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.

本研究旨在探讨microRNA-337（miR-337）对骨髓间充质干细胞成骨分化的抑制作用及其作用机制。在骨髓间充质干细胞 (BMSC) 中进行了 miR-337 的过表达和敲低。使用细胞计数试剂盒-8（CCK-8）评估细胞增殖，通过茜素红染色进行矿化测定，然后测量碱性磷酸酶活性。应用荧光素酶报告基因检测来验证 miR-337 与 Ras 相关蛋白 1A (Rap1A) mRNA 的结合。应用逆转录和定量聚合酶链反应 (RT-qPCR) 检测矮小相关转录因子 2 (Runx2)、碱性磷酸酶 (ALP)、骨钙素 (OCN)、骨桥蛋白 (OPN)、骨形态发生蛋白 (BMP2) 的表达) 和 miR-337。然后通过蛋白质印迹分析确定 Rap1A 的蛋白质水平。高糖抑制成骨分化，但增加了 miR-337 的水平。miR-337 的过表达抑制了高糖处理的 BMSC 中的成骨分化，而 miR-337 的敲低则逆转了这一过程。荧光素酶报告基因检测证实 miRNA-337 的假定配对结合位点位于 Rap1A WT 的 3'-UTR。此外，Rap1A 的敲低明显抑制了成骨分化，这阻断了 miR-337 敲低对高糖处理的 BMSC 中成骨分化的影响。MiR-337 可以抑制直接靶向 Rap1A 的高糖治疗 BMSC 的成骨分化，从而为临床糖尿病骨质疏松症患者提供潜在的治疗策略。