Innate cytokines dictate the fate of CD161^hi CD62L⁺ and CD62L⁻ cells. (A) CD161^hi CD62L⁺ (top) and CD62L⁻ (bottom) cells were stimulated with αCD3/αCD28 and IL-1β (red) or IL-12 (blue) and cultured for 14 days before assessment of Tbet, RORγ, perforin, and granzyme B expression. Data represent at least 4 experiments. (B) CD161^hi CD62L⁺ (top) and CD62L⁻ (bottom) cells were stimulated and cultured for 14 days as in panel A before stimulation with PMA/ionomycin and assessment by cytokine flow cytometry (left panels). The IFN-γ MFI is shown (right panels) for cells cultured in IL-1β (red) or IL-12 (blue). Data represent at least 7 separate experiments. (C) CD161^hi CD62L⁺ (left) and CD62L⁻ (right) cells were stimulated with αCD3/αCD28 and innate cytokines, cultured for 14 days, and restimulated with αCD3 with or without αCD28 for 72 hours before assessment of tritiated thymidine incorporation. All conditions proliferated after stimulation on day 14 with αCD3/αCD28 (data not shown) and only those stimulated with αCD3 alone are shown. Data depict the mean ± SE of at least 5 experiments. ***P < .001 (1-way ANOVA) indicates a significant difference in proliferation of cells cultured in the indicated condition and those in IL-12. (D) Sort-purified subsets were stimulated with αCD3/αCD28 in the presence of IL-1β (red) or IL-12 (blue) and cultured for 14 days before assessment of CD161 expression. Data represent at least 5 experiments. (E) CD161^hi (red) and CD161^lo (blue) CD62L⁺ (left) and CD62L⁻ (right) cells were stimulated with αCD3/αCD28 in the presence of IL-12 and cultured for 14 days before assessment of CD161 expression. Data represent 6 experiments.