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研究生: 林楷涵
Kai-Han Lin
論文名稱: 三環藥物Teroxirone對非小細胞肺癌細胞增生機制的探討
The Mechanistic Study of Growth Inhibition by Teroxirone in Human Non-Small Cell Lung Cancer Cells
指導教授: 方剛
Fang, Kang
學位類別: 碩士
Master
系所名稱: 生命科學系
Department of Life Science
論文出版年: 2012
畢業學年度: 100
語文別: 中文
論文頁數: 78
中文關鍵詞: TeroxironeDNA損傷細胞凋亡
英文關鍵詞: Teroxirone, DNA damage, apoptosis
論文種類: 學術論文
相關次數: 點閱:216下載:10
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  • 肺癌細胞依其組織型態可分為兩類:小細胞肺癌 (small cell lung cancer, SCLC) 以及非小細胞肺癌 (non-small cell lung cancer, NSCLC),而其中大多數屬於非小細胞肺癌。目前在臨床治療上經常使用的抗癌藥物 (例如: camptothecin, doxorubicin, etoposide, cisplatin) 容易產生抗藥性,導致治療的困難,因此本文開發目前臨床上還未徹底了解,但具有抗癌效果的化合物。
    研究目標是發展小分子且低劑量的三環氧化物teroxirone,了解teroxirone如何引發細胞凋亡。實驗以MTT及細胞群落分析 (colony formation assay) 確認teroxirone確實能降低非小細胞肺癌的存活率。接著要確認非小細胞肺癌細胞的存活率下降的確是由於細胞凋亡所造成。再利用單細胞凝膠電泳 (single cell gel electrophoresis (SCGE) / comet assay),偵測teroxirone是否造成DNA 損傷。再利用propidium iodide (PI) 染色觀察teroxirone對細胞週期的影響,再利用二維流式細胞儀及DNA片段化實驗觀察teroxirone所誘導的細胞凋亡。
    實驗結果顯示,teroxirone都會對NSCLC細胞造成不同程度DNA損傷,且細胞凋亡與p53的基因型有關。

    Lung cancer according to biological characteristics can be classified two types: small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC), most of which belong to non-small cell lung cancer. Clinical treatment used anticancer drugs (ex: camptothecin, doxorubicin, etoposide, cisplatin) frequently is easy to become resistant, leading to difficulties of treatment. The purpose of the thesis is to study the effectiveness of potential anticancer drugs.

    The goal of this study is to understand how low doses of teroxirone induce NSCLC cell apoptosis. Teroxirone can reduce the proliferation rate of non-small cell lung cancer on indicated by MTT assay and colony forming assay. To confirm the reduced NSCLC cell growth rate is indeed due to cell death. To be have used the single cell gel electrophoresis (SCGE) / comet assay to detect DNA damage. To be have used propidium iodide (PI) staining to observe that impact of teroxirone on cell cycle and the induced apoptosis by flow cytometry. Double staining with PI and Annexin V – FITC of cell death for apoptosis. At the same time, DNA fragmentation also confirmed teroxirone indeed induced cells apoptosis.

    The experimental results showed that teroxirone caused different degrees of DNA damage in NSCLC cells, but is caused apoptosis only occurred in p53 wild-type cells.

    中文摘要 4 英文摘要 5 圖目錄 6 一.緒論 8 1.文獻評述 8 (1)肺癌. 8 (2)現行使用之肺癌藥物 9 (3)具有抗癌功效的天然三環氧化物 ― Triptolide 11 (4)α-1,3,5-triglycidyl-s-triazinetrione (teroxirone) 相關研究及副作用 12 (5)DNA 損傷會造成細胞週期停滯,導致細胞凋亡 14 (6)p53與DNA 損傷的影響 15 (7)p53在DNA 損傷、 細胞週期停滯及細胞凋亡中所扮演的角色 16 (8)細胞凋亡 17 2.研究目標 20 二.材料與方法 21 1.細胞株 21 2.細胞培養 21 3.新藥物的開發與研究 22 4.3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay 23 5.細胞群落形成分析 (Colony forming assay, CFA) 23 6.Single cell gel electrophoresis (SCGE/Comet assay) 24 7.以propidium iodide (PI) 染色觀察細胞週期分布 25 8.以propidium iodide (PI) 和Annexin V雙染色觀察細胞凋亡 26 9.DNA片段化 (DNA fragmentation) 27 10.以caspase抑制劑抑制細胞凋亡 28 三.結果 30 1.Teroxirone會抑制非小細胞肺癌細胞的增生 30 2.低濃度的teroxirone造成非小細胞肺癌細胞產生不同程度DNA的損傷 32 3.Teroxirone會先讓非小細胞肺癌細胞的細胞週期停滯在S及G2/M期,接著再使sub G1細胞數增加 33 4.Teroxirone會導致有p53基因的非小細胞肺癌細胞產生細胞凋亡 34 5.DEVD-CHO可以抑制由teroxirone所誘導的細胞凋亡 35 四.討論 37 1.Teroxirone抑制非小細胞肺癌細胞的增生和p53基因有關 37 2.低濃度的teroxirone造成非小細胞肺癌細胞產生不同程度DNA的損傷 38 3.Teroxirone會先讓非小細胞肺癌細胞的細胞週期停滯在S及G2/M期,接著再使sub G1細胞數增加 38 4.Teroxirone會導致有p53基因的非小細胞肺癌細胞產生細胞凋亡 39 5.DEVD-CHO可以抑制由teroxirone所誘導的細胞凋亡 40 五.參考文獻 42 六.圖 52

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