FIGURE 1.

CD4⁺CD25⁺ T cells from splenocytes of tumor-bearing mice are elevated, express CTLA-4 and Foxp3, and exhibit suppressor function in vitro. A. Splenocytes from tumor-bearing or age-matched naïve mice were stained by anti-CD4-PETXRED and anti-CD25-PE, and analyzed by flow cytometry. One of two independent experiments with similar results is shown. B. The frequency of CD4⁺CD25⁺ T cells in CD4⁺ T cells in naïve (n=4) or tumor-bearing (n=7) mice is shown. Tumor vs. naïve: p<0.005. The surface (C) or intracellular (D) expression of CTLA-4 in purified tumor Treg or naïve Treg is shown in one of two independent experiments with similar results. E. The expression of Foxp3 in tumor Treg or tumor CD4⁺CD25⁻ T cells is shown in one of three independent experiments with similar results. F. Tumor-primed CD4⁺ T cells were cultured alone, with tumor Treg or tumor CD4⁺CD25⁻ T cells. IL-2 in the culture supernatant was determined by ELISA. CD4 vs. CD4 + tumor Treg: p<0.0005. CD4 vs. CD4 + tumor CD4⁺CD25⁻: NS. G. Tumor-primed CD4⁺ T cells, tumor Ag-loaded DC and naïve CD8⁺ T cells were cultured alone, with tumor Treg or tumor CD4⁺CD25⁻ T cells. IFN-γ in the culture supernatants was determined by ELISA. CD4/DC/CD8 vs. CD4/DC/CD8 + tumor Treg: p<0.0005; CD4/DC/CD8 vs. CD4/DC/CD8 + tumor CD4⁺CD25⁻: NS. The data represents three independent experiments.