Objective:To investigate the effects of the miR-149-5p on osteogenic differentiation of aortic valve interstitial cells(VICs)and explore its possible mechanism.Methods:Normal and calcified aortic valve tissues were collected,sample RNAs were extracted to detect the expression level of miR-149-5p,and correlation analysis was performed with the expression levels of miR-149-5p and calcification related genes Runt related transcription factor 2(RUNX2).Human VICs were cultured and induced by osteogenic differentiation medium(OM).Alizarin staining was used to detect the degree of osteogenic differentiation,and real-time PCR was used to detect the expression levels of miR-149-5p.VICs were divided into three groups:control group,NC+OM group,and miR-149-5p+OM group.The latter two groups were transfected with NC mimic and miR-149-5p mimic,respectively.Real-time PCR was used to detect the effect of miR-149-5p overexpression on the expression of osteogenic genes and target genes.TargetScan and miRDB database were used to predict the target genes of miR-149-5p,dual luciferase reporter system was used to detect the luciferase activity of target genes 3'-UTR,and Results:The expression of miR-149-5p was decreased in calcified aortic valves and was negatively correlated with the expression of RUNX2(P＜0.05).After osteogenic differentiation induction,the positive area of Alizarin staining in human VICs was increased,while the expression of miR-149-5p decreased(P＜0.05).After human VICs were transfected with miR-149-5p mimic,the expression of miR-149-5p in the cells was significantly up-regulated compared with NC+OM group(P＜0.05).At the same time,the calcium content and the expression of OCN and ALPP in human VICs were lower in the miR-149-5p+OM group than that in the NC+OM group(P＜0.05).Pro-osteogenic gene IL6 was predicted as the target genes of miR-149-5p by TargetScan database.miR-149-5p could inhibit the luciferase activity of IL6 3'-UTR(P＜0.05).Real-time PCR results showed that the expression of IL6 in the hVICs of miR-149-5p+OM group was significantly down-regulated(P＜0.05).Conclusions:miR-149-5p,which was decreased in calcified aortic valves and osteogenic differentiated VICs can attenuate osteogenic differentiation.The potential mechanism is inhibiting the expression of pro-calcification gene IL-6.
维普
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