IL-6 induces the production of IL-21 in CD8⁺ T cells through Stat3. (A and B) CD8⁺ and CD4⁺ T cells were activated using anti-CD3/CD28 Abs in the absence or presence of IL-6. IL-21 production was determined by ELISA (n = 3). (C) IL-4 production after activation for 72 h (n = 3). (D) CD69 expression in fresh CD8⁺ T cells (shaded) or activated as in A in the absence (blue) or presence (red) of IL-6. (E) Percentage of live CD8⁺ T cells activated as in A for 2 d by flow cytometry (n = 3). (F) CD8⁺ T cells were activated as in A for 72 h with or without IL-6, as indicated. IL-21 production was determined (n = 3). (G and H) CD8⁺ T cells were activated in the absence (Med) or presence of cytokines. IL-21 (G) and IFN-γ (H) production were determined (n = 3). (I) Relative mRNA levels for IL-21 in CD8⁺ T cells activated with or without IL-6 or IL-2. (J) The levels of Tyr⁷⁰⁵-phosphorylated STAT3 (P-STAT3), total STAT3, or actin in CD8⁺ T cells were activated in the absence or presence of IL-6 for 0 min (Uns), 15 min, 30 min, 1 h, or 3 h. (K and L) Relative IL-21 mRNA levels (K) and IL-21 production (L) in WT or Stat3 T cell conditional KO (Stat3 KO) CD8⁺ T cells activated with or without IL-6 for 72 h (n = 3). Error bars represent the mean ± SD. *, P < 0.05, as determined by Student’s t test (E), one-way (F–H) or two-way ANOVA (A, B, C, and L). u.d., undetectable levels. Results are representative of two to three experiments.