title: High gene expression of inflammatory markers and IL-17A correlates with severity of injection site reactions of Atlantic salmon vaccinated with oil-adjuvanted vaccines.
abstract: BACKGROUND
  Two decades after the introduction of oil-based vaccines in the control of bacterial and viral diseases in farmed salmonids, the mechanisms of induced side effects manifested as intra-abdominal granulomas remain unresolved. Side effects have been associated with generation of auto-antibodies and autoimmunity but the underlying profile of inflammatory and immune response has not been characterized. This study was undertaken with the aim to elucidate the inflammatory and immune mechanisms of granuloma formation at gene expression level associated with high and low side effect (granuloma) indices.Groups of Atlantic salmon parr were injected intraperitoneally with oil-adjuvanted vaccines containing either high or low concentrations of Aeromonas salmonicida or Moritella viscosa antigens in order to induce polarized (severe and mild) granulomatous reactions. The established granulomatous reactions were confirmed by gross and histological methods at 3 months post vaccination when responses were known to have matured. The corresponding gene expression patterns in the head kidneys were profiled using salmonid cDNA microarrays followed by validation by real-time quantitative PCR (qPCR). qPCR was also used to examine the expression of additional genes known to be important in the adaptive immune response.
  
  
  RESULTS
  Granulomatous lesions were observed in all vaccinated fish. The presence of severe granulomas was associated with a profile of up-regulation of innate immunity-related genes such as complement factors C1q and C6, mannose binding protein, lysozyme C, C-type lectin receptor, CD209, Cathepsin D, CD63, LECT-2, CC chemokine and metallothionein. In addition, TGF-beta (p = 0.001), IL-17A (p = 0.007) and its receptor (IL-17AR) (p = 0.009) representing TH17 were significantly up-regulated in the group with severe granulomas as were arginase and IgM. None of the genes directly reflective of T(H)1 T cell lineage (IFN-gamma, CD4) or T(H)2 (GATA-3) responses were differentially expressed.
  
  
  CONCLUSIONS
  Granulomatous reactions following vaccination with oil-based vaccines in Atlantic salmon have the profile of strong expression of genes related to innate immune responses. The expression of TGF-beta, IL-17A and its receptor suggests an involvement of T(H)17 T cell lineage and is in conformity with strong infiltration of neutrophils and macrophages into inflamed areas. Arginase upregulation shows that macrophages in these reactions are alternatively activated, indicating also a T(H)2-profile. To what extent the expression of IL-17A and its receptor reflects an autoimmune vaccine-based reaction remains elusive but would be in conformity with previous observations of autoimmune reactions in salmon when vaccinated with oil-based vaccines.
authors.affiliation: Department of Basic Sciences and Aquatic Medicine, Norwegian School of Veterinary Sciences, Oslo, Norway.
authors.forename: Stephen
authors.initials: S
authors.lastname: Mutoloki
authors.forename: Glenn A
authors.initials: GA
authors.lastname: Cooper
authors.forename: Inderjit S
authors.initials: IS
authors.lastname: Marjara
authors.forename: Ben F
authors.initials: BF
authors.lastname: Koop
authors.forename: Øystein
authors.initials: Ø
authors.lastname: Evensen
mesh_terms: D000276:Adjuvants, Immunologic / Q000009:adverse effects*; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D004306:Dose-Response Relationship, Immunologic; D020869:Gene Expression Profiling; D006099:Granuloma / Q000139:chemically induced / Q000235:genetics* / Q000473:pathology; D007249:Inflammation / Q000235:genetics; D007267:Injections; D020381:Interleukin-17 / Q000235:genetics*; D009821:Oils; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D020317:Salmo salar / Q000235:genetics*; D014611:Vaccination / Q000009:adverse effects*
chemical_list: D000276:Adjuvants, Immunologic; D015415:Biomarkers; D020381:Interleukin-17; D009821:Oils