Up-Regulation of ICAM-1 by Cytokines in Human Tracheal Smooth Muscle Cells Involves an NF-κB-Dependent Signaling Pathway That Is Only Partially Sensitive to Dexamethasone

Abstract

Abstract Although the precise mechanisms by which steroids mediate their therapeutic effects remain unknown, steroids have been reported to abrogate cytokine-induced activation of the transcription factor NF-κB. In some cell types, NF-κB activation is necessary to regulate cytokine-mediated cellular functions. However, compelling evidence suggests that the steroid inhibition of NF-κB is complex and cell specific. Using EMSA, we show that stimulation with TNF-α or IL-1β induces NF-κB DNA-binding activity in human airway smooth muscle cells. TNF-α and IL-1β also increased luciferase activity in airway smooth muscle cells transfected with a reporter plasmid containing κB enhancer elements. Cytokines activated NF-κB by rapidly degrading its cytosolic inhibitor IκBα, which was then regenerated after 60 min. Cytokine-mediated IκBα reappearance was completely blocked by the protein synthesis inhibitor cycloheximide. Inhibition of cytokine-mediated IκBα proteolysis using the protease inhibitors N-tosyl-l-phenylalanine chloromethyl ketone and N-acetyl-l-leucinyl-l-leucinyl-norleucinal also inhibited cytokine-mediated early expression of ICAM-1. Although dexamethasone partially inhibited IL-1β- and TNF-α-induced up-regulation of ICAM-1 at 4 h, dexamethasone had no effect on cytokine-induced ICAM-1 expression at 18–24 h. In addition, neither cytokine-induced degradation or resynthesis of IκBα nor NF-κB DNA-binding activity were affected by dexamethasone. In cells transfected with the luciferase reporter, dexamethasone did not affect TNF-α-induced NF-κB-dependent transcription. Interestingly, cytokine-mediated expression of cyclooxygenase-2 was completely abrogated by dexamethasone at 6 h. Together, these data demonstrate that cytokine-mediated NF-κB activation and ICAM-1 expression involve activation of a steroid-insensitive pathway.