Effects Of Ellagic Acid On Proliferation,migration And Invasion Of Triple Negative Breast Cancer MDA-MB-231 Cells In Silenced And Overexpressed By BRCA1 Gene

ObjectTo investigate the effects and underlying mechanism of ellagic acid(EA)on the biological behaviors of BRCA1 gene silencing and overexpressing in triple negative breast cancer cell line MDA-MB-231.Methods1.Different concentrations of EA(0μg / m L,1 μg / m L,5 μg / m L,10 μg /m L and 20 μg / m L)were used to treat MDA-MB-231 cells for 24 h,48 h and72 h.The inhibition rate of proliferation were assessed by MTT and the IC50 was calculated to select the optimal drug concentration.2.The silenced or overexpressed BRCA1 were transfected into the triple-negative breast cancer cell line MDA-MB-231 cells,respectively,The expressions of BRCA1 was detected by RT-q PCR and western blot,to determine whether silenced or overexpressed BRCA1 could successfully transfected into MDA-MB-231 cells.3.EA was applied at a final concentration of 5 μg /m L for 0 h,24 h,48 h,72 h,96 h.MTT assay was used to detect cell proliferation at different time points.4.Cell migration ability by scratch assay at 24 h and 48 h.5.Changes in tumor cell migration and invasion ability were evaluated by transwell assay at 24 h.6.The expressions of BRCA1 and PARP1 were detected by RT-q PCR and western blot.Results1.Different concentrations of EA(1～20 μg/m L)had a significant inhibitory effect on the proliferation of MDA-MB-231 cells at 24 h,48 h and 72 h.The IC50 were 13.739 μg/m L,10.645 μg/m L,5.344 μg/ m L,respectively.2.SiRNA and overexpressed of BRCA1 were efficiently transfected into tumors cells.The RT-q PCR and western blot were used to test their expression.3.MTT assay showed that the proliferation activity of EA in BRCA1 siRNA group was significantly lower than that of control group,EA group and negative control group in knockdown experiments(P<0.001).The proliferation activity of ellagic acid in EA group was significantly lower than that of control group,BRCA1 OE + EA group and negative control group in overexpression experiments(P<0.001).4.The scratch test showed that the cell migration rate in siRNA+EA group were lower than those of the other three groups in knockdown experiments(P<0.001).The cell migration rate in EA group were lower than those of the other three groups in overexpression experiments(P<0.01).5.The transwell migration and invasion assay showed that the number of transmembrane cells in siRNA+EA group were lower than those of the other three groups in knockdown experiments(P<0.001).The number of transmembrane cells in EA group were lower than those of the other three groups in overexpression experiments(P<0.01).6.The results of RT-q PCR and western blot showed that the expression of PARP1 mRNA and protein in siRNA+EA group was significantly lower than that in the other three groups in knockdown experiments(P<0.001).The results of RT-q PCR and western blot showed that the expression of PARP1 mRNA and protein in EA group was significantly lower than that in the other three groups in overexpression experiments(P<0.001).Conclusion1.The EA can inhibit the proliferation of human triple-negative breast cancer MDA-MB-231 cells in vitro in a dose-dependent manner.2.The EA can inhibit the proliferation,migration and invasion of BRCA1 gene silencing and overexpressing in MDA-MB-231 cells in vitro.3.The EA can inhibit the expression of PARP1 protein in human triple negative breast cancer MDA-MB-231 cells,BRCA1 gene silencing and overexpressing of MDA-MB-231 cells.4.The EA can inhibit BRCA1 gene silencing and overexpressing in MDA-MB-231 and its underlying mechanism may be related to the inhibition of PARP1 expression which a key factor in the DNA repair pathway.