IsoLCA protects from fibrosis by inhibiting IL-17A production by ILC3 and Th17 cells during intestinal inflammation. All data presented in this figure use Salmonella-infected C57BL/6J-congenic mice. (A) Flow cytometry plots of ILC3s in the SI. (B) Absolute number of ILC3 (n = 3–6 mice). (C) MFI of RORγt by live CD45⁺Lineage⁻RORγt⁺ ILC3s (n = 3–6 mice). (D) Flow cytometry plots of ILC3s in the mLN. (E) Flow cytometry plots of IL-17A production by live CD45⁺Lineage⁻CD3⁻ ILC3s in the mLN. (F) Frequency of IL-17A⁺ ILC3s in the mLN (n = 3–5 mice). (G) Number of IL-17A⁺ ILC3 in mLN (n = 3–5 mice). (H) Flow cytometry plots of Th17 cells in the SI. (I) Frequency of Th17 cells in the SI (n = 4–5 mice). (J) Number of IL-17A⁺IFNγ⁻ Th17 cells (n = 4–5 mice). (K) Number of IL-17A⁺IFNγ⁺ pathogenic Th17 cells (n = 4–5 mice). (L) Number of IL-17A⁺IFNγ⁺ Th1 cells. (M)Il17a (IL-17A) transcript levels in ceca. (N) Masson’s trichrome–stained cecal tissue sections. Scale bar = 1,000 μm. (O) Quantification of collagen deposition (n = 4–5 mice). Lineage markers: CD3ε, CD3, CD5, CD8α, TCRβ, Cd11b, CD11c, CD19, B220, Ly6C, Ly6G, Ter119. Data are representative of two independent experiments and 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). 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.