FIGURE 3.

Characterization of FNDC5/irisin deficiency BMSCs and BMMs isolated and cultured in vitro. (a) The mRNA expression of osteogenic markers on Day 5 and Day 7 (n = 3). The BMSCs were treated with either growth medium (GM) or osteoblast differentiation medium (DM) including ascorbic acid (Asc, 50 μg/ml), dexamethasone (Dex, 0.1 μM) and β‐glycerophosphate (β‐Gly, 10 mM) in the presence or absence of irisin. (b) ALP staining of BMSCs from F/I KO and control mice after treated with or without osteogenic supplements for 14 days. (c) Pictures of ARS staining and concentration of melted nodules solution on Day 14 (upper lane) and Day 21 (lower lane). (d) The mRNA expression level of cathepsin K, Mmp9, and Trap in RANKL‐induced osteoclast culture on Day 3 and Day 4 (n = 3). (e) TRAP staining of BMMs from F/I KO and control mice in the presence or absence of RANKL for 4 days. Values are shown as means ± SD. ALP, alkaline phosphatase; ARS, alizarin red S; Asc, ascorbic acid; BMM, bone marrow–derived macrophage; BMSC, bone marrow–derived mesenchymal cell; CON, control; Dex, dexamethasone; DM, differentiation medium; F/I KO, Osx‐Cre:FNDC5/irisin KO mice; FNDC5, fibronectin‐type III domain‐containing 5; GM, growth medium; KO, knockout; Mmp9, matrix metalloproteinase 9; mRNA, messenger RNA; RANKL, receptor activator of nuclear factor‐κΒ ligand; TRAP, tartrate‐resistant acid phosphatase; β‐Gly, β‐glycerophosphate. **p < .01, ***p < .001, ****p < .0001 versus control