NF-κB activation potentiates the degree of IL-1β production and secretion upon inflammasome activation.

<p>THP-1 reporter cells were primed overnight with heat-killed <i>P. aeruginosa</i> and stimulated the next day with live <i>P. aeruginosa</i> or additional heat-killed <i>P. aeruginosa</i> for the times indicated. Cell culture supernatants were assayed for (A) NF-κB/AP-1 activity, (B) IL-8, and (C) IL-1β secretion (n = 3–6 experiments). Using the same stimulation method, THP-1 reporter cells were treated with Bay11-7082 (20 µM) for 1 hour prior to priming with heat-killed PAO1 or live PAO1. Supernatants were assayed at 24 hours for (D) NF-κB/AP-1 activity, (E) IL-8, and (F) IL-1β secretion (n = 3–5). PBMCs from CF patients (n = 11–15) and controls (n = 10–13) were treated with z-YVAD-fmk (20 µM) or Bay11-7082 (10 µM) and stimulated with live PAO1 (MOI = 1), ATP (5 mM), or Poly(dA:dT) (1 µg/ml) according to the schedule in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037689#pone-0037689-g001" target="_blank">Figure 1</a>. (G) IL-1β and (H) IL-8 levels were measured at 24 hours. Statistical analysis was performed using two way ANOVA with Bonferroni correction for multiple comparisons. *, **, and *** signify P<0.05, 0.01, and 0.001.