Fig. 3: IL-33 regulates EGF production.

a Representative imaging of IL-33-GFP small intestine (SI) organoids cultured ex vivo for 48 hs. Arrows indicate GFP⁺ cells within the organoids. b, Relative expression of IL-33 mRNA in WT and IL-33-deficient organoids; data combined from three experiments (n = 6 culture wells/group). c, d WT and IL-33 KO organoid growth after irradiation. c Representative images and quantification of viable WT and IL-33 KO organoids 48 h after irradiation ex vivo; combined from three independent experiments (n = 9 wells/group). d Time course of WT (gray) and IL-33 KO (blue) organoid size following irradiation (n = 3 mice/group). e Size of organoids cultured in EGF-deficient NR media relative to their growth in EGF-containing ENR media (n = 6 wells/group). f Passaged WT and IL-33 KO SI organoids, cultured for five days in NR media without EGF before and after passaging; n = 3 mice/group from three independent experiments. g qPCR analysis of Egf expression in WT and IL-33 KO organoids cultured in ENR or NR media for four days; data combined from two independent experiments (n = 4 mice/group). h Representative western blot images of total p38 and phosphorylated p38 (p-p38) from organoids cultured +/− ST2-Fc (2 µg/ml) for eight hours (representative of 3 independent experiments). i qPCR analysis of Egf mRNA expression in IL-33 KO organoids cultured with IL-33 in NR media for 24 h +/− p38 inhibitor SB202190; n = 5 wells/group, combined from two experiments. j ELISA for EGF protein in homogenized IL-33 KO organoids cultured with IL-33 in NR media for 24 h +/− SB202190; protein level normalized for cellular content by DNA picogreen fluorescence (PF); n = 4 wells/group, combined from two experiments. Comparisons were performed using two-tailed unpaired t-tests (c, f) two-tailed Mann-Whitney U tests (b, g), Kruskal-Wallis multiple comparison testing (i), or ANOVA multiple comparison testing (d, j). Graphs indicate the mean, and error bars (d) indicate SEM; *p < .05, **p < 0.01, ***p < 0.001. Source data for graphs and the full blot for the cropped image in (h) are provided in the Source Data file. The exact p-values are as follows: (b) WT vs KO p = 0.022; (c) unirradiated WT vs unirradiated KO, p = 0.91; 6 Gy WT vs 6 Gy KO, p = 0.006; (d) 6 Gy WT vs KO at 24 h, p < 0.0008; 6 Gy WT vs KO at 48 h, p = 0.0015; (e) WT vs KO in NR, p < 0.0001; (f) organoids/well, WT vs KO, p = 0.0002; area, WT vs KO, p = 0.0002; (g) ENR, WT vs KO, p > 0.99; NR, WT vs KO, p = 0.029; (i) 0 vs 50, p = 0.0137; 50 vs 50 + SB202190, p = 0.0033; (j) 0 vs 50, p = 0.04; 50 vs 50 + SB202190, p = 0.015.