| Summary: | 碩士 === 中山醫學大學 === 醫學分子毒理學研究所 === 99 === Lung cancer is the top ten causes of death worldwide due to its high incidence and mortality rates. In Taiwan, lung cancer had even become the leading cause of death in the last decade. Some target drugs such as Iressa were applied in the treatment of lung cancer patients clinically; the high-priced and severed side-effects of this kind of target drugs, however, limited their medical application. Lots of efforts have been made to pursuit of new compounds which are capable of eliminating lung cancer cells more effectively. Cyclodepsipeptides, a wide variety of cyclic peptides of natural origin, have a wide range of biological activities, such as immunosuppressant, antibiotic, antifungi, antiinflammatory or antitumoral activities. Besides, many of these cyclic depsipeptides represent useful tools for the research of biological processes involved in cellular regulation. Destruxin B, isolated from entomopathogenic fungus, is one of cyclodepsipeptides with insecticidal and phytotoxic activities. Previous report showed that the combination of Iressa with Destruxin B increased the killing effect in lung cancer cells; the effect of destruxin B alone on lung cancer cells, however, has not been reported yet. In this study, the toxic effect and the underlying mechanisms of destruxin B on A549 lung cancer cells was investigated. Our results showed that destruxin B inhibited the growth of HUVEC (Human Umbilical Vein Endothelial Cell), MLF (Mouse Lung Fibroblast), MRC-5 (Human Lung Fibroblast) and A549 (Human Lung Cancer Cells) in both dose- and time-dependent manners, and the IC50 was determined as 7.0 μM、3.3 μM、7.5 μM and 4.9 μM for each cell line at 48 h treatment. Data from flow cytometric analysis showed that destruxin B induced apoptotic cell death evidenced by increasing of subG1 population. Moreover, the activity of caspase - 2, - 3 and -9, but not caspase-8 and -12, were induced by destruxin B-treated A549 cells. Additionally, destruxin B increased the expression levels of PUMA, a pro-apoptotic protein, while decreased Mcl-1 levels, which is recognized as an anti-apoptosis molecule. Interestingly, Bax translocated from cytoplasm to mitochondrial membrane instead of increasing protein levels after administration of destruxin B. Knockdown of Bax by shRNA effectively attenuated destruxin B-triggered apoptosis in A549 cells. These results pointed that the destruxin B increased the expression levels of PUMA and decreased Mcl-1expression, which might subsequently induce mitochondrial translocation of Bax, and then activates the caspase cascade, finally leds to apoptotic cell death in A549 cells. In addition, destruxin B also increase the expression levels of p53, a tumor suppressor. Destruxin B also inhibited the growth however, A549-shp53 and H1299 ( p53 null ) cells as well, indicated that p53 might not involve in destruxin B-induced apoptosis in lung cancer cells. Taken together, our findings suggest that destruxin B-induced apoptosis in A549 lung cancer cells is via Bcl-2 family-dependent mitochondrial pathway.
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