Effect And Mechanism Of Brazilin On Proliferation,Apoptosis And Metastasis Of SKOV3 Cells In Ovarian Cancer

ObjectiveTo investigate the effect and mechanism of Brazilin on cell proliferation,apoptosis and metastasis of SKOV3 cells in human ovarian cancer,and to provide a basis for the clinical use of Brazilin in the treatment of ovarian cancer.Methods1 、Different concentrations of Brazilin were used to treat human ovarian cancer SKOV3 cells for 24 h,and the effect of Brazilin on the proliferation of SKOV3 cells was detected by CCK-8 method,and applicable concentrations were choosed from follow-up experiments.2、Cell grouping and administration method:(1)Divide the experiment into control group(conventionally cultured cells),20,30 and 40μmol/L Brazilin groups.(2)The experiment was divided into control group(conventionally cultured cells),Brazilin group(40μmol/L Brazilin)and Brazilin+DKK1 combined group(40μmo L/L Brazilin+10ng/m L DKK1).3 、 Hoechst33342 staining method to observe the changes of apoptosis morphology of SKOV3 cells after Brazilin treatment under fluorescence microscopy.flow cytometry.5、Cell scratch experiments explored the effect of Brazilin on the migration ability of SKOV3 cells after administration.6 、 Transwell chamber experiment to explore the effect of Brazilin administration on the invasion ability of SKOV3 cells.7、The expression levels of apoptosis related proteins bax,bcl-2,epithelial interstitial transformation(EMT)associated proteins vimentin,E-cadherin and N-cadherin in SKOV3 cells after Brazilin administration.8 、 To explore whether Brazilin regulates apoptosis and metastasis in SKOV3 cells through the Wnt/β-catenin pathway.The expression levels of Wnt/β-catenin pathway associated proteins cyclin D1,c-myc,and β-catenin were determined by Western blot assay using the SKOV3 cells treatment with the Wnt/β-catenin pathway inhibitor,DKK1 combined with Brazilin.Reaults1、Brazilin has the ability to inhibit the proliferation of SKOV3 cells,its 24 h IC50=25.02μmol/L(P<0.05),and it was found that Brazilin’s inhibitory effect on SKOV3 cells was dose-dependent.2 、After SKOV3 cells were treated with different concentrations of Brazilin(20,30,40μmol/L Brazilin),it was observed under fluorescence microscopy that with the increase of Brazilin concentration,nucleus shrinkage gradually increased,chromatin wrinkled obviously,and the blue fluorescence intensity of nuclear membrane fragmentation increased significantly.3 、 Flow cytometry analysis of Brazilin can induce apoptosis in SKOV3cells(P<0.05).4 、Brazilin inhibited the migration of SKOV3 cells compared to the control group(P<0.05).5 、 The results showed that Brazilin significantly inhibited the invasion ability of SKOV3 cells compared with the control group(P<0.05).6 、Western blot analysis showed that the expression of apoptotic protein bax was significantly increased(P<0.05)and bcl-2 was significantly decreased(P<0.05).The expression levels of EMT-related proteins vimentin and Ncadherin were significantly reduced(P<0.05),and the expression levels of Ecadherin were significantly increased(P<0.05).7 、Compared with the Brazilin group,the expression of cyclin D1,c-myc and β-catenin in the combination group of Brazilin and Wnt pathway inhibitor DKK1 was significantly increased(P<0.05).Compared with the control group,the expression of cyclin D1,c-myc and β-catenin in the Brazilin group and the Brazilin+DKK1 combination group was significantly reduced(P<0.05).ConclusionsBrazilin can inhibit the proliferation,migration,invasion and EMT process of SKOV3 cells,and the molecular mechanism of its action may be related to the inhibition of the activation of the Wnt/β-catenin signaling pathway.