Study Of The Combined Effect And Its Mechanism Of Genistein And EGF On Human Breast Cancer Cell MCF-7

Genistein(4'5,7—Trihydroxyisoflavone) is one of the isoflavones rich in plant foods, such as soybean, various vegetables and fruit,etc. There are many biological effects for genistein in vivo or in vitro such as anti-oxidizatione,anti-cancer etc.It has been shown in epidemiological investigations ,experiments in cells and animals that there was chemo-preventive effect for genistein on many cancers such as breast cancer,colon cancer and prostate cancer,etc, whose incidence were higher in western countries, while the mechanism of genistein has not been clear yet.The epidermal growth factor(EGF) is an important factor that promotes cell proliferation. It can combine with EGFR, and play an important role in the regulation of cell proliferation and propergation by signal transduction. As the level was higher in women with breast cancer, and there was few data about the biological effect of genistein on human breast cancer cell MCF-7 under high EGF situation, study of combined effect of genistein and EGF was made to learn its influence to cell proliferation, cell cycle and expression of related proteins, so as to provid a new idea for the study of molecular mechanism of genistein.on cell proliferationPart 1 The combined effect of genistem and EGF on the proliferation of human breast cancer cell MCF-7MTT method, inverted microscope observation and cells cluster formation test were applied to observe the effect of genistein and EGF on the proliferation of human breast cancer cell MCF-7. The MCF-7 cells were treated with 0.0,12.0,25.0,50.0 and 100.0μmol/L genistein respectively. 2 days,4 days and 6 days later, the proliferation status of MCF-7 was observed with MTT method. It was shown that more and more obvious cell proliferation inhibition occurred asthe concentration of genistein increased, and 50.0umol/L of genistein,4 days was the optimal concentration and observation time respectively.The MCF-7 cells were also treated with 0.0,2.04,4.15,8.30,and 12.75nmol/L EGF for 2 days,4 days and 6 days respectively. It was shown in the result of MTT method that more and more obvious cell proliferation promotion appeared as the concentration of EGF increased, and 8.30nmol/L of EGF, 4 days was the optimal concentration and observation time respectively.When the MCF-7 cells were treated with 50.0umol/L genistein and 8.30nmol/L EGF at the same time,the cell proliferation was obviously inhibited and the degree was significantly higher than that in cells treated only with genistein. It was shown in the test of 2x2 factorial-analysis that there was interraction between genistein and EGF in the MCF-7 proliferation.lt could be observed in the inverted microscope that the cells in control group was gland-epidermal like with various capes as in normal status.Gland-tube-like structure was observed before they grew full in a single layer,and iso-colloid substance could be seen inside, and there was a trend for the cells to grow in clusters after they grew full in the single layer. When they were treated with genistein and EGF at the same time, the cells became round, conjunction between the cells reduced greatly, the cells grew in dispersion,and the inhibition degree of the cell density was the highest. Part 2 The combined effect of genistein and EGF on the cycle of human breast cancer ceO MCF-7The flow cytometry analysis was used to observe the effect of genistein and EGF on the cycle of the human breast cancer cell MCF-7. The MCF-7 cells were treated with 0.0,12.0,25.0,50.0 and lOO.Oumol/L genistein and 0.0,2.04, 4.15, 8.30 and 12.75nmol/L EGF for 2days, 4 days and 6 days respectively, the cell cycle was analyzed with flow cytometry analysis. It was shown that cell cycle was blocked in G2-M interval by genistein, and this change became greater and greater as the genistein concentration and exposure time increasd.When the cellswere treated with EGF, there was no G₂-M block in the cell cycle,but the cell percentage in G₁ intreval decreased and the cell percentage in S interval increased as the EGF concentration and exposre increased.When the...