ABSTRACT
BACKGROUND: The natural products have increasing important for the development of anticancer agents. Colchicum baytopiorum C.D. Brickell (C. baytopiorum), an endemic species for Turkey, contains colchicine and its derivatives. Stimulation of apoptotic and autophagy-mediated cell deaths are effective strategy for anticancer therapies. AIM: The aim of the study is to determine the role of the extract on both apoptotic and autophagic cell death in HeLa cell line. METHODS: The cell viability of C. baytopiorum (0.1 mg/ml) was determined by MTT assay. Active caspase-3 and t-Bid expressions were evaluated by immunohistochemical method. The mRNA expression of apoptotic regulatory genes (Bcl-xL, Bid, Bad, PUMA, NOXA, Caspase-3, -8, -9, Fas, FADD, TRADD, TRAF2, TNF, TNFR1), autophagic cell death related genes (Atg5-12, Beclin-1, DAPK), and also both autophagic and apoptotic cell death regulatory genes (Bif-1 and BNIP-3) were investigated by qRT-PCR. RESULTS: We determined that the expressions of both apoptotic and autophagic regulatory genes were significantly increased in the treatment group compared to control group. Also, we showed that C. baytopiorum crude extract induces the cross-connection between apoptotic and autophagic cell deaths in HeLa cells. CONCLUSION: We suggested that this endemic plant extract seems to be a new promising therapeutic approach in cancer.
ABSTRACT
We aim to determine the regulation of apoptosis by paclitaxel-induced and understand cancer dynamics to treatment targets for HeLa cells by identifying decrease/increase genes expression on HeLa cells. In this study, the anti-tumor effects of Paclitaxel (PAC) on HeLa cells have been studied in order to determine the cellular and molecular mechanisms of these effects. PAC has been applied to HeLa cells in 6 different doses (3, 7.5, 15, 30, 60, 120 nM) for 48 hours and the IC50 dose MTT method, has been determined with apoptic index (AI) DAPI. Morphological aspects have been demonstrated using light, phase contrast and fluorescent microscopes, additionally activation of Caspase 3,7 and 10 have been shown using florescent spectroscopy. RT-PCR and qRT-PCR have been used to evaluate pro/anti-apoptotic gene expression. According to the parameters being evaluated; PAC has reduced cell multiplication based on dosage and time (p<0.01). 15 nM has been determined as the IC50 value. AI value has been determined as 42%. In the molecular level analyses in addition to the increase in Caspase3,7,10 activation, RT-PCR results show that bax, bak, bcl-x, bik, mcl-1 genes are expressed in the control group as well as the experimental 15 nM group; whereas bak, bcl-x ve bik genes have a decrease in expression compared to the control group. qRT-PCR results show that Apaf1, Bad, Bax, Bcl2L11, Caspase1, Caspase10, Caspase4, Caspase7, Dffa, Fas, Htra2, Lrdd, NFKB1, NFKB2, PMAIP1, RELA, RELB, TNFRSF10A, TNFRSF10C, TNFRSF10D, TNFRSF1A, TNFRSF21, TNFRSF25 gene expressions have increased significantly. On the other hand, BAG1, BBC3, Bcl2L1, Bcl2L10, Bid, Caspase2, Caspase6, Caspase8, Caspase9, FADD, FAM96A, FasLG, HRK, SOCS3, TNF, TNFSF10, TRAF5, TRAF6 mRNA levels are significantly decreased.
