IL-17A/polyI:C-provoked synergistic induction of proinflammatory cytokines depends on the NF-κB pathway.

<p>BEAS-2B cells were transfected with siRNA for NF-κB p65 (solid bars) or a negative control (shadowed bars) for 24 hours. At 2 days after exposure to siRNA, transfected cells were treated for 24 hours with IL-17A and/or polyI:C, and then the expression of mRNA was evaluated by quantitative real-time PCR. Treatment with p65 siRNA reduced endogenous p65 mRNA levels about 90% as compared with the random oligomer negative control (A). PolyI:C-induced G-CSF and IL-8 mRNA expression was suppressed by p65 siRNA, and the IL-17A/polyI:C-provoked synergistic induction was strongly attenuated (B, C). BEAS-2B cells were pre-incubated in the presence of the IκBα inhibitor BAY11-7082 (0–10 μM) for 1 hour and treated with IL-17A and/or polyI:C in submerged cultures. After 24-hour treatments, total RNA was harvested, and G-CSF (D) and IL-8 (E) mRNA expression was evaluated using quantitative real-time PCR that was normalized to β-actin levels. BAY11-7082 treatment inhibited IL-17A/polyI:C-provoked synergistic G-CSF and IL-8 induction in a concentration-dependent manner. All measurements were averaged from duplicate wells, and the experiment was repeated three times. * p < 0.001, p65 siRNA vs. negative control. † p < 0.01, p65 siRNA vs. negative control in polyI:C treatment. ‡ p < 0.01, p65 siRNA vs. negative control in co-treatment with IL-17A and polyI:C. § p < 0.05, vs. BAY11-7082 absent.