Interleukin-8 up-regulation by neutrophil elastase is mediated by MyD88/IRAK/TRAF-6 in human bronchial epithelium

Cystic fibrosis is characterized in the lungs by neutrophil-dominated infla mmation mediated significantly by neutrophil elastase (NE). Previous work h as shown that NE induces interleukin-8 (IL-8) gene expression and protein s ecretion in bronchial epithelial cells. We sought to determine the intracel lular mechanisms by which NE up-regulates IL-8 in bronchial epithelial cell s. The data show that stimulation of 16HBE14o(-) cells with NE induced IL-8 protein production and gene expression. Both responses were abrogated by a ctinomycin D, indicating that regulation is at the transcriptional level. E lectrophoretic mobility shift assays demonstrated that nuclear factor kappa B (NF kappaB) was activated in 16HBE14o(-) cells stimulated with NE. Wester n blot analysis demonstrated that activation of NF kappaB by NE was precede d by phosphorylation and degradation of I kappaB proteins, principally I ka ppaB beta. In addition, we observed that interleukin-1 receptor-associated kinase (IRAK) was degraded in 16HBE14o(-) cells stimulated with NE. Quantif ication of IL-8 reporter gene activity by luminometry demonstrated that dom inant negative MyD88 (MyD88 Delta) or TRAF-6 (TRAF-6 Delta) inhibited IL-8 reporter gene expression in response to NE. Furthermore, MyD88 Delta inhibi ted NE-induced IRAK degradation. These results show that NE induces IL-8 ge ne up-regulation in bronchial epithelial cells through an IRAK signaling pa thway involving both MyD88 and TRAF-6, resulting in degradation of I kappaB beta and nuclear translocation of NF kappaB. These findings may have impli cations for therapeutic treatments in the cystic fibrosis condition.