The Effects Of Mifepristone On Human Endometrial Cellsand Its Mechanism

Background:Perimenopausal dysfunctional uterine bleeding(referred to as "dub")is a common gynecological disease,is due to abnormal uterine reproductive endocrine axis dysfunction caused by bleeding.20% perimenopausal women can appear anovulatory dysfunctional uterine bleeding.In the past clinical perimenopausal dysfunctional uterine bleeding of drug treatment with progesterone,endometrial shedding of synthetic progesterone endometrial atrophy method and application of anti fibrinolytic drugs,but there are some medication for a long time,dosage,side effects and other issues.Mifepristone(Mifeprielone)is a kind of progesterone drug resistance,can be combined with progesterone receptor,competitive inhibition of progesterone play a role,in addition,also dissolve can inhibit ovulation,corpus luteum and mifepristone influences the development and function of the film inside the uterus,can increase the uterine muscle cells,their sensitivity to prostaglandins,widely used in the termination of pregnancy in clinic,auxiliary induced labor,etc.Several studies have found that mifepristone growth plays an important role in the lining of the uterus,however,as the body's hormone levels and different dosage of different,different effect on the lining of the uterus mifepristone.Human premenopausal women,low doses of mifepristone could inhibit the role of endometrial hyperplasia,and large doses of mifepristone may lead to different degrees of endometrial hyperplasia.Clinical research before we do that,the short course(5 days)low dose(10mg)cycle(the first day of menstruation taking,three consecutive menstrual period,a total of 20 pieces of mifepristone can help achieve the treatment of perimenopausal dysfunctional uterine bleeding patients stop bleeding,establish normal menstrual cycle,to prevent the relapse of the.But the specific mechanism of mifepristone on endometrial growth is not clear.Still need further study to explore ?Purpose and significance:This study was to explore the effect of mifepristone on the human endometrial cells growth,and discussed its mechanism of action,so as to provide theoretical basis for clinical use of mifepristone treatment uterine related diseases and support.Methods:1.Different concentrations of mifepristone treatment human endometrial cells,determined by MTT method to detect different concentration mifepristone treatment time on the influence of human endometrial cells proliferation activity.2.Different concentrations of mifepristone treatment human endometrial cells,under a microscope observation of human endometrial cells form mifepristone influences the way.3.Different concentrations of mifepristone treatment human endometrial cells,after Annexin V + / PI-double dyed by flow cytometry to detect different concentrations of mifepristone impact on the level of human endometrial cells apoptosis.4.Different concentrations of mifepristone treatment human endometrial cells,western blot(western blot)method of detection of mifepristone on apoptosis in human endometrial cells way proteins Bax,pro-caspase3 / caspase3 protein expression level of influence.5.Different concentrations of mifepristone treatment human endometrial cells,western blot(western blot)method of detection of mifepristone for necrosis pathway in human endometrial cells protein expression of p-MLKL,real-time fluorescent quantitative PCR(real time PCR)method for detection of mifepristone RIPK3 necrosis pathway in human endometrial cells factor expression.6.Mifepristone treatment,different concentration of human endometrial cells,real-time fluorescent quantitative PCR(real time PCR)methods to detect estrogen receptor in human endometrial cells mifepristone ESR1 and the effects of progesterone receptor mRNA expression of PGR levels.7.Different concentrations of mifepristone treatment human endometrial cells,real-time fluorescent quantitative PCR(real time PCR)method for detection of mifepristone in human endometrial cells to promote angiogenesis factor angiogenin(Ang)1 and basic fibroblast growth factor(bFGF).Results:1.Different concentrations of mifepristone to separate human endometrial cells for 24 h,48 h and 72 h,determined by MTT,according to the results of mifepristone treatment can inhibit the activity of human endometrial cells,and the toxic effects of mifepristone on human endometrial cells with increased with increasing of the concentration,as the growth of the role of time increases,mifepristone treatment human endometrial cells for 24 h,48 h and 72 h IC50(including 18 g/ml,respectively,including 15 g/ml,including 12 g/ml.2.Dfferent concentrations of mifepristone treatment after human endometrial cells respectively,were observed under optical microscope cell morphology,found that treated with mifepristone significantly impaired cellular structure,characterized by the membrane boundary is not clear,nuclear chromatin density and cell size smaller and round,some cells appear rupture,prompt processing mifepristone damage to human endometrial cells have a certain role.And mifepristone treatment of endometrial cells is unprocessed reduce obviously.3.Different concentrations of mifepristone treatment human endometrial cells,flow cytometry test results showed that mifepristone treatment can promote the apoptosis of human endometrial cells,and the promoting effect of human endometrial cells apoptosis mifepristone increased as the concentration increases.4.Western blot(western blot),according to the results of treatment of mifepristone in human endometrial cells cells and promote apoptosis related proteins Bax,cleaved caspase 3,caspase 3 protein expression levels of the rise significantly.5.Western blot(western blot),according to the results of mifepristone treatment of endometrial cells necrosis pathway in cell protein expression of p-MLKL raised obviously,real-time fluorescent quantitative PCR(real time PCR)test results showed that mifepristone treatment of endometrial cells necrosis pathway in cell factor RIPK3 expression significantly raised.6.Real-time fluorescent quantitative PCR(real time PCR)test results showed that 5,15,20 mu g/mu l mifepristone treatment after human endometrial cells,cells of ESR1 content declined obviously,and 5,10,15,20 mu g/mu l mifepristone treatment of endometrial cells after PGR levels were significantly lower in the cell.7.Real-time fluorescent quantitative PCR(real time PCR)test results showed that mifepristone treatment cell,Ang-1 and the expression of bFGF were significantly reduced.Conclusion:1.Mifepristone can affect the cell morphology of human endometrial cells and inhibit the growth of human endometrial cells,and with dose and time dependent.2.Mifepristone can promote apoptosis related proteins Bax,cleaved by promoting the expression of caspase 3,caspase 3 promote human endometrial cells apoptosis and play the role of inhibition of cell proliferation.3.Mifepristone may increase necrosis protein by the expression of p-MLKL and RIPK3 promote human endometrial cells necrosis and play the role of inhibition of cell proliferation.4.Mifepristone may PGR ESR1 by regulating estrogen receptor and progesterone receptor expression in human endometrial play an important role in growth and development.5.Mifepristone may inhibit angiogenesis in human endometrium by down regulating the expression of Ang-1 and bFGF,which may play an important role in the inhibition of uterine bleeding.This study discusses the influence on the growth of the human endometrial cells mifepristone,found by promoting apoptosis related proteins Bax,mifepristone cleaved the expression of caspase 3,caspase 3 and promote human endometrial cells apoptosis,and increase necrosis protein by the expression of p-MLKL and RIPK3 promote human endometrial cells necrosis and inhibit the growth of the cells.In addition,mifepristone may PGR ESR1 by regulating estrogen receptor and progesterone receptor expression in human endometrial play an important role in growth and development,through regulating the expression of Ang-1 and b FGF? This research provides a theoretical basis for the application of the clinical mifepristone and support.