Characterization of the phenotype and function of Tregs. (A) CD4⁺CD25^hi cells from either wild-type FVB or luciferase-transgenic FVB mice were sorted to purities of more than 99%, with more than 96% expressing Foxp3 on FACS analysis. (B) By quantitative RT-PCR, sorted CD4⁺CD25^hi cells had significantly greater Foxp3 expression compared with CD4⁺CD25⁻ cells (P = .03). Error bars represent standard error. (C) Histopathologic findings in the colon of lethally-irradiated recipients 8 days after transplantation of T-cell–depleted bone marrow alone (i), with Tcons (physiologic ratio of CD4⁺ and CD8⁺ T cells) (ii), or with Tcons and CD4⁺CD25^hi (iii). Goblet-cell (thin black arrow) depletion, lymphocytic infiltration (thick black arrow), and mucosal disruption in colon of animals with Tcons but without CD4⁺CD25^hi. (D) Lethal GvHD induced by FVB Tcons (▪) in Balb/c recipients was inhibited with the cotransfer of FVB Tregs (○) at 1:1 dose ratio (n = 4; P = .007). Recipients receiving only TCD-BMs (▵; n = 4) survived without evidence of GvHD. Data are representative of 3 or more experiments.