MicroRNA-449c-5p inhibits osteogenic differentiation of human VICs through Smad4-mediated pathway

Rongjian Xu,Jianwu Dai,Xianglin Hou,Chunying Shi,Zhifeng Xiao,Jianzhou Liu,Yannan Zhao,Min Zhao,Bing Chen,Yun Yang,Zhuo Huang,Qi Miao

doi:10.1038/s41598-017-09390-z

Abstract

Calcific aortic valve disease (CAVD) is the most common heart valve disorder, yet its mechanism remains poorly understood. Valve interstitial cells (VICs) are the prevalent cells in aortic valve and their osteogenic differentiation may be responsible for calcific nodule formation in CAVD pathogenesis. Emerging evidence shows microRNA (miRNA, or miR) can function as important regulators of many pathological processes, including osteogenic differentiation. Here, we aimed to explore the function of miR-449c-5p in CAVD pathogenesis. In this study, we demonstrated the role of miR-449c-5p in VICs osteogenesis. MiRNA microarray assay and qRT-PCR results revealed miR-449c-5p was significantly down-regulated in calcified aortic valves compared with non-calcified valves. MiR-449c-5p overexpression inhibited VICs osteogenic differentiation in vitro, whereas down-regulation of miR-449c-5p enhanced the process. Target prediction analysis and dual-luciferase reporter assay confirmed Smad4 was a direct target of miR-449c-5p. Furthermore, knockdown of Smad4 inhibited VICs osteogenic differentiation, similar to the effect observed in up-regulation miR-449c-5p. In addition, animal experiments proved indirectly miR-449c-5p could alleviate aortic valve calcification. Our data suggested miR-449c-5p could function as a new inhibitory regulator of VICs osteogenic differentiation, which may act by targeting Smad4. MiR-449c-5p may be a potential therapeutic target for CAVD.

Highlights

Calcific aortic valve disease (CAVD) is associated with significant cardiovascular morbidity and mortality in the elderly and results in severe end-stage cardiovascular dysfunction
Our results showed miR-449c-5p expression in calcified aortic valves were significantly down-regulated than those in adjacent non-calcified valves (Fig. 2b), which suggests miR-449c-5p might participate in the pathogenesis of aortic valve calcification
We for the first time identified the differentially expressed miRNAs in non-calcified and calcified aortic valves derived from 3 CAVD patients using miRNA microarray assay

Summary

Introduction

Calcific aortic valve disease (CAVD) is associated with significant cardiovascular morbidity and mortality in the elderly and results in severe end-stage cardiovascular dysfunction. MicroRNA (miRNA, or miR) can act as fine-tuners in the regulation of diverse biological and pathological processes, including proliferation, apoptosis and differentiation, and miRNA dysregulation often results in impaired cellular function[10,11,12,13]. It has been reported some miRNAs are involved in smooth muscle cell www.nature.com/scientificreports/. (c) Immunohistochemical staining of Vimentin (Left: low magnification; Right: high magnification). Our findings may provide clues for the pharmacological intervention for CAVD

Objectives

Methods

Results

Conclusion