The isolation of intestinal lamina proprial cells and flow cytometry were done as previously described(6 (link)). Splenocytes were made into single cell suspension and CD4+ T cells were purified with CD4+ T cell isolation kit (Miltenyi) with 90–95% purity. 24-well-plates were coated with anti-hamster antibody (MP Biomedical). CD4+ T cells were cultured in IMDM medium (Sigma Aldrich) supplied with soluble hamster-anti-mouse CD3 (0.25 µg/ml unless otherwise indicated in the text), hamster-anti-mouse CD28 (1 µg/ml), Gentamicin (50 µg/ml), anti-mouse IL-4 (11B11, 2 µg/ml, BioXCell), and anti-mouse IFN-γ XMG1.2, 2 µg/ml, BioXCell). For iTreg cell differentiation, 5 ng/ml TGF-β was added to the culture. For Th17 cell differentiation, TGF-β was added at 5 ng/ml and IL-6 was added at 20 ng/ml. In some experiments, FICZ was added at a concentration of 200 nM. For blocking RORγt or Stat3 activity, cells were cultured with 10 µM Digoxin (Sigma) or 15 µM STA21 (Santa Cruz), respectively, with controls of DMSO. For IL-21 neutralization, the cells with cultured with 12.5 µg/ml anti-IL21 (R&D Systems) or control IgG.