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Toll-like receptor (TLR) 2 and TLR5, but not TLR4, are required for Helicobacter pylori-induced NF-kappa B activation and chemokine expression by epithelial cells.

Abstract

Infection with Helicobacter pylori, a Gram-negative, microaerophilic, flagellated bacteria that adheres to human gastric mucosa, is strongly associated with gastric ulcers and adenocarcinoma. The mechanisms through which gastric epithelial cells recognize this organism are unclear. In this study we evaluated the interactions between the Toll-like receptors (TLRs) and H. pylori-mediated NF-kappa B activation and the induction of chemokine mRNA expression. By reverse transcriptase-PCR we determined that MKN45 gastric epithelial cells express low but detectable amounts of TLR2, -4, and -5 but no MD-2. To determine which, if any, TLRs may play a role in the response of epithelial cells to H. pylori, HEK293 cells were cotransfected with the NF-kappa B-Luc reporter, CD14 and MD2 expression plasmids, and expression plasmids for TLR2, TLR4, or TLR5. Infection of the cultures with H. pylori (strain 26695) induced NF-kappa B activity in cells transfected with TLR2 and TLR5, but not TLR4. Consistent with the HEK293 experiments, H. pylori-induced NF-kappa B activation was decreased in MKN45 gastric epithelial cells by transfection of dominant-negative versions of TLR2 and TLR5 but not TLR4. Highly purified lipopolysaccharide from H. pylori strain 26695 activated NF-kappa B in HEK293 via TLR2 but not TLR4. Partially purified flagellin from H. pylori was also capable of inducing NF-kappa B activation in HEK cells transfected with TLR5. Additionally, chemokine gene expression was induced by H. pylori in HEK293 cells following stable transfection with TLR2 or TLR5 expression plasmids. These studies demonstrate that gastric epithelial cells recognize and respond to H. pylori infection at least in part via TLR2 and TLR5. Furthermore, the unique lipopolysaccharide of H. pylori is a TLR2, not a TLR4 agonist.

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  • Authors+Show Affiliations

    ,

    University of Virginia Health System, Digestive Health Center of Excellence and Department of Microbiology, Charlottesville, Virginia 22908-0708, USA. mfs3k@virginia.edu

    , , , , , ,

    Source

    The Journal of biological chemistry 278:35 2003 Aug 29 pg 32552-60

    MeSH

    Cell Line
    Chemokines
    DNA, Complementary
    Enzyme Activation
    Epithelial Cells
    Flagellin
    Genes, Dominant
    Genes, Reporter
    Helicobacter pylori
    Humans
    Lipopolysaccharide Receptors
    Lipopolysaccharides
    Membrane Glycoproteins
    NF-kappa B
    Plasmids
    Protein Binding
    RNA, Messenger
    Receptors, Cell Surface
    Reverse Transcriptase Polymerase Chain Reaction
    Signal Transduction
    Stomach
    Time Factors
    Toll-Like Receptor 2
    Toll-Like Receptor 4
    Toll-Like Receptor 5
    Toll-Like Receptors
    Transfection

    Pub Type(s)

    Journal Article
    Research Support, U.S. Gov't, P.H.S.

    Language

    eng

    PubMed ID

    12807870

    Citation

    Smith, Michael F., et al. "Toll-like Receptor (TLR) 2 and TLR5, but Not TLR4, Are Required for Helicobacter Pylori-induced NF-kappa B Activation and Chemokine Expression By Epithelial Cells." The Journal of Biological Chemistry, vol. 278, no. 35, 2003, pp. 32552-60.
    Smith MF, Mitchell A, Li G, et al. Toll-like receptor (TLR) 2 and TLR5, but not TLR4, are required for Helicobacter pylori-induced NF-kappa B activation and chemokine expression by epithelial cells. J Biol Chem. 2003;278(35):32552-60.
    Smith, M. F., Mitchell, A., Li, G., Ding, S., Fitzmaurice, A. M., Ryan, K., ... Goldberg, J. B. (2003). Toll-like receptor (TLR) 2 and TLR5, but not TLR4, are required for Helicobacter pylori-induced NF-kappa B activation and chemokine expression by epithelial cells. The Journal of Biological Chemistry, 278(35), pp. 32552-60.
    Smith MF, et al. Toll-like Receptor (TLR) 2 and TLR5, but Not TLR4, Are Required for Helicobacter Pylori-induced NF-kappa B Activation and Chemokine Expression By Epithelial Cells. J Biol Chem. 2003 Aug 29;278(35):32552-60. PubMed PMID: 12807870.
    * Article titles in AMA citation format should be in sentence-case
    TY - JOUR T1 - Toll-like receptor (TLR) 2 and TLR5, but not TLR4, are required for Helicobacter pylori-induced NF-kappa B activation and chemokine expression by epithelial cells. AU - Smith,Michael F,Jr AU - Mitchell,Anastasia, AU - Li,Guolian, AU - Ding,Song, AU - Fitzmaurice,Ann Marie, AU - Ryan,Kieran, AU - Crowe,Sheila, AU - Goldberg,Joanna B, Y1 - 2003/06/13/ PY - 2003/6/17/pubmed PY - 2003/10/3/medline PY - 2003/6/17/entrez SP - 32552 EP - 60 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 278 IS - 35 N2 - Infection with Helicobacter pylori, a Gram-negative, microaerophilic, flagellated bacteria that adheres to human gastric mucosa, is strongly associated with gastric ulcers and adenocarcinoma. The mechanisms through which gastric epithelial cells recognize this organism are unclear. In this study we evaluated the interactions between the Toll-like receptors (TLRs) and H. pylori-mediated NF-kappa B activation and the induction of chemokine mRNA expression. By reverse transcriptase-PCR we determined that MKN45 gastric epithelial cells express low but detectable amounts of TLR2, -4, and -5 but no MD-2. To determine which, if any, TLRs may play a role in the response of epithelial cells to H. pylori, HEK293 cells were cotransfected with the NF-kappa B-Luc reporter, CD14 and MD2 expression plasmids, and expression plasmids for TLR2, TLR4, or TLR5. Infection of the cultures with H. pylori (strain 26695) induced NF-kappa B activity in cells transfected with TLR2 and TLR5, but not TLR4. Consistent with the HEK293 experiments, H. pylori-induced NF-kappa B activation was decreased in MKN45 gastric epithelial cells by transfection of dominant-negative versions of TLR2 and TLR5 but not TLR4. Highly purified lipopolysaccharide from H. pylori strain 26695 activated NF-kappa B in HEK293 via TLR2 but not TLR4. Partially purified flagellin from H. pylori was also capable of inducing NF-kappa B activation in HEK cells transfected with TLR5. Additionally, chemokine gene expression was induced by H. pylori in HEK293 cells following stable transfection with TLR2 or TLR5 expression plasmids. These studies demonstrate that gastric epithelial cells recognize and respond to H. pylori infection at least in part via TLR2 and TLR5. Furthermore, the unique lipopolysaccharide of H. pylori is a TLR2, not a TLR4 agonist. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/12807870/Toll_like_receptor__TLR__2_and_TLR5_but_not_TLR4_are_required_for_Helicobacter_pylori_induced_NF_kappa_B_activation_and_chemokine_expression_by_epithelial_cells_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=12807870 DB - PRIME DP - Unbound Medicine ER -