RESUMO
Objective To investigate the effect and mechanism of miR-20a-5p on human nephroblastoma cell line WiT49 transplanted tumor in nude mice.Methods The gene expression chip was downloaded from GEO database,and the differential gene miR-20a-5p was obtained by GEO2R.The NF-κB gene was positively correlated with the expression of miR-20a-5p through cBioPortal database.The target gene of miR-20a-5p was predicted to be NFKBIB of the NF-κB transcription factor suppressor protein family by targetscan database,and was verified by dual luciferase assay.Nephroblastoma cell line WiT49 was cultured in vitro and transfected into WiT49 cells with lentiviral vectors constructed with miR-20a-5p mimics and its suppressor gene.Twelve nude mice were randomly divided into three groups:WiT49 model group,WIT49-miR-20a-5p overexpression group and WIT49-miR-20a-5p knockdown group.The tumor mass and volume of each group were detected by tumor formation experiment in nude mice.real time fluorescent quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-20a-5p,NFKBIB and NF-κB in each group;CCK-8 cell proliferation assay was used to verify the proliferation of tumor cells in each group.Results miR-20a-5p is highly expressed in nephroblastoma and is positively correlated with the expression of NF-κB.miR-20a-5p and NFKBIB have mutual binding sites and binding effects.In the tumor formation experiment of nude mice,the tumor volume and mass of WIT49-miR-20a-5P overexpression group were significantly increased compared with WiT49 model group,and the difference was statistically significant(P<0.05).In the qRT-PCR test,the expressions of miR-20a-5p and NF-κB in the WIT49-miR-20a-5p overexpression group were higher than those in the WiT49 model group,and NFKBIB expression in the WIT49-miR-20a-5p overexpression group was lower than that in the WiT49 model group,with statistical significance(P<0.05).CCK-8 cell proliferation assay showed that the absorbance of WIT49-miR-20a-5p overexpression group at 24 and 48 hours was higher than that of WiT49 model group,and the absorbance of WIT49-miR-20a-5p knockdown group at 24,48 and 72 hours was lower than that of WiT49 model group,and the difference was statistically significant(P<0.05).Conclusion miR-20a-5p may promote the growth of human nephroblastoma cell WiT49 transplanted tumor in nude mice by regulating NFKBIB activation of NF-κB pathway.