Staphylococcus aureus convert neonatal conventional CD4+ T cells into FOXP3+ CD25+ CD127low T cells via the PD‐1/PD‐L1 axis

The gut microbiota provides an important stimulus for the induction of regulatory T (Treg) cells in mice, whether this applies to newborn children is unknown. In Swedish children, Staphylococcus aureus has become a common early colonizer of the gut. Here, we sought to study the effects of bacterial stimulation on neonatal CD4⁺ T cells for the induction of CD25⁺ CD127^low Treg cells in vitro. The proportion of circulating CD25⁺ CD127^low Treg cells and their expression of FOXP3, Helios and CTLA‐4 was examined in newborns and adults. To evaluate if commensal gut bacteria could induce Treg cells, CellTrace violet‐stained non‐Treg cells from cord or peripheral blood from adults were co‐cultured with autologous CD25⁺ CD127^low Treg cells and remaining mononuclear cells and stimulated with S. aureus. Newborns had a significantly lower proportion of CD25⁺ CD127^low Treg cells than adults, but these cells were Helios⁺ and CTLA‐4⁺ to a higher extent than in adults. FOXP3⁺ CD25⁺ CD127^low T cells were induced mainly in neonatal CellTrace‐stained non‐Treg cells after stimulation with S. aureus. In cell cultures from adults, S. aureus induced CD25⁺ CD127^low T cells only if sorted naive CD45RA⁺ non‐Treg cells were used, but these cells expressed less FOXP3 than those induced from newborns. Sorted neonatal CD25⁺ CD127^low T cells from S. aureus‐stimulated cultures were still suppressive. Finally, blocking PD‐L1 during stimulation reduced the induction of FOXP3⁺ CD25⁺ CD127^low T cells. These results suggest that newborns have a higher proportion of circulating thymically derived Helios⁺ Treg cells than adults and that S. aureus possess an ability to convert neonatal conventional CD4⁺ T cells into FOXP3⁺ CD25⁺ CD127^low Treg cells via the PD‐1/PD‐L1 axis.     

Critical stoichiometric ratio of CD4+ CD25+ FoxP3+ regulatory T cells and CD4+ CD25− responder T cells influence immunosuppression in patients with B‐cell acute lymphoblastic leukaemia

Regulatory T (Treg) cells act to suppress activation of the immune system and thereby maintain immunological homeostasis and tolerance to self-antigens. The frequency and suppressing activity of Treg cells in general are high in different malignancies. We wanted to identify the role and regulation of CD4⁺ CD25⁺ FoxP3⁺ Treg cells in B-cell acute lymphoblastic leukaemia (B-ALL). We have included patients at diagnosis (n = 54), patients in clinical remission (n = 32) and normal healthy individuals (n = 35). These diagnosed patients demonstrated a lower number of CD4⁺ CD25⁺ cells co-expressing a higher level of FoxP3, interleukin-10, transforming growth factor-β and CD152/CTLA-4 than the normal population. Treg cells from patients showed a higher suppressive capability on CD4⁺ CD25⁻ responder T (Tresp) cells than normal. The frequency and immunosuppressive potential of CD4⁺ CD25⁺ FoxP3⁺ Treg cells became high with the progression of malignancy in B-ALL. Relative distribution of Tresp and Treg cells was only ˜5 : 1 in B-ALL but ˜35 : 1 in normal healthy individuals, further confirming the elevated immunosuppression in patients. A co-culture study at these definite ex vivo ratios, indicated that Treg cells from B-ALL patients exhibited higher immunosuppression than Treg cells from normal healthy individuals. After chemotherapy using the MCP841 protocol, the frequency of CD4⁺ CD25⁺ cells was gradually enhanced with the reduction of FoxP3, interleukin-10 positivity corresponded with disease presentation, indicating reduced immunosuppression. Taken together, our study indicated that the CD4⁺ CD25⁺ FoxP3⁺ Treg cells played an important role in immunosuppression, resulting in a positive disease-correlation in these patients. To the best of our knowledge, this is the first detailed report on the frequency, regulation and functionality of Treg cells in B-ALL.     

CD4+ CD25+ GARP+ regulatory T cells display a compromised suppressive function in patients with dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a lethal inflammatory heart disease and closely connected with dysfunction of the immune system. Glycoprotein A repetitions predominant (GARP) expressed on activated CD4⁺ T cells with suppressive activity has been established. This study aimed to investigate the frequency and function of circulating CD4⁺   CD25⁺ GARP⁺ regulatory T (Treg) cells in DCM. Forty‐five DCM patients and 46 controls were enrolled in this study. There was a significant increase in peripheral T helper type 1 (Th1) and Th17 number and their related cytokines [interferon‐γ (IFN‐γ), interleukin (IL‐17)], and an obvious decrease in Treg number, transforming growth factor‐β₁ (TGF‐β₁) levels and the expression of forkhead box P3 (FOXP3) and GARP in patients with DCM compared with controls. In addition, the suppressive function of CD4⁺ CD25⁺ GARP⁺ Treg cells was impaired in DCM patients upon T‐cell receptor stimulation detected using CFSE dye. Lower level of TGF‐β₁ and higher levels of IFN‐γ and IL‐17 detected using ELISA were found in supernatants of the cultured CD4⁺ CD25⁺ GARP⁺ Treg cells in DCM patients compared with controls. Together, our results indicate that CD4⁺ CD25⁺ GARP⁺ Treg cells are defective in DCM patients and GARP seems to be a better molecular definition of the regulatory phenotype. Therefore, it might be an attractive stategy to pay more attention to GARP in DCM patients.     

Co‐expression of TNFR2 and CD25 identifies more of the functional CD4+FOXP3+ regulatory T cells in human peripheral blood

Previously, we found that co‐expression of CD25 and TNFR2 identified the most suppressive subset of mouse Treg. In this study, we report that human peripheral blood (PB) FOXP3⁺ cells present in CD25^high, CD25^low and even CD25^– subsets of CD4⁺ cells expressed high levels of TNFR2. Consequently, TNFR2‐expressing CD4⁺CD25 ⁺ Treg included all of the FOXP3⁺ cells present in the CD4⁺CD25^high subset as well as a substantial proportion of the FOXP3⁺ cells present in the CD4⁺CD25^low subset. Flow cytometric analysis of PB identified five‐fold more Treg, determined by FOXP3 expression, in the CD4⁺CD25⁺TNFR2⁺ subset than in the CD4⁺CD25^high subset. In addition, similar levels of FOXP3⁺ cells were identified in both the CD4⁺CD25⁺TNFR2⁺ and CD4⁺CD25⁺CD127^low/? subsets. Furthermore, the CD4⁺CD25⁺TNFR2⁺ subset expressed high levels of CTLA‐4, CD45RO, CCR4 and low levels of CD45RA and CD127, a phenotype characteristic of Treg. Upon TCR stimulation, human PB CD4 ⁺ CD25 ⁺ TNFR2 ⁺ cells were anergic and markedly inhibited the proliferation and cytokine production of co‐cultured T‐responder cells. In contrast, CD4 ⁺ CD25 ⁺ TNFR2 ^– and CD4 ⁺ CD25 ^– TNFR2 ⁺ T cells did not show inhibitory activity. As some non‐Treg express TNFR2, the combination of CD25 and TNFR2 must be used to identify a larger population of human Treg, a population that may prove to be of diagnostic and therapeutic benefit in cancer and autoimmune diseases.     

Depletion of CD4+CD25+CD127lo regulatory T cells does not increase allergen‐driven T cell activation

Background It has been suggested that allergic diseases are caused by defective suppression of allergen‐specific Th2 cells by CD4⁺CD25⁺ regulatory T cells. However, such studies have been hampered by the difficulty in distinguishing regulatory T cells from CD25‐expressing activated T cells. Recently, it was shown that conventional T cells expressed high levels of CD127, whereas regulatory T cells were CD127^lo, allowing discrimination between these distinct T cell subpopulations. Objective The aim of this study was to study whether the putative regulatory subset defined as CD4⁺CD25⁺CD127^lo was involved in grass pollen‐reactive T cell responses. Methods Peripheral blood mononuclear cells (PBMCs) were obtained from allergic donors and non‐atopic controls out of season. Grass pollen‐induced cytokine production and proliferation were compared in cultures of undepleted cells and cells depleted of CD4⁺CD25⁺, CD4⁺CD25⁺CD127^hi or CD4⁺CD25⁺CD127^lo T cells. Results Undepleted cell cultures from allergic patients showed significantly increased proliferation and Th2 cytokine production compared with non‐atopic controls. Depletion of all CD25⁺ T cells did not increase cytokine production or proliferation, and more importantly, no increase in Th2 cytokine production or proliferation was observed in cell cultures depleted of CD4⁺CD25⁺CD127^lo cells (putative regulatory T cells) compared with undepleted PBMCs in both the allergic and the non‐atopic group. Conclusion Our study showed that T cells from grass pollen‐allergic patients and non‐atopic controls responded very differently to grass pollen extract, but this difference could not be explained by differences in regulatory T cell function. Further studies are needed to understand the importance of regulatory T cells in allergy.     

The calcineurin inhibitor tacrolimus allows the induction of functional CD4+CD25+ regulatory T cells by rabbit anti-thymocyte globulins

Rabbit anti-thymocyte globulins (rATG) induce CD4⁺CD25⁺forkhead box P3 (FoxP3⁺) regulatory T cells that control alloreactivity. In the present study, we investigated whether rATG convert T cells into functional CD4⁺CD25⁺FoxP3⁺CD127^−/low regulatory T cells in the presence of drugs that may hamper their induction and function, i.e. calcineurin inhibitors. CD25^neg T cells were stimulated with rATG or control rabbit immunoglobulin G (rIgG) in the absence and presence of tacrolimus for 24 h. Flow cytometry was performed for CD4, CD25, FoxP3 and CD127 and the function of CD25⁺ T cells was examined in suppression assays. MRNA expression profiles were composed to study the underlying mechanisms. After stimulation, the percentage CD4⁺CD25⁺FoxP3⁺CD127^−/low increased (from 2% to 30%, mean, P < 0·01) and was higher in the rATG samples than in control rIgG samples (2%, P < 0·01). Interestingly, FoxP3⁺T cells were also induced when tacrolimus was present in the rATG cultures. Blockade of the interleukin (IL)-2 pathway did not affect the frequency of rATG-induced FoxP3⁺ T cells. The rATG tacrolimus-induced CD25⁺ T cells inhibited proliferative responses of alloantigen-stimulated effector T cells as vigorously as rATG-induced and natural CD4⁺CD25⁺FoxP3⁺CD127^−/low T cells (67% ± 18% versus 69% ± 16% versus 45% ± 20%, mean ± standard error of the mean, respectively). At the mRNA-expression level, rATG-induced CD25⁺ T cells abundantly expressed IL-10, IL-27, interferon (IFN)-γ, perforin and granzyme B in contrast to natural CD25⁺ T cells (all P = 0·03), while FoxP3 was expressed at a lower level (P = 0·03). These mRNA data were confirmed in regulatory T cells from kidney transplant patients. Our findings demonstrate that tacrolimus does not negatively affect the induction, phenotype and function of CD4⁺CD25⁺ T cells, suggesting that rATG may induce regulatory T cells in patients who receive tacrolimus maintenance therapy.     

Increased frequency and suppressive activity of CD127low/− regulatory T cells in the peripheral circulation of patients with head and neck squamous cell carcinoma are associated with advanced stage and nodal involvement

The presence of regulatory T (Treg) cells is thought to be an important mechanism by which head and neck squamous cell carcinoma (HNSCC) successfully evades the immune system. Using multicolour flow cytometry, the frequency and functional capacity of two CD4⁺ CD127^low/− Treg cell populations, separated on the basis of different levels of CD25 expression (CD25^inter and CD25^high), from the peripheral circulation of newly presenting HNSCC patients were assessed with regard to clinicopathological features and healthy controls. The frequency of circulating Treg cells was similar between HNSCC patients and healthy controls, and for patients with HNSCC developing from different subsites (laryngeal compared with oropharyngeal). However, patients with advanced stage tumours and those with nodal involvement had significantly elevated levels of CD4⁺ CD25^high CD127^low/− Treg cells compared with patients who had early stage tumours (P = 0·03) and those without nodal involvement (P = 0·03), respectively. CD4⁺ CD25^high CD127^low/− Treg cells from the entire HNSCC patient cohort and from patients whose tumours had metastasized to the lymph nodes were also shown to suppress the proliferation of effector T cells significantly more, compared with those from healthy controls (P = 0·04) or patients with no nodal involvement (P = 0·04). Additionally, CD4⁺ CD25^inter CD127^low/− Treg cells consistently induced greater suppressive activity than CD4⁺ CD25^high CD127^low/− Treg cells on the proliferation of the effector T-cell populations (CD4⁺ CD25⁻ CD127^−/+ and CD4⁺ CD25⁺ CD127⁺). Peripheral Treg cells, identified by the CD127^low/− phenotype, have been shown to be influenced by a patient''s tumour stage and/or nodal status in HNSCC; suggesting a role in tumour progression that could be manipulated by future immunotherapy.     

Histone methylation mediates plasticity of human FOXP3+ regulatory T cells by modulating signature gene expressions

CD4⁺ FOXP3⁺ regulatory T (Treg) cells constitute a heterogeneous and plastic T-cell lineage that plays a pivotal role in maintaining immune homeostasis and immune tolerance. However, the fate of human Treg cells after loss of FOXP3 expression and the epigenetic mechanisms contributing to such a phenotype switch remain to be fully elucidated. In the current study, we demonstrate that human CD4⁺ CD25^high CD127^low/− Treg cells convert to two subpopulations with distinctive FOXP3⁺ and FOXP3⁻ phenotypes following in vitro culture with anti-CD3/CD28 and interleukin-2. Digital gene expression analysis showed that upon in vitro expansion, human Treg cells down-regulated Treg cell signature genes, such as FOXP3, CTLA4, ICOS, IKZF2 and LRRC32, but up-regulated a set of T helper lineage-associated genes, especially T helper type 2 (Th2)-associated, such as GATA3, GFI1 and IL13. Subsequent chromatin immunoprecipitation-sequencing of these subpopulations yielded genome-wide maps of their H3K4me3 and H3K27me3 profiles. Surprisingly, reprogramming of Treg cells was associated with differential histone modifications, as evidenced by decreased abundance of permissive H3K4me3 within the down-regulated Treg cell signature genes, such as FOXP3, CTLA4 and LRRC32 loci, and increased abundance of H3K4me3 within the Th2-associated genes, such as IL4 and IL5; however, the H3K27me3 modification profile was not significantly different between the two subpopulations. In conclusion, this study revealed that loss of FOXP3 expression from human Treg cells during in vitro expansion can induce reprogramming to a T helper cell phenotype with a gene expression signature dominated by Th2 lineage-associated genes, and that this cell type conversion may be mediated by histone methylation events.     

Identification of FOXP3-negative regulatory T-like (CD4(+)CD25(+)CD127(low)) cells in patients with immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome

Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome is an autoimmune disorder caused by mutations in the FOXP3 gene, which plays a key role in the generation of CD4⁺CD25⁺regulatory T (Treg) cells. We selected CD127 as the surface marker of Treg cells to illustrate the development and function of Treg cells in IPEX syndrome. CD4⁺CD25⁺FOXP3⁺ T cells, the putative Treg cells, were almost completely absent in all patients. Importantly, a substantial number of CD4⁺CD25⁺CD127^low T cells were observed in 3 IPEX patients with hypomorphic mutations in the FOXP3 gene. We demonstrated that CD4⁺CD25⁺CD127^low T cells isolated from these 3 patients exhibited an appreciable suppressive activity on effector T cell proliferation, although less than that displayed by Treg cells from healthy controls. These results suggest that genetically altered FOXP3 can drive the generation of functionally immature Treg cells, but that intact FOXP3 is necessary for the complete function of Treg cells.     

Low expression of CD39+/CD45RA+ on regulatory T cells (Treg) cells in type 1 diabetic children in contrast to high expression of CD101+/CD129+ on Treg cells in children with coeliac disease

Type 1 diabetes (T1D) and coeliac disease are both characterized by an autoimmune feature. As T1D and coeliac disease share the same risk genes, patients risk subsequently developing the other disease. This study aimed to investigate the expression of T helper (Th), T cytotoxic (Tc) and regulatory T cells (T_reg) in T1D and/or coeliac disease children in comparison to healthy children. Subgroups of T cells (Th : CD4⁺ or Tc : CD8⁺); naive (CD27⁺CD28⁺CD45RA⁺CCR7⁺), central memory (CD27⁺CD28⁺CD45RA⁻CCR7⁺), effector memory (early differentiated; CD27⁺CD28⁺CD45RA⁻CCR7⁻ and late differentiated; CD27⁻CD28⁻CD45RA⁻CCR7⁻), terminally differentiated effector cells (TEMRA; CD27⁻CD28⁻CD45RA⁺CCR7⁻) and T_reg (CD4⁺CD25⁺FOXP3⁺CD127⁻) cells, and their expression of CD39, CD45RA, CD101 and CD129, were studied by flow cytometry in T1D and/or coeliac disease children or without any of these diseases (reference group). Children diagnosed with both T1D and coeliac disease showed a higher percentage of TEMRA CD4⁺ cells (P < 0·05), but lower percentages of both early and late effector memory CD8⁺ cells (P < 0·05) compared to references. Children with exclusively T1D had lower median fluorescence intensity (MFI) of forkhead box protein 3 (FoxP3) (P < 0·05) and also a lower percentage of CD39⁺ and CD45RA⁺ within the T_reg population (CD4⁺CD25⁺FOXP3⁺CD127⁻) (P < 0·05). Children with exclusively coeliac disease had a higher MFI of CD101 (P < 0·01), as well as a higher percentage of CD129⁺ (P < 0·05), in the CD4⁺CD25^hi lymphocyte population, compared to references. In conclusion, children with combined T1D and coeliac disease have a higher percentage of differentiated CD4⁺ cells compared to CD8⁺ cells. T1D children show signs of low CD39⁺/CD45RA⁺ T_reg cells that may indicate loss of suppressive function. Conversely, children with coeliac disease show signs of CD101⁺/CD129⁺ T_reg cells that may indicate suppressor activity.     

Loss of regulatory characteristics in CD4+CD25+/hi T cells induced by impaired transforming growth factor beta secretion in pneumoconiosis

Pneumoconiosis is caused by the accumulation of airborne dust in the lung, which stimulates a progressive inflammatory response that ultimately results in lung fibrosis and respiratory failure. It is possible that regulatory cells in the immune system could function to suppress inflammation and possibly slow or reverse disease progression. However, results in this study suggest that in pneumoconiosis patients, the regulatory T cells (Tregs) and B cells are functionally impaired. First, we found that pneumoconiosis patients presented an upregulation of CD4⁺CD25⁺ T cells compared to controls, whereas the CD4⁺CD25⁺ and CD4⁺CD25^hi T cells were enriched with Th1‐ and Th17‐like cells but not Foxp3‐expressing Treg cells and evidenced by significantly higher T‐bet, interferon (IFN)‐γ, and interleukin (IL)‐17 expression but lower Foxp3 and transforming growth factor (TGF)‐β expression. Regarding the CD4⁺CD25^hi T‐cell subset, the frequency of this cell type in pneumoconiosis patients was significantly reduced compared to controls, together with a reduction in Foxp3 and TGF‐β and an enrichment in T‐bet, RORγt, IFN‐γ, and IL‐17. This skewing toward Th1 and Th17 types of inflammation could be driven by monocytes and B cells, since after depleting CD14⁺ monocytes and CD19⁺ B cells, the levels of IFN‐γ and IL‐17 were significantly decreased. Whole peripheral blood mononuclear cells and isolated monocytes and B cells in pneumoconiosis patients also presented reduced capacity of TGF‐β secretion. Furthermore, monocytes and B cells from pneumoconiosis patients presented reduced capacity in inducing Foxp3 upregulation, a function that could be rescued by exogenous TGF‐β. Together, these data indicated a potential pathway for the progression of pneumoconiosis through a loss of Foxp3⁺ Treg cells associated with impaired TGF‐β secretion.     

The imbalance of T helper 17/regulatory T cells and memory B cells during the early post‐transplantation period in peripheral blood of living donor liver transplantation recipients under calcineurin inhibitor‐based immunosuppression

There is limited clinical research regarding the changes in peripheral lymphocyte subsets during the early post-operative period of liver transplantation. Serial changes of T cells and B cells in living donor liver transplantation (LDLT) recipients during the early post-transplantion period were prospectively investigated. From June 2010 to February 2011, 27 consecutive LDLT recipients were enrolled. Percentages of T helper type 1 (Th1; interferon-γ-producing), Th2 (interleukin-4-producing), Th17 (interleukin-17-producing), regulatory T (Treg; CD4⁺ CD25⁺ FoxP3⁺), memory B (CD19⁺ CD24^hi CD38⁻) and mature B (CD19⁺ CD24^int CD38^int) cells were measured using fluorescence-activated cell sorting. Patients were followed up for a median of 9·9 months (range 6·8−15·5 months) after transplantation. Serial monitoring of immunological profiles showed no significant suppression of Th1, Th2, Th17, mature B or memory B cells, whereas frequencies of Treg cells significantly decreased. Interleukin-17 production by central and effector memory cells was not suppressed during the early post-operative period. The continuous production of interleukin-17 by the memory T cells may contribute to the persistence of Th17 cells. This prospective study demonstrated that current immunosuppression maintained the effector T or memory B cells during the early post-transplantation period but significantly suppressed Treg cells. Serial immune monitoring may suggest clues for optimal or individualized immunosuppression during the early post-operative period in clinical practice.     

Loss of FOXP3 expression in natural human CD4+CD25+ regulatory T cells upon repetitive in vitro stimulation

The adoptive transfer of CD4⁺CD25⁺ natural regulatory T cells (Treg) is a promising strategy for the treatment of autoimmune diseases and the prevention of alloresponses after transplantation. Clinical trials exploring this strategy require efficient in vitro expansion of this rare cell population. Protocols developed thus far rely on high‐grade purification of Treg prior to culture initiation, a process still hampered by the lack of Treg cell‐specific surface markers. Depletion of CD127⁺ cells was shown to separate activated conventional T cells from natural Treg cell populations allowing the isolation of highly enriched FOXP3⁺ cells with all functional and molecular characteristics of natural Treg. Here, we demonstrate that upon in vitro expansion, CpG methylation in a conserved region within the FOXP3 gene locus increased in CD4⁺CD25⁺CD127^low Treg, correlating with loss of FOXP3 expression and emergence of pro‐inflammatory cytokines. Further analysis identified CD45RA^?FOXP3⁺ memory‐type Treg as the main source of converting cells, whereas CD45RA⁺FOXP3⁺ Treg from the same donors showed no conversion within 3 wk of in vitro expansion. Thus, Treg cell lineage differentiation does not seem to represent a final fate decision, as natural Treg can lose their cell‐type‐specific characteristics after repetitive TCR stimulation.     

Loss of β‐arrestin 2 exacerbates experimental autoimmune encephalomyelitis with reduced number of Foxp3+ CD4+ regulatory T cells

β-Arrestins are well-known regulators and mediators of G protein-coupled receptor signalling, and accumulating evidence reveals that they are functionally involved in inflammation and autoimmune diseases. Of the two β-arrestins, β-arrestin 1 is documented to play regulatory roles in an animal model of multiple sclerosis (MS), whereas the role of β-arrestin 2 is less clear. Here, we show that β-arrestin 2-deficient mice displayed the exacerbated and sustained symptoms of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. At the cellular level, deficiency of β-arrestin 2 led to a decreased number of Foxp3⁺ CD4⁺ regulatory T (Treg) cells in peripheral lymphoid organs of EAE mice. Consistently, our in vitro observations also revealed that loss of β-arrestin 2 impaired the conversion of Foxp3⁻ CD4⁺ T cells into Foxp3⁺ CD4⁺ inducible Treg cells. Taken together, our data suggest that β-arrestin 2 plays a regulatory role in MS, that is opposite to that of β-arrestin 1, in autoimmune diseases such as MS, which is at least partially through regulation of iTreg cell differentiation.     

CD2‐mediated regulation of peripheral CD4+ CD25+ regulatory T‐cell apoptosis accompanied by down‐regulation of Bim

Extensive studies on CD4⁺ CD25⁺ regulatory T (Treg) cells suggest that they are important in regulating immune responses. However, mechanisms of peripheral Treg cell homeostasis are unknown. We found that stromal cells isolated from secondary lymphoid organs such as spleen and lymph nodes could support the survival of Treg cells. This was dependent on CD2 engagement and a direct interaction between Treg cells and stromal cells. In the presence of stromal cells, Bim, a pro‐apoptotic factor, was partially decreased in Treg cells. This effect could be inhibited by anti‐CD2 blocking antibodies, indicating that stimulation through CD2 on Treg cells regulates Bim expression, which may be relevant to Treg cell apoptosis. Therefore, Treg cell interactions with stromal cells through CD2 may be essential for Treg cell survival. Surprisingly, the expression of CD2 ligands on stromal cells was not detected. Hence, it is not clear how CD2 on Treg cells contributes to a direct interaction with the stromal cells and participates in survival support for Treg cells. Taken together, CD2 stimuli were mandatory for Treg cell survival with reduced Bim expression, but CD2 may not function as a direct receptor for molecules on stromal cells.     

Functional islet‐specific Treg can be generated from CD4+CD25− T cells of healthy and type 1 diabetic subjects

CD4⁺CD25⁺FOXP3⁺ Treg cells require TCR engagement for suppressive function, thus ensuring that suppression occurs only in the presence of specific antigens; however, to date no studies have addressed the function of self‐antigen‐specific Treg in humans. These studies were designed to determine whether peripheral generation and function of islet antigen‐specific adaptive Treg are defective in human subjects with type 1 diabetes (T1D). Islet antigen‐specific adaptive Treg were induced in vitro by activation of CD4⁺FOXP3^? T cells with glutamic acid decarboxylase and islet‐specific glucose‐6‐phosphate catalytic subunit‐related protein peptides in the context of T1D‐associated HLA‐DRβ alleles. Antigen‐specific Treg were characterized using flow cytometry for FOXP3 and class II tetramer and assessed for the ability to inhibit proliferation. These adaptive Treg were then compared with influenza‐specific Treg from the same study population. The function of tetramer⁺ cells that expressed FOXP3 was similar for both influenza and islet antigens generated from control and T1D subjects. In fact, the potency of suppression correlated with FOXP3 expression, not antigen specificity. Thus, these data suggest that development of functional adaptive Treg can occur in response to islet antigens and activation of islet‐specific Treg may potentially be used as a targeted immunotherapy in T1D.     

CD8+ CD122+ regulatory T cells contain clonally expanded cells with identical CDR3 sequences of the T‐cell receptor β‐chain

We identified CD8⁺ CD122⁺ regulatory T cells (CD8⁺ CD122⁺ Treg cells) and reported their importance in maintaining immune homeostasis. The absence of CD8⁺ CD122⁺ Treg cells has been shown to lead to severe systemic autoimmunity in several mouse models, including inflammatory bowel diseases and experimental autoimmune encephalomyelitis. The T‐cell receptors (TCRs) expressed on CD8⁺ CD122⁺ Treg cells recognize the target cells to be regulated. To aid in the identification of the target antigen(s) recognized by TCRs of CD8⁺ CD122⁺ Treg cells, we compared the TCR diversity of CD8⁺ CD122⁺ T cells with that of conventional, naive T cells in mice. We analysed the use of TCR‐Vβ in the interleukin 10‐producing population of CD8⁺ CD122⁺ T cells marked by high levels of CD49d expression, and found the significantly increased use of Vβ13 in these cells. Immunoscope analysis of the complementarity‐determining region 3 (CDR3) of the TCR β‐chain revealed remarkable skewing in a pair of Vβ regions, suggesting the existence of clonally expanded cells in CD8⁺ CD122⁺ T cells. Clonal expansion in Vβ13⁺ cells was confirmed by determining the DNA sequences of the CDR3s. The characteristic TCR found in this study is an important building block for further studies to identify the target antigen recognized by CD8⁺ CD122⁺ Treg cells.     

CD4+CD25highforkhead box protein 3+ regulatory T lymphocytes suppress interferon‐γ and CD107 expression in CD4+ and CD8+ T cells from tuberculous pleural effusions

Tuberculous pleural effusion is characterized by a T helper type 1 (Th1) profile, but an excessive Th1 response may also cause tissue damage that might be controlled by regulatory mechanisms. In the current study we investigated the role of regulatory T cells (T_reg) in the modulation of Th1 responses in patients with tuberculous (TB) pleurisy. Using flow cytometry we evaluated the proportion of T_reg (CD4⁺CD25^highforkhead box protein 3⁺), interferon (IFN)‐γ and interleukin (IL)‐10 expression and CD107 degranulation in peripheral blood (PB) and pleural fluid (PF) from patients with TB pleurisy. We demonstrated that the proportion of CD4⁺CD25⁺, CD4⁺CD25^highFoxP3⁺ and CD8⁺CD25⁺ cells were increased in PF compared to PB samples. Mycobacterium tuberculosis stimulation increased the proportion of CD4⁺CD25^low/negIL‐10⁺ in PB and CD4⁺CD25^low/negIFN‐γ⁺ in PF; meanwhile, CD25^high mainly expressed IL‐10 in both compartments. A high proportion of CD4⁺CD107⁺ and CD8⁺CD107⁺ cells was observed in PF. T_reg depletion enhanced the in‐vitro M. tuberculosis‐induced IFN‐γ and CD4⁺ and CD8⁺ degranulation responses and decreased CD4⁺IL‐10⁺ cells in PF. Our results demonstrated that in TB pleurisy T_reg cells effectively inhibit not only IFN‐γ expression but also the ability of CD4⁺ and CD8⁺ cells to degranulate in response to M. tuberculosis.