Abstract： The aim of this study was to investigate the effects of hirsutine on apoptosis of breast cancer cells and its possible mechanism.The MCF-10A,MCF-7 and MDA-MB-231 cells were treated with hirsutine at different concentrations for 48 h or incubated with 160 μmol/L hirsutine for 24,48,and 72 h.The MCF-10A cell line is a non-tumorigenic epithelial cell line,and the MCF-7 and MDA-MB-231 are human breast adenocarcinoma cell lines.CCK-8 assay was employed to detect the cell viability.Flow cytometry was used to assay the apoptosis and mitochondrial membrane potential (MMP).The protein expressions of Bcl-2,Bax,cleavedcaspase 9,cleaved-caspase 3 and cytochrome C (Cyt C) in the MDA-MB-231 cells were detected by Western blotting.The results showed that hirsutine remarkably reduced the viability of MCF-7 and MDA-MB-231 cells in a time-and dose-dependent manner (P ＜0.05) with IC50 values of 447.79 and 179.06 μmol/L,respectively.In the MDA-MB-231 cells,hirsutine induced apoptosis and depolarization of MMP (P ＜ 0.05),released Cyt C from mitochondria (P ＜ 0.05),and activated caspase 9 and caspase 3 (P ＜ 0.05).However,these effects induced by hirsutine were all inhibited by cyclosporin A (CsA) (P ＜ 0.05),a specific inhibitor of mitochondrial permeability transition pore (MPTP).In addition,hirsutine down-regulated the protein level of Bcl-2 and up-regulated the protein level of Bax (P ＜ 0.05).These results suggest that hirsutine may induce apoptosis of human breast cancer MDA-MB-231 cells through decreasing the ratio of Bcl-2 to Bax,opening MPTP,releasing Cyt C from mitochondria,and activating caspase 9 and caspase 3.