The Effect Of MiR-101 On The Sensitivity Of SKOV3/DDP Via MAPK/ERK Signaling Pathway

Objectives To explore the effect of miR-101 on cisplatin sensitivity of SKOV3/DDP cells and its underlying mechanism.Methods In this study,we adopted the method of in vitro experiment,and up-regulated miR-101 expression level of the cisplatin-resistant cell line SKOV3/DDP by cell transfection.Meanwhile,RT-PCR assay was applied for the expression of miR-101 and BDNF m RNA,and the cisplatin sensitivity and proliferation activity of SKOV3/DDP cells were measured using CCK-8.The migration ability of SKOV3/DDP cells were tested by wound heal assay and the invasion ability of SKOV3/DDP cells were tested by Transwell assay.In addition,the ultrastructure changes of SKOV3/DDP cells in each group were detected by transmission electron microscopy,and the apoptosis rate of SKOV3/DDP cells were detected by V-FITC/PI Annexin flow cytometry.The expression of c AMP and p-PKA were detected by enzyme linked immunosorbent assay,and the expression level of p-ERK,p-CREB,BDNF protein were detected by western-blot.The results were statistically analyzed by SPSS13.0 software,and used Graphpad prism 6software for statistical mapping.Results The expression level of miR-101 m RNA in SKOV3/DDP cells were significantly higher than that in control group,liposome group and blank plasmid group SKOV3/DDP cells（F=470.41,P<0.05）.The IC₅₀ of cisplatin of SKOV3/DDP cells in miR-101 group was significantly decreased when compare with blank control group,liposome group and blank plasmid group（F=200.23,P<0.05）.In miR-101 group,the proliferation,migration and invasion of SKOV3/DDP cells were significantly weaker than those in blank control group,liposome group and blank plasmid group（F=10.05,P<0.05）（F=11.86,P<0.05）（F=47.37,P<0.05）.In addition,the apoptosis rate of miR-101-SKOV3/DDP group was significantly higher than that of control group,liposome group and blank plasmid group（F=142.31,P<0.05）.The expression of c AMP and pPKA in miR-101-transfected group were significantly decreased（F=30.674,P<0.05）（F=65.267,P<0.05）,and the expression level of p-ERK and p-CREB in miR-101-transfected group were lower than in blank control group,liposome group and blank plasmid group（F=43.611,P<0.05）（F=35.541,P<0.05）.Furthermore,BDNF m RNA and protein expression level in SKOV3/DDP cells were decreased after miR-101 tansfection（F=173.342,P<0.05）（F=47.957,P<0.05）.Conclusions 1 After transfected with miR-101 plasmid,the expression level of miR-101 in epithelial ovarian cancer cisplatin-resistant SKOV3/DDP cells was high.2Overexpressed miR-101 in SKOV3/DDP cells significantly decreased cisplatin IC50.3Overexpressed miR-101 can significantly reduce the proliferation,migration and invasion of SKOV3/DDP cells and increase the apoptosis rate of SKOV3/DDP cells.4Overexpressed miR-101 can inhibit the activity of c AMP-PKA-ERK1/2-CREB-BDNF signal pathway in SKOV3/DDP cells and increase the sensitivity of SKOV3/DDP cells to cisplatin.