Supplementary Figure 5: IL-1, but not IL-23, overrides RA-mediated repression of T_H17 development.

(a) FACS-sorted naïve CD4⁺ T cells from Il17f ^{Thy-1.1 1} mice were activated with plate-bound αCD3 and soluble αCD28 under Th17 polarizing conditions, with or without addition of IL-1β (20 ng/ml) or RA (1 nM). Cells recovered at day 3-4 were stained for surface CD4 and IL-17F (Thy1.1) and intracellular Foxp3 and analyzed by flow cytometry. Numbers are percentages of cells in the each quadrant. (b) Naïve CD4⁺ T cells from Il17f ^Thy-1.1 mice were activated under Th17 conditions as above with or without IL-1β or IL-23. Cells recovered at day 5 were stained for surface expression of CD4 and Thy1.1 (IL-17F) and intracellular expression of IL-17A, IFN-γ and Foxp3 and analyzed by flow cytometry. Numbers are percentages of cells in the each quadrant. (c) Pooled data from b, including pooled frequencies of IFN-γ⁺ cells, showing frequencies of cells expressing indicated cytokines or Foxp3. Data are representative of one of three similar experiments (a,b) or three independent experiments with 6 samples per group in c (means and s.e.m. in c). **p<0.01 (two-tailed unpaired T-test).