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. Author manuscript; available in PMC: 2016 Aug 1.
Published in final edited form as: J Bone Miner Res. 2015 May 21;30(8):1508–1522. doi: 10.1002/jbmr.2472

Figure 3. The pro-inflammatory cytokines IL-17A and TNFα suppress TcREG induction by osteoclasts.

Figure 3

A. TcREG induction by osteoclasts in the presence of IL-17: TcREG were induced in vitro using OT-I CD8 T-cells co-cultured with bone marrow cells-derived osteoclasts, pulsed with SIINFEKL ovalbumin peptide. TcREG induction was observed in OT-I T-cells in the absence of (0 ng/ml) but significantly reduced in the presence of 10 ng/ml IL-17A. B. TcREG induction by osteoclasts in the presence of TNFα: TNFα also reduced TcREG induction in a dose dependent manner. Similar levels of suppression of TcREG induction was observed at 50 or 100 ng/ml RANKL each dose of TNFα indicating that the ratio of RANKL to TNFα is not critical. ANOVA analysis to determine the variation due to TNFα and RANKL concentration indicates that TNFα accounted for 85.5% of the total variance (P < 0.0001) and RANKL accounted for 0.65% of the total variance (P < 0.001). The interaction term accounted for remaining 9.3% of the total variance indicating that TNFα had a strong effect at each concentration of RANKL tested. The results represent mean ± standard deviation (error bars) from 4 experiments (n = 4 wells/group/experiment). P values were calculated using Mann-Whitney test; **** = P < 1×104 and ** = P < 0.01 relative to no (0 ng/ml of) TNFα.