miR-26b-5p promotes osteogenesis of bone mesenchymal stem cells via suppressing FGF21

Abstract

Background: miR-26b-5p actively participates in the osteogenic differentiation of bone mesenchymal stem cells (BMSCs). The database showed that fibroblast growth factor (FGF)-21 is a potential binding site of miR-26b-5p. This study aimed to investigate the molecular osteogenic mechanisms of miR-26b-5p targeting FGF21. Methods: Bone marrow was aspirated from the anterior superior iliac spine during bone marrow puncture. BMSCs were used to establish an in vitro cell model, and BMSCs markers were analyzed by flow cytometry. miR-26b-5p were overexpressed for 48 hours, and then placed in an osteogenic induction medium for osteogenic induction culture, the expression of RNA was detected using RT-qPCR. On day 7 of induction, RT-qPCR was used to measure Runx2, Osterix (Osx), and target gene FGF21 expression levels in each group. RT-qPCR, the dual-luciferase reporter gene system and western blot were used to verify that FGF21 was a direct target of miR-26b-5p. Results: BMSCs were identified according to the antigenic characteristics. miR-26b-5p expression was significantly upregulated after the expression of miR-26b-5p mimics, and FGF21 expression was downregulated; in miR-26b-5p inhibitor, the opposite results were revealed. After overexpression of miR-26b-5p, the alkaline phosphatase activity and nodules of Alizarin red S in the culture medium was increased; the opposite results were revealed in miR-26b-5p inhibitor. The expressions of Runx2 and Osx in the miR-26b-5p group were also significantly higher; in the miR-26b-5p inhibitor group, the opposite results were revealed. Luciferase reporter assays demonstrated that FGF21 was a direct target of miR-26b-5p. The western blotting analysis showed that FGF21 expression was significantly downregulated in the miR-26b-5p overexpressed group. Finally, the expressions of the characteristic osteogenic factors in the miR-26b-5p control + FGF21 group was significantly lower, but then increased significantly in the miR-26b-5p mimics + FGF21 group; the expressions of the characteristic osteogenic factors in the miR-26b-5p control + si-FGF21 group was significantly higher. Conclusions: miR-26b-5p can regulate the osteogenic differentiation of BMSCs and participate in PMOP pathogenesis via suppressing FGF21.