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 pro inflammatory functions in both tissue injury and allergic inflammation of the airway mucosa and submucosa, To understand the mechanisms by w hich 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 in crease in immunoreactive IL-1 beta, detectable within 2 h in both secr eted and cell-associated forms, BK-induced IL-1 beta synthesis was inh ibited by a B2-type BK receptor antagonist and by treatment of the cel ls with pertussis toxin, indicating the involvement of a BK receptor t hat couples to the G(i)/G(o) class of heterotrimeric G proteins. Where as cycloheximide and actinomycin D both inhibited BK-induced IL-1 beta synthesis, results from Northern blot and nuclear run-on assays sugge sted that BK acted primarily at the transcription level which led to t he accumulation of IL-1 beta message in stimulated cells, Gel mobility shift assays were used with nuclear extracts from stimulated WI-38 ce lls to examine the transcription mechanism for BK-induced IL-1 beta ex pression. A DNA binding activity specific for the decameric KB enhance r was detected and was found to contain the p50 and p65 subunits of th e NF-kappa B/rel protein family, BK-induced NF-kappa B activation corr elated with IL-1 beta message upregulation with respect to agonist con centration, time course, sensitivity to bacterial toxins, and blockade by the B2 receptor antagonist, After BK stimulation, a significant in crease in the activity of chloramphenicol acetyltransferase was observ ed in WI-38 cells transfected with a reporter plasmid bearing the kapp a B enhancers from the IL-1 beta gene, Deletion of the kappa B enhance r sequence significantly reduced BK-induced chloramphenicol acetyltran sferase activity, These findings suggests a novel function of BK in th e activation of NF-kappa B and the induction of cytokine gene expressi on.