MiR-155 Inhibits the Expression of MMPs in HK2 Cells by Targeting KLF4

Abstract

To determine whether miR -155 inhibits the expression of matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9) in HK2 cells by targeting Kruppel-like factor 4 (KLF4). Methods MiR -155-mimic, miR -155-NC empty plasmid, and culture medium were transfected into renal tubular epithelial cells, respectively. Six hours later, the expression of miR -155 was detected by real time-PCR; the expressions of MMP-2, MMP-9 and KLF4 were detected by Western blot; and the activity of MMP-2 and MMP-9 in the cells was detected by gelatin zymography. Because KLF4 was predicted as the target gene of miR -155 by bioinformatics. The miR-155 overexpressed HK2 cells were transfected with KLF4 overexpression plasmid or empty plasmid. Six hours later, the expressions of MMP-2, MMP-9 and KLF4, and the activity of MMP-2 and MMP-9 were measured again. Finally, cells containing luciferase plasmids with KLF4-3′-UTR wild type (WT) or mutant (MUT) sequence were constructed and transfected with miR -155-mimic or empty plasmid. Luciferase assay was used to confirm whether KLF4 -3′-UTR was the binding site in targeting miR -155．Results Compared with the cells transfected with empty plasmid, the expression of miR -155 was up-regulated and the expressions of MMP-2, MMP-9 and KLF4 were down-regulated in the cells transfected with miR -155-mimic. Compared with the cells transfected with miR -155 mimic or miR -155 mimic+empty plasmid, the expressions of MMP-2, MMP-9 and KLF4 were up-regulated in the KLF4+miR -155 transfected cells. Luciferase assays confirmed that miR -155 binds to KLF4 , and KLF4 -3′-UTR is the target gene of miR -155. Conclusion MiR-155 inhibits the expressions of MMP-2 and MMP-9 in HK2 cells by targeting KLF4 -3′-UTR.

Keywords: Chronic kidney disease, MiR -155 Kruppel-like factor 4, Matrix metalloproteinase