18β-glycyrrhetinic acid-induced apoptosis and changed intracellular Ca~(2+) concentration in human breast carcinoma cells

Background and purpose:18β-glycyrrhetinic acid(GA) is one of the important components of glycyrrhiza.Recent years,studies showed that GA has the effect of proliferation inhibition in human acute lymphoblastic leukemia cells,human liver carcinoma and lung cancer cells.We investigated the effects of GA on induction of apoptosis in human breast carcinoma(MCF7) cells.The previous studies have demonstrated that the dynamic change of intracellular free Ca~(2+) concentration([Ca~(2+)]i)plays important roles in many links of apoptosis-induced process.Therefore we also researched the relationship between GA-induced apoptosis and [Ca~(2+)]i in MCF-7 cells.Methods:After MCF-7 cells were treated with 50-250 μmol/L GA for 24 h,cell viability for proliferation was assessed by MTT assay.MCF-7 cells treated with 100 μmol/L and 150 μmol/L GA for 24 h,the apoptotic rates in MCF7 cells were examined by terminal deoxynucleotide transferase mediated dUTP nick-end-labeling method,flow cytometry with Annexin V/ propidium iodide fluorescent stain and single cell gel electrophoresis assay(SCGE).For cells treated with 150 μmol/L GA for 24 h,i was measured by Fure-2 fluorescein load method.For cells treated with 150 μmol/L GA combined with 100 μmol/L BAPTA-AM or 0.5 mmol/L EGTA for 24 h,cell apoptosis were examined by SCGE.Results:For cells treated with GA from 100 μmol/L to 250 μmol/L,the rate of proliferative inhibition was increased significantly(P0.01 and P0.05).The degrees of inhibition were dose-dependent.IC50 of GA-inhibited proliferation was 234.33 μmol/L.For cells treated with 100 μmol/L and 150 μmol/L GA,the rates of cell apoptosis rose remarkably(P0.01 and P0.05).[Ca~(2+)]i after treatment with 150 μmol/L GA was much higher than that of control(P0.05).The rate of cell apoptosis after treatment with the combination of GA and BAPTA-AM or EGTA was markedly lower than that treated with a single GA(P0.05and P0.01).Conclusions:GA has the effects of proliferation inhibition and apoptosis induction in MCF-7 cells.GA-inducted apoptosis plays main role in inhibiting MCF-7 cell proliferation.The apoptosis induced by GA depends on rise of [Ca~(2+)]i.It is the part cause of the rise of [Ca~(2+)]i that extracellular free Ca~(2+) flowed into MCF-7 cell.