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Fig. 3 | BMC Immunology

Fig. 3

From: Lipopolysaccharide- TLR-4 Axis regulates Osteoclastogenesis independent of RANKL/RANK signaling

Fig. 3

Evaluation of the involvement of TLR4 signaling in RANKL-induced osteoclastogenesis. a Immunoblotting analyses with antibodies against TLR4 (~ 95 kDa) and GAPDH (loading control; ~ 37 kDa) are shown. Protein levels were quantified by densitometry, corrected for the sample load based on GAPDH expression, and expressed as a fold increase relative to the control lane (−). The results represent one of three experiments performed. b Representative images of TRAP stained osteoclasts in response to treatment with RANKL and TAK-242 (5 μM/mL). TRAP stained osteoclasts in panels A and B were obtained with a 4× objective (magnification: 40X), while those in panels C and D were obtained with a 10× objective (magnification: 100X). c The number of TRAP-positive multinucleated osteoclasts were counted in both groups (n = 3). Statistical analysis was performed to compare the number of osteoclasts in the RANKL+TAK-242 group with the control group (RANKL). The t-test was applied; the difference between groups is not statistically significant. d TRAP stained osteoclasts in response to treatment with LPS (5 μg/mL) and LPS/TAK-242 (5 μM/mL) are shown. TRAP stained osteoclasts in panels A and C were obtained with a 4× objective (magnification: 40X), while those in panels B and D were obtained with a 10× objective (magnification: 100X). e The number of TRAP-positive multinucleated osteoclasts were counted in both groups (n = 3). Statistical analysis was performed to compare the number of osteoclasts in the LPS + TAK-242 group with the control group (LPS) using the t-test. **P < 0.01 vs LPS group. f Effect of TAK-242 on LPS-induced TNF-α production from RAW cells-derived osteoclast. ELISA determined the concentrations of TNF-α in the culture medium. One-way ANOVA was applied, and values are expressed as mean ± standard deviation (SD). **P < 0.01 vs. LPS group. Scanned uncropped autoradiograms are presented in Additional file 5, Figure S5. Corresponding immunoblots are shown in panel A

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