Intestinal ILC2s constrain type 3 immune response during fibrosis. (A) Experimental schematic depicting tamoxifen administration and analysis timeline. (B) Representative flow cytometry plots showing live CD45⁺Lineage⁻Il17rb-eGFP⁺ ILC2s in the SI of Salmonella-infected mice. (C) Frequency of ILC2s in the SI (n = 3–4 mice). (D) Absolute number of ILC2s in the SI and mLNs (n = 3–5 mice). (E) CFU counts of Salmonella isolated from the cecum (n = 5–7 mice). (F) Representative flow cytometry plots of live CD45⁺Lineage⁻RORγt⁺ ILC3s. (G) Absolute number of ILC3s in the SI (n = 3–4 mice). (H) Frequency of IL-17A⁺ live CD45⁺Lineage⁻RORγt⁺ ILC3s in the SI (n = 4–5 mice). (I) Frequency of IL-17A⁺CD45⁺Lineage⁻RORγt⁺ ILC3s in the mLN (n = 4–5 mice). (J) Flow cytometry plots showing live CD45⁺Lineage⁻RORγt⁺ ILC3s in the mLN. (K) Absolute number of ILC3s in the mLN (n = 5–6 mice). (L) Frequency of Lineage⁻RORγt⁺ ILC3s in the mLN (n = 5–6 mice). (M) Frequency of Lineage⁺CD3⁺IL-17A⁺ Th17 cells in the SI (n = 5–7 mice). (N) Frequency of Lineage⁺CD3⁺IL-17A⁺ Th17 cells in the mLN (n = 4–5 mice). (O)Il17a relative transcript levels in the cecum (n = 5–6 mice). (P) Flow cytometry plots showing live CD45⁺Lineage⁻GM-CSF⁺ ILC3s in the SI. (Q) Frequency of GM-CSF⁺ ILC3s in the SI (n = 4–6 mice). (R) Frequency of GM-CSF⁺ ILC3s in the mLN (n = 4–5 mice). (S) Frequency of GM-CSF⁺ Lineage⁺CD3⁺ Th17 cells in the SI (n = 4–5 mice). (T) Representative flow cytometry plots of CD64⁺MHCII⁺ macrophages in the SI. (U) Frequency of macrophages in the SI (n = 4–5 mice). (V) MHCII expression levels on macrophages in the SI (n = 4–5 mice). (W)Il22 relative transcript levels in the cecum (n = 5 mice). (X) Quantification of epithelial shedding score (n = 5–7 mice). (Y) Representative Masson’s trichrome–stained cecal tissue sections. Scale bar = 1,000 μm. (Z) Quantification of collagen deposition and Col3a transcript levels (n = 4 mice). Data are representative of two to three independent experiments. Rora^iIL17Rb refers to Il17rb-CreERT2-eGFP × Rora^fl/fl mice. WT mice refer to Il17rb-CreERT2-eGFP mice (both groups were tamoxifen-treated at the same time). Data are presented as the mean ± SEM. Statistical significance was determined using an unpaired two-tailed t test. *P < 0.05, **P < 0.01; ns, not significant. Lineage markers: CD3ε, CD3, CD5, CD8α, TCRβ, Cd11b, CD11c, CD19, B220, Ly6C, Ly6G, Ter119. For the detection of intracellular cytokines, isolated cells were stimulated with 50 ng/ml PMA (Sigma-Aldrich) and 750 ng/ml ionomycin (Sigma-Aldrich) for 4–6 h in the presence of 10 μg/ml brefeldin A (Sigma-Aldrich) in complete RPMI-1640 medium (containing 10% FBS, 50 mM 2-mercaptoethanol, 1 mM L-glutamine, 100 U/ml penicillin, and 100 μg/ml streptomycin) to evaluate their cytokine production. PMA, phorbol 12-myristate 13-acetate.