Blockade of IL-27 signaling augments reconstitution of Foxp3-expressing Tregs. (A) Balb/c mice were transplanted with B6 Foxp3^EGFP BM and spleen cells and then treated with either an isotype control (●) or p28 (□) antibody. Animals were killed 14 days after transplantation and the absolute number of CD4⁺ and CD8⁺ Foxp3⁺ T cells in the spleen, liver, lung, and colon are shown. (B) Sorted CD4⁺ or CD8⁺ Foxp3^EGFP− T cells were cultured with anti-CD3/28 antibodies, IL-2, and TGF-β alone, or together with graded doses of IL-27. The percentage of CD4⁺ and CD8⁺ Foxp3⁺ T cells is depicted. (C) Balb/c mice were transplanted with B6 Rag-1 BM alone or together with sorted CD4⁺ (0.4 × 10⁶) and CD8⁺ (0.2 × 10⁶) Foxp3^EGFP− T cells and then treated with either an isotype control (●) or p28 (□) antibody. Animals were killed 14 days posttransplantation and the absolute number of CD4⁺ and CD8⁺ Foxp3⁺ T cells in spleen, liver, lung, and colon is shown. (D) Balb/c mice were transplanted with B6 Foxp3^GFP BM and spleen cells and then treated with either an isotype control (▪, n = 10), anti-IL-6R (▲, n = 10), p28 (▼, n = 10), or combination of p28 and anti-IL-6R (♦, n = 10) antibodies. The absolute number of CD4⁺ and CD8⁺ Foxp3⁺ T cells in spleen, liver, lung, and colon 2 weeks posttransplantation is depicted. (E) Balb/c mice transplanted with B6 Rag-1 BM alone (●, n = 5) or with 0.5 × 10⁶ CD4⁺EGFP⁻ and 0.3 × 10⁶ CD8⁺EGFP⁻ T cells from Foxp3^ΔEGFP animals that had been reconstituted with 40 × 10⁶ to 60 × 10⁶ spleen cells from B6.SJL Foxp3^EGFP (CD45.1) animals 1 to 2 days after birth to prevent the development of lethal autoimmunity. Mice transplanted with these T cells were treated with either isotype (▪, n = 10) or p28 antibody (□, n = 10) on days 0 and 6 posttransplantation. Overall survival is depicted. Data are from 2 experiments for each panel. *P < .05, **P < .01, ***P < .001. EGFP, enhanced green fluorescent protein; TGF, transforming growth factor.