Abstract
We studied the effect of leukotriene D4 (LTD4) on a human bronchial epithelial cell line (16HBE) overexpressing the cysteinyl leukotriene (CysLT) 1 receptor (HBECysLT1R), looking at the associated signal transduction mechanisms as well as at effects on inflammatory cell adhesion. The results obtained showed that LTD4 increases the phosphorylation of extracellular signal-regulated protein kinase (ERK) 1/2 and of the signal transducer and activator of transcription 1 (STAT-1) in serine 727 (STAT-1Ser727), resulting in increased eosinophil adhesion to HBECysLT1R, associated with enhanced surface expression of intercellular adhesion molecule (ICAM) 1. Pretreatment with a CysLT1R-selective antagonist or with a selective inhibitor of protein kinase C (PKC) or with a selective inhibitor of the mitogen-activated protein kinase kinase (MEK) successfully suppressed both LTD4-induced STAT-1Ser727 phosphorylation and the associated increase in eosinophil adhesion. The use of the MEK inhibitor and of the selective CysLT1R antagonist in electrophoretic mobility shift assay experiments showed that LTD4 promotes the nuclear translocation of STAT-1 through the activation of ERK1/2 pathway. The key role of STAT-1 in leukotriene D4 transduction signaling was confirmed by RNA interference experiments, where silencing of STAT-1 expression abolished the effect of leukotriene D4 on eosinophil adhesion. In conclusion, for the first time, we provide evidence of the involvement of STAT-1 in the signal transduction mechanism of the CysLT1 receptor; phosphorylation of STAT-1, through PKC and ERK1/2 activation, causes enhanced ICAM-1 surface expression and eosinophil adhesion. Effective CysLT1R antagonism may therefore contribute to the control of the chronic inflammatory condition that characterizes human airways in asthma.
Footnotes
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This study was supported in part by the European Community VIth Framework Program (Grant LSHM CT 2004 005033; to A.S.) and by an unrestricted research grant by Merck Co.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.107.131649.
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ABBREVIATIONS: CysLT, cysteinyl leukotriene; GPCR, G protein-coupled receptor; CysLT1R, CysLT1 receptor; LTD4, leukotriene D4; MAPK, mitogen-activated protein kinase; ERK, extracellular signal-regulated protein kinase; PKC, protein kinase C; STAT, signal transducer and activator of transcription; IFN, interferon; ICAM, intercellular adhesion molecule; FBS, fetal bovine serum; GF109203X, 3-[1-[3-(dimethylaminopropyl]-1H-indol-3-yl]-4-(1H-indol-3-yl)-1H-pyrrole-2,5-dione monohydrochloride; PD98059, 2′-amino-3′-methoxyflavone; MEK, mitogen-activated protein kinase kinase; PBS, phosphate-buffered saline; EMSA, electrophoretic mobility shift assay; siRNA, small interfering RNA; ANOVA, analysis of variance.
- Received September 13, 2007.
- Accepted February 26, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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