Abstract
BackgroundHuman aortic valve interstitial cells (hAVICs) are a key factor in the pathogenesis of calcific aortic valve disease (CAVD). This research examines the role and mechanism of microRNA miR-138-5p in osteogenic differentiation of hAVICs.MethodsRT-qPCR analysis was applied for detecting miR-138-5p and RUNX2 expression in valve tissues of CAVD patients and controls. On completion of induction of osteogenic differentiation of hAVICs, and after overexpression or interference of miR-138-5p expression, the condition of osteogenic differentiation and calcification of hAVICs was confirmed using alkaline phosphatase staining and alizarin red staining. Subsequently, western blot was utilized to detect the expression of osteogenesis-related proteins OPN and ALP, and Wnt/β-catenin signaling pathway-related proteins. Finally, the relationship between miR-138-5p and RUNX2 was validated by dual-luciferase reporter assay and Pearson’s correlation test.ResultsDown-regulation of miR-138-5p was found in CAVD patients and during osteogenic differentiation of hAVICs. Overexpression of miR-138-5p contribute to the inhibition of osteoblast differentiation and calcium deposition in hAVICs, and of ALP and OPN protein expression. RUNX2 was a target gene of miR-138-5p, and it was negatively correlated with miR-138-5p in CAVD. Additionally, overexpression of RUNX2 could reverse the inhibitory effect of miR-138-5p on osteogenic differentiation of hAVICs.ConclusionmiR-138-5p can act as a positive regulator of osteogenic differentiation in CAVD patients to involve in inhibiting valve calcification, which is achieved through RUNX2 and Wnt/β-catenin signaling pathway.
Highlights
Calcific aortic valve disease (CAVD), with the prevalence ranking first among valvular heart diseases clinically, is characterized by valve thickening and calcification, aortic hemodynamic disorders and heart failure, which is mainly responsible for valve replacement failure
Results miR‐138‐5p affects osteogenic differentiation of Human aortic valve interstitial cells (hAVICs) First, by using RT-qPCR for determining miR-138-5p expression in the valve tissues of CAVD patients, significantly lower miR-138-5p was identified in the CAVD patients compared with the controls (Fig. 1A)
We found that miR138-5p negatively regulates the process of osteogenic differentiation of human calcified aortic valve by targeting RUNX2
Summary
Calcific aortic valve disease (CAVD), with the prevalence ranking first among valvular heart diseases clinically, is characterized by valve thickening and calcification, aortic hemodynamic disorders and heart failure, which is mainly responsible for valve replacement failure. MicroRNAs (miRNAs) refer to small non-coding RNA molecules which are highly conserved and single-stranded They mediate the levels of their target genes through transcriptional repression or mRNA degradation, regulating a variety of physiological and pathological processes [5], including cell proliferation, apoptosis [6], inflammation [7], and cancer [8]. Osteogenic differentiation of human mesenchymal stem cells (hMSCs) can be negatively regulated by miR-138-5p; this miRNA mediates AK-ERK1/2 activity, thereby decreasing Runx expression and achieving the negative regulation [11]. By contrast, this process can be promoted by miR138-5p knockdown and the resulting up-regulation of FOXC1 [12]. This research examines the role and mechanism of microRNA miR-138-5p in osteogenic differentiation of hAVICs
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