Dendritic cell-expanded, islet-specific CD4+ CD25+ CD62L+ regulatory T cells restore normoglycemia in diabetic NOD mice

Most treatments that prevent autoimmune diabetes in nonobese diabetic (NOD) mice require intervention at early pathogenic stages, when insulitis is first developing. We tested whether dendritic cell (DC)-expanded, islet antigen-specific CD4+ CD25+ suppressor T cells could treat diabetes at later stages of disease, when most of the insulin-producing islet β cells had been destroyed by infiltrating lymphocytes. CD4+ CD25+ CD62L+ regulatory T cells (T reg cells) from BDC2.5 T cell receptor transgenic mice were expanded with antigen-pulsed DCs and IL-2, and were then injected into NOD mice. A single dose of as few as 5 × 104 of these islet-specific T reg cells blocked diabetes development in prediabetic 13-wk-old NOD mice. The T reg cells also induced long-lasting reversal of hyperglycemia in 50% of mice in which overt diabetes had developed. Successfully treated diabetic mice had similar responses to glucose challenge compared with nondiabetic NOD mice. The successfully treated mice retained diabetogenic T cells, but also had substantially increased Foxp3+ cells in draining pancreatic lymph nodes. However, these Foxp3+ cells were derived from the recipient mice and not the injected T reg cells, suggesting a role for endogenous T reg cells in maintaining tolerance after treatment. Therefore, inoculation of DC-expanded, antigen-specific suppressor T cells has considerable efficacy in ameliorating ongoing diabetes in NOD mice

CD25+ CD4+ T cells, expanded with dendritic cells presenting a single autoantigenic peptide, suppress autoimmune diabetes

In the nonobese diabetic (NOD) mouse model of type 1 diabetes, the immune system recognizes many autoantigens expressed in pancreatic islet β cells. To silence autoimmunity, we used dendritic cells (DCs) from NOD mice to expand CD25+ CD4+ suppressor T cells from BDC2.5 mice, which are specific for a single islet autoantigen. The expanded T cells were more suppressive in vitro than their freshly isolated counterparts, indicating that DCs from autoimmune mice can increase the number and function of antigen-specific, CD25+ CD4+ regulatory T cells. Importantly, only 5,000 expanded CD25+ CD4+ BDC2.5 T cells could block autoimmunity caused by diabetogenic T cells in NOD mice, whereas 105 polyclonal, CD25+ CD4+ T cells from NOD mice were inactive. When islets were examined in treated mice, insulitis development was blocked at early (3 wk) but not later (11 wk) time points. The expanded CD25+ CD4+ BDC2.5 T cells were effective even if administered 14 d after the diabetogenic T cells. Our data indicate that DCs can generate CD25+ CD4+ T cells that suppress autoimmune disease in vivo. This might be harnessed as a new avenue for immunotherapy, especially because CD25+ CD4+ regulatory cells responsive to a single autoantigen can inhibit diabetes mediated by reactivity to multiple antigens

Direct expansion of functional CD25+ CD4+ regulatory T cells by antigen-processing dendritic cells

An important pathway for immune tolerance is provided by thymic-derived CD25+ CD4+ T cells that suppress other CD25- autoimmune disease-inducing T cells. The antigen-presenting cell (APC) requirements for the control of CD25+ CD4+ suppressor T cells remain to be identified, hampering their study in experimental and clinical situations. CD25+ CD4+ T cells are classically anergic, unable to proliferate in response to mitogenic antibodies to the T cell receptor complex. We now find that CD25+ CD4+ T cells can proliferate in the absence of added cytokines in culture and in vivo when stimulated by antigen-loaded dendritic cells (DCs), especially mature DCs. With high doses of DCs in culture, CD25+ CD4+ and CD25- CD4+ populations initially proliferate to a comparable extent. With current methods, one third of the antigen-reactive T cell receptor transgenic T cells enter into cycle for an average of three divisions in 3 d. The expansion of CD25+ CD4+ T cells stops by day 5, in the absence or presence of exogenous interleukin (IL)-2, whereas CD25- CD4+ T cells continue to grow. CD25 + CD4+ T cell growth requires DC-T cell contact and is partially dependent upon the production of small amounts of IL-2 by the T cells and B7 costimulation by the DCs. After antigen-specific expansion, the CD25 + CD4+ T cells retain their known surface features and actively suppress CD25- CD4+ T cell proliferation to splenic APCs. DCs also can expand CD25+ CD4+ T cells in the absence of specific antigen but in the presence of exogenous IL-2. In vivo, both steady state and mature antigen-processing DCs induce proliferation of adoptively transferred CD25+ CD4+ T cells. The capacity to expand CD25+ CD4+ T cells provides DCs with an additional mechanism to regulate autoimmunity and other immune responses

Beyond cDC1: Emerging Roles of DC Crosstalk in Cancer Immunity

Dendritic cells (DCs) efficiently process and present antigens to T cells, and by integrating environmental signals, link innate and adaptive immunity. DCs also control the balance between tolerance and immunity, and are required for T-cell mediated anti-tumor immunity. One subset of classical DCs, cDC1, are particularly important for eliciting CD8 T cells that can kill tumor cells. cDC1s are superior in antigen cross-presentation, a process of presenting exogenous antigens on MHC class I to activate CD8+ T cells. Tumor-associated cDC1s can transport tumor antigen to the draining lymph node and cross-present tumor antigens, resulting in priming and activation of cytotoxic T cells. Although cross-presenting cDC1s are critical for eliciting anti-tumor T cell responses, the role and importance of other DC subsets in anti-tumor immunity is not as well-characterized. Recent literature in other contexts suggests that critical crosstalk between DC subsets can significantly alter biological outcomes, and these DC interactions likely also contribute significantly to tumor-specific immune responses. Therefore, antigen presentation by cDC1s may be necessary but not sufficient for maximal immune responses against cancer. Here, we discuss recent advances in the understanding of DC subset interactions to maximize anti-tumor immunity, and propose that such interactions should be considered for the development of better DC-targeted immunotherapies

CD25+ CD4+ T Cells, Expanded with Dendritic Cells Presenting a Single Autoantigenic Peptide, Suppress Autoimmune Diabetes

In the nonobese diabetic (NOD) mouse model of type 1 diabetes, the immune system recognizes many autoantigens expressed in pancreatic islet β cells. To silence autoimmunity, we used dendritic cells (DCs) from NOD mice to expand CD25+ CD4+ suppressor T cells from BDC2.5 mice, which are specific for a single islet autoantigen. The expanded T cells were more suppressive in vitro than their freshly isolated counterparts, indicating that DCs from autoimmune mice can increase the number and function of antigen-specific, CD25+ CD4+ regulatory T cells. Importantly, only 5,000 expanded CD25+ CD4+ BDC2.5 T cells could block autoimmunity caused by diabetogenic T cells in NOD mice, whereas 105 polyclonal, CD25+ CD4+ T cells from NOD mice were inactive. When islets were examined in treated mice, insulitis development was blocked at early (3 wk) but not later (11 wk) time points. The expanded CD25+ CD4+ BDC2.5 T cells were effective even if administered 14 d after the diabetogenic T cells. Our data indicate that DCs can generate CD25+ CD4+ T cells that suppress autoimmune disease in vivo. This might be harnessed as a new avenue for immunotherapy, especially because CD25+ CD4+ regulatory cells responsive to a single autoantigen can inhibit diabetes mediated by reactivity to multiple antigens

Loss of Zbtb32 in NOD mice does not significantly alter T cell responses. [version 2; referees: 2 approved]

Background: We previously identified the transcriptional regulator Zbtb32 as a factor that can promote T cell tolerance in the Non-Obese Diabetic (NOD) mouse, a model of Type 1 diabetes. Antigen targeted to DCIR2+ dendritic cells (DCs) in vivo inhibited both diabetes and effector T cell expansion in NOD mice. Furthermore, Zbtb32 was preferentially induced in autoreactive CD4 T cells stimulated by these tolerogenic DCIR2+ DCs, and overexpression of Zbtb32 in islet-specific T cells inhibited the diabetes development by limiting T cell proliferation and cytokine production. Methods: To further understand the role of Zbtb32 in T cell tolerance induction, we have now used CRISPR to target the Zbtb32 gene for deletion directly in NOD mice and characterized the mutant mice. We hypothesized that the systemic loss of Zbtb32 in NOD mice would lead to increased T cell activation and increased diabetes pathogenesis. Results: Although NOD.Zbtb32-/- male NOD mice showed a trend towards increased diabetes incidence compared to littermate controls, the difference was not significant. Furthermore, no significant alteration in lymphocyte number or function was observed. Importantly, in vitro stimulation of lymphocytes from NOD.Zbtb32-/- mice did not produce the expected hypersensitive phenotype observed in other genetic strains, potentially due to compensation by homologous genes. Conclusions: The loss of Zbtb32 in the NOD background does not result in the expected T cell activation phenotype

Dendritic cells with TGF-β1 differentiate naïve CD4 +CD25- T cells into islet-protective Foxp3+ regulatory T cells

CD4+CD25+Foxp3+ regulatory T cells (T regs) are important for preventing autoimmune diabetes and are either thymic-derived (natural) or differentiated in the periphery outside the thymus (induced). Here we show that β-cell peptide-pulsed dendritic cells (DCs) from nonobese diabetic (NOD) mice can effectively induce CD4 +CD25+Foxp3+ T cells from naïve islet-specific CD4+CD25- T cells in the presence of TGF-β1. These induced, antigen-specific T regs maintain high levels of clonotype-specific T cell receptor expression and exert islet-specific suppression in vitro. When cotransferred with diabetogenic cells into NOD scid recipients, T regs induced with DCs and TGF-β1 prevent the development of diabetes. Furthermore, in overtly NOD mice, these cells are able to significantly protect syngeneic islet grafts from established destructive autoimmunity. These results indicate a role for DCs in the induction of antigen-specific CD4+CD25+Foxp3+ T cells that can inhibit fully developed autoimmunity in a nonlymphopoenic host, providing an important potential strategy for immunotherapy in patients with autoimmune diabetes

The role of dendritic cell subsets and innate immunity in the pathogenesis of type 1 diabetes and other autoimmune diseases

Dendritic cells (DCs) are key antigen presenting cells that have an important role in autoimmune pathogenesis. DCs control both steady-state T cell tolerance and activation of pathogenic responses. The balance between these two outcomes depends on several factors, including genetic susceptibility, environmental signals that stimulate varied innate responses, and which DC subset is presenting antigen. Although the specific DC phenotype can diverge depending on the tissue location and context, there are 4 main subsets identified in both mouse and human: conventional cDC1 and cDC2, plasmacytoid DCs, and monocyte-derived DCs. In this review, we will discuss the role of these subsets in autoimmune pathogenesis and regulation, as well as the genetic and environmental signals that influence their function. Specific topics to be addressed include: impact of susceptibility loci on DC subsets, alterations in DC subset development, the role of infection- and host-derived innate inflammatory signals, and the role of the intestinal microbiota on DC phenotype. The effects of these various signals on disease progression and the relative effects of DC subset composition and maturation level of DCs will be examined. These areas will be explored using examples from several autoimmune diseases but will focus mainly on type 1 diabetes