Blockade of the LFA-1 molecule resulted in increased CD4⁺CD25⁺FoxP3⁺ Tregs frequency in the LNs. (A) B6 recipients of BALB/c SG were treated with anti–LFA-1 alone, CTLA-4 Ig alone, or a combination of anti–LFA-1 plus CTLA-4 Ig, and graft draining LNs and peripheral blood were isolated 9 days after skin grafting. Cells were stained with anti-CD4, anti-CD25, and anti-FoxP3, and analyzed by flow cytometry. Data shown are gated on CD4⁺ T cells are representative of 2-3 experiments (n = 6-9 mice/group). (B) Summary data from 2-3 independent experiments with a total of 6-9 mice per group are shown. Frequencies of CD25⁺ FoxP3⁺ cells in the LNs were significantly increased in animals treated with anti–LFA-1 alone (P = .0043) and in those treated with anti–LFA-1 plus CTLA-4 Ig (P = .0022) compared with untreated control animals. Frequencies of CD25⁺Foxp3⁺ T cells in the peripheral blood were not affected by treatment with anti–LFA-1.