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Ellen J Lee, Paige Snow, Joao M Furtado, Ruth Napier, Emily E Vance, Phyllis Silver, Justine Smith, Rachel R Caspi, Holly Lallman Rosenzweig; Deficiency in Nod2 is associated with dysregulation of Th17-related responses in the eye. Invest. Ophthalmol. Vis. Sci. 2017;58(8):539.
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Mutation in NOD2 results in Blau syndrome, which is characterized by granulomatous uveitis in association with dermatitis and arthritis. NOD2 belongs to a family of innate immune receptors that function in host defense, but we recently identified a novel suppressive role for Nod2 in experimental autoimmune uveitis (EAU), a T cell-dependent model of uveitis. Nod2 deficiency resulted in greater expansion, but similar proportion, of CD4+ T cells in eyes. Here we sought to study the cellular mechanisms behind modulation of EAU by Nod2.
Mice deficient in Nod2 or congenic wild type (WT) C57BL/6J controls were immunized for EAU with a mixture of interphotoreceptor retinoid-binding protein (IRBP) and IRBP1-20 peptide. Flow cytometric analysis of CD4+ T cells isolated from whole inflamed eyes harvested d21 post-immunization was used to evaluate Th1 (IFNγ+) and Th17 (IL-17+) effector subsets. Single cell suspensions isolated from inflamed eyes were re-stimulated in vitro with IRBP1-20 peptide to assess antigen-recall cytokine responses by flow cytometry after intracellular cytokine staining for IL-17A, TNF, IFNγ, GM-CSF, and IL-22. Relative contributions of IL-17 and IFNγ to disease in vivo were evaluated by fundus imaging and histopathology following antibody neutralization (100 μg/dose), on days -1,0,2,5,9, and 13 relative to immunization.
Nod2 deficiency resulted in increased percentages of ocular CD4+ T cells that produced IL-17 (6.1% vs. 1.52% in WT) or IFNγ (9.38% vs. 1.57% in WT). Antigen-recall studies of purified ocular cells revealed a dramatic increase in the proportion of IL-17+CD4+ cells in Nod2 KO compared to WT mice (12.3% vs. 3.5%). Further evaluation of IRBP-stimulated cytokine production of IL-17+CD4+ cells revealed enhanced production of all cytokines tested, including IFNγ (i.e. IL-17+IFNγ+ cells). Blockade of IL-17 in vivo significantly diminished EAU severity for both genotypes, but to a greater extent in Nod2 KO mice. In contrast, blockade of IFNγ significantly worsened EAU severity in WT mice, but did not alter disease severity in Nod2 KO mice.
These data suggest that the exacerbated uveitis accompanying Nod2 deficiency is mediated primarily through dysregulation of the Th17-related rather than Th1-related response.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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