We show here that the anti-T lymphocyte immunoglobulin (ATG) can induce Treg cells following 24-h incubation in human peripheral blood mononuclear cells (PBMCs). The ATG-induced Treg cells express known cell surface markers (e.g., CD25, FoxP3) and suppress the proliferation of autologous responder PBMCs, stimulated with allogeneic PBMCs, when added into the mixed lymphocyte culture (MLC) at zero time point or 48 h later. We expanded the characteristics of the ATG-induced human Treg cells by showing that they express a novel biomarker designated "activated CD44". ATG-induced Treg cells retain their suppressor function after freezing and thawing or irradiation. Suppression of MLC by ATG-induced Treg cells is consistently seen when the Treg cells and the responder cells were derived from the same donor, but not when they derived from different donors. Finally, patients undergoing stem cell transplantation and conditioned with ATG generate in vivo Treg cells that suppress MLC.

This study is the first to show that PBMCs of patients with GO substantially increase Treg cells in both frequency and potency after in vitro incubation with rATG.