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S. D. Vogt, S. R. Barnum, R. W. Read; Flow Cytometric Analysis of Leukocytes Infiltrating the Eyes of Complement Inhibitor Transgenic and Wild Type Mice Induced for Experimental Autoimmune Uveitis. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5142.
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© ARVO (1962-2015); The Authors (2016-present)
We previously demonstrated that CNS-targeted production of soluble complement receptor-1 related protein Y (sCrry) ameliorated disease severity in experimental autoimmune uveitis (EAU) when compared to wild type controls. The goal of the present study was to profile infiltrating leukocytes isolated from the eyes of sCrry transgenic and wild type mice induced for EAU to determine if differences in cell populations existed between the two strains.
Mice expressing sCrry under the control of the GFAP promoter and wild type C57BL/6 mice were induced for EAU with IRBP peptide 1-20 and sacrificed day 21 post induction. Eyes were enucleated from perfused animals and digested with collagenase and DNase. Enriched single cell suspensions of leukocytes were produced using a density gradient. Retinal inflammatory cells were phenotyped ex vivo using 3 and 4 color flow cytometric analysis of the cell surface markers CD4, CD8, CD11b, CD25, and CD69 and intracellular cytokines INFγ, TNFα, IL-4 and IL-10. Statistical analyses were carried out using the student's t-test.
Percentages of cells expressing selected surface markers and cytokines are provided in Table 1. A higher percentage of CD4+ cells were seen in wild type mice, a higher percentage of IFN-gamma in sCrry mice and a higher percentage of CD25+ and CD69+ cells in sCrry mice. However, none of these findings were statistically significant. A possible trend was seen for higher percentages of CD4+ cells infiltrating wild type mouse eyes as compared to sCrry mice.
No significant differences were observed between retinal infiltrating cell populations of wild type and sCrry transgenic lines. A possible trend was identified toward a higher percentage of CD4+ cells in wild type mice, consistent with the prior finding of more severe inflammation in wild type mice.
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