Impaired Th17 accumulation in the gut in the absence of IL-1R1 signaling. (A) C57BL/6 Rag1^−/− mice were transferred with 4 × 10⁵ CD4⁺ CD25⁻ CD45RB^Hi T cells from C57BL/6 WT or Il1r1^−/− mice. At sacrifice, CD4⁺ T cells in the colon, MLN and spleen were evaluated by FACS analysis. Data are shown as mean ± SEM (n = 20–21 from 3 pooled independent experiments). (B) Total CD4⁺ T cells were isolated from the spleen of WT or Il1r1^−/− mice and cultured in the presence of 2.5 ng/ml TGF-β and 50 ng/ml IL-6, plus plate-bound αCD3 (2 µg/ml) and αCD28 (1 µg/ml). After 3 d, the frequency of IL-17A–producing cells was analyzed by intracellular staining. Data are shown as mean ± SD from 2 pooled independent experiments (n = 4), together with representative FACS plots (gated on CD4⁺TCRβ⁺ cells). (C–E) C57BL/6 Rag1^−/− mice were transferred with 4 × 10⁵ CD4⁺ CD25⁻ CD45RB^Hi T cells from C57BL/6 WT or Il1r1^−/− mice. (C and D) At sacrifice, colonic lamina propria leukocytes were isolated and analyzed by intracellular FACS analysis. (C) Frequency of indicated T helper subsets among CD4⁺ T cells. In the representative FACS plot shown on the right, data are gated on CD4⁺TCRβ⁺ cells. (D) Accumulation of effector CD4⁺ T cell populations in the colon. Data are shown as mean ± SEM (n = 20–21 from 3 pooled independent experiments). (E) CD4⁺ T cells were FACS sorted from the colon of recipients of Il1r1^−/− or WT naive CD4⁺ T cells and restimulated overnight with PMA and ionomycin. Cytokine secretion in the supernatants was assessed by FlowCytomix (n = 12–14 from 3 pooled independent experiments). (F) C57BL/6 Rag1^−/− mice were transferred with 4 × 10⁵ CD4⁺ CD25⁻ CD45RB^Hi T cells from C57BL/6 WT or csf2^−/− mice and culled at development of clinical signs of disease. Colonic inflammation score was assessed histologically (n = 7–10). *, P < 0.05; **, P < 0.01; ***, P < 0.001.