Tolerogenic semimature dendritic cells suppress experimental autoimmune thyroiditis by activation of thyroglobulin-specific CD4+CD25+ T cells

Ex vivo treatment of bone marrow-derived dendritic cells (DCs) with TNF-alpha has been previously shown to induce partial maturation of DCs that are able to suppress autoimmunity. In this study, we demonstrate that i.v. administration of TNF-alpha-treated, semimature DCs pulsed with thyrogloblin (Tg), but not with OVA Ag, inhibits the subsequent development of Tg-induced experimental autoimmune thyroiditis (EAT) in CBA/J mice. This protocol activates CD4(+)CD25(+) T cells in vivo, which secrete IL-10 upon specific recognition of Tg in vitro and express regulatory T cell (Treg)-associated markers such as glucocorticoid-induced TNFR, CTLA-4, and Foxp3. These CD4(+)CD25(+) Treg cells suppressed the proliferation and cytokine release of Tg-specific, CD4(+)CD25(-) effector cells in vitro, in an IL-10-independent, cell contact-dependent manner. Prior adoptive transfer of the same CD4(+)CD25(+) Treg cells into CBA/J hosts suppressed Tg-induced EAT. These results demonstrate that the tolerogenic potential of Tg-pulsed, semimature DCs in EAT is likely to be mediated through the selective activation of Tg-specific CD4(+)CD25(+) Treg cells and provide new insights for the study of Ag-specific immunoregulation of autoimmune diseases.     

Oral tolerance induction with antigen conjugated to cholera toxin B subunit generates both Foxp3+CD25+ and Foxp3-CD25- CD4+ regulatory T cells

Oral administration of Ag coupled to cholera toxin B subunit (CTB) efficiently induces peripheral immunological tolerance. We investigated the extent to which this oral tolerance is mediated by CD25+CD4+ regulatory T cells (T(reg)). We found that total T(reg), KJ1-26+ T(reg) and CTLA-4+ T(reg) were all increased in Peyer's patches, mesenteric lymph nodes, and, to a lesser extent, in spleen of mice after intragastric administration of OVA/CTB conjugate, which also increased TGF-beta in serum. This could be abolished by co-administering cholera toxin or by treatment with anti-TGF-beta mAb. CD25+ T(reg), but also CD25-CD4+ T cells from OVA/CTB-treated BALB/c or DO11.10 mice efficiently suppressed effector T cell proliferation and IL-2 production in vitro. Following adoptive transfer, both T cell populations also suppressed OVA-specific T cell and delayed-type hypersensitivity responses in vivo. Foxp3 was strongly expressed by CD25+ T(reg) from OVA/CTB-treated mice, and treatment also markedly expanded CD25+Foxp3+ T(reg). Furthermore, in Rag1(-/-) mice that had adoptively received highly purified Foxp3-CD25-CD4+ OT-II T cells OVA/CTB feeding efficiently induced CD25+ T(reg) cells, which expressed Foxp3 more strongly than naturally developing T(reg) and also had stronger ability to suppress effector OT-II T cell proliferation. A remaining CD25- T cell population, which also became suppressive in response to OVA/CTB treatment, did not express Foxp3. Our results demonstrate that oral tolerance induced by CTB-conjugated Ag is associated with increase in TGF-beta and in both the frequency and suppressive capacity of Foxp3+ and CTLA-4+ CD25+ T(reg) together with the generation of both Foxp3+ and Foxp3-CD25- CD4+ T(reg).     

Experience-driven development: effector/memory-like alphaE+Foxp3+ regulatory T cells originate from both naive T cells and naturally occurring naive-like regulatory T cells

Naturally occurring Foxp3+CD25+CD4+ regulatory T cells (Treg) have initially been described as anergic cells; however, more recent in vivo studies suggest that Tregs vigorously proliferate under both homeostatic as well as inflammatory conditions. We have previously identified a subset of murine CD4+ Tregs, which is characterized by expression of the integrin alphaEbeta7 and which displays an effector/memory-like phenotype indicative of Ag-specific expansion and differentiation. In the present study, the alphaE+ Treg subset was found to contain a large fraction of cycling cells under homeostatic conditions in healthy mice. Using an adoptive transfer system of Ag-specific T cells, we could demonstrate that the vast majority of transferred natural, naive-like CD25+CD4+ Tregs acquired expression of the integrin alphaEbeta7 upon tolerogenic application of Ag via the oral route. In addition, using the same system, Foxp3+ Tregs could be de novo induced from conventional naive CD25-CD4+ T cells, and this conversion was associated with concomitant expression of alphaE. These findings suggest that Tregs expressing the integrin alphaE are effector/memory Tregs with a high turnover rate that can develop in the periphery upon Ag contact under tolerogenic conditions, both from thymic-derived CD25+CD4+ Tregs with a naive-like phenotype as well as from conventional naive T cells.     

Selective expansion of foxp3-positive regulatory T cells and immunosuppression by suppressors of cytokine signaling 3-deficient dendritic cells

Dendritic cells (DCs) induce immunity and immunological tolerance as APCs. It has been shown that DCs secreting IL-10 induce IL-10(+) Tr1-type regulatory T (Treg) cells, whereas Foxp3-positive Treg cells are expanded from naive CD4(+) T cells by coculturing with mature DCs. However, the regulatory mechanism of expansion of Foxp3(+) Treg cells by DCs has not been clarified. In this study, we demonstrated that suppressors of cytokine signaling (SOCS)-3-deficient DCs have a strong potential as Foxp3(+) T cell-inducing tolerogenic DCs. SOCS3(-/-) DCs expressed lower levels of class II MHC, CD40, CD86, and IL-12 than wild-type (WT)-DCs both in vitro and in vivo, and showed constitutive activation of STAT3. Foxp3(-) effector T cells were predominantly expanded by the priming with WT-DCs, whereas Foxp3(+) Treg cells were selectively expanded by SOCS3(-/-) DCs. Adoptive transfer of SOCS3(-/-) DCs reduced the severity of experimental autoimmune encephalomyelitis. Foxp3(+) T cell expansion was blocked by anti-TGF-beta Ab, and SOCS3(-/-) DCs produced higher levels of TGF-beta than WT-DCs, suggesting that TGF-beta plays an essential role in the expansion of Foxp3(+) Treg cells. These results indicate an important role of SOCS3 in determining on immunity or tolerance by DCs.     

Dendritic cells genetically engineered to express IL-10 induce long-lasting antigen-specific tolerance in experimental asthma

Dendritic cells (DCs) are professional APCs that have a unique capacity to initiate primary immune responses, including tolerogenic responses. We have genetically engineered bone marrow-derived DCs to express the immunosuppressive cytokine IL-10 and tested the ability of these cells to control experimental asthma. A single intratracheal injection of OVA-pulsed IL-10-transduced DCs (OVA-IL-10-DCs) to naive mice before OVA sensitization and challenge prevented all of the cardinal features of airway allergy, namely, eosinophilic airway inflammation, airway hyperreactivity, and production of mucus, Ag-specific Igs, and IL-4. OVA-IL-10-DCs also reversed established experimental asthma and had long-lasting and Ag-specific effects. We furthermore showed, by using IL-10-deficient mice, that host IL-10 is required for mediating the immunomodulatory effects of OVA-IL-10-DCs and demonstrated a significant increase in the percentage of OVA-specific CD4(+)CD25(+)Foxp3(+)IL-10(+) regulatory T cells in the mediastinal lymph nodes of OVA-IL-10-DC-injected mice. Finally, adoptive transfer of CD4(+) mediastinal lymph node T cells from mice injected with OVA-IL-10-DCs protected OVA-sensitized recipients from airway eosinophilia upon OVA provocation. Our study describes a promising strategy to induce long-lasting Ag-specific tolerance in airway allergy.     

Rapamycin-conditioned dendritic cells are poor stimulators of allogeneic CD4+ T cells, but enrich for antigen-specific Foxp3+ T regulatory cells and promote organ transplant tolerance

The ability of dendritic cells (DC) to regulate Ag-specific immune responses via their influence on T regulatory cells (Treg) may be key to their potential as therapeutic tools or targets for the promotion/restoration of tolerance. In this report, we describe the ability of maturation-resistant, rapamycin (RAPA)-conditioned DC, which are markedly impaired in Foxp3(-) T cell allostimulatory capacity, to favor the stimulation of murine alloantigen-specific CD4(+)CD25(+)Foxp3(+) Treg. This was distinct from control DC, especially following CD40 ligation, which potently expanded non-Treg. RAPA-DC-stimulated Treg were superior alloantigen-specific suppressors of T effector responses compared with those stimulated by control DC. Supporting the ability of RAPA to target effector T and B cells, but permit the proliferation and suppressive function of Treg, an infusion of recipient-derived alloantigen-pulsed RAPA-DC followed by a short postoperative course of low-dose RAPA promoted indefinite (>100 day) heart graft survival. This was associated with graft infiltration by CD4(+)Foxp3(+) Treg and the absence of transplant vasculopathy. The adoptive transfer of CD4(+) T cells from animals with long-surviving grafts conferred resistance to rejection. These novel findings demonstrate that, whereas maturation resistance does not impair the capacity of RAPA-DC to modulate Treg, it profoundly impairs their ability to expand T effector cells. A demonstration of this mechanism endorses their potential as tolerance-promoting cellular vaccines.     

Cutting edge: in vitro-generated tolerogenic APC induce CD8+ T regulatory cells that can suppress ongoing experimental autoimmune encephalomyelitis

APC exposed to TGFbeta2 and Ag (tolerogenic APC) promote peripheral Ag-specific tolerance via the induction of CD8(+) T regulatory cells capable of suppressing Th1 and Th2 immunity. We postulated that tolerogenic APC might reinstate tolerance toward self-neuronal Ags and ameliorate ongoing experimental autoimmune encephalomyelitis (EAE). Seven days after immunization with myelin basic protein (MBP), mice received MBP-specific tolerogenic APC, and EAE was evaluated clinically. To test for the presence and the phenotype of T regulatory cells, CD4 and/or CD8 T cells from tolerogenic APC-treated mice were transferred to naive mice before their immunization with MBP. The MBP-specific tolerogenic APC decreased both the severity and incidence of ongoing EAE. Tolerance to self-neuronal Ags was induced in naive recipient mice via adoptive transfer of CD8(+), but not CD4(+) T cells. Rational use of in vitro-generated tolerogenic APC may lead to novel therapy for autoimmune disease.     

Generation of antigen-specific, Foxp3-expressing CD4+ regulatory T cells by inhibition of APC proteosome function

We tested the hypothesis that immature APC, whose NF-kappaB-signaling pathway and thus maturation was blocked by the proteosome inhibitor benzyloxycarbonyl-isoleucyl-glutamyl(O-tert-butyl)-alanyl-leucinal (PSI), could be a source of Ag-specific regulatory T (Treg) cells. DO11.10 CD4(+) T cells that were incubated with Ag- and PSI-pulsed APC proliferated poorly, produced less IL-2, IFN-gamma, and IL-10 in secondary cultures, and inhibited the response of both naive and memory CD4(+) T cells stimulated by Ag-pulsed APC. The generation of PSI-APC Treg cells required IL-10 production by APC. PSI-APC Treg cell inhibition required cell-cell contact but not IL-10 or TGF-beta. Addition of IL-2 did not reverse, but Ab to CTLA-4 did reverse partially the inhibitory effect. Depletion of CD25(+) T cells before initial culture with PSI-APC did not affect Treg generation. PSI-APC Treg cells expressed high levels of Foxp3, inhibited proliferation of naive DO11.10 T cells in vivo, and abrogated colitis driven by a memory Th1 response to bacterial-associated Ag. We conclude that NF-kappaB-blocked, immature APC are able to induce the differentiation of Treg cells that can function in vitro and in vivo in an Ag-specific manner.     

Expression of dual TCR on DO11.10 T cells allows for ovalbumin-induced oral tolerance to prevent T cell-mediated colitis directed against unrelated enteric bacterial antigens

The triggering Ag for inflammatory bowel disease and animal models of colitis is not known, but may include gut flora. Feeding OVA to DO11.10 mice with OVA-specific transgenic (Tg) TCR generates Ag-specific immunoregulatory CD4(+) T cells (Treg) cells. We examined the ability of oral Ag-induced Treg cells to suppress T cell-mediated colitis in mice. SCID-bg mice given DO11.10 CD4(+)CD45RB(high) T cells developed colitis, and cotransferring DO11.10 CD45RB(low)CD4(+) T cells prevented CD4(+)CD45RB(high) T cell-induced colitis in the absence of OVA. The induction and prevention of disease by DO11.10 CD4(+) T cell subsets were associated with an increase in endogenous TCRalpha chain expression on Tg T cells. Feeding OVA to SCID-bg mice reconstituted with DO11.10 CD4(+)CD45RB(high) attenuated the colitis in association with increased TGF-beta and IL-10 secretion, and decreased proliferative responses to both OVA and cecal bacteria Ag. OVA feeding also attenuated colitis in SCID-bg mice reconstituted with a mix of BALB/c and DO11.10 CD45RB(high) T cells, suggesting that OVA-induced Treg cells suppressed BALB/c effector cells. The expression of endogenous non-Tg TCR allowed for DO11.10-derived T cells to respond to enteric flora Ag. Furthermore, feeding OVA-induced Treg cells prevented colitis by inducing tolerance in both OVA-reactive and non-OVA-reactive T cells and by inducing Ag-nonspecific Treg cells. Such a mechanism might allow for Ag-nonspecific modulation of intestinal inflammation in inflammatory bowel disease.     

CpG-DNA aided cross-priming by cross-presenting B cells

Covalent linkage of immunostimulatory CpG-DNA to OVA (CpG-OVA complex) results in CpG-DNA-aided cross-presentation of OVA by dendritic cells (DCs). In this study, we analyzed the thesis that CpG-OVA complexes may be cross-presented by B cells to route internalized Ag into the class I MHC presentation pathway. First, we describe that conjugation of CpG-DNA to OVA enhances up to 40-fold internalization of OVA by B cells, which in turn generate the CD8 T cell epitope SIINFEKL complexed to MHC class I, albeit less efficiently than DCs. Furthermore, upon internalization, CpG-DNA conjugated to OVA stimulates B cells to up-regulate costimulatory molecules and cytokines including IL-12. Adoptive transfer of CpG-OVA complex-loaded wild-type B cells cross-primes naive CD8 T cells both in wild-type mice and in MyD88-deficient mice. Overall, these findings disclose attributes of B cells, including cross-presentation of exogenous Ag and cross-priming of naive CD8 T cells that hitherto have been considered as hallmarks restricted to DCs.     

Rapid and selective expansion of nonclonotypic T cells in regulatory T cell-deficient, foreign antigen-specific TCR-transgenic scurfy mice: antigen-dependent expansion and TCR analysis

Foreign Ag-specific TCR-transgenic (Tg) mice contain a small fraction of T cells bearing the endogenous Vbeta and Valpha chains as well as a population expressing an intermediate level of Tg TCR. Importantly, these minor nonclonotypic populations contain > or = 99% of the CD4(+)Foxp3(+) regulatory T cells (Treg) and, despite low overall Treg expression, peripheral tolerance is maintained. In the OT-II TCR (OVA-specific, Vbeta5(high)Valpha2(high)) Tg scurfy (Sf) mice (OT-II Sf) that lack Treg, nonclonotypic T cells markedly expanded in the periphery but not in the thymus. Expanded T cells expressed memory/effector phenotype and were enriched in blood and inflamed lungs. In contrast, Vbeta5(high)Valpha2(high) clonotypic T cells were not expanded, displayed the naive phenotype, and found mainly in the lymph nodes. Importantly, Vbeta5(neg) T cells were able to transfer multiorgan inflammation in Rag1(-/-) recipients. T cells bearing dual TCR (dual Vbeta or dual Valpha) were demonstrated frequently in the Vbeta5(int) and Valpha2(int) populations. Our study demonstrated that in the absence of Treg, the lack of peripheral expansion of clonotypic T cells is due to the absence of its high-affinity Ag OVA. Thus, the rapid expansion of nonclonotypic T cells in OT-II Sf mice must require Ag (self and foreign) with sufficient affinity. Our study has implications with respect to the roles of Ag and dual TCR in the selection and regulation of Treg and Treg-controlled Ag-dependent T cell expansion in TCR Tg and TCR Tg Sf mice, respectively.     

Regulatory role of B cells in a murine model of allergic airway disease

Mice sensitized to OVA and subjected to acute OVA aerosol exposures develop allergic airway disease (AAD). However, chronic continuous Ag exposure results in resolution of AAD and the development of local inhalational tolerance (LIT). Because we have previously observed the persistence of B cells in the bronchoalveolar lavage (BAL) and hilar lymph nodes (HLN) at the resolution stage of this model, we investigated the role of B cells in the modulation of AAD. Although B cell-deficient mice developed LIT, adoptive transfer of HLN B cells from LIT mice to OVA-sensitized recipients resulted in attenuated AAD following subsequent OVA aerosol exposure, as determined by reduced BAL leukocytosis and eosinophilia, decreased tissue inflammation, and absent methacholine hyper-responsiveness. In similar adoptive transfer studies, HLN B cells from AAD mice were without effect. The protection transferred by LIT HLN B cells was Ag specific and was associated with accumulation of Foxp3(+) T regulatory cells regionally in BAL and HLN, but not systemically in the spleen. Fluorescent labeling of LIT HLN B cells before adoptive transfer demonstrated that these cells had the capacity to migrate to local inflammatory sites. In vitro assessment demonstrated that the LIT HLN B cells exerted this regulatory effect via TGF-beta induced conversion of CD4(+)CD25(-) T effector cells into functionally suppressive CD4(+)CD25(+)Foxp3(+) T regulatory cells. These findings illustrated a novel regulatory role for regional B cells in AAD and suggested a possible contributory role of B cells, along with other cell types, in the establishment of LIT.     

Antigen targeting to plasmacytoid dendritic cells via Siglec-H inhibits Th cell-dependent autoimmunity

Plasmacytoid dendritic cells (PDCs) have been shown to present Ags and to contribute to peripheral immune tolerance and to Ag-specific adaptive immunity. However, modulation of adaptive immune responses by selective Ag targeting to PDCs with the aim of preventing autoimmunity has not been investigated. In the current study, we demonstrate that in vivo Ag delivery to murine PDCs via the specifically expressed surface molecule sialic acid binding Ig-like lectin H (Siglec-H) inhibits Th cell and Ab responses in the presence of strong immune stimulation in an Ag-specific manner. Correlating with sustained low-level MHC class II-restricted Ag presentation on PDCs, Siglec-H-mediated Ag delivery induced a hyporesponsive state in CD4(+) T cells leading to reduced expansion and Th1/Th17 cell polarization without conversion to Foxp3(+) regulatory T cells or deviation to Th2 or Tr1 cells. Siglec-H-mediated delivery of a T cell epitope derived from the autoantigen myelin oligodendrocyte glycoprotein to PDCs effectively delayed onset and reduced disease severity in myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis by interfering with the priming phase without promoting the generation or expansion of myelin oligodendrocyte glycoprotein-specific Foxp3(+) regulatory T cells. We conclude that Ag delivery to PDCs can be harnessed to inhibit Ag-specific immune responses and prevent Th cell-dependent autoimmunity.     

Transforming growth factor-beta enhances the in vivo effector function and memory phenotype of antigen-specific T helper cells in experimental autoimmune encephalomyelitis.

Transforming growth factor-beta (TGF-beta) had a profound effect on the in vitro phenotypic development of Ag-activated Th cells and enhanced the in vivo effector function of these cells upon adoptive transfer. Previous studies have shown that there are two types of Th cell populations found in unimmunized animals, naive helper cells, which are short-lived and express low levels of CD44 and high levels of CD45R and Mel-14, and memory helper cells, which have a long life span and express high levels of CD44 and low levels of CD45R and Mel-14. Culturing of Ag-specific murine Th cell lines and clones in the presence of TGF-beta greatly enhanced both the memory phenotype of the cultured cells and the effector function upon adoptive transfer in experimental autoimmune encephalomyelitis. Histologic evaluation of spinal cords from recipients receiving passively transferred murine T cell lines cultured with TGF-beta revealed large demyelinated plaques (multiple sclerosis-like) that were not present in animals receiving cells cultured with Ag alone. TGF-beta also enhanced the capability of myelin basic protein-specific Lewis rat T cell lines to transfer experimental autoimmune encephalomyelitis and potentiated a purified protein derivative-specific rat helper cell line to transfer delayed type hypersensitivity. Thus, the effects of TGF-beta did not appear to be limited by species specificity, Ag specificity, or in vivo T cell function. This is the first study showing that TGF-beta can potentiate the development and maintenance of the Th cell memory phenotype in vitro and enhance their in vivo effector function in an animal disease model.     

Intestinal epithelial antigen induces mucosal CD8 T cell tolerance, activation, and inflammatory response

Intestinal autoimmune diseases are thought to be associated with a breakdown in tolerance, leading to mucosal lymphocyte activation perhaps as a result of encounter with bacterium-derived Ag. To study mucosal CD8(+) T cell activation, tolerance, and polarization of autoimmune reactivity to self-Ag, we developed a novel (Fabpl(4x at -132)-OVA) transgenic mouse model expressing a truncated form of OVA in intestinal epithelia of the terminal ileum and colon. We found that OVA-specific CD8(+) T cells were partially tolerant to intestinal epithelium-derived OVA, because oral infection with Listeria monocytogenes-encoding OVA did not elicit an endogenous OVA-specific MHC class I tetramer(+)CD8(+) T cell response and IFN-gamma-, IL-4-, and IL-5-secreting T cells were decreased in the Peyer's patches, mesenteric lymph nodes, and intestinal mucosa of transgenic mice. Adoptive transfer of OVA-specific CD8(+) (OT-I) T cells resulted in their preferential expansion in the Peyer's patches and mesenteric lymph nodes and subsequently in the epithelia and lamina propria but failed to cause mucosal inflammation. Thus, CFSE-labeled OT-I cells greatly proliferated in these tissues by 5 days posttransfer. Strikingly, OT-I cell-transferred Fabpl(4x at -132)-OVA transgenic mice underwent a transient weight loss and developed a CD8(+) T cell-mediated acute enterocolitis 5 days after oral L. monocytogenes-encoding OVA infection. These findings indicate that intestinal epithelium-derived "self-Ag" gains access to the mucosal immune system, leading to Ag-specific T cell activation and clonal deletion. However, when Ag is presented in the context of bacterial infection, the associated inflammatory signals drive Ag-specific CD8(+) T cells to mediate intestinal immunopathology.     

Antigen-driven interactions with dendritic cells and expansion of Foxp3+ regulatory T cells occur in the absence of inflammatory signals

Foxp3+ regulatory T cells (Tregs) play a pivotal role in the maintenance of peripheral T cell tolerance and are thought to interact with dendritic cells (DC) in secondary lymphoid organs. We analyzed here the in vivo requirements for selective expansion of Ag-specific Treg vs CD4+CD25- effector T cells and engagement of Ag-specific Treg-DC interactions in secondary lymphoid organs. Using i.v. Ag delivery in the absence of inflammation, we found that CD4+CD25+Foxp3+ Tregs undergo vigorous expansion and accumulate whereas naive CD4+CD25-Foxp3- T cells undergo abortive activation. Quantifying directly the interactions between Tregs and CD11c+ DC, we found that Tregs establish cognate contacts with endogenous CD11c+ DC in spleen and lymph nodes at an early time point preceding their expansion. Importantly, we observed that as few as 10(3) Tregs selectively expanded by i.v. Ag injection are able to suppress B and T cell immune responses in mouse recipients challenged with the Ag. Our results demonstrate that Tregs are selectively mobilized by Ag recognition in the absence of inflammatory signals, and can induce thereafter potent tolerance to defined Ag targets.     

CD8+ CD205+ splenic dendritic cells are specialized to induce Foxp3+ regulatory T cells

Foxp3(+)CD25(+)CD4(+) regulatory T cells (Treg) mediate immunological self-tolerance and suppress immune responses. A subset of dendritic cells (DCs) in the intestine is specialized to induce Treg in a TGF-beta- and retinoic acid-dependent manner to allow for oral tolerance. In this study we compare two major DC subsets from mouse spleen. We find that CD8(+) DEC-205/CD205(+) DCs, but not the major fraction of CD8(-) DC inhibitory receptor-2 (DCIR2)(+) DCs, induce functional Foxp3(+) Treg from Foxp3(-) precursors in the presence of low doses of Ag but without added TGF-beta. CD8(+)CD205(+) DCs preferentially express TGF-beta, and the induction of Treg by these DCs in vitro is blocked by neutralizing Ab to TGF-beta. In contrast, CD8(-)DCIR2(+) DCs better induce Foxp3(+) Treg when exogenous TGF-beta is supplied. In vivo, CD8(+)CD205(+) DCs likewise preferentially induce Treg from adoptively transferred, Ag-specific DO11.10 RAG(-/-) Foxp3(-)CD4(+) T cells, whereas the CD8(-)DCIR2(+) DCs better stimulate natural Foxp3(+) Treg. These results indicate that a subset of DCs in spleen, a systemic lymphoid organ, is specialized to differentiate peripheral Foxp3(+) Treg, in part through the endogenous formation of TGF-beta. Targeting of Ag to these DCs might be useful for inducing Ag-specific Foxp3(+) Treg for treatment of autoimmune diseases, transplant rejection, and allergy.     

Immunodominance in the TCR repertoire of a [corrected] TCR peptide-specific CD4+ Treg population that controls experimental autoimmune encephalomyelitis

Immunodominance in self-Ag-reactive pathogenic CD4(+) T cells has been well established in several experimental models. Although it is clear that regulatory lymphocytes (Treg) play a crucial role in the control of autoreactive cells, it is still not clear whether immunodominant CD4(+) Treg clones are also involved in control of autoreactivity. We have shown that TCR-peptide-reactive CD4(+) and CD8(+) Treg play an important role in the spontaneous recovery and resistance from reinduction of experimental autoimmune encephalomyelitis in B10.PL mice. We report, by sequencing of the TCR alpha- and beta-chain associated with CD4(+) Treg, that the TCR repertoire is limited and the majority of CD4(+) Treg use the TCR Vbeta14 and Valpha4 gene segments. Interestingly, sequencing and spectratyping data of cloned and polyclonal Treg populations revealed that a dominant public CD4(+) Treg clonotype expressing Vbeta14-Jbeta1.2 with a CDR3 length of 7 aa exists in the naive peripheral repertoire and is expanded during the course of recovery from experimental autoimmune encephalomyelitis. Furthermore, a higher frequency of CD4(+) Treg clones in the naive repertoire correlates with less severity and more rapid spontaneous recovery from disease in parental B10.PL or PL/J and (B10.PL x PL/J)F(1) mice. These findings suggest that unlike the Ag-nonspecific, diverse TCR repertoire among the CD25(+)CD4(+) Treg population, TCR-peptide-reactive CD4(+) Treg involved in negative feedback regulation of autoimmunity use a highly limited TCR V-gene repertoire. Thus, a selective set of immunodominant Treg as well as pathogenic T cell clones can be targeted for potential intervention in autoimmune disease conditions.     

Foxp3+ regulatory T cells impede the priming of protective CD8+ T cells

T cell activation is controlled by incompletely defined opposing stimulation and suppression signals that together sustain the balance between optimal host defense against infection and peripheral tolerance. In this article, we explore the impacts of Foxp3(+) regulatory T cell (Treg) suppression in priming Ag-specific T cell activation under conditions of noninfection and infection. We find the transient ablation of Foxp3(+) Tregs unleashes the robust expansion and activation of peptide-stimulated CD8(+) T cells that provide protection against Listeria monocytogenes infection in an Ag-specific fashion. By contrast, Treg ablation had nonsignificant impacts on the CD8(+) T cell response primed by infection with recombinant L. monocytogenes. Similarly, nonrecombinant L. monocytogenes administered with peptide stimulated the expansion and activation of CD8(+) T cells that paralleled the response primed by Treg ablation. Interestingly, these adjuvant properties of L. monocytogenes did not require CD8(+) T cell stimulation by IL-12 produced in response to infection, but instead were associated with sharp reductions in Foxp3(+) Treg suppressive potency. Therefore, Foxp3(+) Tregs impose critical barriers that, when overcome naturally during infection or artificially with ablation, allow the priming of protective Ag-specific CD8(+) T cells.