title: Alteration of TLR3 pathways by glucocorticoids may be responsible for immunosusceptibility of human corneal epithelial cells to viral infections.
abstract: OBJECTIVE
  The toll-like receptor 3 (TLR3) recognizes viral double-stranded RNA and its synthetic analog polyriboinosinic-polyribocytidylic acid (poly(I:C)), and the activation of TLR3 is known to induce the production of type I interferon (IFN) and inflammatory cytokines/chemokines. The purpose of this study was to determine the role played by innate responses to a herpes simplex virus 1 (HSV-1) infection of the corneal epithelial cells. In addition, we determined the effects of immunosuppressive drugs on the innate responses.
  
  
  METHODS
  Cultured human corneal epithelial cells (HCECs) were exposed to poly(I:C), and the expressions of the mRNAs of the cytokines/chemokines macrophage-inflammatory protein 1 alpha (MIP1-alpha), macrophage-inflammatory protein 1 beta (MIP1-beta), interleukin-6 (IL-6), interleukin-8 (IL-8), regulated on activation, normal T cell expressed and secreted (RANTES), Interferon-beta (IFN-beta), and TLR3 were determined using real-time reverse transcription-polymerase chain reaction (RT-PCR). The effects of dexamethasone (DEX, 10(-6) or 10(-5) M) and cyclosporine A (CsA, 10(-6) or 10(-5) M) on the expression of these cytokines and TLR3 were also determined using real-time RT-PCR. Levels of MIP1-alpha, MIP1-beta, IL-6, IL-8, RANTES, and IFN-beta were measured using the enzyme-linked immunosorbent assay (ELISA). The activation of nuclear factor kappa B (NFkappaB) and interferon regulatory factor 3 (IRF3) in HCECs was assessed by immunohistochemical staining. The effects of DEX and CsA on HCECs exposed to HSV-1 (McKrae strain) were also examined.
  
  
  RESULTS
  The expressions of MIP1-alpha, MIP1-beta, IL-6, IL-8, RANTES, IFN-beta, and TLR3 were up-regulated in HCECs exposed to poly(I:C). The poly(I:C)-induced expressions of IL-6 and IL-8 were down-regulated by both DEX and CsA, while the expressions of IFN-beta and TLR3 were suppressed by DEX alone. Similarly, the poly(I:C)-induced activation of NFkappaB was decreased by both DEX and CsA, and the activation of IRF3 was reduced by DEX alone. When HCECs were inoculated with HSV-1, DEX led to a decrease in the expression of IL6, IFN-beta, and TLR3, and an extension of plaque formation.
  
  
  CONCLUSIONS
  These results indicate that DEX may increase the susceptibility of HCECs to viral infections by altering the TLR3 signaling pathways.
authors.affiliation: Department of Ophthalmology, Ehime University School of Medicine, Shitsukawa, Japan. yukoabc@m.ehime-u.ac.jp
authors.forename: Yuko
authors.initials: Y
authors.lastname: Hara
authors.forename: Atsushi
authors.initials: A
authors.lastname: Shiraishi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Kobayashi
authors.forename: Yuko
authors.initials: Y
authors.lastname: Kadota
authors.forename: Yuji
authors.initials: Y
authors.lastname: Shirakata
authors.forename: Koji
authors.initials: K
authors.lastname: Hashimoto
authors.forename: Yuichi
authors.initials: Y
authors.lastname: Ohashi
mesh_terms: D002478:Cells, Cultured; D016572:Cyclosporine / Q000494:pharmacology; D016207:Cytokines / Q000378:metabolism; D003627:Data Interpretation, Statistical; D003907:Dexamethasone / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000187:drug effects* / Q000378:metabolism / Q000821:virology*; D015870:Gene Expression / Q000187:drug effects; D020869:Gene Expression Profiling; D005938:Glucocorticoids / Q000494:pharmacology*; D018259:Herpesvirus 1, Human / Q000502:physiology*; D006801:Humans; D050838:Interferon Regulatory Factor-3 / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D011070:Poly I-C / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects; D051196:Toll-Like Receptor 3 / Q000378:metabolism*
chemical_list: D016207:Cytokines; D005938:Glucocorticoids; C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D016328:NF-kappa B; D012333:RNA, Messenger; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3; D003907:Dexamethasone; D016572:Cyclosporine; D011070:Poly I-C