title: MicroRNA-9 regulates steroid-resistant airway hyperresponsiveness by reducing protein phosphatase 2A activity.
abstract: BACKGROUND
  Steroid-resistant asthma is a major clinical problem that is linked to activation of innate immune cells. Levels of IFN-γ and LPS are often increased in these patients. Cooperative signaling between IFN-γ/LPS induces macrophage-dependent steroid-resistant airway hyperresponsiveness (AHR) in mouse models. MicroRNAs (miRs) are small noncoding RNAs that regulate the function of innate immune cells by controlling mRNA stability and translation. Their role in regulating glucocorticoid responsiveness and AHR remains unexplored.
  
  
  OBJECTIVE
  IFN-γ and LPS synergistically increase the expression of miR-9 in macrophages and lung tissue, suggesting a role in the mechanisms of steroid resistance. Here we demonstrate the role of miR-9 in IFN-γ/LPS-induced inhibition of dexamethasone (DEX) signaling in macrophages and in induction of steroid-resistant AHR.
  
  
  METHODS
  MiRNA-9 expression was assessed by means of quantitative RT-PCR. Putative miR-9 targets were determined in silico and confirmed in luciferase reporter assays. miR-9 function was inhibited with sequence-specific antagomirs. The efficacy of DEX was assessed by quantifying glucocorticoid receptor (GR) cellular localization, protein phosphatase 2A (PP2A) activity, and AHR.
  
  
  RESULTS
  Exposure of pulmonary macrophages to IFN-γ/LPS synergistically induced miR-9 expression; reduced levels of its target transcript, protein phosphatase 2 regulatory subunit B (B56) δ isoform; attenuated PP2A activity; and inhibited DEX-induced GR nuclear translocation. Inhibition of miR-9 increased both PP2A activity and GR nuclear translocation in macrophages and restored steroid sensitivity in multiple models of steroid-resistant AHR. Pharmacologic activation of PP2A restored DEX efficacy and inhibited AHR. MiR-9 expression was increased in sputum of patients with neutrophilic but not those with eosinophilic asthma.
  
  
  CONCLUSIONS
  MiR-9 regulates GR signaling and steroid-resistant AHR. Targeting miR-9 function might be a novel approach for the treatment of steroid-resistant asthma.
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Jing Jing
authors.initials: JJ
authors.lastname: Li
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Hock L
authors.initials: HL
authors.lastname: Tay
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Steven
authors.initials: S
authors.lastname: Maltby
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia; Department of Physiology, Xiangya School of Medicine, Central South University, Changsha, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Xiang
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Fiona
authors.initials: F
authors.lastname: Eyers
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, Discipline of Paediatrics and Child Health, School of Medicine and Public Health, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Luke
authors.initials: L
authors.lastname: Hatchwell
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Hong
authors.initials: H
authors.lastname: Zhou
authors.affiliation: School of Chemistry, University of New South Wales, Sydney, Australia.
authors.forename: Hamish D
authors.initials: HD
authors.lastname: Toop
authors.affiliation: School of Chemistry, University of New South Wales, Sydney, Australia.
authors.forename: Jonathan C
authors.initials: JC
authors.lastname: Morris
authors.affiliation: Firestone Institute for Respiratory Health, St Joseph's Healthcare & Department of Medicine, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Parameswaran
authors.initials: P
authors.lastname: Nair
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, Discipline of Paediatrics and Child Health, School of Medicine and Public Health, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia.
authors.forename: Joerg
authors.initials: J
authors.lastname: Mattes
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia. Electronic address: Paul.Foster@newcastle.edu.au.
authors.forename: Paul S
authors.initials: PS
authors.lastname: Foster
authors.affiliation: Priority Research Centre for Asthma and Respiratory Disease, School of Biomedical Sciences and Pharmacy, Faculty of Health, University of Newcastle and Hunter Medical Research Institute, Callaghan, Australia. Electronic address: Ming.Yang@newcastle.edu.au.
authors.forename: Ming
authors.initials: M
authors.lastname: Yang
mesh_terms: D000818:Animals; D001249:Asthma / Q000139:chemically induced / Q000188:drug therapy / Q000235:genetics* / Q000276:immunology; D016535:Bronchial Hyperreactivity / Q000139:chemically induced / Q000188:drug therapy / Q000235:genetics* / Q000276:immunology; D003907:Dexamethasone / Q000494:pharmacology; D004195:Disease Models, Animal; D021181:Egg Hypersensitivity / Q000188:drug therapy / Q000209:etiology / Q000235:genetics* / Q000276:immunology; D004804:Eosinophils / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics / Q000276:immunology; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D051379:Mice; D008807:Mice, Inbred BALB C; D035683:MicroRNAs / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000276:immunology; D009504:Neutrophils / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism; D010047:Ovalbumin; D061251:Primary Cell Culture; D054648:Protein Phosphatase 2 / Q000235:genetics* / Q000276:immunology; D011965:Receptors, Glucocorticoid / Q000235:genetics* / Q000276:immunology; D015398:Signal Transduction
chemical_list: D005938:Glucocorticoids; D008070:Lipopolysaccharides; C551804:MIRN9 microRNA, mouse; D035683:MicroRNAs; D009841:Oligonucleotides; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D007371:Interferon-gamma; D010047:Ovalbumin; D008156:Luciferases; D054648:Protein Phosphatase 2
keywords: MicroRNA; innate immunity; protein phosphatase 2A; pulmonary macrophages; steroid-resistant airway hyperresponsiveness