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研究生: 陳建宏
Jian-Hong Cheng
論文名稱: 氧化壓迫與p53參與三氯乙烯及四氯乙烯誘導人類肺癌細胞凋亡之研究
指導教授: 黃榮南
Rong-Nan Huang
口試委員:
學位類別: 碩士
Master
系所名稱: 生醫理工學院 - 生命科學系
Department of Life Science
畢業學年度: 89
語文別: 中文
論文頁數: 65
中文關鍵詞: 三氯乙烯四氯乙烯氧化壓迫脂質過氧化
外文關鍵詞: p53, apoptosis, oxidative stress, TCE
相關次數: 點閱:15下載:0
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  • 目錄 ---------------------------------------------------------------- I 『圖』目錄 --------------------------------------------------------IV 中英文名詞對照表 ------------------------------------------------V 縮寫表 ---------------------------------------------------------------VI 中文摘要 ---------------------------------------------------------- VII 英文摘要 --------------------------------------------------------- VIII 第一章緒論 ------------------------------------------------------ 1 第一節 碳氫化物之簡介 ---------------------------------------------------1 第二節 三氯乙烯及四氯乙烯之毒理作用 ------------------------------2 第三節 活性氧物種(Reactive Oxygen Species)與氧化壓迫(oxidative stress)之生成與影響 -------------------------------------------- 5 壹、自由基(Free Radicals)和活性氧物種(ROS)之生成及特性 --------------5 貳、活性氧物種與細胞凋亡(Apoptosis) --------------------------------------6 第四節 抑癌基因(Tumor suppressor gene)之功能探討 ---------------6 壹、p53之簡介 -------------------------------------------------- ------------6 貳、p53蛋白之調控機轉探討 -----------------------------------------------7 參、p53蛋白之功能---------------------------------------------------------- 8 第五節 細胞凋亡(Apoptosis) ------------------------------------------ 9 壹、細胞凋亡的標誌(Apoptotic markers) ------------------------------------ 9 貳、細胞凋亡之訊息傳導路徑之探討 --------------------------------------10 參、細胞凋亡與癌症 --------------------------------------------------------11 第六節 研究動機與目的 -------------------------------------------------11 第二章 材料與方法 ---------------------------------------------------13 1.細胞株------------------------------------------------------------- 13 2.細胞培養---------------------------------------------------------- 14 3.藥劑處理法(氣相處理法) ------------------------------------- 14 4.脂質過氧化測定(Malondialdehyde assay) ----------------15 5.DCF螢光的測定(DCF assay) -------------------------------- 16 6.Sub-G1之檢測與DNA含量分析 -------------------------- 16 7.細胞凋亡蛋白脢Caspase-3活性分析 --------------------- 17 8.細胞存活率分析(SRB assay) --------------------------------- 17 9.西方點墨法(Western blotting) -------------------------------- 18 10.蛋白質定量 ----------------------------------------------------- 18 第三章 結果 -------------------------------------------------------------19 第一節 三氯乙烯及四氯乙烯誘引細胞活性氧物種之形成--------- 19 壹、三氯乙烯及四氯乙烯對細胞內過氧化氫濃度之影響-------------------19 貳、抗氧化劑(Antioxidants)抑制細胞脂質過氧化作用 -------------------- 19 參、抑制GSH濃度對脂質過氧化作用之影響------------------------------ 20 第二節 活性氧物種對人類肺癌細胞H460及H1299之影響-------20 壹、三氯乙烯及四氯乙烯誘引H460細胞Sub-G1峰之形成----------------20 貳、活性氧物種直接影響細胞凋亡之產生 --------------------------------21 第三節 三氯乙烯及四氯乙烯誘引H460細胞凋亡之機制探討 --- 21 壹、Caspase-3之活性表現----------------------------------------------------21 貳、p53及其下游之蛋白質Bax表現情形-----------------------------------22 參、抑制Bcl-2蛋白質間接造成細胞凋亡 ----------------------------------22 肆、p53基因轉殖株細胞(H1299-T)之存活率比較 ------------------------22 第四章 討論---------------------------------------------------------36 參考文獻 ------------------------------------------------------------------ 41 附錄 -------------------------------------------------------------------------- 49 『圖』目錄 圖一、三氯乙烯及四氯乙烯對細胞內過氧化氫濃度之影響。 圖二、抗氧化劑(D-Mannitol)抑制細胞脂質過氧化作用。 圖三、抗氧化劑(Uric acid)抑制細胞脂質過氧化作用。 圖四、抗氧化劑(Sodium selenite)抑制細胞脂質過氧化作用。 圖五、GSH對脂質過氧化作用之影響。 圖六、三氯乙烯誘引人類肺癌細胞H460之Sub-G1峰之形成。 圖七、抗氧化劑(D-Mannitol)抑制細胞凋亡之形成 圖八、三氯乙烯及四氯乙烯誘引人類肺癌細胞H460之Sub-G1峰形成之量化圖。 圖九、三氯乙烯及四氯乙烯誘引人類肺癌細胞H460內caspase-3活性增加。 圖十、三氯乙烯及四氯乙烯誘引人類肺癌細胞H460內p53和Bax 蛋白質表現。 圖十一、三氯乙烯及四氯乙烯抑制人類肺癌細胞H460內Bcl-2之蛋白質表現。 圖十二、p53基因轉殖株細胞 (H1299-T) 之存活率比較。 圖十三、三氯乙烯及四氯乙烯誘導人類肺癌細胞H460細胞凋亡之可能機制。

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