Nuclear factor-kappa B activation pathway in intestinal epithelial cells is a major regulator of chemokine gene expression and neutrophil migration induced by<i>Bacteroides fragilis</i>enterotoxin

Abstract

Although intestinal epithelial cells are known to up-regulate the expression of several chemokine genes in response to the stimulation with B. fragilis enterotoxin (BFT), there has been little understanding on the cellular mechanisms of BFT-induced mucosal inflammation. To test whether nuclear transcriptional factor-kappa B (NF-kappaB) is involved in the process, we stimulated intestinal epithelial cells with BFT, and evaluated the signalling NF-kappaB pathways. BFT increased signals of NF-kappaB in HT-29 and T84 epithelial cell lines as well as primary human colon epithelial cells. NF-kappaB molecules activated by BFT stimulation were composed of p65 and p50 heterodimers. In contrast, BFT decreased the signals of IkappaBalpha and IkappaB epsilon, as assessed by immunoblot. Super-repressors of IkappaBalpha, IkappaB kinase (IKK)beta, and NF-kappaB inducing kinase (NIK) inhibited an up-regulated transcription of downstream target gene (CXCL8) of NF-kappaB. Moreover, blocking the activation of NF-kappaB by MG-132 or antisense p50 oligonucleotide transfection resulted in down-regulated expression of chemokines such as CXCL1, CXCL8, and CCL2 in BFT-stimulated HT-29 cells. In addition, NF-kappaB inhibition suppressed the BFT-induced neutrophil transepithelial migration in T84 cells. These results indicate that NF-kappaB can be a central regulator of chemokine gene expression in BFT-stimulated intestinal epithelial cells and may be an important regulator of neutrophil migration.