Figure 5.

Neutrophils from IKKβ^Δmye mice produce increased IL-1β following urethane injection. Expression of cytokines by A) mRNA and B) protein in the lungs of WT and IKKβ^Δmye mice harvested 1 week after urethane (n=10-11 mice per group; *p < 0.05 compared with WT). C) Concentration of IL-1β in the conditioned media following 12-hour culture of lung Ly6G⁺ neutrophils, Ly6C⁺ monocytes, or Ly6G/C^neg macrophages isolated from IKKβ^Δmye mice at 1 week after urethane injection (n=3; *p < 0.05 compared with Ly6C⁺ and Ly6G/C^neg). D) Concentration of IL-1β in the conditioned media following 12-hour culture of equal numbers of lung Ly6G⁺ neutrophils from WT and IKKβ^Δmye mice isolated at 1 week after urethane injection (n=8 mice per group). E) IL-1β protein levels in lung homogenates at 1 week after urethane in the lungs of IKKβ^Δmye mice treated with liposomal clodronate or PBS on day 5 following urethane injection (n=6 mice per group). F) IL-1β protein levels at 1 week after urethane in the lungs of IKKβ^Δmye mice treated with anti-Ly6G antibodies (100 μg) or control IgG antibodies by IP injection on days −1, 2, and 5 relative to the day of urethane injection (n=3-5 mice per group; *p < 0.05 compared with IKKβ^Δmye mice treated with control IgG antibodies). Lung Ly6G⁺ neutrophils were isolated from WT and IKKβ^Δmye mice at 1 week after urethane injection. G) IL-1β concentration in the conditioned media after culture with inhibitors (all 100 M) of caspase-1 (Ac-YVAD-CMK), neutrophil elastase and proteinase-3 (MeOSuc-APPV-CMK), or cathepsin G (Z-GLP-CMK) (n=3-8 replicates per group; #p>0.05 compared to WT Unt; *p<0.05 compared to either WT or IKKβ^Δmye Unt). H) mRNA expression of cathepsin G in lung Ly6G⁺ neutrophils isolated from WT and IKKβ^Δmye mice at 1 week after urethane injection.