Immune cells in the tumor and draining lymph nodes in IL-17–deficient mice. MC38 cells were inoculated into IL-17–deficient mice and normal wild-type mice as we described. Twenty-five days after tumor inoculation, single-cell (s.c.) suspensions were made from tumor tissues, tumor-draining lymph nodes (TDLN) and spleens, or non–tumor-bearing IL-17–deficient mice and wild-type mice. (A-E) Phenotype and cytokine profile of each immune cell subsets were analyzed by fluorescence-activated cell sorter. Results are the percentage of the targeted cell subset in a particular immune cell population. One representative of 5 experiments is shown in panels A to E (n = 13 mice per group in panels A-E). (A) T-cell subsets in tumor. Single-cell suspensions were made from tumor tissues. The levels of CD4⁺ and CD8⁺ T cells, and FOXP3⁺ T cells were quantified with LSR II by gating on CD3⁺ cells. The levels of these 3 T-cell subsets were comparable in IL-17–deficient mice and wild-type mice (P > .05 for all). (B) IL-10⁺ T cells and Treg cells in tumor. Single-cell suspensions were made from tumor tissues. The levels of IL-10⁺CD4⁺ T cells and CD4⁺FOXP3⁺ T cells were quantified with LSR II by gating on CD3⁺CD4⁺ cells. The levels of these 2 T-cell subsets were comparable in IL-17–deficient mice and wild-type mice (P > .05 for all). (C) IFN-γ⁺ T cells in tumor. Single-cell suspensions were made from tumor tissues. The levels of IFN-γ⁺CD4⁺ T cells and IFN-γ⁺CD8⁺ T cells were quantified with LSR II by gating on CD3⁺ cells. The levels of these 2 T-cell subsets were significantly lower in IL-17–deficient mice than wild-type mice (P < .01 for all). (D) NK cells in TDLNs. Single-cell suspensions were made from TDLNs. The levels of CD49b⁺NK1.1⁺ NK cells were quantified with LSR II by gating on CD3⁻ cells. The levels of NK cells were significant lower in IL-17–deficient mice than wild-type mice (P < .01). (E) IFN-γ⁺ cells in TDLNs. Single-cell suspensions were made from TDLNs. The levels of IFN-γ⁺ NK cells, IFN-γ⁺CD4⁺ T cells, and IFN-γ⁺CD8⁺ T cells were quantified with LSR II by gating on NK cells and CD4⁺ and CD8⁺ T cells. The levels of these 3 IFN-γ⁺ cell subsets were significantly lower in IL-17–deficient mice than wild-type mice (P < .01 for all). (F) NK- and T-cell activities in tumor-bearing mice (top panels) and non–tumor-bearing mice (bottom panels). TDLN T cells were stimulated with irradiated MC38, and non–TDLN T cells were stimulated with phorbol-12-myristate-13-acetate and ionomycin. Spleen NK cells were stimulated with IL-2 and irradiated YAC-1 cells. The expression of IFN-γ was detected by ELISPOT assay in these stimulated T and NK cells. Results are mean ± SEM values of IFN-γ⁺ spots in triplicate wells. □ represent wild-type mice; and ■, IL-17–deficient mice. n = 5 mice per group. *P < .01. **P < .05.