肝細胞癌是一種常見的惡性腫瘤，為全球最好發的癌症之一。而且在臺灣，最新的統計結果顯示男性是排在第二位，女性則是在第四位。在本篇研究中，我們的數據顯示 ABT-751 能夠誘導肝癌細胞株 Hep-3B 其自噬作用以及細胞凋亡的發生。本篇研究的目的，在於探討以 ABT-751 在 Hep-3B 裡誘導自噬作用以及細胞凋亡的作用機制。ABT-751 是一種新型的口服抗微管蛋白的藥物，在過去的研究中顯示 ABT-751 可以透過和 beta-tubulin 的結合，而達到抑制微管的效果。在本實驗室之前的研究中，已經於另一株肝癌細胞株 Huh-7 中證實 ABT-751 會導致細胞起始自噬作用，最終會導致細胞凋亡的發生，且自噬作用是扮演延緩細胞凋亡的角色。本篇研究一開始，藉由 MTT assay，我們決定以 2 μM 做為作用濃度。實驗中發現 ABT-751 會透過影響微管蛋白而去抑制 Hep-3B 細胞的分裂。此外，透過 Cyto-ID、Acridine orange staining 以及西方墨點法試驗分析顯示，在 24、48 小時，ABT-751 會誘導 Hep-3B 啟動自噬作用。為了得知自噬作用在本篇研究中是扮演何種角色，我們利用了自噬作用的抑制劑 3-Methyladenine (3-MA)以及 bafilomycin A1 (BafA1)。結果顯示，當 ABT-751所誘導的自噬作用被抑制時，Hep-3B 細胞其細胞凋亡的比例有所增加。總結實驗結果顯示，ABT-751 除了可以透過影響微管蛋白而去抑制 Hep-3B 細胞的分裂外，還可以誘導自噬作用的發生。而當自噬作用被抑制時，其造成細胞凋亡的比例將會有所提升。

One of the most prone to diagnosed cancer in the world, Hepatocellular carcinoma (HCC) is a common malignant tumor. Besides, the latest statistics show that HCC is the second most common cancer in men and forth in women in Taiwan. In this study, data show that in hepatocellular carcinoma cell line, Hep-3B, ABT-751 can induce autophagy and apoptosis. This study is aim to investigate the mechanism of ABT-751-induced autophagy and apoptosis in Hep-3B.ABT-751 is a novel oral administration of anti-microtubule drugs. In previous studies, it has been demonstrated that ABT-751 can bond with β-tubulin to inhibit microtubules. In our research, we decided to use 2 μM as the application concentration by MTT test. ABT-751 is found to inhibit the splitting of Hep-3B cells by affecting microtubules. Besides, results of Cyto-ID, Acridine orange staining and western blotting show that ABT-751 induced autophagy at 24 and 48 hours. By using the autophagy inhibitors 3-Methyladenine (3-MA) and bafilomycin A1 (BafA1), we tend to investigate the role of autophagy during this study, results show that when autophagy is inhibited, the percentage of apoptotic cell would increase. In all, results indicated that ABT-751 inhibited splitting of Hep-3B by affecting microtubules, still, ABT-751 induced the occurrence of autophagy. Additionally, the proportion of apoptotic cells will increase, when the autophagy is inhibited.

論文審定書 i
致謝 ii
中文摘要 iii
英文摘要 iv
圖次 vi
表次 vii
壹、緒論 (Introduction) 1
貳、實驗與材料方法 (Materials and methods) 5
參、結果 (Results) 20
肆、討論 (Discussion) 34
伍、參考文獻 (References) 37
陸、Supplementary Data 42

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