Effects of miR-195-5p on the CORM-3-induced osteogenic differentiation of rat BMSCs
A successful transfection with miR-195-5p mimics, or miR-195-5p inhibitor was confirmed by RT- qPCR (Fig. 3A). mRNA expressions of the Runx2 and OPN were assessed by RT-qPCR at different time points during osteogenic differentiation. After the overexpression of miR-195-5p, mRNA levels of both Runx2 and OPN were significantly lower than that in the CORM-3 group on the 3rd and 7th day of osteogenic induction (P< 0.05) (Fig. 3B) . While miR-195-5p was down-regulated, the mRNA expressions of Runx2 and OPN were increased by 1.4- and 1.5-fold, respectively, on the 3rd day (P < 0.05)), and both 1.3-fold on the 7th day of osteogenic induction, compared with CORM-3 group (P < 0.05) (Fig. 3C). In consistence with the regulatory effect of miR-195-5p on the mRNA expression of Runx2 and OPN, the protein expressions of Runx2 and OPN after the osteogenic induction were significantly decreased in miR-195-5p overexpressed cells, but increased in miR-195-5p deficient cells (P < 0.05) (Fig. 3D-E). The results from the alizarin red staining and semi quantitative analysis demonstrated that mineralization on 14 days of osteogenic induction after miR-195-5p mimics transfection was decreased obviously (P < 0.05), however, the opposite effect was not observed in miR-195-5p-deficient cells (Fig. 3F).