Characterisation of the Immunophenotype of Dogs with Primary Immune-Mediated Haemolytic Anaemia

Abstract

BackgroundImmune-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.AimsTo 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.Animals19 dogs with primary IMHA, 22 dogs with inflammatory diseases and 32 healthy control dogs.MethodsResidual 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.ResultsThere 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.ConclusionsThe 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.

The utility of diagnostic tests for immune‐mediated hemolytic anemia

“…Interleukin‐15 (IL‐15), IL‐18, granulocyte‐monocyte colony stimulating factor, and monocyte chemoattractant protein‐1 concentrations were increased in animals with IMHA that died ≤ 30 days after hospital admission . Similarly, IL‐2, IL‐6, and tumor necrosis factor‐α were present at higher concentrations in dogs with primary IMHA (n = 19) when compared with dogs that had other inflammatory diseases (n = 22) or healthy dogs (n = 32) . In question 4, none of the survey respondents indicated that they recommended these protein assays to help diagnose patients with IMHA.…”

Immunsuppressive Therapie bei Hunden und Katzen. Eigenschaften von Wirkstoffen und ihre Anwendung bei verschiedenen immunvermittelten Erkrankungen

“…Beim Menschen konnte eine klare Verbindung zwischen einer reduzierten Kontrollfunktion der regulatorischen T-Zellen und allergischen sowie autoimmunen Erkrankungen hergestellt werden (117). Die Studien zur Rolle regulatorischer T-Zellen bei immunvermittelten Erkrankungen von Hunden erbrachten allerdings unterschiedliche Ergebnisse (14,49,54,68,104,110). Volkmann et al (110) wiesen eine Reduktion der regulatorischen T-Zellen bei Hunden mit immunvermittelter Thrombozytopenie (IMTP) und chronischen Enteropathien nach.…”

“…Volkmann et al (110) wiesen eine Reduktion der regulatorischen T-Zellen bei Hunden mit immunvermittelter Thrombozytopenie (IMTP) und chronischen Enteropathien nach. Für die IMHA konnte ein veränderter Immunphänotyp im Vergleich zu gesunden Hunden und Hunden mit einer chronisch entzündlichen Erkrankung ermittelt werden (104). In Bezug auf regulatorische T-Zellen ergab sich zwischen den Gruppen allerdings kein signifikanter Unterschied.…”

Farmacocinética y efectos adversos de la doxiciclina en el tratamiento de la Ehrlichiosis: fundamentos teóricos para ensayos clínicos en caninos

“…La Infección con Ehrlichia canis cursa con cuadros de leucopenia, trombocitopenia, anemia, hiperproteinemia, hiperglobulinemia, hipergamaglobulinemia e hipoalbuminemia (Figura 1) (22). Luego de la infección, los pacientes caninos pueden sufrir anemia aguda grave la cual está relacionada con anticuerpos específicos para moléculas que se encuentran naturalmente en la pared de los eritrocitos provocando la activación de la cascada de complemento, resultando en la lisis intravascular y opsonización de los glóbulos rojos con el fin de facilitar la fagocitosis por parte de monocitos en el hígado y el bazo (anemia hemolítica inmunomediada) (24,25).…”

“…The Ehrlichia canis infection presents leukopenia, thrombocytopenia, anemia, hyperproteinemia, hyperglobulinemia, hypergammaglobulinemia and hypoalbuminemia ( Figure 1) (22). After infected, canine patients may suffer from severe acute anemia, which is related to specific antibodies that adhere to molecules found naturally in the erythrocyte wall causing the activation of the cascade complement, resulting in intravascular lysis and opsonization of the Red blood cells in order to facilitate phagocytosis by monocytes in the liver and spleen (immunemediated hemolytic anemia) (24,25).…”