title: Common and distinct signalling cascades in the production of tumour necrosis factor-alpha and interleukin-13 induced by lipopolysaccharide in RBL-2H3 cells.
abstract: BACKGROUND
  Activation of mast cells by lipopolysaccharide (LPS) results in the production of TNF-alpha and IL-13. TNF-alpha and IL-13 are key mediators in the development of neutrophilic and allergic inflammation, respectively. LPS-induced TNF-alpha and IL-13 production in mast cells has been reported to be mediated by Toll-like receptor 4 (TLR4) signalling, but differences in signal transduction mechanisms leading to the production of these cytokines are not clearly defined.
  
  
  OBJECTIVE
  We investigated the molecular mechanisms responsible for LPS-induced TNF-alpha and IL-13 production in mast cells.
  
  
  METHODS
  TNF-alpha and IL-13 production by LPS was assessed by transfecting RBL-2H3 cells with dominant-negative (DN) expression vectors.
  
  
  RESULTS
  Transfection of RBL-2H3 cells with plasmids encoding DN mutants of myeloid differentiation protein (MyD88) and TNFR-associated factor (TRAF6) inhibited both LPS-induced TNF-alpha and IL-13 production. IkappaBalpha-DN inhibited LPS-induced production of TNF-alpha, but not IL-13. We also found that inhibition of p38 kinase suppressed both TNF-alpha and IL-13 induction by LPS, and inhibition of JNK reduced IL-13 production, but not TNF-alpha. Furthermore, we found that protein kinase R (PKR) was activated by LPS in these cells. Treatment with 2-aminopurine, a PKR inhibitor, attenuated LPS-induced nuclear factor-kappaB activation and TNF-alpha production, whereas inhibition of PKR had little effect on IL-13 production.
  
  
  CONCLUSIONS
  These findings indicate that the production of TNF-alpha and IL-13 by LPS required TLR4/MyD88/TRAF6 signalling as a common pathway of mast cell-mediated inflammation. We furthermore found that TNF-alpha and IL-13 production were differentially regulated by signalling cascades through PKR and mitogen-activated protein kinases downstream of TRAF6 in mast cells.
authors.affiliation: Division of Molecular Cell Immunology and Allergology, Nihon University School of Medicine, Tokyo, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Gon
authors.forename: S
authors.initials: S
authors.lastname: Nunomura
authors.forename: C
authors.initials: C
authors.lastname: Ra
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000943:Antigens, Differentiation / Q000276:immunology; D002478:Cells, Cultured; D006967:Hypersensitivity / Q000276:immunology; D020672:I-kappa B Proteins / Q000276:immunology; D018793:Interleukin-13 / Q000096:biosynthesis*; D048031:JNK Mitogen-Activated Protein Kinases / Q000276:immunology; D008070:Lipopolysaccharides / Q000276:immunology*; D048670:MAP Kinase Kinase 4; D008407:Mast Cells / Q000276:immunology*; D020929:Mitogen-Activated Protein Kinase Kinases / Q000276:immunology; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B / Q000276:immunology; D011971:Receptors, Immunologic / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology*; D048029:TNF Receptor-Associated Factor 6 / Q000276:immunology; D014162:Transfection / Q000379:methods; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis*; D019892:eIF-2 Kinase / Q000276:immunology; D048051:p38 Mitogen-Activated Protein Kinases / Q000276:immunology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; D020672:I-kappa B Proteins; D018793:Interleukin-13; D008070:Lipopolysaccharides; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D011971:Receptors, Immunologic; D048029:TNF Receptor-Associated Factor 6; D014409:Tumor Necrosis Factor-alpha; D019892:eIF-2 Kinase; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases