The absence of DNAM-1 on donor cells attenuates GVHD in a T_reg-dependent fashion after lethal and sublethal conditioning. (A,B) All donor mice were treated with 500 μg CD25 antibody (Ab; PC61) for T_reg depletion or rat IgG1 (isotype control; MAC49, T_reg replete) intraperitoneally on days −3 and −1. (A) Lethally irradiated B6C3F1 mice were transplanted with 5 × 10⁶ BM from CD25 Ab– or IgG–treated B6.WT mice and 2 × 10⁶ MACS-purified CD4⁺ T cells from CD25 Ab– or IgG–treated B6.WT or B6.DNAM-1^−/− mice on day 0. TCD BM from B6.WT donors was transplanted into irradiated B6C3F1 recipients as non-GVHD controls. Representative fluorescence activated cell sorting (FACS) dots of T_reg depletion with CD25 Ab or isotype control (left) and survival data (right) are shown. P = .77, CD25 Ab–treated B6.WT vs B6.DNAM-1^−/−. **P = .0024, IgG-treated B6.WT vs B6.DNAM-1^−/−. Data combined from 4 experiments (n = 26 to 28 per T-cell–deplete group, n = 18 per T-cell–replete group, and n = 6 to 8 in TCD control). (B) Sublethally irradiated (500 cGy) B6D2F1 mice were transplanted with 50 × 10⁶ splenocytes from CD25 Ab– or IgG–treated B6.WT or B6.DNAM-1^−/− mice on day 0. T-cell depleted (TCD) grafts from IgG-treated B6.WT donors were transplanted as non-GVHD controls. P = .0002, IgG-treated B6.WT vs B6.DNAM-1^−/−; P = .065, CD25 Ab–treated B6.WT vs B6.DNAM-1^−/−. Data combined from 2 replicate experiments (n = 12 per T-cell–replete group and n = 8 in TCD control).