title: Roflumilast N-oxide prevents cytokine secretion induced by cigarette smoke combined with LPS through JAK/STAT and ERK1/2 inhibition in airway epithelial cells.
abstract: Cigarette smoke is a major cause of chronic obstructive pulmonary disease (COPD). Airway epithelial cells and macrophages are the first defense cells against cigarette smoke and these cells are an important source of pro-inflammatory cytokines. These cytokines play a role in progressive airflow limitation and chronic airways inflammation. Furthermore, the chronic colonization of airways by Gram-negative bacteria, contributes to the persistent airways inflammation and progression of COPD. The current study addressed the effects of cigarette smoke along with lipolysaccharide (LPS) in airway epithelial cells as a representative in vitro model of COPD exacerbations. Furthermore, we evaluated the effects of PDE4 inhibitor, the roflumilast N-oxide (RNO), in this experimental model. A549 cells were stimulated with cigarette smoke extract (CSE) alone (0.4% to 10%) or in combination with a low concentration of LPS (0.1 µg/ml) for 2 h or 24 h for measurement of chemokine protein and mRNAs and 5-120 min for protein phosphorylation. Cells were also pre-incubated with MAP kinases inhibitors and Prostaglandin E2 alone or combined with RNO, before the addition of CSE+LPS. Production of cytokines was determined by ELISA and protein phosphorylation by western blotting and phospho-kinase array. CSE did not induce production of IL-8/CXCL8 and Gro-α/CXCL1 from A549 cells, but increase production of CCL2/MCP-1. However the combination of LPS 0.1 µg/ml with CSE 2% or 4% induced an important production of these chemokines, that appears to be dependent of ERK1/2 and JAK/STAT pathways but did not require JNK and p38 pathways. Moreover, RNO associated with PGE2 reduced CSE+LPS-induced cytokine release, which can happen by occur through of ERK1/2 and JAK/STAT pathways. We report here an in vitro model that can reflect what happen in airway epithelial cells in COPD exacerbation. We also showed a new pathway where CSE+LPS can induce cytokine release from A549 cells, which is reduced by RNO.
authors.affiliation: UMR991 INSERM/Université de Rennes 1, Rennes, France.
authors.forename: Tatiana
authors.initials: T
authors.lastname: Victoni
authors.affiliation: UMR991 INSERM/Université de Rennes 1, Rennes, France.
authors.forename: Florence
authors.initials: F
authors.lastname: Gleonnec
authors.affiliation: Laboratório de Reparo Tecidual, DHE/IBRAG/UERJ, Rio de Janeiro, Brasil.
authors.forename: Manuella
authors.initials: M
authors.lastname: Lanzetti
authors.affiliation: Nycomed GmbH, Konstanz, Germany.
authors.forename: Hermann
authors.initials: H
authors.lastname: Tenor
authors.affiliation: Laboratório de Reparo Tecidual, DHE/IBRAG/UERJ, Rio de Janeiro, Brasil.
authors.forename: Samuel
authors.initials: S
authors.lastname: Valença
authors.affiliation: Laboratório de Reparo Tecidual, DHE/IBRAG/UERJ, Rio de Janeiro, Brasil.
authors.forename: Luis Cristovão
authors.initials: LC
authors.lastname: Porto
authors.affiliation: UMR991 INSERM/Université de Rennes 1, Rennes, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Lagente
authors.affiliation: UMR991 INSERM/Université de Rennes 1, Rennes, France.
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Boichot
mesh_terms: D000631:Aminopyridines / Q000494:pharmacology*; D001549:Benzamides / Q000494:pharmacology*; D002460:Cell Line; D018932:Chemokine CCL2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D054360:Chemokine CXCL1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D045424:Complex Mixtures / Q000302:isolation & purification / Q000494:pharmacology*; D003521:Cyclopropanes / Q000494:pharmacology; D015232:Dinoprostone / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D016209:Interleukin-8 / Q000037:antagonists & inhibitors / Q000378:metabolism; D053612:Janus Kinases / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D019950:Mitogen-Activated Protein Kinase 1 / Q000235:genetics / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000235:genetics / Q000378:metabolism; D058988:Phosphodiesterase 4 Inhibitors / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020545:Respiratory Mucosa / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D050791:STAT Transcription Factors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D012906:Smoke / Q000032:analysis; D014026:Nicotiana / Q000737:chemistry*
chemical_list: D000631:Aminopyridines; D001549:Benzamides; C111835:CCL2 protein, human; C475829:CXCL1 protein, human; D018932:Chemokine CCL2; D054360:Chemokine CXCL1; D045424:Complex Mixtures; D003521:Cyclopropanes; D016209:Interleukin-8; D008070:Lipopolysaccharides; D058988:Phosphodiesterase 4 Inhibitors; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D050791:STAT Transcription Factors; D012906:Smoke; D053612:Janus Kinases; C535150:MAPK1 protein, human; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C517734:roflumilast N-oxide; D015232:Dinoprostone