Figure 6
Detection of Th17 cells in experimental autoimmune uveitis (EAU) donors. Normal mice (n = 20) were immunized with IRBP1-20. The spleen cells (n = 1) and intraocular cells (n = 10) were collected from EAU donors. (A) By flow cytometric analysis, fresh cells from EAU eyes (upper histogram), fresh cells from EAU spleen (middle histogram), and fresh cells from the spleen of a normal mouse (lower histogram) were stained with anti-mouse IL-17 abs and anti-mouse CD4 Abs after permeabilization. The numbers in the histograms indicate the percentages of cells that were double-positive for IL-17/CD4. (B) The intraocular T cells were collected from EAU eyes (n = 10) and evaluated by the IRBP retinal antigen-specific assay. The EAU T cells (1 × 105/well) were co-cultured with anti-TNF-α antibody (1 μg/ml) in the presence of antigen-presenting cells (20 Gy x-irradiated spleen cells: 1 × 104/well) plus mouse IRBP peptide (10 μg/ml) for 48 hours. As a control, T cells were prepared in the absence of peptide and antibody (T cells alone). ELISA was used to measure the IL-17 cytokine concentration in the supernatants of the T-cell cultures. **P < 0.005, between two groups. Abs, antibodies; IRBP, interphotoreceptor retinoid-binding protein.