Treg-mediated suppression of T-cell proliferation is impaired at the site of autoimmune inflammation. (A-B) PBMCs and SFMCs were stained for CD4 and FOXP3 expression by flow cytometry. (A) Dotplots showing the percentage of FOXP3⁺ cells within CD4⁺ cells in paired PBMCs (left panel) and SFMC (right panel) of JIA patients, 1 representative of n = 16. (B) Accumulative data of the percentage of CD4⁺FOXP3⁺cells in PBMCs of HCs and paired PBMCs and SFMCs from JIA patients (n = 16), ***P < .001. (C-E) CD4⁺CD25⁺CD127^low Tregs were sorted by flow cytometry and co-cultured with CFSE-labeled effector cells. At day 4, proliferation of CFSE⁺ effector cells was analyzed. (C) Proliferation of CD4⁺ (let panel) and CD8⁺ (right panel) SFMCs in the absence (open histograms) or presence of Tregs at 1 to 4 ratio (closed histograms). Percentages indicate the percentage of proliferating cells, 1 representative of n = 3. (D-E) Suppression of CD4⁺ (D) and CD8⁺ T-cell proliferation (E) in the presence of Tregs at a 1:8 and 1:4 ratio or additional effector cells (+eff) at a 1:4 ratio for PBMCs from HC (black bars), PBMCs from JIA patients (gray bars) and SFMCs from JIA patients (white bars). The results show percentage of suppression in the presence of Tregs or additional effector cells relative to effector cells cultured alone. Bars represent mean ± SEM of n = 6 PBMCs HCs, n = 2 PBMCs JIA, and n = 3 SFMCs JIA, *P < .05, ***P < .001 compared with PBMCs (HCs).