Fig. 5

MAPK1 downregulation promotes osteoclast differentiation by decreasing FoxO1 expression. A Downregulation of FoxO1 in Raw264.7 cells with stable overexpression of miR-92a-1-5p. GAPDH: internal control. B Marked downregulation of FoxO1 mRNA expression following transfection with Mapk1 siRNAs. C-D Ctsk and Trap expression determined by qPCR following transfection with FoxO1 siRNAs. E–F Trap + osteoclasts were identified (scale bar: 50 μm) and quantified in Raw264.7 cells (E) and BMMs (F) in the presence of RANKL. G-H Immunofluorescent staining (scale bar: 100 μm) and quantification of osteoclast markers CTSK and TRAP at 48 h post-transfection in the presence of RANKL. I Trap activity was detected at 6 days post-transfection with indicated siRNAs. J Comparison of Trap activity in Raw264.7 cells transfected with indicated siRNAs or siRNAs + FoxO1 fusion protein. Data were analyzed using one-way ANOVA with multiple-comparisons test. *, P < 0.05; **, P < 0.01; ***, P < 0.001