Extended Data Fig. 1: Characterization of skin immune cells upon IL-23 induction.
From: Skin-resident innate lymphoid cells converge on a pathogenic effector state
a, Increase in ear skin thickness is significantly greater in response to IL-23 treatment than PBS vehicle. Increase in ear thickness (mean ± s.d.) following treatment with IL-23 (red, n = 10 mice) or PBS vehicle (black, n = 10 mice). Data from 2 independent experiments; repeated measures two-way ANOVA with Geisser–Greenhouse correction, Bonferroni adjusted. b, c, Immune cell composition and skin phenotype in different mouse genotypes (corresponding to the experiment in Fig. 1b). b, Left three panels: number of cells producing IL-22 or IL-17 among ILCs (black dots, bars), αβ T cells (blue squares, bars) and γδ T cells (pink triangles, bars) in wild-type, Tcrd−/− (lacking γδ T cells), and Rag1−/− (lacking all T cells and B cells) mice. Right two panels: number of total ILCs or total CD45+ cells in wild-type, Tcrd−/−, Rag1−/− and Rag2−/−Il2rg−/− (lacking T cells, B cells, and ILCs) mice. n = 5 WT, n = 3 Rag2−/−Il2rg−/−, n = 2 Tcrd−/−, n = 2 Rag1−/− mice; statistics given for WT, two-way ANOVA, Bonferroni adjustment. Experiments were repeated with similar results. c, Haematoxylin and eosin (H&E) staining of ear sections from wild-type, Tcrd−/−, Rag1−/− and Rag2−/−Il2rg−/− mice. Arrows indicate acanthosis. Representative micrographs of two independent experiments. d, Increase in skin thickness (mean ± s.d.) over time in Rag2−/−Il2rg−/− mice with (blue) or without (black) intravenously transferred ILCs. n = 4 mice for each group, pooled from 3 experiments; repeated measures two-way ANOVA with Geisser–Greenhouse correction, Bonferroni adjusted. e, IL-23-induced inflammation is dependent on Rorc. Increase in ear thickness (mean ± s.d.) following treatment with IL-23 (blue, n = 9 mice) or PBS vehicle (black, n = 8 mice) in Rorc−/− mice. Data pooled from 2 experiments; repeated measures two-way ANOVA with Geisser–Greenhouse correction, Bonferroni adjusted. f, FTY720 blocks white blood cell circulation. Total circulatory white blood cell (WBC) numbers (mean ± s.d.) in untreated (non-Tx) and FTY720-treated (FTY720-Tx) wild-type (n = 3 mice for each group) and Rag1−/− mice (n = 2 mice non-Tx, n = 4 mice FTY720-Tx). Unpaired two-tailed Welch t-test for wild-type mice. g, IL-23-dependent increase in ear skin thickness does not require circulating cells. Increase in skin thickness (mean ± s.d.) over time following IL-23 treatment in FTY720-treated (red) and untreated (black, non-Tx) Rag1−/− mice (Methods). n = 4 Rag1−/− non-Tx, n = 8 Rag1−/− FTY720-Tx mice; data from 2 independent experiments; difference not significant; repeated measures two-way ANOVA with Geisser–Greenhouse correction, Bonferroni adjusted. h, A secondary challenge with IL-23 increases susceptibility. Increase in skin thickness (mean ± s.d.) owing to primary (white) or secondary (blue) challenges by either IL-23 (n = 14 mice) or saline control (PBS) (n = 5 mice). Data from 2 independent experiments; repeated measures two-way ANOVA, Bonferroni adjusted. i, FTY720 treatment does not impact increased susceptibility to a secondary IL-23 challenge. Increase in skin thickness (mean ± s.d.) over time in mice treated initially with either IL-23 (black) or IL-23 and FTY720 (red) and subsequently with IL-23 (Methods). Bottom bars (grey): period of primary (left) and secondary (right) challenges. n = 6 IL-23 only, n = 6 IL-23 and FTY720. j, Gating strategy on CD45+ cells for sorting total skin ILCs for scRNA-seq and IL-5 fate mapping experiments. ILCs are defined as CD90.2+ and Lin− (CD3e, CD4, CD8, TCRβ, CD11b, CD11c, CD19, B220, NK1.1, Ter119, Gr1, FcεRIa), followed by exclusion of TCRγδ+ cells.
