ABSTRACT
AIM: To investigate the effects of microRNA (miRNA)-483-3p on the growth and migration of human glioma cell line A172 and its potential mechanisms.METHODS: The abundance of miRNA-483-3p in human embryonic kidney 293 cells and different human glioma cell lines (A172, U251 and SHG44) was measured by RT-qPCR.After down-regulation of miRNA-483-3p by transfection of inhibitor in the A172 cells, the cell viability, cell cycle distribution and cell migration were detected by CCK-8 assay, flow cytometry and Transwell assay, respectively.Furthermore, the protein levels of cell cycle-related molecules and epithelial-mesenchymal transition markers were measured by Western blot.Luciferase reporter assay was used to predict and verify the target gene of miRNA-483-3p.RESULTS: miRNA-483-3p was highly expressed in human glioma cells.Knockdown of miRNA-483-3p inhibited A172 cell viability, arrested cell cycle and decreased cell migration rate.Furthermore, the protein levels of cyclin D1, cyclin-dependent kinase 4, phoshorylated retinoblastoma protein, N-cadherin and vimentin were significantly decreased after knockdown of miRNA-483-3p, accompanied with the up-regulation of E-cadherin and β-catenin protein expression.Luciferase reporter assay demonstrated that Smad4 was a potential target gene of miRNA-483-3p.Down-regulation of Smad4 in the A172 cells transfected with miRNA-483-3p inhibitor partially reversed the effect of miRNA-483-3p on cell viability and migration.CONCLUSION: Knockdown of miRNA-483-3p restrains the growth and migration of A172 cells by targeting Smad4.
ABSTRACT
Background:Suppression of tumor suppressor genes plays a key role in the pathogenesis and progress of tumors. Some microRNAs may contribute to tumorigenesis by regulating tumor suppressor genes. Aims:To investigate the targeted regulatory effect of miR-483-3p on deleted in liver cancer 1(DLC1)gene in colorectal cancer. Methods:Sixteen patients with colorectal cancer admitted from October 2012 to April 2013 at Nanjing Drum Tower Hospital were enrolled. Expression of DLC1 in cancerous and adjacent noncancerous tissues was determined by Western blotting,and expression of miR-483-3p was determined by qRT-PCR. Dual luciferase reporter gene plasmid containing the 3’untranslated region(3’UTR)of DLC1 was constructed to validate the regulation of DLC1 by miR-483-3p in human colon cancer cell line HCT116. MiR-483-3p mimic was transfected into HEK293T cells and expression of DLC1 was determined by Western blotting;MiR-483-3p mimic was transfected into HCT116 cells and cell proliferation was measured by CCK-8 assay. Results:Expression of DLC1 was significantly lower in cancerous tissue than in noncancerous tissue,while expression of miR-483-3p was significantly higher in cancerous tissue than in noncancerous tissue(P<0. 05). MiR-483-3p mimic reduced the expression of DLC1 through directly binding to the 3’UTR of DLC1. Transfection of miR-483-3p mimic enhanced the proliferation of HCT116 cells significantly(P<0. 05). Conclusions:DLC1 is a target gene of miR-483-3p. MiR-483-3p might promote the development of colorectal cancer by down-regulating DLC1 expression at post-transcriptional level.