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研究生(外文):Mei-Ling Chan
論文名稱(外文):Pristimerin induces apoptosis in the human breast cancer cell line MDA-MB-231
指導教授(外文):Chin-Chung WuYang-Chang Wu
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Pristimerin是一種triterpenoid,對多種細胞株具有細胞毒殺活性,但是其明確的作用機轉尚未被研究。在本研究中,pristimerin會抑制乳癌細胞MDA-MB-231的生長,其IC50值約為0.61μM。Pristimerin會增加subG1比例,活化caspase-3,並且造成poly (ADP-ribose) polymerase (PARP) 斷裂。以非選擇性caspase 抑制劑 (Z-VAD-fmk)前處理後,會明顯抑制pristimerin所引起的subG1增加,表示pristimerin引起的細胞凋亡和caspase有關。然而pristimerin不會影響pro-apoptotic proteins,Bax和Bad以及anti-apoptotic proteins,Bcl-2和Bcl-xL的表現,顯示pristimerin並非藉由調節Bcl-2 family proteins的表現來引發細胞凋亡。Cyclosporin A是一種粒線體mitochondrial permeability transition pore (PTP)抑制劑,會抑制pristimerin引起的cytochrome c釋放; 另外,在intact cell中,也會抑制PARP cleavage。因此,我們認為pristimerin是經由直接作用在粒線體,使PTP打開,cytochrome c釋出,進而活化caspase cascade,最後引起細胞凋亡。

Pristimerin, a triterpenoid from plants, has been shown to cause cytotoxicity in several cancer cell lines. However, the mechanism for the cytotoxic effect of pristimerin was never explored. In the present study, pristimerin inhibited the growth of MDA-MB-231, a human breast cancer cell line, with an IC50 value of 0.61 mM. Pristimerin treatment increased the subG1 cell population of MDA-MB-231 cells, and caused the activation of caspase-3 and the cleavage of poly(ADP) ribose polymerase (PARP) in a concentration- and time-dependent manner. Furthermore, cells pretreated with a pan-caspase inhibitor Z-VAD-FMK markedly prevented pristimerin-induce apoptosis. These results suggest that pristimerin could induce caspase-dependent apoptosis in MDA-MB-231 cells. Interestingly, pristimerin did not significantly affect the levels of pro-apoptotic proteins, Bax and Bad, and anti-apoptotic proteins, Bcl-2 and Bcl-xL. Therefore, pristimerin seems to induce apoptosis in MDA-MB-231 cells through a novel mechanism which is independent of Bcl-2 family proteins. Cyclosporin A, a mitochondrial mitochondrial permeability transition pore (PTP) inhibitor, inhibited the pristimerin-induced cytochrome c release in a cell-free system and PARP cleavage in the intact cells. These suggested that pristimerin caused the opening of mitochondrial permeability transition pores, which resulted in cytochrome c release and following caspase-dependent apoptosis.

實驗材料與方法…………… 28
結果………………………… 36
圖表………………………… 42
討論與結論………………… 58
參考文獻…………………… 64

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