title: Characterisation of the Immunophenotype of Dogs with Primary Immune-Mediated Haemolytic Anaemia.
abstract: BACKGROUND
  Immune-mediated haemolytic anaemia (IMHA) is reported to be the most common autoimmune disease of dogs, resulting in significant morbidity and mortality in affected animals. Haemolysis is caused by the action of autoantibodies, but the immunological changes that result in their production have not been elucidated.
  
  
  OBJECTIVE
  To investigate the frequency of regulatory T cells (Tregs) and other lymphocyte subsets and to measure serum concentrations of cytokines and peripheral blood mononuclear cell expression of cytokine genes in dogs with IMHA, healthy dogs and dogs with inflammatory diseases.
  
  
  METHODS
  19 dogs with primary IMHA, 22 dogs with inflammatory diseases and 32 healthy control dogs.
  
  
  METHODS
  Residual EDTA-anti-coagulated blood samples were stained with fluorophore-conjugated monoclonal antibodies and analysed by flow cytometry to identify Tregs and other lymphocyte subsets. Total RNA was also extracted from peripheral blood mononuclear cells to investigate cytokine gene expression, and concentrations of serum cytokines (interleukins 2, 6 10, CXCL-8 and tumour necrosis factor α) were measured using enhanced chemiluminescent assays. Principal component analysis was used to investigate latent variables that might explain variability in the entire dataset.
  
  
  RESULTS
  There was no difference in the frequency or absolute numbers of Tregs among groups, nor in the proportions of other lymphocyte subsets. The concentrations of pro-inflammatory cytokines were greater in dogs with IMHA compared to healthy controls, but the concentration of IL-10 and the expression of cytokine genes did not differ between groups. Principal component analysis identified four components that explained the majority of the variability in the dataset, which seemed to correspond to different aspects of the immune response.
  
  
  CONCLUSIONS
  The immunophenotype of dogs with IMHA differed from that of dogs with inflammatory diseases and from healthy control dogs; some of these changes could suggest abnormalities in peripheral tolerance that permit development of autoimmune disease. The frequency of Tregs did not differ between groups, suggesting that deficiency in the number of these cells is not responsible for development of IMHA.
authors.affiliation: Department of Clinical Science and Services, Royal Veterinary College, Hawskhead Lane, North Mymms, Hatfield, Hertfordshire, United Kingdom.|Immune Regulation Laboratory, Royal Veterinary College, London, United Kingdom.
authors.forename: James W
authors.initials: JW
authors.lastname: Swann
authors.affiliation: Immune Regulation Laboratory, Royal Veterinary College, London, United Kingdom.
authors.forename: Kelly
authors.initials: K
authors.lastname: Woods
authors.affiliation: Immune Regulation Laboratory, Royal Veterinary College, London, United Kingdom.
authors.forename: Ying
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Department of Clinical Science and Services, Royal Veterinary College, Hawskhead Lane, North Mymms, Hatfield, Hertfordshire, United Kingdom.
authors.forename: Barbara
authors.initials: B
authors.lastname: Glanemann
authors.affiliation: Department of Clinical Science and Services, Royal Veterinary College, Hawskhead Lane, North Mymms, Hatfield, Hertfordshire, United Kingdom.|Immune Regulation Laboratory, Royal Veterinary College, London, United Kingdom.
authors.forename: Oliver A
authors.initials: OA
authors.lastname: Garden
mesh_terms: D000744:Anemia, Hemolytic, Autoimmune / Q000097:blood / Q000276:immunology / Q000662:veterinary*; D000818:Animals; D016207:Cytokines / Q000097:blood; D004283:Dog Diseases / Q000097:blood / Q000276:immunology*; D004285:Dogs; D005260:Female; D016130:Immunophenotyping; D018836:Inflammation Mediators / Q000097:blood; D016753:Interleukin-10 / Q000097:blood; D018655:Lymphocyte Count; D008297:Male; D050378:T-Lymphocytes, Regulatory / Q000276:immunology
chemical_list: D016207:Cytokines; D018836:Inflammation Mediators; D016753:Interleukin-10