Helicobacter pylori-dependent ceramide production may mediate increased interleukin 8 expression in human gastric cancer cell lines

Background & Aims: Helicobacter pylori adheres to gastric epithelial cells, activates nuclear factor KB (NF-KB), and stimulates interleukin (IL)-8 pro duction, but the responsible molecular mechanisms remain largely unknown. B ecause several studies have shown that sphingolipids are involved in a numb er of signaling pathways, including NF-KB activation, we investigated the p ossible role of sphingolipids in the regulation of IL-8 expression in Kato III and AGS cells. Methods: IL-8 production in the conditioned media was qu antified by enzyme immunoassay. Induction of messenger RNA (mRNA) was asses sed by Northern blot analysis. Activation and binding activity of transcrip tion factors were examined by luciferase assay and electrophoretic mobility shift assay, respectively. Intracellular levels of ceramide were quantifie d by diacylglycerol kinase assay. Results: A cell-permeable ceramide analog ue (C-2-ceramide) increased IL-8 expression with comparable mRNA induction. This effect was mimicked by sphingomyelinase, but not by phospholipase A(2 ) OF phospholipase C. C-2-ceramide induced IL-8 gene transcription mainly t hrough activation of NF-KB because mutation of the NF-KB-binding site compl etely abrogated the induction of luciferase activity. Direct contact of liv e H. pylori with epithelial cells increased the intracellular concentration of ceramide. Conclusions: The results suggest a novel role of the sphingom yelin-ceramide pathway, at least in part through NF-KB, in IL-8 production induced by H. pylori infection in gastric epithelial cells.