miR-628-3p regulates osteoblast differentiation by targeting RUNX2: Possible role in atrophic non-union.

ABSTRACT

Atrophic non-union is a serious complication of fractures. The underlying biological mechanisms involved in its pathogenesis are not yet completely understood. MicroRNAs (miRNAs or miRs) are a type of endogenous small non-coding RNA, which participate in various physiological and pathophysiological processes. In this study, differentially expressed miRNAs were screened in patients with atrophic non-union. In total, 4 miRNAs (miR­149*, miR­221, miR­628-3p and miR­654-5p) were upregulated and 7 miRNAs (let-7b*, miR­220b, miR­513a-3p, miR­551a, miR­576-5p, miR­1236 and kshv-miR­K12-6-5p) were downregulated at the fracture sites in patients with atrophic non-union. Among the upregulated miRNAs, miR­628-3p and miR­654-5p expression was found to be persistently decreased during osteoblast differentiation, indicating their possible inhibitory effect on osteogenesis. Gain-of-function experiment demonstrated that miR­628-3p, but not miR­654-5p, attenuated osteoblast differentiation. Further, in silico analysis revealed that runt-related transcription factor 2 (RUNX2), the master transcript factor for osteoblast differentiation, was the target of miR-628-3p, which had two binding site-condense regions in the 3' untranslated region. The exact binding site of miR-628-3p was further identified with luciferase reporter assay. In addition, the overexpression of miR­628-3p appeared to be associated with the suppression of RUNX2 expression at both the mRNA and protein level, suggesting that miR­628-3p inhibits osteoblast differentiation via RUNX2. On the whole, the findings of this study provide evidence of the upregulation of miR­628-3p in patients with atrophic non-union and that miR­628-3p may exert an inhibitory effect on osteogenesis via the suppression of its target gene, RUNX2. The study provides valuable insight into the pathogenesis of atrophic non-union and suggests new potential therapeutic targets for the treatment of this disorder.