Fig. 4. IL-33-driven tuft and goblet cell expansion requires stromal IL-13.

a Distal colonic sections from Spry2^FF and Spry2^IEKO mice were probed for Il13 by in situ RNAscope analysis and counted for IL-13⁺ cells per 100 crypts (dashed line around colonic crypt at base of epithelium); n = 7 FF, 6 KO mice; *p = 0.0301. Scale bar = 25 µm. b Epithelial scrapings from Spry2^FF and Spry2^IEKO mice were measured for IL-13 levels by ELISA; n = 5 per group; *p = 0.0338. c Number of Gata3⁺ cells was quantified in distal colonic tissue. Gata3 = red, E-cadherin = green; n = 3 per group; *p = 0.0409. Scale bar = 25 µm. d Gene set enrichment analysis (GSEA) using a published intestinal ILC2 gene signature from bulk tissue³¹ was performed on RNA sequencing of distal colonic homogenates; n = 6 per group. Colonoids generated from WT mice were treated with e IL-33 (100 ng/ml, n = 5 cultures per group) or f IL-13 (10 ng/ml, n = 8 cultures per group) for 24 h, and analyzed for tuft (Dclk1) and goblet cell (Muc2, Tff3) marker expression. Each colonoid culture was from a different mouse. *p < 0.05; **p < 0.01. g WT and IL-13^−/− mice were i.p. injected with PBS or IL-33 for 4 days, and distal colonic expression of tuft cell markers (Dclk1, Trpm5, and Pou2f3) was assayed by qPCR, revealing a requirement for IL-13 in IL-33 mediated tuft cell induction in the colon; n = 7 WT + PBS, 8 WT + IL-33, 8 IL-13KO + PBS, and 10 IL-13KO + IL-33. *p < 0.05; **p < 0.01. Data are presented as mean ± SEM. Analyzed by a–f two-sided t test or g one-way ANOVA with Tukey post hoc test.