Induction of apoptosis by chalepin through phosphatidylserine externalisations and DNA fragmentation in breast cancer cells (MCF7)

Chalepin, a naturally occurring compound isolated from Ruta angustifolia have been shown to exert a promising anticancer activity through various mechanisms. Hence, the need to investigate the apoptotic inducing ability of chalepin in MCF7 cells by various detection assays. Cytotoxicity screening of chalepin against MCF7 cells was conducted using SRB assay. Apoptosis induction was examined by established morphological and biochemical assays including phase contrast and Hoechst/PI staining fluorescence microscope. Similarly, Annexin-V/FITC and TUNEL assays were conducted using flow cytometry whereas caspase-3 activity was evaluated using microplate reader. The result indicates remarkable cytotoxic activity against MCF7 cells, whereas it shows moderate cytotoxic activity against MDA-MB231 cells. Interestingly, chalepin did not present any toxicity against MRC5 normal cell line. Morphological examination using both phase contrast and fluorescence microscope displays typical apoptotic features such as membrane blebbing, DNA fragmentation, chromatin condensation and apoptotic bodies' formation following chalepin treatment against MCF7 cells at different concentration for 48 h. Apoptosis induction is significantly associated with externalisation of phosphatidylserine, and DNA fragmentation in MCF7 cells chalepin treated cells when compared with control. The protein expressions of caspase-8, 9 and cleaved PARP1 were upregulated which correlated well with increased caspase-3 activity. From our recent findings, chalepin was able to induced apoptosis in MCF7 cells and therefore, could be evaluated further as a potential source of anticancer agent for cancer treatment such as breast cancer.