Up-regulation of inducible nitric oxide synthase and nitric oxide in Helicobacter pylori-infected human gastric epithelial cells: possible role of interferon-gamma in polarized nitric oxide secretion.
BACKGROUND
Nitric oxide (NO) generated by nitric oxide synthase (NOS) is known to be an important modulator of the mucosal inflammatory response. In this study, we questioned whether Helicobacter pylori infection could up-regulate the epithelial cell inducible NOS (iNOS) gene expression and whether NO production could show polarity that can be regulated by immune mediators.


METHODS
Human gastric epithelial cell lines were infected with H. pylori, and the iNOS mRNA expression was assessed by quantitative RT-PCR. NO production was assayed by determining nitrite/nitrate levels in culture supernatants. To determine the polarity of NO secretion by the H. pylori-infected epithelial cells, Caco-2 cells were cultured as polarized monolayers in transwell chambers, and NO production was measured.


RESULTS
iNOS mRNA levels were significantly up-regulated in the cells infected with H. pylori, and expression of iNOS protein was confirmed by Western blot analysis. Increased NO production in the gastric epithelial cells was seen as early as 18 hours postinfection, and reached maximal levels by 24 hours postinfection. The specific MAP kinase inhibitors decreased H. pylori-induced iNOS and NO up-regulation. After H. pylori infection of polarized epithelial cells, NO was released predominantly into the apical compartment, and IL-8 was released predominantly into basolateral compartment. The addition of IFN-gamma to H. pylori-infected polarized epithelial cells showed a synergistically higher apical and basolateral NO release.


CONCLUSIONS
These results suggest that apical NO production mediated by MAP kinase in H. pylori-infected gastric epithelial cells may influence the bacteria and basolateral production of NO and IL-8 may play a role in the tissue inflammation.
Kim
Jung Mogg
JM
Department of Microbiology and Institute of Biomedical Science, Hanyang University College of Medicine, Seoul 133-791, Korea.
Kim
Joo Sung
JS
Jung
Hyun Chae
HC
Song
In Sung
IS
Kim
Chung Yong
CY
D018938:Caco-2 Cells; D016764:Cell Polarity / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000201:enzymology / Q000382:microbiology; D005753:Gastric Mucosa / Q000166:cytology / Q000201:enzymology / Q000382:microbiology; D015870:Gene Expression / Q000276:immunology; D016481:Helicobacter Infections / Q000276:immunology* / Q000378:metabolism; D016480:Helicobacter pylori; D006801:Humans; D066298:In Vitro Techniques; D007371:Interferon-gamma / Q000276:immunology* / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009566:Nitrates / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D009573:Nitrites / Q000378:metabolism; D015854:Up-Regulation / Q000276:immunology
D004791:Enzyme Inhibitors; D009566:Nitrates; D009573:Nitrites; D009569:Nitric Oxide; D007371:Interferon-gamma; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D020928:Mitogen-Activated Protein Kinases; D019331:NG-Nitroarginine Methyl Ester