Figure 2: Compromised Th17 cell differentiation in ICER/CREM^−/− mice.

Naive CD4⁺ T cells from B6.ICER/CREM^+/+ mice or B6.ICER/CREM^−/− mice were polarized for 3 days as indicated condition. (a) Representative flow plots of intracellular expressions of IFNγ or IL-4 and IL-17A (left), the percentages of cells (right) and polarized in indicated condition were measured by flow cytometry (*P<0.05; mean±s.e.m., n=3). See Supplementary Fig. 2A for FACS gating strategy. (b) Real-time PCR analysis of indicated gene expressions in those differentiated cells, relative to β-actin (n=3) (*P<0.05; mean±s.e.m., n=3). (c) The percentage of IL-17-producing cells in Th17-polarized T cells with different concentrations of IL-6. Cumulative results of four independent experiments are shown (****P<0.0001, *P<0.05; mean±s.e.m., two-way analysis of variance (ANOVA)). (d–f) Real-time PCR analysis of (d) Il17a, (e) Il17f and (f) Il23r in Th17-polarized T cells on day 3. A profile representative of three mice is shown (*P<0.05; mean±s.e.m., n=3). (g) Empty vector (Empty), ICER-expressing (ICER) or ICERγ-expressing (ICERγ) plasmids were transfected to Th17-polarized T cells from indicated strains of IL-17A reporter mice on day1, and the percentage of IL-17-producing cells was measured by flow cytometry. A profile representative of three mice is shown on the right (*P<0.05; mean±s.e.m., two-way ANOVA, Bonferroni, #P<0.05 two-tailed t-test, n=3). See Supplementary Fig. 2B for FACS gating strategy.