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. 2015 Aug 14;290(39):23738–23750. doi: 10.1074/jbc.M115.667535

FIGURE 6.

FIGURE 6.

Mutation of motif 2 and motif 3 blocks TNF/IL-1-mediated osteoclastogenesis. A, schematic of hFas-WT and hFas-M23. ECD, extracellular domain; TM, transmembrane domain; ICD, intracellular domain. B, flow cytometric analysis of BMMs expressing GFP, hFas-WT, or hFas-M23. C, BMMs expressing GFP, hFas-WT, or hFas-M23 were treated with M-CSF (44 ng/ml) and α-Fas (10 ng/ml), M-CSF (44 ng/ml) and α-Fas (10 ng/ml) plus TNF (5 ng/ml), or M-CSF (44 ng/ml) and α-Fas (10 ng/ml) plus IL-1 (5 ng/ml) for 5 days. BMMs expressing GFP or hFas-WT were used as negative and positive controls, respectively. The cultures were then stained for TRAP activity on day 5. Quantification of the osteoclast formation assays is shown under the images as mean number of multinucleated TRAF-positive cells (>3 nuclei) per well. Data are mean ± S.D. **, p < 0.001; NS, not significant. Scale bars = 200 μm.