Licochalcone B Arrests Cell Cycle Progression And Induces Apoptosis In Human Breast Cancer MCF-7 Cells

Background:Licochalcone B(LCB)as an anti-inflammatory agent has been obtained patent licenses.Emerging evidence shows that LCB may be a promising alternative compound with anti-cancer activities.However,the anticancer mechanism of LCB in MCF-7 cells has not been fully investigated.Objective:The study aimed to investigate the effects of licochalcone B(LCB)on the cell cycle and apoptosis in human breast cancer MCF-7 cells and to expound the mechanism of apoptosis of MCF-7 cells induced by LCBMethods:Cell proliferation inhibition was determined by sulforhodamine B assay.In order to observe the morphology of apoptosis of MCF-7 cells which treated with different concentration of LCB dyed by Hoechst33258 fluorescent.We used annexin V fluorescein isothiocyanate(FITC)and propidium iodide(PI)labeling to measure the apoptosis in MCF-7 cells.Cell-cycle progression unduced by LCB was examined by Flow cytometry.Total intracellular reactive oxygen species(ROS)levels was detected by reactive oxygen species assay kit.Mitochondrial membrane potential(MMP)was measured by flow cytometry after treatment of mitochondrial membrane potential detection kit staining.The mRNA and protein levels of cell cycle-related proteins(cyclin A,CDK2,CDC25A,P21)and apoptosis-associated proteins(Caspase3,Caspase9,Bcl-2,Bax,cytochrome C,P53)were examined by RT-qPCR or western blot,respectively.Results:LCB inhibited MCF-7 cells proliferation in a concentration-and time-dependent manner.The morphological changes of MCF-7 cells treated with LCB showed typical morphological features.The apoptotic rate was increased and showed concentration-dependent.In addition,treatment with LCB after 24h showed that the cell progression in S phase increased and declined in G1 or G2/M phase.LCB-treatment led to S phase arrest in MCF-7 cells,which could be elucidated by the decreased mRNA and protein levels of Cyclin A,Cdk2 and Cdc25 A,and the increased protein level of p21.Licochalcone B treatment induced the total ROS level increased in a concentration-and time-dependent mannar.Moreover,with LCB concentration increased,it led to the loss of MMP,resulting in the release of cytochrome C.The above apoptotic events were supported by the fact that LCB upregulated the mRNA and protein levels of Caspase 3,Caspase 9 and Bax,and downregulated the mRNA and protein level of Bcl-2,which was triggered by the increased p53 protein level in LCB-treated MCF-7 cells.Conclusion:In summary,this study demonstrated that LCB could induce apoptosis through cell cycle arrest and mitochondrialinduced apoptotic pathway in MCF-7 cells.Treatment with LCB could activate p53 and its downstream effector p21,and then induced S-phase arrest.Moreover,the activation of p53 and p21 can also induce cytochrome c release from mitochondria to cytosol,promoting the activation of caspase-9 and caspase-3 and the down-regulation of Bcl-2.Together,our findings provide the substantial evidence of anticancer activities of LCB,and reveal the underlying mechanism of LCB in human breast cancer MCF-7cells.