang-Ku treatment. Our study demonstrated the scientific evidence that Niu-Chang-Ku extract induced apoptosis of the human promeylocytic leukemia cell. This also provided a solid evidence for the anticancer bioactivity of Niu-Chang-Ku.
目 錄
摘 要 …………………………… Ⅰ-ⅡAbstract …………………………… Ⅲ-Ⅳ
誌 謝 ………………………… Ⅴ-Ⅵ
目 錄 …………………………… Ⅶ
圖表索引 …………………………… Ⅷ-Ⅸ
Introduction ………………………… 2- 4
Literature Review …………………………… 5-20
Material and Methods …………………………… 21-29
Results …………………………… 30-47
Discussion …………………………… 48-51
References …………………………… 52-60
附錄一 …………………………… 61
學會及期刊發表之論文 …………………………… 62-63
作者簡介 …………………………… 64
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