pH60/Surv vaccine reduces the percentage of CD4⁺CD25⁺ T cells in Peyer patches without their activation. (A) Percentage of CD4⁺CD25⁺ cells in Peyer patches. ^*P < .05 compared with the pBud control group. (B) Expression of CD25 (left panel), CD69 (middle panel), and GITR (right panel) on CD3⁺CD4⁺CD25⁺ cells. Shaded histograms represent the pBud control group; open histograms, the pH60/Surv group. Experiment was repeated once and yielded similar results. (C) Inhibition of CD3-induced splenocyte proliferation by CD4⁺CD25⁺ cells. CFSE-labeled splenocytes were cultured with anti-CD3 antibody in the absence (shaded area) or presence of unlabeled CD3⁺CD4⁺CD25⁺ cells (purity greater than 99%, as determined by flow cytometry) at a 100:1 ratio. CD3⁺CD4⁺CD25⁺ cells were isolated from pBud control mice (black dotted line) or pH60/Surv-vaccinated mice (gray line).