TNF-induced osteoclastogenesis is restrained by RBP-J. BMMs derived from Rbpj^+/+ and Rbpj^ΔM/ΔM littermates were stimulated with RANKL or TNF in the presence of M-CSF. After 4 d, TRAP staining was performed (A), and the number of TRAP-positive MNCs per well was counted (B). TRAP-positive cells appear red in the photographs. Bar, 200 µm. *, P < 0.05; **, P < 0.01. (C) Quantitative real-time PCR analysis of mRNA expression of Acp5 (encoding TRAP) and Ctsk (encoding cathepsin K) in BMMs from Rbpj^+/+ and Rbpj^ΔM/ΔM littermates treated with TNF or RANKL in the presence of M-CSF for the indicated times. *, P < 0.05; **, P < 0.01. (D) Fluorescent images of actin ring staining from cell cultures of BMMs from Rbpj^+/+ and Rbpj^ΔM/ΔM littermates treated with TNF in the presence of M-CSF on dentin slices for 4 d. Bar, 100 µm. (E) Toluidine blue–stained dentin resorption pits formed by the osteoclasts derived from Rbpj^+/+ and Rbpj^ΔM/ΔM BMMs treated with RANKL or TNF in the presence of M-CSF for 7 d. Bar, 200 µm. (F) Quantitative real-time PCR analysis of RBPJ mRNA in human CD14-positive monocytes transfected with human RBPJ-specific siRNAs (RBPJ) or nontargeting control siRNAs (Control), and cultured for 2 d in the presence of M-CSF. **, P < 0.01. (G) Osteoclastogenesis of human osteoclast precursors transfected with RBP-J siRNAs or nontargeting control siRNAs in the presence of TNF with M-CSF for 6 d. The number of TRAP-positive MNCs per well was counted. **, P < 0.01. Data are representative of at least 20 independent experiments (A and B), or at least 3 independent experiments in (C–G).