Bradykinin stimulates NF-kappa B activation and interleukin 1 beta gene expression in cultured human fibroblasts

Bradykinin (BK), a pluripotent nonameric peptide, is known for its proinflammatory functions in both tissue injury and allergic inflammation of the airway mucosa and submucosa, To understand the mechanisms by which BK serves as an inflammatory mediator, the human lung fibroblast cell line WI-38 was stimulated with BK and the expression of IL-1 beta gene was examined, BK at nanomolar concentrations induced a marked increase in immunoreactive IL-1 beta, detectable within 2 h in both secreted and cell-associated forms, BK-induced IL-1 beta synthesis was inhibited by a B2-type BK receptor antagonist and by treatment of the cells with pertussis toxin, indicating the involvement of a BK receptor that couples to the G(i)/G(o) class of heterotrimeric G proteins. Whereas cycloheximide and actinomycin D both inhibited BK-induced IL-1 beta synthesis, results from Northern blot and nuclear run-on assays suggested that BK acted primarily at the transcription level which led to the accumulation of IL-1 beta message in stimulated cells, Gel mobility shift assays were used with nuclear extracts from stimulated WI-38 cells to examine the transcription mechanism for BK-induced IL-1 beta expression. A DNA binding activity specific for the decameric KB enhancer was detected and was found to contain the p50 and p65 subunits of the NF-kappa B/rel protein family, BK-induced NF-kappa B activation correlated with IL-1 beta message upregulation with respect to agonist concentration, time course, sensitivity to bacterial toxins, and blockade by the B2 receptor antagonist, After BK stimulation, a significant increase in the activity of chloramphenicol acetyltransferase was observed in WI-38 cells transfected with a reporter plasmid bearing the kappa B enhancers from the IL-1 beta gene, Deletion of the kappa B enhancer sequence significantly reduced BK-induced chloramphenicol acetyltransferase activity, These findings suggests a novel function of BK in the activation of NF-kappa B and the induction of cytokine gene expression.