title: Ciliated cultures from patients with primary ciliary dyskinesia do not produce nitric oxide or inducible nitric oxide synthase during early infection.
abstract: BACKGROUND
  The mechanism behind why patients with primary ciliary dyskinesia (PCD) exhibit low nasal and exhaled nitric oxide (NO) remains unknown. One hypothesis is that reduced NO biosynthesis is caused by a defect in one or more NO synthases (NOSs). In healthy cells, the biosynthesis of NO is increased following exposure to respiratory pathogens. Here, we aimed to investigate whether ciliated epithelial cells from patients with PCD increase NO production following pneumococcal infection.
  
  
  METHODS
  Human respiratory epithelium was cultured to a basal or ciliated cell phenotype using submerged or air-liquid interface cultures, respectively. Cells were exposed to media or pneumococci until cells became damaged (&lt; 4 h). Apical fluids were collected prior and following infection, and NO production was determined using chemiluminescence. NOS gene expression was determined using real-time quantitative polymerase chain reaction.
  
  
  RESULTS
  Levels of NO and NOS2 gene expression increased significantly following infection of healthy ciliated epithelial cells but not basal cells. No increase in NO was seen in ciliated cell cultures from patients with PCD, and NOS2 gene expression remained unchanged from baseline.
  
  
  CONCLUSIONS
  These results suggest that the biosynthesis of NO in ciliated cells from patients with PCD is abnormal following early bacterial challenge, suggesting an abnormality in the function of inducible NOS in PCD.
authors.affiliation: Department of Respiratory Medicine, Portex Unit, Institute of Child Health, UCL, and Great Ormond Street Hospital for Children NHS Foundation Trust, London; Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Claire M
authors.initials: CM
authors.lastname: Smith
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Mina J
authors.initials: MJ
authors.lastname: Fadaee-Shohada
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Rounak
authors.initials: R
authors.lastname: Sawhney
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Norman
authors.initials: N
authors.lastname: Baker
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Gwyneth
authors.initials: G
authors.lastname: Williams
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Hirst
authors.affiliation: Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England.
authors.forename: Peter W
authors.initials: PW
authors.lastname: Andrew
authors.affiliation: Department of Respiratory Medicine, Portex Unit, Institute of Child Health, UCL, and Great Ormond Street Hospital for Children NHS Foundation Trust, London; Department of Infection, Immunity and Inflammation, University of Leicester, Leicester, England. Electronic address: c.ocallaghan@ucl.ac.uk.
authors.forename: Christopher
authors.initials: C
authors.lastname: O'Callaghan
mesh_terms: D002478:Cells, Cultured; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D007619:Kartagener Syndrome / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000096:biosynthesis*; D052247:Nitric Oxide Synthase Type II / Q000096:biosynthesis* / Q000235:genetics; D012313:RNA / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D012141:Respiratory Tract Infections / Q000201:enzymology* / Q000473:pathology
chemical_list: D009569:Nitric Oxide; D012313:RNA; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II