CD103⁺ Treg cells induced apoptosis of activated CD4⁺ T cells from chronic GVHD recipients. (A) Twenty days after infusion of CD103⁺ Treg cells, the spleen cells of the untreated and treated recipients as well as control donor DBA/2 mice were stained for CD5.1 versus CD4. The CD5.1⁺CD4⁺ donor-type T cells were gated. One representative of 4 recipients is shown. The mean (± SE) of the percentage of CD5.1⁺CD4⁺ T cells among spleen mononuclear cells of untreated and treated recipients are 16.8% (± 3.6%) and 8.5% (± 1.6%), respectively. (B) Gated FoxP3⁻CD4⁺ T cells were shown in CD44 versus CD62L. The activated CD44^hiCD62L^loCD4⁺ T cells were gated, and the percentage of the cells among total FoxP3⁻CD4⁺ T cells was shown above the gated box. One representative of 4 recipients is shown. The means (± SE) of the percentage of CD44^hiCD62L^loCD4⁺ T cells among total FoxP3⁻CD4⁺ T cells of untreated and treated recipients are 71.0% (± 4.7%) and 38.5% (± 6.7%), respectively. (C) CD103⁻CD4⁺ T cells from chronic GVHD recipients or normal DBA/2 donors (10⁶ each) were cocultured with CD103⁺ Treg cells from chronic GVHD recipients for 8 hours. Thereafter, the cells were stained for annexin V versus DAPI. The percentage of annexin V⁺DAPI⁺ cells among CD103⁻CD4⁺ T cells is shown beside the gating box. One representative of 4 experiments is shown. The means (± SE) of annexin V⁺DAPI⁺ cell among CD103⁻CD4⁺ T cells from chronic GVHD recipients with or without CD103⁺ Treg cells in coculture are 25.3% (± 3.3%) versus 4.9% (± 1.4%). The means (± SE) of annexin V⁺DAPI⁺ cells among donor CD103⁻CD4⁺ T cells with or without CD103⁺ Treg cells in coculture are 4.7% (± 1.1%) versus 4.2% (± 0.8%).