Inhibition of osteoclastogenesis by GM-CSF.

The c-Fms⁺RANK⁻ cells were cultured in the presence of M-CSF and sRANKL with or without GM-CSF. After 6 days of cultivation, cells were subjected to a TRAP activity assay (A) and TRAP staining (B). The inhibitory effect of GM-CSF on osteoclast differentiation was dose dependent and abolished by addition of neutralizing antibody against GM-CSF (A). In the absence of GM-CSF, multinuclear TRAP-positive cells were formed (B). Only mononuclear TRAP-positive cells and TRAP-negative cells were observed in the presence of 0.01 ng/mL GM-CSF, and cell clusters that were TRAP-negative were detected in the presence of 0.1 ng/mL GM-CSF (B). Bar = 25 μm. (C) The c-Fms⁺RANK⁻ cells were cultured in the presence of M-CSF and sRANKL, and 1 ng/mL GM-CSF was added to the culture on days 0, 1, 3, and 5. Inhibition of osteoclastogenesis was observed only when GM-CSF was added on days 0 and 1. Control = no addition of GM-CSF.