The Bidirectional Effects Of Genistein And Estradiol On Human Breast Cancer Cells And Its Mechanism

Objective:Breast cancer is the most popular carcinoma in women, and it is one of the greatest threats to the health of women. Genistein is one of the main protective factors of soybean isoflavones family, and as an estrogen analogs, genistein with endogenous hormone estrogen has a co-effect on the the key points of breast cancer prevention and control. Epidemiology and in vitro research have showed that genistein had effects on the prevention and treatment of breast cancer, but the influence was different, if women began to eat soy products since puberty, genistein could inhibit the occurrence of breast cancer, and the soy isoflavone intake and risk of breast cancer is significant negative correlation before menopause, but after menopause, genistein may stimulate the growth of breast cancer, so it suggests that endogenous estrogen level could influence results of genistein on the prevention and treatment of breast cancer. Recent studies all focused on the effect of genistein on the breast cancer cells, and the endogenous estrogen was ignored, so the results were contradictory. In addition, genistein combined with the different estrogen receptor, and the consequence was different. The debate about the effects of the soy isoflavones on breast cancer is sharper. In this study, breast cancer cells MDA-MB-231 (ER beta) and breast cancer cells T47D (ER alpha + ER beta) were chosen as research subjects, and MTT and flow cytometry were used to study the influence of genistein on the proliferation, apoptosis and cell cycle of the two breast cancer, with different level of estradiol (premenopausal and postmenopausal), and genome-wide expression profile chip will be used as a high-throughput gene expression analysis tool to obtain differentially expressed genes spectrum with the different treatments.Methods:In MTT, the dose of estradiol was respectively 0 pmol/l,80 pmol/l, 800 pmol/l,1000 pmol/l,1600 pmol/l, and the dose of genistein was respectively 0 μmol/1,5 μmol/1,10 μmol/1,20 μmol/l,40 μmol/1. the MDA-MB-231 (ER beta) and the T47D (ER alpha+ER beta) were treated with them for 96 hours, and then the MTT was used to test OA value. In the flow cytometry test, the genistein concentration was 20 p.mol/1, and the estradiol level was respectively 80 pmol/1 and 800 pmol/l. In gene chip test, the dose of genistein and estradiol was same as that of in the flow cytometry test. Agilent single standard expression of gene chip was used to screen differentially expressed genes and difference gene EGFR was randomly chosen and real time quantitative PCR was applied to verify the results of gene chip.Results:In MTT, the results showed that,when the dose of the estradiol was respectively 0 pmol/1 and 80 pmol/1,20μmol/l genistein could effectively inhibit the proliferation of MDA-MB-231 (ER beta), and when the estradiol was 800 pmol/1,10μmol/l genistein can significantly inhibit the proliferation of MDA-MB-231 (ER beta), and when the estradiol was 0 pmol/1, the 10μmol/l genistein could significantly inhibit the proliferation of T47D (ER alpha + ER beta), and when the estradiol level was 80 pmol/L and 800 pmol/L,20μmol/L genistein could inhibit the proliferation of T47D (ER alpha+ER beta). In the flow cytometry test, when when the estradiol level was 80 pmol/L and 800 pmol/L,20μmol/L genistein could make MDA-MB-231 (ER beta) cell cycle arrest in G0/G1 phase and S phase, and T47D (ER alpha+ER beta) cell cycle arrest in G2/M phase. When the genistein concertration was 20μmol/L,80 pmol/L and 800 pmol/L estradiol all could make apoptosis rate of MDA-MB-231 and T47D increase significantly, compared with blank control group, and in T47D, with the estradiol concertration increasing, the apoptosis rate decreased significantly. It indicated that genistein could induce obvious apoptosis in T47D cells with low estradiol concertration. In gene chip test, there were 49 differentially expressed genes in MDA-MB-231 cells, and among them,35 were up and 14 were down. There were 84 differentially expressed genes in T47D cells, and among them,69 were up and 15 were down. The results of GO showed that in the MDA-MB-231 cells, the 49 differentially expressed genes were maily related with the biological processes and molecular function of the cell, and in T47D cells,, the 84 differentially expressed genes were maily related with biological process and cell components of the cell. The RT-qPCR results showed that the expression of EGFR was consistented with that of the chip.Conclusion:The effects of Genistein on proliferation, cell cycle, apoptosis differentially expressed genes spectrum in breast cancer cell MDA-MB-231 (ER beta) and T47D (ER alpha + ER beta) were related with the concentration of estradiol and the type of estrogen receptor, the specific mechanism remained to be further researched.