Background: Treg cells represent important viral reservoirs during chronic HIV infection. CD39 is closely involved in Treg-mediated immunosuppressive effects. However, CD39 expression on nTregs and mTregs and a relationship with HIV DNA levels during HIV infection is still unclear. In this study, we analyzed the distribution of HIV DNA in Treg subsets and the association between HIV DNA and CD39 expression on Treg subsets. Methods: Sixty-two HIV-infected patients with different HIV stages and 14 uninfected individuals were enrolled. nTregs (CD4+CD25+CD127lowCD45RO-) and mTregs (CD4+CD25+CD127lowCD45RO+) were isolated by magnetic selection and flow cytometric sorting. HIV DNA was quantified by real-time polymerase chain reaction (PCR). CD39 expression on nTregs and mTregs was analyzed by flow cytometry. Results: Higher levels of HIV DNA were detected in mTregs than those in nTregs during chronic HIV infection. The frequency of CD39+ nTregs and HIV DNA levels in nTregs were increased in patients with advanced HIV infection. Furthermore, HIV DNA levels in nTregs correlated positively with CD39+ nTreg frequency. CD39+ nTreg frequency was also increased in immune non-responders. Conclusions: mTregs and nTregs are both important reservoirs of virus during chronic HIV infection and HIV DNA levels increase in nTregs in patients with advanced HIV infection. We observed increased frequency of CD39+ nTregs and HIV DNA levels in nTregs in patients with advanced HIV infection. HIV DNA levels in nTregs correlated positively with CD39+ nTreg frequency.

BACKGROUND: The immune mechanism depends on CD4+ T cells for its regular function, and altered T cell function leads to microbial disease progression.
OBJECTIVE: The present study aimed to determine the role of naturally induced T-regulatory (nTreg) cells (CD4+ CD25+ Fox P3+) in periodontal disease pathogenesis.
METHODS: A total of 30 patients attending the out-patient clinic of the Department of Periodontology and Implantology, Faculty of Dental Sciences, Sri Ramachandra University (SRU), Chennai, India were recruited for the study. They were categorized in three groups as healthy individuals, individuals with chronic gingivitis, and individuals with chronic periodontitis gingival tissues. nTreg (CD4+ CD25+ Fox P3+) cells were isolated using flow cytometry. Different conjugated, isolated cells were then gated in the order of CD4+, CD25+, and Fox P3+ cells.
RESULTS: The results of our study showed an increase in the proportions of Treg cells in individuals with chronic periodontitis compared to individuals with gingivitis and healthy individuals.
CONCLUSIONS: Further elucidation of cellular and molecular processes underlying Treg cells will help unravel the complexity behind periodontal disease pathogenesis besides paving the way in developing newer treatment strategies.

Arsenic contamination of drinking water is a matter of global concern. Arsenic intake impairs immune responses and leads to a variety of pathological conditions including cancer. In order to understand the intricate tuning of immune responses elicited by chronic exposure to arsenic, a mouse model was established by subjecting mice to different environmentally relevant concentrations of arsenic in drinking water for 30days. Detailed study of the thymus, a primary immune organ, revealed arsenic-mediated tissue damage in both histological specimens and scanning electron micrographs. Analysis of molecular markers of apoptosis by Western blot revealed a dose-dependent activation of the apoptotic cascade. Enzymatic assays supported oxidative stress as an instigator of cell death. Interestingly, assessment of inflammatory responses revealed disparity in the NF-κB/IL-6/STAT3 axis, where it was found that in animals consuming higher amounts of arsenic NF-κB activation did not lead to the classical IL-6 upregulation response. This deviation from the canonical pathway was accompanied with a significant rise in numbers of CD4+ CD25+ FoxP3 expressing cells in the thymus. The cytokine profile of the animals exposed to higher doses of arsenic also indicated an immune-suppressed milieu, thus validating that arsenic shapes the immune environment in context to its dose of exposure and that at higher doses it leads to immune-suppression. Our study establishes a novel role of arsenic in regulating immune homeostasis in context to its dose, where, at higher doses, arsenic related upregulation of NF-κB cascade takes on an alternative role that is correlated with increased immune-suppression.

To study the development and function of \"natural-arising\" T regulatory (nTreg) cells, we developed a novel nTreg model on pure nonobese diabetic background using epigenetic reprogramming via somatic cell nuclear transfer. On RAG1-deficient background, we found that monoclonal FoxP3(+)CD4(+)Treg cells developed in the thymus in the absence of other T cells. Adoptive transfer experiments revealed that the thymic niche is not a limiting factor in nTreg development. In addition, we showed that the T-cell receptor (TCR) β-chain of our nTreg model was not only sufficient to bias T-cell development toward the CD4 lineage, but we also demonstrated that this TCR β-chain was able to provide stronger TCR signals. This TCR-β-driven mechanism would thus unify former per se contradicting hypotheses of TCR-dependent and -independent nTreg development. Strikingly, peripheral FoxP3(-)CD4(+)T cells expressing the same TCR as this somatic cell nuclear transfer nTreg model had a reduced capability to differentiate into Th1 cells but were poised to differentiate better into induced nTreg cells, both in vitro and in vivo, representing a novel peripheral precursor subset of nTreg cells to which we refer to as pre-nTreg cells.

MicroRNAs (miRNAs) regulate T cell development and function and the disruption of miRNAs in natural regulatory CD4(+) FOXP3(+) T cells (nTreg) leads to autoimmune disease in mice. To investigate miRNA expression in relation to autoimmune disease risk in humans we sequenced them in purified CD4(+) T cell subsets from individuals at high risk of type 1 diabetes (pre-T1D), as well as other healthy individuals. Differences in miRNA expression patterns were observed between specific T cell subsets and, within subsets, between pre-T1D and healthy individuals. Compared to healthy, naive CD4(+) T cells in pre-T1D displayed 32 differentially expressed miRNAs, potentially a template for altered miRNA expression in effector memory T cells in T1D. Naive nTreg in pre-T1D displayed two differentially expressed miRNAs, Let-7c and miR-15a. In contrast, nTreg activated in vivo displayed a large number of differentially expressed miRNAs, revealing a pro-inflammatory and FOXP3-repressive signature. Differential expression of specific miRNAs was also a signpost to altered T cell function. For example, in pre-T1D, increased expression of miR-26a in nTreg activated in vivo or in vitro was associated with decreased expression of its target, the histone methyltransferase EZH2. Chemical inhibition of EZH2 decreased the number of activated naïve nTreg and their expression of nTreg signature genes FOXP3 and TIGIT. Our findings demonstrate that miRNAs differentially expressed in CD4(+) T cell subsets are markers of risk and T cell dysfunction in T1D.

Natural regulatory T cells (nTreg) can suppress different immune-cell responses and maintain the balance between tolerance and immunity in the individual. These cells are defined by CD4+ , CD25hi, and FOXP3+ expression, although a variety of other nTreg-associated markers have been reported to be expressed at different levels (e.g., HLA-DR, CTLA-4, GARP, Helios, CD39, etc.), presumably reflecting different functional stages of the heterogeneous nTreg population. Several markers show low/negative expression (i.e., CD127, CD49d, and CD26), but none of these markers are specific to nTreg. CD25hi expression has been a useful surface marker to identify/isolate nTreg; however, CD25 is also expressed on \"adaptive\" or \"induced\" Treg, as well as in activated conventional T cells. In addition, the fact that FOXP3 is also found in CD25 low/negative CD4+ T cells, and in a subset of CD8+ T cells, further complicates the definition of a specific nTreg marker. Although CD4+, CD25hi, and CD127low/negative markers characterize the majority of nTreg, it is still imperative to find additional surface-marker combinations that improve the identification/isolation of nTreg and their subsets. Herein, we present evidence that CD4+ CD25hi CD6(lo/-) nTreg have high expression of FOXP3and exhibit in vitro suppressive activity on CD8+ T-cell proliferation. Dot-plot analyses of CD4+ cells, with CD6, CD127, CD49d, or CD26 reveal that a higher enrichment yield of CD25hi FOXP3+ cells can be achieved in the combined CD6(lo/-) CD127(lo/-) population. We conclude that FOXP3+ nTreg cells are characterized by CD6(lo/-) expression, providing a new tool for the identification of nTreg cells without recourse to intracellular staining, and for the purification of these cells by negative selection.

Regulatory T cells play a critical role in the immune response to vaccination, but there is only a limited understanding of the response of regulatory T cells to aluminum adjuvants and the vaccines that contain them. Available studies in animal models show that although induced T regulatory cells may be induced concomitantly with effector T cells following aluminum-adjuvanted vaccination, they are unable to protect against sensitization, suggesting that under the Th2 immune-stimulating effects of aluminum adjuvants, Treg cells may be functionally compromised. Allergic diseases are characterized by immune dysregulation, with increases in IL-4 and IL-6, both of which exert negative effects on Treg function. For individuals with a genetic predisposition, the beneficial influence of adjuvants on immune responsiveness may be accompanied by immune dysregulation, leading to allergic diseases. This review examines aspects of the regulatory T cell response to aluminum-adjuvanted immunization and possible genetic susceptibility factors related to that response.

BACKGROUND: The mechanisms contributing to clinical immune tolerance remain incompletely understood. This study provides evidence for specific immune mechanisms that are associated with a model of operationally defined clinical tolerance.
OBJECTIVE: Our overall objective was to study laboratory changes associated with clinical immune tolerance in antigen-induced T cells, basophils, and antibodies in subjects undergoing oral immunotherapy (OIT) for peanut allergy.
METHODS: In a phase 1 single-site study, we studied participants (n = 23) undergoing peanut OIT and compared them with age-matched allergic control subjects (n = 20) undergoing standard of care (abstaining from peanut) for 24 months. Participants were operationally defined as clinically immune tolerant (IT) if they had no detectable allergic reactions to a peanut oral food challenge after 3 months of therapy withdrawal (IT, n = 7), whereas those who had an allergic reaction were categorized as nontolerant (NT; n = 13).
RESULTS: Antibody and basophil activation measurements did not statistically differentiate between NT versus IT participants. However, T-cell function and demethylation of forkhead box protein 3 (FOXP3) CpG sites in antigen-induced regulatory T cells were significantly different between IT versus NT participants. When IT participants were withdrawn from peanut therapy for an additional 3 months (total of 6 months), only 3 participants remained classified as IT participants, and 4 participants regained sensitivity along with increased methylation of FOXP3 CpG sites in antigen-induced regulatory T cells.
CONCLUSIONS: In summary, modifications at the DNA level of antigen-induced T-cell subsets might be predictive of a state of operationally defined clinical immune tolerance during peanut OIT.

IL-4 is thought to promote induction of transplantation tolerance and alloantigen-specific CD4(+)CD25(+) T regulatory cells (Treg). This study examined the effect of IL-4 on the induction and maintenance of the CD4(+) T regulatory cells (Treg) that mediate transplantation tolerance. Tolerance was induced in DA rats with PVG heterotopic cardiac allografts by a short course of cyclosporine. Naïve and tolerant lymphocytes, including the CD4(+) and CD4(+)CD25(+) T cell subsets, were assayed in mixed lymphocyte cultures with or without recombinant (r)IL-4 or other cytokines. The proliferation, cell surface and cytokine phenotype of these cells was examined, as was their capacity to adoptively transfer tolerance. rIL-4 enhanced the proliferation of naïve and tolerant lymphoid cells, including CD4(+) and CD4(+)CD25(+) T cells, but this was not alloantigen specific. Naïve or tolerant CD4(+) T cells cultured with rIL-4 and donor PVG antigen effected rapid graft rejection, even though before culture tolerant CD4(+) T cells transferred antigen-specific tolerance. These rIL-4 cultured CD4(+) T cells had a phenotype consistent with activated CD4(+)CD25(+)FoxP3(-) Th2 cells. While naïve natural CD4(+)CD25(+) T cells (nTreg) cultured with alloantigen and rIL-4 had enhanced proliferation and capacity to suppress rejection in vivo, the culture of tolerant CD4(+)CD25(+) T cells with alloantigen and rIL-4 could not sustain their proliferation against specific donor, nor their capacity to transfer tolerance to specific donor allograft. Thus, IL-4 promotes both regulatory and effector T cells early in the immune response, but once alloimmune tolerance is established, IL-4 promoted the activation of effector cells to mediate rejection and did not support alloantigen-specific Treg that could transfer specific tolerance.

Complement is undeniably quintessential for innate immunity by detecting and eliminating infectious microorganisms. Recent work, however, highlights an equally profound impact of complement on the induction and regulation of a wide range of immune cells. In particular, the complement regulator CD46 emerges as a key sensor of immune activation and a vital modulator of adaptive immunity. In this review, we summarize the current knowledge of CD46-mediated signalling events and their functional consequences on immune-competent cells with a specific focus on those in CD4(+) T cells. We will also discuss the promises and challenges that potential therapeutic modulation of CD46 may hold and pose.