Fig. 6

miRNA-5106 targets SIK2 and SIK3 to induce osteoblastic activity in vitro. a Luciferase reporter constructs containing either a WT SIK2 and SIK3 3′ UTR (WT 3′ UTR) or this same region after site-directed mutagenesis (3′ UTR-Mut). Luc, luciferase. Endogenous miRNA-5106 effects in BMSCs on WT SIK2 and SIK3 3′ UTR (luc-UTR), the SIK2 and SIK3 3′ UTR mutant (luc-UTR-Mut), as assessed following agomiR-NC or agomiR-5106 treatment; b The levels of mRNA SIK2 and SIK3 were western blotting analysis in agomiR-5106 treated BMSCs following transfection using lipofecamine control, 200 µM agomiR-NC, agomiR-5106, antagomiR-NC or antagomiR-5106 for 48 h; c The expression of SIK2 and SIK3 was measured by qRT-PCR analysis; d, e Western blotting and qRT-PCR analysis were used following control, siRNA-NC, siRNA-SIK2, and siRNA-SIK3 transfection to assess SIK2 and SIK3 expression, with a scrambled siRNA utilized as a negative control (siRNA-NC); f, g The levels of osteogenic genes were measured by western qRT-PCR and blotting analysis; h, i SIK2 and SIK3 expressions in mice callus decreased significantly on day 14 after receiving the injections of agomiR-5106 compared with PBS and antagomiR-5106 groups; j, k SIK2 and SIK3 expressions were measures by qRT-PCR analysis following different treatments; l Osteogenic related genes level was assessed by qRT-PCR analysis; m Alizarin red-mediated calcium staining and n ALP staining in BMSCs following differently transfected, Scale bar = 10 mm. Data are mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001