Alteronol induced apoptosis of mouse melanoma B16-F0 cells

Aim To evaluate the mechanism of apoptosis induced by the alteronol in mouse melanoma cell line B16F0.Methods 3-(4,5-Dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide(MTT) method was used to test cell viability;Trypan blue exclusion method was used to determine cell fatality rates of B16F0 cells;Acridine orange / ethidium bromide(AO / EB) and Hoechst 33258 staining was used to observe cell morphological changes;Apoptotic was determined by staining cells with annexin V fluorescein isothiocyanate(FITC) and propidium iodide(PI) double staining of Flow Cytometry was used to determine the cell apoptotic rate;Changes of Caspase-9 and Caspase-3 were measured by Caspase-9 /3 kit and Real-Time fluorescence quantitative method to detect the expression of Bax / Bcl-2.Results Alteronol was able to inhibit the proliferation of B16F0 cells in a dose-dependent manner(P 0.05 or P 0.01);The typical morphological changes in apoptotic B16F0 were observed under the fluorescent microscrope;With the increase of the concentration of alteronol,the number of apoptotic cells increased in a concentration dependent way(P 0.05 or P 0.01);Caspase-3 and Caspase-9 activities in B16F0 cells elevated(P 0.05 or P 0.01);The level of the expression of Bax / Bcl-2 was up-regulated(P 0.01).Conclusion These findings demonstrate that alterornol may induce apoptosis of B16F0 cells through up-regulating the level of the expression of Bax / Bcl-2,while activating Caspase-9 and Caspase-3.In summary,the results suggest that atteronol induces B16F0 apoptosis through a mitochondrial regulation mediated by endogenous pathways.