This study aims to investigate the inhibitory effect of microRNA-337 (miR-337) on osteogenic differentiation in bone marrow mesenchymal stem cells and its action of mechanisms. Overexpression and knockdown of miR-337 were performed in bone marrow mesenchymal stem cells (BMSCs). Cell proliferation was assessed by using a cell counting kit-8 (CCK-8), mineralization assay was performed by alizarin red staining, and alkaline phosphatase activity was then measured. Luciferase reporter assay was applied to verify miR-337 binding to Ras-related protein 1A (Rap1A) mRNA. Reverse transcription and quantitative polymerase chain reaction (RT-qPCR) was applied to measure the expressions of runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), osteocalcin (OCN), osteopontin (OPN), bone morphogenetic protein (BMP2), and miR-337. Then the protein level of Rap1A was determined by western blot analysis. High glucose inhibited osteogenic differentiation but increased the level of miR-337. Overexpression of miR-337 inhibited osteogenic differentiation in high glucose-treated BMSCs, while the knockdown of miR-337 reversed this process. Luciferase reporter assay confirmed that the presumed pairing binding site of miRNA-337 was in the 3'-UTR of the Rap1A WT. In addition, the knockdown of Rap1A distinctly repressed osteogenic differentiation, which blocked the effect of miR-337-knockdown on osteogenic differentiation in high glucose-treated BMSCs. MiR-337 could repress osteogenic differentiation in high glucose-treated BMSCs directly targeting Rap1A, thus provide a potential therapeutic strategy for patients with diabetic osteoporosis in clinic.
Keywords: High glucose; MicroRNA-337; Osteogenic differentiation; Rap1A.