FIGURE 4. IL-1β counteracts RA-driven IL-2/STAT5–dependent repression of T_H17 development.

(a) Expression of Thy1.1 (IL-17F) and Foxp3 from naïve OTII-Tg CD4 T cells (CD4⁺CD25⁻CD62L^hiCD44^lo) from OTII.Il17f^thy1.1 mice activated with OVA peptide and IL-1β–deficient DCs for 4 days under T_H17 polarizing conditions, with or without IL-1β addition, and with or without addition of the indicated doses of rhIL-2 at day 4. (b) Frequencies of Foxp3 and IL-17F single-producers from OTII-Tg CD4⁺ T cells polarized as in a at indicated concentrations of rhIL-2 (c) Quantitative IL-2 ELISA from supernatants of naïve CD4⁺ T cells from Il17f^thy1.1 mice that were activated with plate-bound anti-CD3 and soluble anti-CD28 under T_H17 polarizing conditions in absence (left) or presence (right) of IL-1β addition, with or without addition of addition of RA at the indicated concentrations (1–100 nM). (d) Expression of Cd25 and Cd122 transcripts by quantitative RT-PCR from naïve CD4⁺ T cells from Il17f^thy1.1 mice that were activated with plate-bound anti-CD3 and soluble anti-CD28 under T_H17 polarizing conditions in absence or presence of IL-1β, with or without addition of RA (1 nM) at 60 h. Expression values are normalized to T_H0 controls. (e) Expression of IL-17A and Foxp3 from MACS-purified CD8⁻CD4⁺ thymocytes isolated from WT B6 or Stat5^fl/fl.Cd4-Cre mice that were activated with soluble anti-CD3 and Il1b^−/− splenic DCs under T_H17 polarizing condition (IL-6+TGF-β with or without the indicated additions of RA (1 nM), anti-IL-2 (10 μg/ml) and IL-1β (20 ng/ml) for 4 days. (f) Frequencies of IL-17A⁺ and Foxp3⁺ from CD4⁺ T cells from WT B6 or Stat5^fl/fl.Cd4-Cre mice as treated in (e). Data are: representative of one of three similar independent experiments a; pooled from three independent experiments with nine samples per group (n = 9) b; pooled from two independent experiments with six samples (n = 6) c; representative of two independent experiments with 6 samples per group (n = 6) where individual data points represent each sample d; representative of one of three similar independent experiments e; or pooled from three independent experiments with ten samples (n = 10) per group f. Data are means and s.e.m. in b,c,d,f). **P < 0.01 (two-tailed unpaired T-test).