title: Leukotriene B(4) enhances tumour necrosis factor-alpha-induced CCL27 production in human keratinocytes.
abstract: BACKGROUND
  A chemokine CCL27 recruits skin-homing T cells. CCL27 production by epidermal keratinocytes is dependent on nuclear factor-kappaB (NF-kappaB) activity and is enhanced in lesions with atopic dermatitis or allergic contact dermatitis. A lipid mediator leukotriene B(4) (LTB(4)) may be involved in the development of these allergic dermatoses. LTB(4) acts on cell surface G-protein-coupled receptors, BLT1 and BLT2.
  
  
  OBJECTIVE
  The aim of this study was to investigate the in vitro effects of LTB(4) on CCL27 production in human keratinocytes.
  
  
  METHODS
  Keratinocytes were incubated with TNF-alpha and LTB(4). CCL27 secretion and mRNA levels were analysed by ELISA and RT-PCR, respectively. NF-kappaB activities were analysed by luciferase assays. Protein levels or phosphorylation status were analysed by cell-based ELISA.
  
  
  RESULTS
  LTB(4) alone did not enhance CCL27 production and modestly enhanced NF-kappaB activity in human keratinocytes. However, LTB(4) potently enhanced TNF-alpha-induced CCL27 secretion and mRNA expression and NF-kappaB activity. LTB(4) alone or together with TNF-alpha, induced phosphorylation and degradation of inhibitory NF-kappaB alpha (IkappaBalpha) and phosphorylation of NF-kappaB p65. These effects of LTB(4) were suppressed by BLT1 antagonist U75302, pertussis toxin, phosphoinositide-3 kinase (PI3K) inhibitor LY294002 and extracellular signal-regulated kinase (ERK) kinase inhibitor U0126, but not by BLT2 antagonist LY255283. LTB(4) induced phosphorylation of ERK and Akt, downstream kinase of PI3K; LY294002 suppressed phosphorylation of both kinases while U0126 suppressed only the former.
  
  
  CONCLUSIONS
  These results suggest that LTB(4) may enhance TNF-alpha-induced CCL27 production by activating NF-kappaB via the BLT1/G(i/o)/PI3K/ERK pathway in human keratinocytes. LTB(4) may contribute to the enhanced CCL27 production of keratinocytes in lesions with atopic dermatitis or allergic contact dermatitis.
authors.affiliation: Department of Dermatology, School of Medicine, Teikyo University, Tokyo, Japan. nmk@med.teikyo-u.ac.jp
authors.forename: N
authors.initials: N
authors.lastname: Kanda
authors.forename: S
authors.initials: S
authors.lastname: Watanabe
mesh_terms: D002478:Cells, Cultured; D054425:Chemokine CCL27; D019742:Chemokines, CC / Q000235:genetics / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D015603:Keratinocytes / Q000201:enzymology / Q000378:metabolism*; D007975:Leukotriene B4 / Q000378:metabolism*; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D018102:Receptors, Leukotriene B4 / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D051996:Transcription Factor RelA / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D015854:Up-Regulation
chemical_list: C492276:CCL27 protein, human; D054425:Chemokine CCL27; D019742:Chemokines, CC; D020672:I-kappa B Proteins; C513240:LTB4R protein, human; C485878:LTB4R2 protein, human; C000606204:NFKBIA protein, human; D012333:RNA, Messenger; D018102:Receptors, Leukotriene B4; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D007975:Leukotriene B4; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D020930:MAP Kinase Kinase Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go