Figure 1

Induced Foxo1 Expression in Th9 Cells. (A,B) Foxo1 expression comparison in T helper cells. Foxo1 was measured by Immunoblot (A) and Taqman PCR (B) showing elevated Foxo1 expression in Th9 cells. Naïve CD4⁺ T cells were polarized under Th1, Th2, Th9, Th17 or iTreg (TGF-β1) cell conditions for 4 days and Foxo1 expression was measured by Western blot and Taqman PCR. For the Western blot, β-actin was used as loading control. (C) Temporal Foxo1 expression in Th9 cells. Naïve CD4⁺ T cells were polarized under Th9 cell condition for 1–3 days and Foxo1 expression was measured by Western blot. (D) Flow cytometry of Th9 and Th17 cells (day 4) analyzed for IL-9 and Foxo1 or IL-17A and Foxo1 expression by intracellular staining. (E,F) Induced Foxo1 expression in Th9 cells is TGF-β1/Smad3-dependent. (E) Naïve CD4⁺ T cells were TCR-activated in the presence of IL-4, TGF-β1 or combined together for 4 days and Foxo1 expression was measured by Western blot. (F) Naïve CD4⁺ T cells were differentiated under Th9 cell condition or in the presence of TGF-β1 for 4 days in the presence or absence of Smad3 inhibitor SIS3 (10 μM). Foxo1 expression was measured by Taqman PCR. Data are representative of two independent experiments. **p < 0.01 by Unpaired Student t test.