Figure 4.

The inhibitory effect of TNF‐α on osteoclastogenesis was abolished by seeding monocytes on bone. Micrographs of cultures on bone of the three subsets. (a) condition 1 (control) with 30 ng/ml M‐CSF and 20 ng/ml RANKL for 6 days; (b) condition 2, with 30 ng/ml M‐CSF and 20 ng/ml RANKL with 10 ng/ml TNF‐α for 6 days; and (c) condition 3, with 30 ng/ml M‐CSF and 20 ng/ml RANKL first cultured for 3 days followed by 3 days culture with 30 ng/ml M‐CSF, 20 ng/ml RANKL, and 10 ng/ml TNF‐α. Scale bar = 100 μm. (d) Total number of TRAcP⁺ multinucleated cells (>2 nuclei). (n = 6, *p < 0.05). (e–g) Micrographs of the TRAcP⁺ cells formed on plastic next to bone slices of the same cultures. Cells were visualized by TRAcP staining (purple) and nuclei were counterstained by DAPI (blue). Note the presence of multinucleated TRAcP⁺ cells in monocyte cultures in condition 2. Scale bar = 100 μm