Fig. 4: IL-1R activation drives the CRC EGC phenotype.

a, b Top ligands from orthotopic CRC tumor-infiltrating immune cells predicted by NicheNet to be inducing CRC EGC signature. NicheNet analysis was performed considering the ligands expressed by murine orthotopic CRC tumor-infiltrating immune cells, data extracted from CRC orthotopic murine CRC dataset (see Supplementary Fig. 4a) and considering the differentially expressed genes between 24 h healthy conditioned medium (H-CM) EGCs and tumor microenvironment (TME)-CM EGCs as target genes, data extracted from in vitro CRC EGCs bulk RNAseq dataset (see Fig. 2). Schematic representation of TME-derived ligand-EGC interplay (left) and top 5 predicted ligands with their Pearson correlation (right) (a). Heatmap of ligand-target pairs showing regulatory potential scores between top ligands and target genes among the differentially expressed genes between in vitro H-CM EGCs and TME-CM EGCs (b). c Protein level of IL-1β in H-CM and TME-CM (n = 6 H-CM and TME-CM). d, e Primary adult neurosphere-derived EGCs were isolated from WT C57BL/6J mice and treated with or without recombinant (r)IL-1β (10 ng/mL) for 24 h. Protein concentration in the culture supernatants was determined by liquid chromatography/mass spectrometry (n = 4). Schematic experimental representation (d) and heatmap of differentially expressed proteins between vehicle and rIL-1β-treated EGCs (e). f, g Primary embryonic neurosphere-derived EGCs were stimulated for 24 h with H-CM or TME-CM in the presence or absence of IgG or anti-IL-1R (5 µg/mL each) (n = 3 H-CM and TME-CM). Schematic representation of experimental setup (f). Relative mRNA levels for Lcn2, Timp1, Ccl2, and Il6 normalized to the housekeeping gene Rpl32 (g). h WT C57BL/6J mice were intracolonically injected at day(d)0 with MC38 cells, and both stromal and immune cells were assessed for IL-1β and IL-1α expression at d21. Data are presented as the frequency of total live IL-1β⁺ or IL-1α⁺ cells (n = 5 mice). Data are represented as mean ± SEM (c, g, h). Statistical analysis: unpaired two-tailed Mann-Whitney test (c), unpaired one-way Anova test with multiple comparison correction (g). **p < 0.005, ***p < 0.0005, ****p < 0.00005. Source data and exact p values are provided as a Source Data file.