Roles of miR-200a in renal fibrosis through regulating ZEB1 and ZEB2

Objective: This study aims to investigate the role and mechanism of miR-200a in the epithelial-mesenchymal transition (EMT) of kidney epithelial cells. Methods: Through treatment with transforming growth factor-beta 1 (TGF beta 1), renal epithelial cells HKC were induced to EMT. MiR-200a expression was detected by qRT-PCR and EMT related proteins E-Cadeherin and Vimentin were detected by Western blot. Bioinformatics methods were used to predict the target genes of miR-200a. After transfection with miR-200a mimics or siRNA of ZEB1 and ZEB2, the expressions of ZEB1, ZEB2 and other EMT related proteins were detected by Western blot. The HKC cells with stable expression of ZEB1 and ZEB2 were selected by G418 pressure screening. EMT changes were evaluated for HKC cells. Dual luciferase assay was applied to identify the target of miR-200a. Results: MiR-200a expression was significantly decreased in EMT processes in HKC cells induced by TGF beta 1, and E-Cadherin expression was down-regulated while Vimentin expression was up-regulated after TGF beta 1 treatment. The results indicated that TGF beta 1 can induce the occurrence of EMT. In vitro results showed that miR-200a induced EMT in HKC cells. Over-expressed miR-200a down-regulated the expressions of ZEB1 and ZEB2, and EMT of HKC cells were significantly repressed. After RNAi interference for ZEB1 and ZEB2, the EMT process in HKC cells was inhibited. HKC cells with stable expression of ZEB1 and ZEB2 were selected by G418 pressure screening, and the selected cells stably expressed green fluorescence. Increased expression of ZEB1 and ZEB2 significantly promoted EMT process in HKC cells. These results of dual luciferase assay indicated that miR-200a regulated the expression of ZEB1 and ZEB2 through complementary binding. Conclusion: MiR-200a expression was significantly down-regulated in EMT cell model induced by TGF beta 1. MiR-200a can inhibit EMT process in HKC cells through directly target ZEB1 and ZEB2, which indicates that miR-200a may be a potential therapeutic target for renal fibrosis.