Figure 7

IL-10 neutralizing antibody reversed suppressive activities of T _reg cells. (A) CD4⁺CD8^-CD25⁺CD44⁺ and CD4⁺CD8^- CD25⁺CD44^- cells were sorted from splenocytes from C57BL/6 mice and cultured with naïve CD4⁺CD25^- T cells in the presence of anti-CD3/anti-CD28 for 72 h. In some cases, anti-IL-10 or isotype control Ab was added. Cell proliferation was measured by [³H]-thymidine incorporation. Data are presented as counts for min, and shown are the mean ± S.D. from three independent experiments (** p < 0.01, *** p < 0.001). (B) Splenocytes from naïve mice were treated with either the IL-10 neutralizing antibodies or an isotype control (rat IgG1) in the presence or absence of Con A (5 μg/ml) for 24 h. Cells were stained with antibodies against CD4, CD8, CD25, CD44, and Foxp3 and analyzed by flow cytometry. (C) CD4⁺CD25^high cells were sorted from splenocytes of C57BL/6 mice and cultured with anti-IL-10 or isotype control Abs for 24 h. Cells were stained with antibodies against Foxp3 and CD4 and analyzed by flow cytometry. Data are presented as the percentage of total splenocytes and shown are representative results of one of four independent experiments.