Enhanced activation of cyclooxygenase-2 downregulates Th1 signaling pathway in Helicobacter pylori-infected human gastric mucosa.

BACKGROUND: Evidence suggests that an impaired T-cell response against Helicobacter pylori plays a role in the pathogenesis of H. pylori-related diseases. Cyclooxygenase (COX) 2 has been shown to inhibit the production of T-helper (Th) 1 cytokines. This study aimed to ascertain whether COX-2 downregulates Th1 signaling pathway in human gastric mucosa colonized by H. pylori. METHODS: COX-2 expression and prostaglandin E(2) (PGE(2)) production were determined in total proteins extracted from freshly obtained gastric biopsies of H. pylori-infected and uninfected patients by Western blotting and enzyme-linked immunosorbent assay (ELISA). Phosphorylated (p)STAT4, pSTAT1, T-bet, and pSTAT6 expression and interleukin (IL)-12, interferon (IFN)-gamma, and IL-4 production were also determined by Western blotting and ELISA, respectively, in total protein extracts from gastric biopsy cultures of H. pylori-infected patients treated without and with COX-2 inhibitor NS-398. RESULTS: Enhanced expression of COX-2 and production of PGE(2) was found in H. pylori-infected compared to uninfected patients. COX-2 inhibition significantly increased expression of Th1 transcription factors along with production of IL-12 and IFN-gamma. By contrast, no changes in the expression of STAT6 and production of IL-4 were found. CONCLUSION: This study provides a mechanism by which H. pylori may actually interfere with normal T-cell activation in human gastric mucosa, possibly enhancing its pathogenicity. The use of COX-2 selective inhibitors as immunomodulators in the course of H. pylori infection deserves investigations.

Interleukin-12 drives the Th1 signaling pathway in Helicobacter pylori-infected human gastric mucosa.

In this study we examined mechanisms that regulate T-helper lymphocyte 1 (Th1) commitment in Helicobacter pylori-infected human gastric mucosa. The levels of gamma interferon (IFN-gamma), interleukin-4 (IL-4), and IL-12 in total extracts of gastric biopsies taken from H. pylori-infected and uninfected patients were determined by an enzyme-linked immunosorbent assay. The levels of signal transducer and activator of transcription 4 (STAT4), STAT6, and T-box expressed in T cells (T-bet) in total proteins extracted from gastric biopsies were determined by Western blotting. Finally, the effect of a neutralizing IL-12 antibody on expression of Th1 transcription factors and the levels of IFN-gamma in organ cultures of H. pylori-infected biopsies was examined. Increased levels of IFN-gamma and IL-12 were found in gastric biopsy samples of H. pylori-infected patients compared to the levels in uninfected patients. In addition, H. pylori-infected biopsies exhibited high levels of expression of phosphorylated STAT4 and T-bet. Higher levels of IFN-gamma and expression of Th1 transcription factors were associated with greater infiltration of mononuclear cells in the gastric mucosa. By contrast, production of IL-4 and expression of phosphorylated STAT6 were not associated with the intensity of mononuclear cell infiltration. In ex vivo organ cultures of H. pylori-infected biopsies, neutralization of endogenous IL-12 down-regulated the expression of phosphorylated STAT4 and T-bet and reduced IFN-gamma production. Our data indicated that IL-12 contributes to the Th1 cell commitment in H. pylori-infected human gastric mucosa.

Extracellular signal-regulated protein kinase mediates interleukin 17 (IL-17)-induced IL-8 secretion in Helicobacter pylori-infected human gastric epithelial cells.

Helicobacter pylori-induced mucosal inflammation results in high production of interleukin 17 (IL-17), a potent inducer of IL-8 in gastric epithelial cells. The aim of this study was to investigate signaling pathways by which IL-17 regulates IL-8 production in human gastric epithelial cells. Activation of mitogen-activated protein (MAP) kinases in both IL-17-stimulated MKN28 cells and epithelial cells isolated from H. pylori-colonized gastric mucosa was assessed by Western blotting. In IL-17-stimulated MKN28 cells the activation of activatior protein 1 (AP-1), nuclear factor (NF)-IL-6, and NF-kappaB was also assessed by electrophoretic mobility shift assay. IL-8 production was evaluated by reverse transcription-PCR and enzyme-linked immunosorbent assay (ELISA) both for IL-17-stimulated MKN28 cells treated with specific MAP kinase inhibitors and gastric biopsy cultures treated with a neutralizing IL-17 antibody. Serum from H. pylori-infected patients was tested for immunoglobulin G response to CagA by ELISA. Treatment of MKN28 cells with IL-17 caused activation of extracellular signal-regulated protein kinase 1/2 (ERK 1/2) but not other MAP kinases and had the downstream effects of AP-1 and NF-kappaB activation and IL-8 synthesis. Blocking ERK 1/2 activity inhibited AP-1-mediated, but not NF-kappaB-mediated, IL-8 induction. Enhanced activation of ERK 1/2 was seen in gastric epithelial cells isolated from H. pylori-infected patients in comparison to uninfected controls, and this was associated with high IL-8. These effects were even more pronounced in patients seropositive for CagA than in seronegative ones. In gastric biopsy cultures, the addition of a neutralizing IL-17 antibody decreased ERK 1/2 activation, thus resulting in a significant inhibition of IL-8. In H. pylori-colonized gastric epithelial cells, IL-17-induced IL-8 synthesis is associated with and depends at least in part on the activation of ERK 1/2 MAP kinase.