IL-6 reprograms CD8⁺ T cells to become B cell helpers through their production of IL-21 in vitro. (A) Culture supernatants (SN) were collected from WT and IL-21 KO CD8⁺ T cells activated in absence (Med SN, CD8) or presence (IL-6 SN, CD8) of IL-6 for 3 d. IgG1 production by WT and IL-21R KO B cells activated for 6 d with LPS and anti-CD40 Abs using these SN was determined (n = 3). (B) WT or IL-21R KO B cells were activated using LPS+IL-4 for 6 d in SN from WT CD8⁺ T cells activated for 3 d in the presence of IL-6. IgG1 production was determined (n = 3). (C–E) WT or IL-21R KO B cells were activated using LPS (C), anti-IgM Ab (D), or anti-CD40 Ab (E) in the presence of SN obtained from WT CD8⁺ T cells activated as in A. After 6 d, IgG1 production was determined (n = 3). (F and G) In co-culture system, WT or IL-21R KO B cells were activated with LPS while WT CD8⁺ T cells were activated using anti-CD3/CD28 Abs with or without IL-6. After 6 d, the levels of IgG1 (F) and IgM (G) in the co-cultures were determined (n = 3). (H) IgG1 levels in 6 d co-cultures of WT or IL-6 KO B cells with WT CD8⁺ T cells activated as in F (n = 3). (I) IgG1 levels in 6-d co-cultures of IL-6 KO B cells with WT or IL-6R KO CD8⁺ T cells activated as in F (n = 3). (J) Co-cultures of B cells and CD8⁺ T cells were established as described in F, but a combination of an anti-IgM Ab and CpG was used to activate B cells. IgG1 levels in the co-cultures were determined (n = 3). (K) IgG1 levels in 6 d co-cultures of WT B cells with WT or IL-21 KO CD8⁺ T cells activated as in J (n = 3). Error bars represent the mean ± SD. *, P < 0.05, as determined by Student’s t test (B), one-way ANOVA (H), or two-way ANOVA test (A, C, D, E–G, and I–K). Results are representative of two to three experiments.