| Background and Aim:Vascular injury and pathological remodeling plays a central role in the pathogenesis of vascular diseases such as atherosclerosis and post-angioplasty restenosis. Both vascular smooth muscle cell (VSMC) differentiation from stem/progenitor cells and VSMC phenotypic modulation have been implicated in such abnormal vascular remodeling. Our group has recently reported that microRNA-34a (miR-34a) promotes VSMC differentiation from stem cells in vitro and in vivo. However, little is known about the functional involvements of miR-34a in VSMC functions and vascular injury-induced neointima formation. In the present study, we aimed to reveal the functional role of miR-34a in VSMC proliferation, migration and neointimal hyperplasia, as well as its underlying mechanism.Methods:Primary VSMCs were isolated from mouse aorta and cultured in vitro. To establish the in vitro VSMC phenotypic switching model (from contractile to synthetic or proliferative), VSMCs were serum starved and then treated with various pathological stimuli. miR-34a expression was evaluated during VSMC phenotypic switching. Using gain/loss-of-function assay, the effects of miR-34a on VSMC proliferation, migration and apoptosis were assessed. Then, functional target of miR-34a in VSMCs was predicted by various bioinformatics software and verified by luciferase assay. Finally, mouse wire-induced femoral artery injury model was performed and the impact of miR-34a on injury-induced neointima formation was determined by perivascular delivery of agomiR-34a.Results:miR-34a was significantly down-regulated in VSMC phenotypic modulation upon various pathological stimuli. miR-34a over-expression in VSMCs dramatically inhibited VSMC proliferation and migration, while knockdown of miR-34a significantly increased VSMC proliferation and migration, respectively. However, VSMC apoptosis wasn’t affected by miR-34a. Notch homolog 1 (Notchl), a well-known regulator in VSMC functions and neointima formation, was predicted as one of the top targets of miR-34a by using several computational prediction tools. Unsurprisingly, Notchl was negatively regulated by miR-34a on both mRNA and protein level in VSMCs. Luciferase assay showed that miR-34a substantially repressed Notch1-3’-UTR-luciferase activity in VSMCs, but not miR-34a binding sites mutant Notchl-3’-UTR-luciferease reporter, validating the Notchl is the functional target of miR-34a in VSMCs. Co-transfection of miR-34a and Notchl experiments also uncovered that miR-34a inhibited VSMC proliferation and migration through regulating Notchl expression. Finally and importantly, perivascular over-expression of miR-34a significantly decreased Notchl expression levels, reduced VSMC proliferation, and attenuated neointima formation in wire-injured arteries.Conclusion:Our data have demonstrated that miR-34a inhibits VSMC proliferation and migration, and ameliorates injury-induced neointima hyperplasia, suggesting its potential therapeutic application for vascular diseases. |
