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CD80 and CD86 are expressed on antigen presenting cells and are required to engage their shared receptor, CD28, for the costimulation of CD4 T cells. It is unclear why two stimulatory ligands with overlapping roles have evolved. CD80 and CD86 also bind the regulatory molecule CTLA-4. We explored the role of CD80 and CD86 in the homeostasis and proliferation of CD4+FoxP3+ regulatory T cells (Treg), which constitutively express high levels of CTLA-4 yet are critically dependent upon CD28 signals. We observed that CD86 was the dominant ligand for Treg proliferation, survival, and maintenance of a regulatory phenotype, with higher expression of CTLA-4, ICOS, and OX40. We also explored whether CD80-CD28 interactions were specifically compromised by CTLA-4 and found that antibody blockade, clinical deficiency of CTLA-4 and CRISPR-Cas9 deletion of CTLA-4 all improved Treg survival following CD80 stimulation. Taken together, our data suggest that CD86 is the dominant costimulatory ligand for Treg homeostasis, despite its lower affinity for CD28, because CD80-CD28 interactions are selectively impaired by the high levels of CTLA-4. These data suggest a cell intrinsic role for CTLA-4 in regulating CD28 costimulation by direct competition for CD80, and indicate that that CD80 and CD86 have discrete roles in CD28 costimulation of CD4 T cells in the presence of high levels of CTLA-4. [ABSTRACT FROM AUTHOR]

Type 1 diabetes (T1D) is caused by immune-mediated destruction of insulin-producing β-cells. Insufficient control of autoreactive T cells by regulatory T cells (Tregs) is believed to contribute to disease pathogenesis, but changes in Treg function are difficult to quantify because of the lack of Treg-exclusive markers in humans and the complexity of functional experiments. We established a new way to track Tregs by using a gene signature that discriminates between Tregs and conventional T cells regardless of their activation states. The resulting 31-gene panel was validated with the NanoString nCounter platform and then measured in sorted CD4(+)CD25(hi)CD127(lo) Tregs from children with T1D and age-matched control subjects. By using biomarker discovery analysis, we found that expression of a combination of six genes, including TNFRSF1B (CD120b) and FOXP3, was significantly different between Tregs from subjects with new-onset T1D and control subjects, resulting in a sensitive (mean ± SD 0.86 ± 0.14) and specific (0.78 ± 0.18) biomarker algorithm. Thus, although the proportion of Tregs in peripheral blood is similar between children with T1D and control subjects, significant changes in gene expression can be detected early in disease process. These findings provide new insight into the mechanisms underlying the failure to control autoimmunity in T1D and might lead to a biomarker test to monitor Tregs throughout disease progression. [ABSTRACT FROM AUTHOR]

T regulatory cells (Tregs) control immune homeostasis by preventing inappropriate responses to self and nonharmful foreign antigens. Tregs use multiple mechanisms to control immune responses, all of which require these cells to be near their targets of suppression; however, it is not known how Treg-to-target proximity is controlled. Here, we found that Tregs attract CD4+ and CD8+ T cells by producing chemokines. Specifically, Tregs produced both CCL3 and CCL4 in response to stimulation, and production of these chemokines was critical for migration of target T cells, as Tregs from Ccl3-/- mice, which are also deficient for CCL4 production, did not promote migration. Moreover, CCR5 expression by target T cells was required for migration of these cells to supernatants conditioned by Tregs. Tregs deficient for expression of CCL3 and CCL4 were impaired in their ability to suppress experimental autoimmune encephalomyelitis or islet allograft rejection in murine models. Moreover, Tregs from subjects with established type 1 diabetes were impaired in their ability to produce CCL3 and CCL4. Together, these results demonstrate a previously unappreciated facet of Treg function and suggest that chemokine secretion by Tregs is a fundamental aspect of their therapeutic effect in autoimmunity and transplantation. [ABSTRACT FROM AUTHOR]

Objective. Granulocyte-macrophage colony stimulating factor (GM-CSF) is a potent inflammatory mediator that is responsible for recruitment and activation of innate immune cells. Recent data from murine studies have identified Th17 cells as a key source of GM-CSF and suggest that T cell-derived GM-CSF is instrumental in the induction of autoimmune disease. The present study was undertaken to analyze the expression of T cell-derived GM-CSF in the joints of patients with juvenile idiopathic arthritis (JIA) and to investigate the differentiation of Th17 cells and how this relates to GM-CSF+ T helper cells. Methods. Synovial fluid (SF) and peripheral blood (PB) samples from 24 patients with JIA were analyzed, by flow cytometry and reverse transcription-polymerase chain reaction, for expression of GM-CSF and the Th17 marker CD161. A cytokine capture assay was used to purify Th17 cells and test the plasticity of cytokine production in response to interleukin-12 (IL-12) and IL-23. Results. The frequency of GM-CSF-producing T helper cells was significantly enriched in SF mononuclear cells compared to PB mononuclear cells from the patients with JIA (24.1% of CD4+ T cells versus 2.9%) and closely correlated with the erythrocyte sedimentation rate (r2 = 0.91, P < 0.001). Synovial GMCSF + T cells were predominantly CD161+ and coexpressed interferon-γ (IFNγ), but not IL-17. Culture of Th17 cells in the presence of IL-12 led to rapid upregulation of GM-CSF and IFNγ, recapitulating the phenotype of GM-CSF-expressing cells within the joint. Conclusion. Our results identify a novel outcome of Th17 plasticity in humans that may account for the enrichment of GM-CSF-expressing T cells in the joints of patients with JIA. The association of GM-CSF expression with systemic inflammation highlights the potential role of Th17-related cytokines in the pathology of JIA. [ABSTRACT FROM AUTHOR]

Regulatory FoxP3+CD4+ T cells (Treg) are vital for maintaining the balance between tolerance, adequate immune response, and autoimmunity. Despite this immunoregulatory role, it has been shown that Treg may also produce proinflammatory cytokines. Here we present a distinct population of Treg, defined by CD161 expression, as the major source of FoxP3+ Treg-derived proinflammatory cytokines. CD161 + Treg can be followed through- out development, from thymus and cord blood to healthy child and adult samples. CD161 + Treg display anergy, are suppressive in cocultures with conventional T cells (Tconv), and possess a predominantly demethylated Treg-specific demethylated region of the FOXP3 locus. In addition to the production of interleukin(IL) 17A, interferon γ, and IL-2, CD161+ FoxP3+ cells share markers with Tconv, including expression of the transcription factors retinoic acid-related orphan receptor Cv2 (RORCv2) and T-cell-specific T-box transcription factor (Tbet). Expression of CD161 and enrichment for cytokine production are stable characteristics of CD161 + Treg upon both short- and longer-term culture in vitro. Additionally, CD161+ Treg are highly enriched within the inflammatory environment of childhood arthritis, suggesting a role in disease. Our data therefore demonstrate that CD161 +FoxP3+ T cells are a novel Treg subset, found in health and disease, which display high proinflammatory potential but also exhibit hallmark Treg characteristics. [ABSTRACT FROM AUTHOR]

Regulatory T cell (Treg) therapy is a potential curative approach for a variety of immune-mediated conditions, including autoimmunity and transplantation, in which there is pathological tissue damage. In mice, IL-33R (ST2)--expressing Tregs mediate tissue repair by producing the growth factor amphiregulin, but whether similar tissue-reparative Tregs exist in humans remains unclear. We show that human Tregs in blood and multiple tissue types produced amphiregulin, but this was neither a unique feature of Tregs nor selectively upregulated in tissues. Human Tregs in blood, tonsil, synovial fluid, colon, and lung tissues did not express ST2, so ST2+ Tregs were engineered via lentiviral-mediated overexpression, and their therapeutic potential for cell therapy was examined. Engineered ST2+ Tregs exhibited TCR-independent, IL-33--stimulated amphiregulin expression and a heightened ability to induce M2-like macrophages. The finding that amphiregulin-producing Tregs have a noneffector phenotype and are progressively lost upon TCR-induced proliferation and differentiation suggests that the tissue repair capacity of human Tregs may be an innate function that operates independently from their classical suppressive function. [ABSTRACT FROM AUTHOR]