Objective:To elucidate the effect of miRNA-451 on the expression of inducible nitric oxide synthase(iNOS)gene in the mesangial cells of mice.Methods:Two template DNA sequences were designed based on mmu-miR-451 sequence in miRBase database.The mmu-miR-451 expression plasmid and a control plasmid

named pGenesil-1-miRNA-451 and pGeneil-1-control respectively

were generated by the cloning of annealed oligonucleotides into pGenesil-1

then transfected into mesangial cells

which were stably selected by G418 to establish mesangial cell lines stablely expressing pGenesil-1-miRNA-451 and pGenesil-1-control.The living cells were counted by MTT assay and cell growth curves were drew to analyze the cell proliferation.The iNOS mRNA level was assessed by RT-PCR.The expression of iNOS protein was determined by Western blot and immunocytochemistry.Results:The recombinant vectors were verified by sequencing.The cell growth curves indicated the proliferation of cells transfected with pGenesil-1-miRNA-451 were inhibited significantly

compared with that of cells transfected with pGenesil-control and cells without treatments.RT-PCR analysis showed the levels of iNOS mRNA were almost unchanged in the cells with or without treatments.Western blot and immunocytochemistry demonstrated a significant decrease of iNOS protein levels in the cells transfected with pGenesil-1-miRNA-451

but not in the cells transfected with pGenesil-1-control

when compared with mesangial cells.Conclusion:miRNA-451 repressed the expression of iNOS gene translationly

and significantly inhibited cell growth.