Melatonin enhances sensitivity of colorectal cancer cells to 5-fluorouracil by regulating miR-532-3p/β-catenin pathway
Objective To investigate the role of melatonin(MLT)in enhancing the sensitivity of colorectal cancer(CRC)to 5-fluorouracil(5-FU)and its molecular mechanism.Methods CRC cell lines SW480 and HCT116 were treated with MLT at different concentrations(0,0.125,0.25,0.5,1.0 or 2.0 mmol/L)and 5-FU at different concentrations(0,1.25,2.5,5,10,20 or 40 μmol/L)for 48 h.According to the inhibition rate of CRC cell activity,1 mmol/L MLT was selected for follow-up experiments.SW480 and HCT116 cells were divided into dimethyl sulfoxide(DMSO)group,MLT group,and 5-FU group,and treated with DMSO,1 mmol/L MLT and 15 μmol/L 5-FU for 48 h,respectively.SW480 and HCT116 cells were divided into miR-532-3p mimic group,miR-532-3p inhibitor group,mimic control group and inhibitor control group,which were transfected with 50 nmol/L miR-532-3p mimic,miR-532-3p inhibitor and their negative controls,respectively.The cell activity was detected by Cell Counting Kit-8(CCK-8)assay.Transwell assay was used to detect cell migration and invasion.The apoptosis was detected by flow cytometry.Real-time quantitative polymerase chain reaction(RT-qPCR)was used to detect the expression of miR-532-3p.The expression of β-catenin protein was detected by Western blot.Results Treatment with 1 mmol/L MLT for 48 h inhibited the proliferation,migration and invasion of SW480 and HCT116 cells,and promoted apoptosis(all P＜0.01).When treated with 5-FU at different concentrations(0,1.25,2.5,5,10,20 or 40 μmol/L),compared with 5-FU alone,the inhibition of the cell activity of SW480 and HCT116 cells treated with both 5-FU and 1 mmol/L MLT was enhanced(all P＜0.05).According to the value of half maximal inhibitory concentration(IC50)of 5-FU in SW480 and HCT116 cells,15 μmol/L 5-FU was selected for subsequent experiments.RT-qPCR results showed that miR-532-3p was underexpressed in SW480 and HCT116 cells.After treating SW480 and HCT116 cells with 1 mmol/L MLT for 48 h,the expression of miR-532-3p was increased(both P＜0.01).After co-treatment with 1 mmol/L MLT and dif-ferent concentrations of 5-FU(0,1.25,2.5,5,10,20 or 40 μmol/L)for 48 h,the inhibition of cell activity of SW480 and HCT116 cells in the miR-532-3p inhibitor group was significantly reduced compared with that in the negative control group(all P＜0.05),and the IC50 val-ues of 5-FU were increased(both P＜0.01).After treating SW480 and HCT116 cells with 1 mmol/L MLT for 48 h,the apoptosis rate of the miR-532-3p inhibitor group was lower than that of the negative control group(both P＜0.01).Western blot results showed that,compared with DMSO groups,the expression of β-catenin in SW480 and HCT116 cells in the MLT groups decreased(both P＜0.01).After treating SW480 and HCT116 cells with 1 mmol/L MLT for 48 h,the expression of β-catenin in the miR-532-3p inhibitor group was higher than that in the negative control group(both P＜0.01).Conclusions MLT enhances the sensitivity of CRC cells to 5-FU and inhibits tumor growth by regulating miR-532-3p/β-catenin pathway.
万方数据
维普
