Bacteroides fragilis enterotoxin (BFT) induces intestinal epithelial cell secretion of interleukin-8 (IL-8) through a nuclear factor-kB, mitrogen activated protein kinase (MAPK) co-activation pathway

Abstract

BACKGROUND: Recent studies have demonstrated that BFT induces IL-8 expression in human intestinal epithelial cells in vitro and implied a significant association between detection of the bft gene in stool specimens of patients with inflammatory bowel disease (IBD) and disease activity. These data suggest that colonization with enterotoxigenic Bacteroides fragilis may stimulate intestinal inflammation in humans. AIMS: To evaluate the kinetics of IL-8 induction stimulated by BFT in intestinal epithelial cells, and to invesugate the intracellular signaling events leading to increased IL-8 level. METHODS: Human colon epithelial cell lines, T84 and HT29/C1, were stimulated by BFT under polarized or subconfluent growth conditions in the presence or absence of (1) p38 and/or ERK MAP kinase inhibitors (SB203580, U126), or (2) the tyrosine kinase inhibitor (Geinstein). ELISA and semi-quantitative RT-PCR were used to evaluate the production of IL-8. Activation of the p38 MAP kinase pathway was confirmed by detection of the active form of p38 kinase with Western blot. NF-kB pathway activation was examined by EMSA and RelA nuclear translocation using confocal microscopy. RESULTS: IL-8 mRNA and protein are induced by biologically active BFT in a concentration-dependent and serum-independent manner. IL-8 is mainly secreted from the basolateral membranes of T84 monolayers. Basolaterally applied BFT is 25 times more potent than apical BFT in stimulating IL-8 secretion. In HT29/ C1 cells, SB203580 (10 p,M) and U126 (5 IzM) abolish 90% and 70% of 1k-8 production, respectively. Genlstein (100 IzM) completely blocks BFT-stimulated 1L-8 production. BFT stimulated nuclear translocation of NF-kB is not affected by blocking of p38 kinase and ERK pathways. CONCLUSION: BFT induces 1L-8 production in epithelial cells through coactivation of NF-kB and MAP kinase signaling pathways.