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ABSTRACT: Nuclear factor kappaB (NF-kappaB) plays a key regulatory role in host cell responses to Helicobacter pylori infection in humans. Although mice are routinely used as a model to study H. pylori pathogenesis, the role of NF-kappaB in murine cell responses to helicobacters has not been studied in detail. We thus investigated the abilities of different Helicobacter isolates to induce NF-kappaB-dependent responses in murine gastric epithelial cells (GECs) and in transgenic mice harboring an NF-kappaB-responsive lacZ reporter gene. H. pylori and Helicobacter felis strains up-regulated the synthesis in mouse GECs of the NF-kappaB-dependent chemokines KC (CXCL1) and MIP-2 (CXCL2). These responses were cag pathogenicity island (cagPAI) independent and could be abolished by pretreatment with a pharmacological inhibitor of NF-kappaB. Consistent with the in vitro data, experimental Helicobacter infection of transgenic mice resulted in increased numbers of GECs with nuclear beta-galactosidase activity, which is indicative of specific NF-kappaB activation. The numbers of beta-galactosidase-positive cells in mice were significantly increased at day 1 postinoculation with wild-type H. pylori strains harboring or not harboring a functional cagPAI, compared to naive animals (P = 0.007 and P = 0.04, respectively). Strikingly, however, no differences were observed in the levels of gastric NF-kappaB activation at day 1 postinoculation with H. felis or at day 30 or 135 postinoculation with H. pylori. This work demonstrates for the first time the induction of NF-kappaB activation within gastric mucosal cells during acute H. pylori infection. Furthermore, the data suggest that helicobacters may be able to regulate NF-kappaB signaling during chronic infection.
Infection and immunity 03/2008; 76(2):551-61. · 4.21 Impact Factor
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ABSTRACT: While pleiotropic adaptive mutations are thought to be central for evolution, little is known on the downstream molecular effects allowing adaptation to complex ecologically relevant environments. Here we show that Escherichia coli MG1655 adapts rapidly to the intestine of germ-free mice by single point mutations in EnvZ/OmpR two-component signal transduction system, which controls more than 100 genes. The selective advantage conferred by the mutations that modulate EnvZ/OmpR activities was the result of their independent and additive effects on flagellin expression and permeability. These results obtained in vivo thus suggest that global regulators may have evolved to coordinate activities that need to be fine-tuned simultaneously during adaptation to complex environments and that mutations in such regulators permit adjustment of the boundaries of physiological adaptation when switching between two very distinct environments.
PLoS Genetics 02/2008; 4(1):e2. · 8.69 Impact Factor
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Bernadette Begue,
Harald Wajant, Jean-Christophe Bambou,
Laurent Dubuquoy,
Daniela Siegmund,
Jean-François Beaulieu,
Danielle Canioni,
Dominique Berrebi,
Nicole Brousse,
Pierre Desreumaux,
Jacques Schmitz,
Michael J Lentze,
Oliver Goulet,
Nadine Cerf-Bensussan,
Frank M Ruemmele
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ABSTRACT: Few data exist on the molecular events causing intestinal epithelial destruction during inflammatory processes, such as inflammatory bowel disease (IBD). In this work, we analyzed the potential implication of tumor necrosis factor (TNF)-related apoptosis inducing ligand (TRAIL) in these inflammatory lesions.
TRAIL and TRAIL-receptor expression were analyzed in normal, inflammatory ileum/colon and human intestinal epithelial cell (IEC) lines (HIEC), Caco-2, and HT-29 using RNase protection assay, real-time and reverse-transcription polymerase chain reaction (RT-PCR), immunohistochemistry, and Western blot analysis. TRAIL-induced activation of NF-kappaB was determined by electrophoretic mobility shift assay. Caspase-recruitment domain (CARD)15 expression and interleukin-(IL)8 production were studied by RT-PCR and enzyme-linked immunosorbent assay. Apoptosis was monitored using Annexin-V/caspase-3 assays.
Normal mature IEC expressed low TRAIL levels, whereas, in inflammatory lesions, TRAIL messenger RNA and protein were markedly up-regulated in IEC and lamina propria lymphocytes at levels comparable with trinitrobenzene sulfonic acid-induced colitis. Interferon-gamma and TNF-alpha potently induced TRAIL in IEC. In vitro analyses revealed a dual biologic effect of TRAIL on HIEC: Under noninflammatory conditions, TRAIL up-regulated via nuclear factor-kappaB CARD15 and IL-8, whereas, under inflammatory conditions, TRAIL became a potent inducer of apoptosis in HIEC, which was confirmed ex vivo using ileal organ cultures. TNF-alpha markedly increased the expression of the proapoptotic receptor TRAIL-R2. TRAIL-induced IEC apoptosis required a functional caspase cascade.
TRAIL is a new inflammatory mediator implicated in the homeostasis of intestinal epithelial barrier functions. TRAIL is highly up-regulated in IEC in inflammatory ileum and colon. It may augment in an auto-/paracrine fashion the elimination of IEC via apoptosis.
Gastroenterology 07/2006; 130(7):1962-74. · 11.68 Impact Factor
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ABSTRACT: The intestine is colonised by a vast population of resident bacteria which have established mutualistic relationships with their host throughout evolution, progressing from commensalism to symbiotic interactions. Intestinal bacteria benefit from resources available in their host, but reciprocally provide advantages to their host, by supplying enzymatic activities not encoded in the host genome, by promoting maturation of the intestine and of the gut associated immune system as well as by modifying the host metabolism. The commensal bacteria, although deprived of pathogenic attributes, might however become a danger for the host in case of translocation, acquisition of pathogenic features or via the inappropriate activation of intestinal inflammation. Remarkably, the commensal flora promotes the onset of innate and adaptive immune defences which, in turn, allow to set up a subtle balance between the host and the flora that promotes the symbiosis.
Journal de la Société de Biologie 02/2006; 200(2):113-20.
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ABSTRACT: The capacity of non-pathogenic enteric bacteria to induce a pro-inflammatory response is under debate in terms of its effect on the symbiosis between the mammalian host and its commensal gut microflora. Activation of NF-kappaB and induction of interleukin-8 (IL-8) and CCL-20 by the commensal Escherichia coli strain MG1655 were first studied in vitro in the human intestinal epithelial cell (IECs) lines HT29-19A and Caco-2, transfected or not with plasmids encoding dominant negative Toll-like receptor (TLR) 5 and myeloid differentiation factor-88 (MyD88) adaptor protein. The response of enterocytes in situ was then assessed using murine ileal biopsies mounted in Ussing chambers. Commensal E. coli induced NF-kappaB DNA binding, NF-kappaB transcriptional activity, CCL-20 expression, and IL-8 secretion in the human IEC lines. E. coli MG1655 flagellin was necessary and sufficient to trigger this pro-inflammatory pathway via its interaction with TLR5 and the subsequent recruitment of the adaptor protein MyD88. Following epithelial cell polarization, signaling could be induced by live E. coli and flagellin on the apical side of HT29-19A. The in vivo relevance of our findings was confirmed, because immunohistochemical staining of murine ileum demonstrated expression of TLR5 in the apical part of enterocytes in situ. Furthermore, flagellin added on the mucosal side of murine ileal biopsies mounted in Ussing chambers induced a basolateral production of KC, a functional murine homolog of human IL-8. These findings provide strong evidence that flagellin released by flagellated commensal bacteria in the intestinal lumen can induce a pro-inflammatory response in enterocytes in vivo.
Journal of Biological Chemistry 11/2004; 279(41):42984-92. · 4.77 Impact Factor
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ABSTRACT: Helicobacter pylori has been reported to induce interleukin-6 (IL-6) production in monocytes/macrophages and in chronically inflamed gastric tissues. The mechanism by which H. pylori induces IL-6 production in macrophages, however, has not been investigated. To identify the H. pylori factor responsible for this activity, we fractionated soluble proteins from H. pylori strain 26695 by ion exchange and size exclusion chromatography and screened the fractions for IL-6-inducing activity on RAW 264.7 macrophages. A single protein was purified and identified by mass spectrometry as H. pylori heat shock protein 60 (HSP60). Consistent with the observed IL-6-inducing activity of H. pylori HSP60, soluble protein extracts of H. pylori 26695 and SS1 strains that were depleted of this protein by affinity chromatography had dramatically reduced IL-6-inducing activities. The immunopurified HSP60 stimulated IL-6 production in macrophages. When stimulated with H. pylori HSP60 or intact bacteria, peritoneal macrophages from mice deficient in Toll-like receptor (TLR)-2, TLR-4, TLR-2/TLR-4, and myeloid differentiation factor 88 produced the same amount of IL-6 than macrophages from wild-type mice, demonstrating the independence of H. pylori HSP60 responses from these signaling molecules. H. pylori HSP60-induced IL-6 mRNA expression, and NF-kappaB activation in RAW 264.7 cells was abrogated in the presence of MG-132, a proteasome inhibitor. In contrast, inhibitors of protein kinase A or C, mitogen-activated protein kinase kinase, and phosphoinositide 3-kinase had no effect on IL-6 mRNA levels. This study demonstrates the induction of innate immune responses by H. pylori HSP60, thereby implicating this highly conserved protein in the pathophysiology of chronic gastritis.
Journal of Biological Chemistry 02/2004; 279(1):245-50. · 4.77 Impact Factor
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ABSTRACT: The intestine is colonised by a vast population of resident bacteria which have established mutualistic relationhips with their host throughout evolution, progressing from commensalism to symbiotic interactions. Intestinal bacteria benefit from resources available in their host, but reciprocally provide advantages to their host, by supplying enzymatic activities not encoded in the host genome, by promoting maturation of the intestine and of the gut associated immune system as well as by modifying the host metabolism. The commensal bacteria, although deprived of pathogenic attributes, might however become a danger for the host in case of translocation, acquisition of pathogenic features or via the inappropriate activation of intestinal inflammation. Remarkably, the commensal flora promotes the onset of innate and adaptive immune defences which, in turn, allow to set up a subtle balance between the host and the flora that promotes the symbiosis. L'intestin est colonisé par une masse considérable de bactéries avec lesquelles se sont établies au cours de l'évolution des relations mutualistes, progressant du commensalisme à une véritable symbiose. En effet, si les bactéries utilisent à leur profit les ressources disponibles chez l'hôte, il apparaît de plus en plus clairement qu'elles exercent sur l'hôte un effet bénéfique, en apportant des activités enzymatiques non codées par le génome de l'hôte, en favorisant la maturation de l'intestin et celle du système immunitaire, mais aussi en modifiant le métabolisme de l'hôte. Ces bactéries, dépourvues de facteurs de pathogénicité, représentent cependant un danger potentiel pour l'hôte, susceptibles en cas de translocation d'infecter des organes vitaux ou de provoquer un choc septique, mais aussi d'acquérir des facteurs de pathogénicité les rendant agressives, ou d'induire des réactions inflammatoires délétères pour l'intestin. De façon remarquable, la flore commensale favorise la mise en place des défenses immunitaires innées et adaptives dans l'intestin de l'hôte. Celles-ci permettent en retour d'établir une situation d'équilibre entre l'hôte et sa flore qui permet la symbiose.
http://dx.doi.org/10.1051/jbio:2006014.
