We show that the lymphoid hyperplasia observed in IL-2Rα- and IL-2-deficient mice is due to the lack of a population of regulatory cells essential for CD4 T cell homeostasis. In chimeras reconstituted with bone marrow cells from IL-2Rα-deficient donors, restitution of a population of CD25+CD4+ T cells prevents the chaotic accumulation of lymphoid cells, and rescues the mice from autoimmune disease and death. The reintroduction of IL-2-producing cells in IL-2-deficient chimeras establishes a population of CD25+CD4+ T cells, and restores the peripheral lymphoid compartments to normal. The CD25+CD4+ T cells regulated selectively the number of naive CD4+ T cells transferred into T cell-deficient hosts. The CD25+CD4+/naive CD4 T cell ratio and the sequence of cell transfer determines the homeostatic plateau of CD4+ T cells. Overall, our findings demonstrate that IL-2Rα is an absolute requirement for the development of the regulatory CD25+CD4+ T cells that control peripheral CD4 T cell homeostasis, while IL-2 is required for establishing a sizeable population of these cells in the peripheral pools.

We recently characterized a CD4+ T cell population expressing the IL-2R alpha chain (CD25), producing IL-10 and resisting clonal deletion induced by viral superantigen (vSAG) encoded by mouse mammary tumor virus [MMTV(SW)]. We now report that these apoptosis-resistant cells are generated in the thymus but not from the immature CD4+ CD8+ thymocytes. They migrate from the thymus and are found in the periphery from at least the 10th day of life, after which they expand with the same kinetics in normal and MMTV(SW)-infected mice. Their strong capacity for expansion in the periphery makes this population insensitive to thymectomy in adulthood. CD4+ CD25+ cells were totally dependent on exogenous IL-2 for growth in vitro and in vivo, and were missing in IL-2 knockout (KO) mice. The absence of this population and/or an inability to produce IL-10 may be the missing link between IL-2R alpha KO, IL-2 KO and IL-10 KO mice, which all die of inflammatory bowel disease.

IntroductionOral tolerance has long been recognized as a physiologic mechanism of immune unresponsiveness to dietary antigens and bacterial microflora antigens, which maintain tissue integrity by preventing harmful delayed-type hypersensitivity (DTH) responses in the intestine and may also limit the efficiency of oral vaccination. Indeed, antigen encounter in the intestine triggers an active inhibitory process preventing the onset of CD4 ϩ and CD8 ϩ T-cell antigen-specific immune responses to subsequent systemic immunization with the same antigen (reviewed in Garside and Mowat 1 ). Several mechanisms have been proposed to explain peripheral tolerance induced by antigen feeding. These include anergy 2,3 or deletion of antigen-specific T cells, 4,5 immune deviation to Th2-biased immune response, and induction of regulatory Th3 (transforming growth factor ␤ [TGF␤]-producing) cells. 6,7 The naturally occurring regulatory subset of CD4 ϩ CD25 ϩ T cells accounting for 5% to 10% of peripheral CD4 ϩ T cells has been extensively reported to exert potent immunosuppressive function in vivo and in vitro toward CD4 ϩ T-cell effectors 8 and may represent regulatory T cells responsible for orally induced peripheral tolerance. Indeed, CD4 ϩ CD25 ϩ T cells, which arise from the thymus as early as day 3 of life, 9 are characterized by a memory phenotype; low proliferative capacity and interleukin-2 (IL-2) production; secretion of high levels of the immunosuppressive cytokines IL-10 and TGF-␤; and expression of cytotoxic Tlymphocyte antigen 4 (CTLA-4), 10-13 a molecule that contributes to orally induced tolerance. 14 These cells have been described in a variety of experimental models to protect from autoimmune diseases, as well as colitis and allograft rejection. 8 Reminiscent to these cells, IL-10-producing ovalbumin (OVA)-specific CD4 ϩ T clones (Tr1) generated in vitro after repeated stimulation with antigen in the presence of IL-10 were shown to prevent colitis when cotransferred with naive CD4 ϩ CD45RB high T cells in OVAfed immunocompromised severe combine immunodeficiency (SCID) or Nude mice. 15 Interestingly, mice genetically deficient for either IL-2, IL-2R, T-cell receptor ␣␤ (TcR␣␤), TGF␤, IL-10, or major histocompatibility complex (MHC) class II were shown to develop spontaneous colitis, [16][17][18][19][20] compatible with a shared physical or functional defect in the regulatory CD4 ϩ CD25 ϩ subset. Although recent studies in TcR transgenic models have reported that oral antigen delivery can induce activation and/or differentiation of regulatory CD4 ϩ CD25 ϩ T cells, 21,22 evidence that they are instrumental for in vivo induction of oral tolerance has not been provided. Moreover, evidence as to whether CD4 ϩ CD25 ϩ cells are responsible for peripheral suppression of antigen-specific CD8 ϩ cytotoxic T-lymphocyte (CTL) responses is still sparse. 23 In this study we examined whether CD4 ϩ CD25 ϩ T cells contribute to oral tolerance in normal nonlymphopenic hosts, using a pathophysiologic model of antigen-specific skin in...

We have previously shown that regulatory CD25+CD4+ T cells are resistant to clonal deletion induced by viral superantigen in vivo. In this work we report that isolated CD25+CD4+ T cells activated in vitro by anti-CD3 Ab are resistant to Fas-induced apoptosis, in contrast to their CD25−CD4+ counterparts. Resistance of CD25+CD4+ T cells to Fas-dependent activation-induced cell death is not linked to their inability to produce IL-2 or to their ability to produce IL-10. The sensitivity of both populations to Fas-induced apoptosis can be modulated in vitro by changing the CD25+CD4+:CD25−CD4+ T cell ratio. The sensitivity of CD25−CD4+ T cells to apoptosis can be reduced, while the sensitivity of CD25+CD4+ T cells can be enhanced. Modulation of Fas-dependent apoptosis is associated with changes in cytokine production. However, while CD25−CD4+ T cell apoptosis is highly dependent on IL-2 (production of which is inhibited by CD25+CD4+ T cells in coculture), modulation of CD25+CD4+ T cell apoptosis is IL-2 independent. Taken together, these results suggest that CD25+CD4+ and CD25−CD4+ T cell sensitivity to Fas-dependent apoptosis is dynamically modulated during immune responses; this modulation appears to help maintain a permanent population of regulatory T cells required to control effector T cells.