Fig. 2
RvD1 inhibits osteoclast activation and recruitment as well as LPS-induced TRAP, cathepsin k, PGE2, TNF-α, and RANK expression and concurrently enhances IL-10 release in LPS-activated RAW 264.7 macrophages. Macrophages were stimulated with LPS (50 ng/ml) with or without RvD1 treatment (0–500 nM) for 72 h. Cell extracts were collected, and then TRAP and cathepsin k protein expression was assessed by western blot (a). The same treatment was conducted after incubation or not of cells with sc-siRNA or siRNA against FPR2 (50 nM) for 24 h. TRAP enzymatic staining was performed, and TRAP-positive cells were observed with an inverted microscope (× 200). Arrows show osteoclasts with three nuclei or more (b). Western blot was employed to confirm FPR2 silencing by siRNA (b). In parallel, cell media was collected and then PGE2 (c), TNF-α (d), IL-10 (e), and RANK (f) expression was assessed by commercial kits. Data are means ± SEM for n = 3, and one-way ANOVA was performed to compare the results. #p < 0.05, ##p < 0.01, ###p < 0.001, and ####p < 0.0001 compared to non-stimulated cells; *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001 compared to LPS-activated cells