miR-199a-5p participates in the regulation of liver fibrosis by targeting TGF-β2

ObjectiveTo explore the mechanism of miR-199a-5p in the occurrence of liver cirrhosis.

MethodsThe miR-199a-5p mimics and miR-199a-5p inhibitor sequences were designed and synthesized to study the overexpression and inhibition of miR-199a-5p.The effects of overexpression of miR-199a-5p on the proliferation activity of Lx-2 cells were detected by CCK8, 5EdU, and Western blotting methods.Using Lx-2 cell cDNA as template, the wild type primer sequence of TGF-β2 3'UTR and mutant primer sequence of miR-199a-5p potential binding site were amplified by PCR to obtain wild type TGF-β2 3'UTR sequence and mutant TGF-β2 3'UTR sequence, respectively.The regulation process of TGF-β expression was revealed by double fluorescence reporter gene experiment.

ResultsIn vitro experiments, the results showed that the vitality of Lx-2 cells transfected with miR-199a-5p mimics significantly increased compared with the control group(P < 0.05 and P < 0.01), and the α-SMA protein levels was significantly higher than that of control group (P < 0.05).After 8 h, 12 h and 24 h of transfection with miR-199a-5p inhibitor, the cell vitality significantly decreased (P < 0.05 and P < 0.01), and the level of α-SMA significantly decreased compared with the control group (P < 0.05).The results of dual luciferase report showed that after transfecting the miR-199a-5p mimics, the expression of luciferase plasmid containing wild-type TGF-β2 3'UTR sequence (WT) significantly decreased compared with the control group (P < 0.01).The expression of luciferase plasmid containing miR-199a-5p binding site mutation sequence (MUT) had no significant changes (P>0.05).

ConclusionsUpregulation of miR-199a-5p can promote the proliferation of Lx-2 cell line, and the overexpression of miR-199a-5p can promote the expression of matrix protein α-SMA, and activate the progression of Lx-2 toward fibrosis.The miR-199a-5p plays a role in promoting liver fibrosis and cirrhosis in the process of liver cirrhosis.TGF-β2 is one of the main regulatory factors of liver fibrosis, and miR-199a-5p can promote liver fibrosis and cirrhosis by regulating the expression of downstream TGF-β2.