Supplementary MaterialsSupplementary File. affinity to respond overtly to host DCs: bidirectional T/DC interaction ensues, leading to progressive DC activation and reciprocal strong proliferation of T cells accompanied by peripheral Treg (pTreg) formation. Likewise, high-affinity CD4 T cells proliferate vigorously and form pTregs when cultured with autologous DCs in vitro in the absence of nTregs: this anti-self response is MHCII/peptide dependent and elicited by the raised level EP of B7 on cultured DCs. The data support a model in which self tolerance is imposed via modulation of CD28 signaling and explains the pathological effects of superagonistic CD28 antibodies. Tolerance to self components involves a combination of intrathymic deletion (negative selection) of T cells with overt self reactivity and suppression by a subset of CD4 T regulatory cells (Tregs) expressing the transcription factor Foxp3 (1, 2). Absence or mutation of causes a lethal syndrome of uncontrolled T cell proliferation and lymphadenopathy, as seen in scurfy mice and diphtheria toxin (DT)-treated Foxp3-DTR mice; in humans, mutation of leads to immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (3). Tregs suppress the activation and effector function of conventional CD4 and CD8 T cells through release of inhibitory cytokines, such as IL-10 and TGF, and by regulating costimulatory molecule expression on dendritic cells (DCs) (4, 5). Typical Tregs are generated in the thymus [natural Tregs (nTregs)] through recognition of MHC II/self peptide ligands in the presence of IL-2 and display strong suppressive function for responses of normal T cells (6). However, optimal suppression requires an additional population of Foxp3+ Tregs generated from conventional CD4 T cells in the peripheral lymphoid tissues (7). Most peripherally induced Tregs (pTregs) are induced in the lamina propria of the small and large intestine through recognition of dietary and commensal microbial antigens in the presence of TGF and retinoic acid synthesized by mucosal DCs (8C10), while some pTregs may be generated by tolerogenic DCs in lymph nodes (LNs) draining the skin (11). Collectively, these findings imply that the primary function of pTregs is to suppress immune responses to microbial antigens, whereas effective self tolerance may require Phen-DC3 the combined action of nTregs and pTregs (7). The stimulus for the onset of T cell proliferation in the absence of Tregs is unclear. Uncontrolled responses to commensal microbiota could be involved, but this possibility is unlikely because lymphoproliferative disease still occurs in DT-treated Foxp3-DTR mice maintained in a germ-free (GF) environment (12). This finding does not rule out a response to food antigens. However, it does raise the possibility that lymphoproliferation in the absence of Tregs could be directed largely to Phen-DC3 self antigens. Although direct evidence on this notion is sparse, culturing T cells with autologous antigen-presenting cells (APCs) in vitro leads to low-level proliferation of naive CD4 T cells; this phenomenon is termed the auto-mixed lymphocyte reaction (auto-MLR) and represents the background response for T cell responses to allogeneic APCs (13C15). This response is enhanced in the absence of Tregs (14) and associated with APC activation and Phen-DC3 up-regulation of costimulatory molecules (16), implying a dysregulated response to self antigens. Under in vivo conditions, proliferation of CD4 T cells in syngeneic irradiated hosts is weak (17) and is largely a reflection of slow MHC-dependent homeostatic proliferation induced by the elevated levels of IL-7 in lymphopenic hosts (18, 19). Far stronger proliferation occurs when naive CD4 T cells are transferred to syngeneic T cell-deficient SCID or hosts (20, 21). Such fast T cell proliferation is more intense in specific-pathogenCfree (SPF) than GF hosts, implying that much of the proliferation is directed to commensal microbiota (20). Nevertheless, even in GF hosts, a proportion of donor CD4 T cells does undergo rapid proliferation. In SPF hosts, levels of B7 (CD80, CD86) on DCs are higher than in normal mice and.