Important role of Hes1 in IL-17–producing γδ T cells in the periphery. (A) Vγ4⁺ IL-17–producing γδ T cells in the thymi and spleens of adult mice. Single-cell suspensions of thymocytes and splenocytes from adult mice (6 weeks old) were stimulated with PMA and ionomycin and analyzed for IL-17–producing cells. Representative dot plots are shown after gating on γδ TCR⁺ cells. (B) Sorted Vγ4⁺ CD27^high and CD27^low γδ T cells from spleens were analyzed for the relative expression of Notch1. Notch1 expression of CD27⁺ γδ T cells was set to 1. (C-E) Hes1 is selectively involved in IL-17 production by γδ T cells in the periphery. Splenocytes from HES1^flox/flox (flox/flox Cre⁻) and HES1^flox/floxMX1-Cre (flox/flox Cre⁺) mice (4 weeks old) were prepared 4 days after the last poly I:C injection. (C-D) IFN-γ and IL-17 production by Vγ4⁺ γδ T cells was analyzed after stimulation with PMA and ionomycin. (C) Representative dot plots are shown after gating on Vγ4⁺ γδ T cells. The numbers in the respective quadrants indicate the percentage of cells. (D) Percentage of IL-17⁺ cells within CD27⁻ Vγ4⁺ γδ T cells are shown. Data shown are the means ± SD of 3 mice. (E) Absolute numbers of IFN-γ- or IL-17–producing Vγ4⁺ γδ T cells are shown. Data shown are the means ± SD of 5 mice. *P < .05. N.S. indicates statistically not significant between groups. Data are representative of 3 independent experiments.