Induction Of Apoptosis In Human Breast Cancer MCF-7 Cells By The Acetoacetate Extract Of The Vitex Negundo Seeds And Its Molecular Mechanism

Objective To investigate the effects of the acetoacetate extract of the Vitex negundo seeds(EVn-50) on apoptosis of human breast cancer MCF-7 cells,and explore its molecular mechanism.Methods MCF-7 cells were cultured in vitro. Fluorescence microscopy after AO/EB double staining was used to observe change of apoptotic cell morphology.Flow cytometry with FITC labeled annexin V/PI staining was used to determine the percentage of apoptotic cells.DNA agarose gel electrophoresis was used to observe cell gene DNA ladder.Caspase-3 activity in MCF-7 cells was determined by colorimetry.Western Blotting was used to analyze the expresson of Mcl-1,Bax and Bcl- 2 protein in MCF-7 cells.Result AO/EB fluorescence microscope double staining results showed that: Evn-50 at 3.0μg/ml,72h typical apoptotic morphology induced MCF-7 cells increased (P <0.05).FITC-Annexin-V/PI double staining flow cytometry results showed that: Evn-50 induced apoptosis in MCF-7 cell percentage increased (P <0.05),at 3.0μg/ml, 48h most significant effect.DNA agarose gel electrophoresis found:Evn-50 can induce apoptosis in MCF-7 cells produced DNA between nucleosomes broken ladder. Caspase-3 activity test showed that:Evn-50 at 3.0μg/ml,48h strongest activation of caspase-3 activity (P <0.05).Western blotting analysis revealed that:EVn-50 effectively reduced Bcl-2,Mcl-1 protein expression and upregulation of Bax protein expression (P <0.05).Conclusions Evn-50 effectively induces apoptosis of MCF-7 cells,and its mechanism is associated with downregulation of Bcl-2 and Mcl-1 expression, upregulation of Bax protein expression, and activation of caspase-3.