title: Synthetic double-stranded RNA induces multiple genes related to inflammation through Toll-like receptor 3 depending on NF-kappaB and/or IRF-3 in airway epithelial cells.
abstract: BACKGROUND
  We hypothesized that synthetic double-stranded (ds)RNA may mimic viral infection and induce expression of genes related to inflammation in airway epithelial cells.
  
  
  OBJECTIVE
  We analysed what gene was up-regulated by synthetic dsRNA poly I : C and then focused this study on the role of Toll-like receptor 3 (TLR3), a receptor of dsRNA and its transcriptional pathway.
  
  
  METHODS
  Airway epithelial cell BEAS-2B and normal human bronchial epithelial cells were cultured in vitro. Expression of targets RNA and protein were analysed by PCR and ELISA. Localization of TLR3 expression in the cells was analysed with flow cytometry. To analyse the role of TLR3 and transcription factors, knockdown of these genes was performed with short interfering RNA (siRNA).
  
  
  RESULTS
  Real-time PCR revealed that poly I : C significantly increased the expression of mRNAs for chemokines IP-10, RANTES, LARC, MIP-1alpha, IL-8, GRO-alpha and ENA-78 and cytokines IL-1beta, GM-CSF, IL-6 and the cell adhesion molecule ICAM-1 in both cell types. Increases in protein levels were also observed. Expression of these genes was significantly inhibited in BEAS-2B cells in which TLR3 expression was knocked down. However, pre-treatment with anti-TLR3 mAb, which interferes with the function of TLR3 expressed on the cell surface, did not inhibit the genes expression and these data were concordant with the results that TLR3 was expressed inside airway epithelial cells. The study of siRNA for NF-kappaB and IRF3 showed that they transduce the signal of poly I : C, but their roles were different in each target gene.
  
  
  CONCLUSIONS
  TLR3 is expressed inside airway epithelial cells and transduces synthetic dsRNA signals. These signals may increase expression of inflammatory cytokines, chemokines and ICAM-1 through activation of transcription factors NF-kappaB and/or IRF3 in airway epithelial cells.
authors.affiliation: First Department of Internal Medicine, Showa University School of Medicine, Tokyo, Japan. smatsuku@med.showa-u.ac.jp
authors.forename: S
authors.initials: S
authors.lastname: Matsukura
authors.forename: F
authors.initials: F
authors.lastname: Kokubu
authors.forename: M
authors.initials: M
authors.lastname: Kurokawa
authors.forename: M
authors.initials: M
authors.lastname: Kawaguchi
authors.forename: K
authors.initials: K
authors.lastname: Ieki
authors.forename: H
authors.initials: H
authors.lastname: Kuga
authors.forename: M
authors.initials: M
authors.lastname: Odaka
authors.forename: S
authors.initials: S
authors.lastname: Suzuki
authors.forename: S
authors.initials: S
authors.lastname: Watanabe
authors.forename: H
authors.initials: H
authors.lastname: Takeuchi
authors.forename: T
authors.initials: T
authors.lastname: Kasama
authors.forename: M
authors.initials: M
authors.lastname: Adachi
mesh_terms: D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000998:Antiviral Agents / Q000276:immunology / Q000494:pharmacology*; D001980:Bronchi / Q000276:immunology* / Q000378:metabolism; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D015870:Gene Expression; D006801:Humans; D050838:Interferon Regulatory Factor-3 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016328:NF-kappa B / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D011070:Poly I-C / Q000276:immunology / Q000494:pharmacology*; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051196:Toll-Like Receptor 3 / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015854:Up-Regulation; D014777:Virus Diseases / Q000276:immunology* / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D000998:Antiviral Agents; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D016328:NF-kappa B; D051196:Toll-Like Receptor 3; D011070:Poly I-C