CRP-induced osteoclast differentiation from CD14+ monocytes isolated from peripheral blood. (A) CD14+ monocytes, which were isolated from peripheral blood, were cultured with 25 ng/mL of M-CSF and 0 to 1.0 μg/mL of CRP in the presence or absence of anti-RANKL. After 21 days of culture, TRAP-positive multinucleated cells were counted. TRAP+ multinucleated osteoclasts were differentiated from the monocytes in a dose-dependent manner with a maximal effect at 1.0 μg/mL of CRP and anti-RANKL partially reduced the CRP-induced osteoclastogenesis. The figures represent one of three independent experiments (original magnification: 200×). (B) Monocytes were cultured on a bone-coating plate with M-CSF, CRP and anti-RANKL for 21 days. The number of pits formed by bone resorption on the plate was counted. CRP significantly increased the bone resorbing function and this CRP-induced bone resorption was incompletely inhibited by neuralization of RANKL. The figures represent one of three independent experiments (original magnification: 200×). (C) The gene expressions of osteoclast markers such as TRAP, cathepsin K, CTR, MMP-9, and RANK were measured from differentiated osteoclasts using real-time PCR. The gene expressions of TRAP, cathepsin K, MMP-9, and RANK increased significantly with CRP stimulation. The data represents the mean ± SEM for three independent experiments; *
P <0.05 and **
P <0.01. CRP, C-reactive protein; M-CSF, macrophage colony-stimulating factor; MMP, matrix metalloproteinases; RANKL, receptor activator of nuclear factor kappa-B ligand; SEM, standard error of the mean; SF, synovial fluid; TRAP, tartrate-resistant acid phosphatase.