Figure 3: Suppressive effects of LGK974 on LPS-induced inflammatory response in bronchial epithelial cells.

(A,C,E,G) BEAS-2B human bronchial epithelial cells were pretreated with 10 nM LGK974 for 2 hours, followed by 0.1 μg/ml LPS stimulation for various time periods of 0.25–2 hours. Western blotting was used to assess protein levels and phosphorylation of NF-κB and IκB. Beta-actin and TBP were used as endogenous controls for total cell lysate and nuclear fraction, respectively (A). The binding of nuclear NF-κB for its target DNA sequence, 5′-GGGACTTTCC-3′, was measured by ELISA (C). The expression of pro-inflammatory cytokines, IL-6 and IL-8, was measured by real time-qPCR (E,G). *p < 0.05, **p < 0.01, ***p < 0.001 compared with LPS-stimulated cells without LGK974 pretreatment at the same treatment hour. (B,D,F,H) Cells were pretreated with 0.001–10 nM of LGK974 for 2 hours, followed by 0.1 μg/ml of LPS stimulation for 2 hours. Western blotting was used to assess protein levels and phosphorylation of NF-κB and IκB (B). The binding of nuclear NF-κB for its target DNA sequence was measured by ELISA (D). The expression of pro-inflammatory cytokines was measured by real time-qPCR (F, H). *p < 0.05, **p < 0.01, ***p < 0.001 compared with LPS-stimulated cells without LGK974 pretreatment. ^###p < 0.001 compared with cells without LPS stimulation pretreatment.