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| 題名: | 2-Acetyl-4,8-dihydrobenzo-dithiophene- 4,8-dione-19 (ADB-19) 對HL-60血癌細胞之抗癌作用與機制研究;Anticancer Activities of 2-Acetyl-4,8-dihydrobenzo- dithiophene-4,8-dione-19 (ADB-19)on HL-60 Leukemia Cells |
| 作者: | 林育賢;Yu-Hsien Lin |
| 貢獻者: | 中國醫藥大學醫學研究所 |
| 關鍵詞: | 抗癌藥物;血癌細胞;Anticancer Drug;Leukemia Cells |
| 日期: | 2004 |
| 上傳時間: | 2010-01-20 16:53:03 (UTC+8) |
| 摘要: | 先前郭盛助博士等人的研究中指出，ADB-19是2-acetyl-4,8-dihydrobenzodithiophene-4,8-dione (ADB) 系列衍生物中對人類癌細胞株最具毒殺性。在本篇論文中，主要的研究目的是探討ADB-19如何抑制血癌細胞株HL-60增生及其機制。 首先我們分析ADB-19 對血癌細胞株 (HL-60，U937與K562)及健康人類周邊單核血球細胞 (PBMC, peripheral blood mononuclear cells) 之細胞存活率、細胞週期與凋亡的影響。HL-60，U937與K562細胞經ADB-19處理24小時後，細胞存活率呈現劑量與時間依存性的抑制，其50 % 抑制生長濃度 (IC50) 分別為 0.4、0.23 與? 0.24 μM， 且ADB-19對健康人之細胞毒殺作用低於HL-60細胞。細胞週期分析顯示ADB-19明顯地促使sub-G1增加，DAPI染色觀察亦確證ADB-19處理的細胞核呈現碎裂狀，此外，藉由DNA電泳分析證實ADB-19會促使核DNA呈梯度斷裂。ADB-19處理後1小時細胞內ROS量顯著上升，而12小時後粒線體膜電位顯著地降低。Caspases -3, -8, 以及 -9 的活性亦在ADB-19 處理2個小時後逐漸上升，而且Caspase專一性抑制劑能抑制 各Caspase 的活性及減少細胞凋亡。西方墨點法分析顯示ADB-19誘導細胞凋亡過程中，除了Caspase-9、-3蛋白質的活化型態與其受質PARP 的裂解增加外，Bcl-2與Survivin 蛋白質量降低，而Bax蛋白質量提昇。在抗氧化劑前處理的實驗中發現，Vit.C能夠抑制ADB-19誘發之ROS生成，並且提升HL-60細胞存活率，顯示ADB-19所導致的細胞凋亡可能是由ROS所引發的。綜合上述，ADB-19誘導細胞凋亡可能經由粒線體主導的路徑所致，這過程中caspase家族、Bcl-2家族與Survivin蛋白質均參與其中。; Previously studies from Dr. Kuo Sheng-Chu reported that ADB-19 was the most potent 2-acetyl-4,8-dihydrobenzodithiophene-4,8-dione (ADB) derivative with significant cytotoxicity against several human tumor cell lines. In this study, the cytotoxic effects and mechanisms exerted by ADB-19 on HL-60、U937、K562 leukemia cells and PBMC (peripheral blood mononuclear cells) were investigated. After 24 hours of treatment with ADB-19, concentration- and time-dependent decreases in the viability of HL-60, U937 and K562 cells were observed and the IC50 values were estimated to be 0.4、0.23 and 0.24 μM respectively. The cytotoxic effect of ADB-19 on PBMC was less significant than that on HL-60 cells. After 24 hr-treatment of ADB-19, cell cycle analysis showed that ADB-19 induced significant sub-G1 peak arrest, which was consistent with fragmented nuclei by DAPI staining. The ADB-19-induced apoptosis was further confirmed by DNA fragmentation assay. ADB-19 also significantly decreased mitochondrial membrane potential in the 12 hr-treated cells. Increase of ROS (reactive oxygen species) production was observed in cells with 1 hr treatment. Caspases -3, -8, and -9 were activated at 2 hr after treatment. Induction of apoptosis in ADB-19-treated HL-60 cells was accompanied by an apparent down-regulation of Bcl-2 and survivin, and an up-regulation of Bax. The significant reduction of caspase activities and apoptosis by caspase inhibitors indicated that the ADB-19-induced apoptosis was mainly mediated by activation of caspases-9 and -3. In addition, results from anti-oxidant pre-treatment experiment showed that Vit.C elevated the viability of ADB-19 treated cells. These results suggest that ADB-19 may induce ROS production, and subscequently result in mitochrondrial-mediated apoptosis in HL-60 cells. |
| 顯示於類別: | [醫學研究所] 博碩士論文
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