Sunitinib Regulates MiR-143 To Inhibit Proliferation And Invasion Of Human Renal Cancer 786-O Cell

Background:Renal carcinoma is a common urological malignancy with a low median progression-free survival.Sunitinib is one of the most effective drugs for advanced kidney cancer,although sunitinib has been considered a routine treatment for renal cancer,overall survival rates have not improved significantly due to side effect and drug resistance.Therefore,the search for new and more effective ways to cure kidney cancer is of great significance.Objective:In this study,renal cancer 786-O cells were used as the research object.First,changes in cell migration,proliferation and invasion ability were detected after lentivirus mediated over-expression of miR-143.Then,the change in miR-143expression was detected after sunitinib treated.Furthermore,to study the effect of sunitinib regulated miR-143 on the migration,proliferation and invasion of renal carcinoma cells and its potential mechanism.Methods:After dilution of miR-143mimics and miR-NC lentivirus by 10,100,1000and 10000 times,infected 293T cells,and 72h later,fluorescent cells were counted under fluorescence microscope,and the virus titer was calculated by combining the viral dilution ratio.MiR-143mimics and miR-NC lentiviruses were stably transfected into renal carcinoma 786-O cells,and the relative expression of miR-143 was detected by qRT-PCR.786-O cells were treated with sunitinib at 0,2,4,6,8,10,12μmol/L for 48h,IC50 of sunitinib on 786-O cells for 48h was calculated,this value was selected as the dosing concentration in subsequent experiments.RNA was extracted from 786-O cells after treated with sunitinib with IC50 value for 48h,and the expression of miR-143was detected by qRT-PCR.After 786-O cells stabilized transfected with miR-143mimics,and changes in migration,proliferation and invasion ability of renal cell carcinoma were detected by wound healing experiment,thiazole blue colorimetry and transwell experiment respectively,the expression of DNMT3B RNA,DNMT3B and p-akt proteins were detected by real-time fluorescence quantitative PCR and western blot.After treated with sunitinib with IC50 concentration for 6,12,24h,the migration ability of 786-O cells were detected by wound healing assay.After treated with sunitinib with IC50 concentration for 48h,and changes in proliferation and invasion ability of renal cell carcinoma were detected by thiazole blue colorimetry and transwell experiment,the expression of DNMT3B RNA,DNMT3B and p-akt proteins were detected by real-time fluorescence quantitative PCR and western blot.Results:The titers of miR-143mimics and miR-NC lentivirus were all more than1×10~8TU/ml,which met the experimental requirement.MiR-143mimics stable 786-O cell line was successfully constructed(P<0.05).The IC50 of sunitinib on 786-O cells for 48h was 6μmol/L.Sunitinib up-regulated the expression of miR-143 in 786-O cells(P<0.05).After stably transfected with miR-143 mimics and treated with sunitinib,the migration,proliferation and invasion of 786-O cells were decreased,and the expressions of DNMT3B RNA,DNMT3B protein and P-Akt protein were decreased,and the effect was more obvious when the two acted together(P<0.05).Conclusion:Sunitinib inhibited migration,proliferation and invasion of renal cancer786-O cell by upregulating miR-143.Sunitinib down-regulated the expression of DNMT3B and p-akt proteins in 786-O cell by upregulating miR-143 to play an anti-tumor effect.