Estrogen Regulates ABCG2-mediated Chemotherapy Sensitivity Of MCF-7

Objective To explore the interaction between ER and GPER in the process of estradiol-mediated regulation of ABCG2 in MCF-7 cells.Methods(1) The concentration gradient was set as follows, 17-β estradiol(E2)(100, 10, 1nmol/L), G1(specificity agonists of GPER)(10, 100, 1000 nmol/L), TAM(10, 100, 1000 nmol/L). The reagents were added to MCF-7 cells treated with serum starvation 24 h. Intervention effects was evaluated by Western blot by testing the expression of ABCG2 protein. And pick out the most appropriate concentration of each reagent by analysing statistical differences between treated groups and control group.(2) Western blot and immunoflurescent were used to detect the expression and localization of ABCG2 treated with E2, G1 respectively or combined with inhibitors. The intervented cells mentioned above was treated doxorubicin(Dox), then the change of drug sensitivity was test by flow cytometry analysis and CCK8 method.Results(1) ABCG2 localizes at both of plasma membrane and cytoplasm. The expression of ABCG2 in cytoplasm was higher than that on plasma membrane in the wild type.(2) When E2 was at a concentration of 10 and 100 nmol/L, it strongly upregulated the expression of ABCG2(P<0.05) and the concentration of 10 nmol/L act more strongly. A concentration of 10 nmol/L was too low to make difference. TAM downregulated the expression of ABCG2 protein. The higher the concentration of TAM( 10, 100, 1000 nmol/L), the stronger the inhibitory effect was. The concentrations of G1 as 10, 100, 1000 nmol/L all inhibited ABCG2 express, but there were no significant difference across groups(P>0.05).(3) E2(10nmol/L) upregulated the level of ABCG2 protein and translocates it from cytoplasm to plasma membrane, which result in inhibiting the effect of chemotherapy drugs Dox. The relative protein expression of ABCG2 in E2 treatment group was higher than those of the control group(P<0.05). Above-mentioned changes were blocked by E2 antagonist(TAM)(P<0.05). The expression of ABCG2 was both downregulated in TAM(1000 nmol/L) and GPER specific angonist(G1)(10nmol/L) treatment groups(P<0.05). Cytotoxic of Dox was improved in those groups as well(P<0.05). The relative protein expression of ABCG2 were lower, which were blocked by GPER specific antagonist(G15).Conclusion(1) E2, TAM, G1 regulated the expression of ABCG2 in MCF-7 cells and it was concentrate-dependent manner.(2) Estradiol upregulates the expression of ABCG2 and weakens the effect of chemotherapy drugs in the case of activate ER and GPER simultaneously. On the contrary, acivating GPER specifically have an opposite result which improves the efficacy of chemotherapy.