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Helicobacter bilis gamma-glutamyltranspeptidase enhances inflammatory stress response via oxidative stress in colon epithelial cells.

ABSTRACT: Helicobacter bilis (H. bilis) infection is associated with cases of inflammatory bowel Disease, thyphlocolitis, hepatitis and cholecystitis. However, little is known about the bacterial virulence determinants or the molecular mechanisms involved. Recently, H. bilis ?-glutamyltranspeptidase (HBgGT) was shown to be a virulence factor decreasing host cell viability. Bacterial gGTs play a key role in synthesis and degradation of glutathione and enables the bacteria to utilize extracellular glutamine and glutathione as sources of glutamate. gGT-mediated loss of cell viability has so far been linked to DNA damage via oxidative stress, but the signaling cascades involved herein have not been described. In this study, we identified enhanced ROS production induced by HBgGT as a central factor involved in the activation of the oxidative stress response cascades, which finally activate CREB, AP-1 and NF-?B in H. bilis infected colon cancer cells. IL-8, an important pro-inflammatory chemokine that is a common downstream target of these transcription factors, was up-regulated upon H. bilis infection in an HBgGT dependent manner. Moreover, the induction of these signaling responses and inflammatory cytokine production in host cells could be linked to HBgGT-mediated glutamine deprivation. This study implicates for the first time HBgGT as an important regulator of signaling cascades regulating inflammation in H. bilis infected host epithelial cells that could be responsible for induction of inflammatory disorders by the bacterium.

SUBMITTER: Javed S

PROVIDER: S-EPMC3751837 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Helicobacter bilis gamma-glutamyltranspeptidase enhances inflammatory stress response via oxidative stress in colon epithelial cells.

Javed Sundus S Mejías-Luque Raquel R Kalali Behnam B Bolz Christian C Gerhard Markus M

PloS one 20130823 8

Helicobacter bilis (H. bilis) infection is associated with cases of inflammatory bowel Disease, thyphlocolitis, hepatitis and cholecystitis. However, little is known about the bacterial virulence determinants or the molecular mechanisms involved. Recently, H. bilis γ-glutamyltranspeptidase (HBgGT) was shown to be a virulence factor decreasing host cell viability. Bacterial gGTs play a key role in synthesis and degradation of glutathione and enables the bacteria to utilize extracellular glutamine ...[more]

PMID: 24009737

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Project description:BACKGROUND:Helicobacter saguini is a novel enterohepatic Helicobacter species isolated from captive cotton top tamarins with chronic colitis and colon cancer. Monoassociated H. saguini infection in gnotobiotic IL-10-/- mice causes typhlocolitis and dysplasia; however, the virulent mechanisms of this species are unknown. Gamma-glutamyltranspeptidase (GGT) is an enzymatic virulence factor expressed by pathogenic Helicobacter and Campylobacter species that inhibits host cellular proliferation and promotes inflammatory-mediated gastrointestinal pathology. The aim of this study was to determine if H. saguini expresses an enzymatically active GGT homologue with virulence properties. EXPERIMENTAL PROCEDURES:Two putative GGT paralogs (HSGGT1 and HSGGT2) identified in the H. saguini genome were bioinformatically analysed to predict enzymatic functionality and virulence potential. An isogenic knockout mutant strain and purified recombinant protein of HSGGT1 were created to study enzymatic activity and virulence properties by in vitro biochemical and cell culture experiments. RESULTS:Bioinformatic analysis predicted that HSGGT1 has enzymatic functionality and is most similar to the virulent homologue expressed by Helicobacter bilis, whereas HSGGT2 contains putatively inactivating mutations. An isogenic knockout mutant strain and recombinant HSGGT1 protein were successfully created and demonstrated that H. saguini has GGT enzymatic activity. Recombinant HSGGT1 protein and sonicate from wild-type but not mutant H. saguini inhibited gastrointestinal epithelial and lymphocyte cell proliferation without evidence of cell death. The antiproliferative effect by H. saguini sonicate or recombinant HSGGT1 protein could be significantly prevented with glutamine supplementation or the GGT-selective inhibitor acivicin. Recombinant HSGGT1 protein also induced proinflammatory gene expression in colon epithelial cells. CONCLUSIONS:This study shows that H. saguini may express GGT as a potential virulence factor and supports further in vitro and in vitro studies into how GGT expression by enterohepatic Helicobacter species influences the pathogenesis of gastrointestinal inflammatory diseases.

Project description:The confounding consequences of Helicobacter bilis infection in experimental mice populations are well recognized, but the role of this bacterium in human diseases is less known. Limited data are available on virulence determinants of this species. In Helicobacter pylori, ?-glutamyltranspeptidase (?GT) contributes to the colonization of the gastric mucosa and to the pathogenesis of peptic ulcer. The role of ?GT in H. bilis infections remains unknown. The annotated genome sequence of H. bilis revealed two putative ggt genes and our aim was to characterize these H. bilis ?GT paralogues. We performed a phylogenetic analysis to understand the evolution of Helicobacter ?GTs and to predict functional activities of these two genes. In addition, both copies of H. bilis ?GTs were expressed as recombinant proteins and their biochemical characteristics were analysed. Functional complementation of Esherichia coli deficient in ?GT activity and deletion of ?GT in H. bilis were performed. Finally, the inhibitory effect of T-cell and gastric cell proliferation by H. bilis ?GT was assessed. Our results indicated that one gene is responsible for ?GT activity, while the other showed no ?GT activity due to lack of autoprocessing. Although both H. bilis and H. pylori ?GTs exhibited a similar affinity to L-Glutamine and ?-Glutamyl-p-nitroanilide, the H. bilis ?GT was significantly less active. Nevertheless, H. bilis ?GT inhibited T-cell proliferation at a similar level to that observed for H. pylori. Finally, we showed a similar suppressive influence of both H. bilis and H. pylori ?GTs on AGS cell proliferation mediated by an apoptosis-independent mechanism. Our data suggest a conserved function of ?GT in the Helicobacter genus. Since ?GT is present only in a few enterohepatic Helicobacter species, its expression appears not to be essential for colonization of the lower gastrointestinal tract, but it could provide metabolic advantages in colonization capability of different niches.

Project description:BACKGROUND:Interleukin-33 (IL-33) is a cytokine belonging to the IL-1 family, and its possible involvement in the pathophysiology of COPD and viral-induced exacerbations has been demonstrated. IL-33 has been shown to be increased in the airway epithelial cells from COPD patients, but the regulating mechanism of IL-33 expression in airway epithelial cells remains largely unknown. In the current study, we examined whether oxidative stress, which participates in the pathogenesis of COPD, affects the expression of IL-33 in airway epithelial cells and also evaluated the effect during viral infection. METHODS:The involvement of oxidative stress in the expression of IL-33, and its signal pathway was examined after stimulation with hydrogen peroxide (H2O2), with or without stimulation by polyinosinic-polycytidylic acid [poly (I:C)], a synthetic analogue of dsRNA that mimics viral infection, or rhinovirus infection in NCI-H292 cells and primary human bronchial epithelial cells (HBECs). In addition, the effect of antioxidant, N-acetylcysteine (NAC) in the expression of IL-33 was compared between HBECs from healthy subjects and those from COPD patients. RESULTS:Treatment with H2O2 significantly potentiated IL-33 expression in NCI-H292 cells, and the potentiation was reversed by NAC treatment. Mitogen-activated protein kinase (MAPK) inhibitors, but not nuclear factor-kappa B inhibitors, also significantly decreased the H2O2-potentiated IL-33 expression. In addition, H2O2 significantly potentiated the poly (I:C)- or rhinovirus-stimulated IL-33 expression. In HBECs from healthy subjects, H2O2-potentiated IL-33 expression and its reversal by NAC was also confirmed. Under the condition without H2O2-stimulation, treatment with NAC significantly decreased the expression of IL-33 in HBECs from COPD patients, but not in those from healthy subjects. CONCLUSIONS:These results demonstrate that oxidative stress involves in the expression of IL-33 in airway epithelial cells via MAPK signal pathway and it augments IL-33 expression during viral infection. This mechanism may participate in the regulation of IL-33 expression in airway epithelial cells in COPD and the viral-induced exacerbations. Modulation of this pathway could become a therapeutic target for viral-induced exacerbations of COPD.

Dataset Information

Helicobacter bilis gamma-glutamyltranspeptidase enhances inflammatory stress response via oxidative stress in colon epithelial cells.

Publications

Helicobacter bilis gamma-glutamyltranspeptidase enhances inflammatory stress response via oxidative stress in colon epithelial cells.

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