GSK5182 inhibits RANKL-induced osteoclast formation. We cultured BMMs with the indicated concentrations of ERRγ modulators GSK5182 (A) or GSK4716 (B) for 24 h. We used immunoblotting to detect the ERRγ protein contents. Graphs represent the results of quantitative analysis of ERRγ protein levels from three independent experiments. (C) We did real-time PCR to assess ERRγ mRNA levels. (D-F) BMMs were cultured at the indicated GSK5182 concentrations in the presence of RANKL (20 ng/ml) and M-CSF (20 ng/ml) for four days. (D) The cells were fixed and stained for TRAP. (E) TRAP-positive multinucleated cells (MNCs) were counted. (F) TRAP activity in the culture medium generated by cells in (D) was quantified at 405 nm. (G) We cultured BMMs with the indicated concentrations of GSK5182 in the presence of M-CSF (30 ng/ml) for three days. We evaluated cell viability by MTS assay. The data presented in (A-C, E, and F) are expressed as the mean ± SD. *P < 0.05, **P < 0.001 vs. the vehicle-treated control (Con).