Figure 6.

Exogenous IL-2 rescues WKO nTreg cell dysfunction. (A) WKO nTreg cells do not proliferate upon CD3-mediated stimulation and respond to exogenous IL-2. [³H]thymidine uptake of CD4⁺CD25⁺ nTreg cells isolated from the spleens of WT (black bars) or WKO (white bars) mice 3 d after stimulation with mitomycin-treated T cell–depleted splenocytes and 0.5 μg/ml anti-CD3ɛ or 0.5 μg/ml anti-CD3ɛ plus 10 μg/ml IL-2. Shown are averages ± SD of one representative of three independent experiments performed in triplicate. (B) IL-2 and antigen receptor–mediated preactivation of WKO nTreg cells substantially rescues their proliferation defects. [³H]thymidine uptake of CD4⁺CD25⁻ T cells isolated from the spleens of WT mice and cocultured with CD4⁺CD25⁺ cells from WT (black curves) or WKO (gray curves) mice at different CD25⁺/CD25⁻ ratios. Dashed curves represent suppressive effects of WT or WKO nTreg cells after a prior 3-d activation with 10 μg/ml of plate-bound anti-CD3ɛ and 25 μg/ml IL-2. Shown are results of one out of three independent experiments performed in triplicate. (C) Decreased IL-10 secretion of stimulated WKO nTreg cells. IL-10 concentrations were determined by ELISA in the supernatant of nTreg cells or CD4⁺CD25⁻ splenocytes isolated from WT or WKO mice that were unstimulated or stimulated for 3 d with 10 μg/ml anti-CD3ɛ and 25 μg/ml IL-2, or PMA and ionomycin. Shown are averages of one representative of four independent experiments, each performed in duplicate. Error bars represent standard deviations.