Inhibition effect of microRNA-124 on rat vascular smooth muscle cell proliferation and its molecular mechanism

Abstract

Objective To investigate the inhibition effect and molecular mechanism of microRNA-124 (miR-124) on rat vascular smooth muscle cell proliferation. Methods We applied 10 male 11-week-old spontaneously hypertensive rats (SHR) as experimental group and 10 age matched Wistar Kyoto (WKY) rats as control group. Aortic smooth muscle cells were isolated from the medial layer of thoracic aorta of SHR and WKY rats and cultured in Dulbecco's modified eagles medium (DMEM). Quantitative real-time PCR (RT-PCR) was used to detect the expression of miR-124 in SHR and WKY aortic smooth muscle cells. The SHR aortic smooth muscle cells were transfected with either miR-124 mimics or NC mimics control. MTT assay was used to explore the proliferation of SHR aortic smooth muscle cells in vitro. The targeted gene of miR-124 was predicted by bioinformatics, and verified by dual luciferase reporter assay. The over-expression of miR-124 in SHR aortic smooth muscle cells was analyzed by the mRNA RT-PCR and protein expression of Meox2 by Western blot analyses. Finally, SHR aortic smooth muscle cells proliferation was also detected using MTT assay after treated with siMeox2. Results The expression of miR-124 in SHR aortic smooth muscle cells was 0.22 times of WKY aortic smooth muscle cells (P<0.01). Over-expression of miR-124 could significantly inhibit SHR aortic smooth muscle cells proliferation. Bioinformatics analysis and dual luciferase reporter assay demonstrated that Meox2 was a target gene of miR-124. The mRNA level of Meox2 in SHR aortic smooth muscle cells after treated with miR-124 mimics was 0.29 times of transfected NC mimics control (P<0.01). Western blot showed that Meox2 protein was also decreased after treated with miR-124 mimics. Knock-down Meox2 could also arrest the SHR aortic smooth muscle cells proliferation. Conclusions The expression of miR-124 is down-regulated in SHR, which may be mediated by Meox2 regulation of SHR aortic smooth muscle cells proliferation and inhibit the occurrence of hypertensive disease. Key words: miR-124; Rats, inbred SHR; Aorta; Myocytes, smooth muscle; Proliferation; Meox2