Frequency Of Foxp3 Research Articles

Adenosine Generated by Regulatory T Cells Induces CD8+ T Cell Exhaustion in Gastric Cancer through A2aR Pathway.

Background Adenosine, derived from the degradation of ATP via ectonucleotidases CD39 and CD73, is a critical immunosuppressive metabolite in the hypoxic microenvironment of tumor tissue. Adenosine signaling via A2aR can inhibit the antitumor immune response of CD8+ T cells. CD39 and CD73 high-expressing Tregs play a critical role in tumor immune evasion of gastric cancer (GC). The present study investigated the underlying mechanism by which Tregs suppress antitumor immune responses in GC. Materials and Methods Fifty-two GC samples were collected, and the frequency of FoxP3+ Tregs and CD8+ T cells and density ratios of A2aR+/CD8+ T cells, CD39+/FoxP3+ Tregs, and CD73+/FoxP3+ Tregs in GC were assessed with multiplex immunofluorescence. The expression of FoxP3 and A2aR in GC tissues was also detected by the immunoblotting assay. We next investigated the relationship between density of FoxP3+ Tregs, ratio of A2aR+/CD8+ T cells, and clinicopathological parameters. At the same time, Tregs and CD8+ T cells were isolated from peripheral blood of five GC patients, and the antagonists of CD39 and CD73 were used to assess the ability of Tregs to decompose ATP into adenosine. In addition, we cocultured CD8+ T cells and Tregs with antagonists of A2aR and A2bR in order to examine the alterations in immune function of CD8+ T cells. Results The density of both FoxP3+ Tregs and A2aR+/CD8+ T cells was higher in GC tissue compared to peritumoral normal tissue and significantly correlated with the TNM stage, lymph node metastasis, and distant metastasis of GC. The process of Treg hydrolysis of ATP into adenosine was blocked by the antagonists of CD39 and CD73. In addition, Tregs could induce apoptosis and inhibit proliferation of CD8+ T cells, while this effect could be obviously reduced by applying the antagonist of A2aR or A2aR+A2bR. Moreover, IFN-γ, TNF-α, and perforin generated by CD8+ T cells could also be inhibited through the adenosine A2aR pathway. Conclusions The FoxP3+ Tregs and A2aR+/CD8+ T cells were excessively infiltrated in GC tissue. Tregs from GC can decompose ATP to adenosine and in turn induce apoptosis and inhibit the proliferation of CD8+ T cells through the A2aR pathway, further leading to immune escape of GC.

A novel method for efficient generation of antigen-specific effector T-cells using dendritic cells transduced with recombinant adeno-associated virus and p38 kinase blockade

BackgroundThe inefficacy of standard therapeutic strategies for ovarian cancer is reflected by the enduring poor prognosis of this malignancy. Due to the potential for exquisite specificity, sensitivity and long-term memory, immunotherapy offers an alternative modality for durable control of the disease, provided appropriate antigens can be identified andpresented in the right context.MethodsWe tested a novel dendritic cell vaccine formulation to reprogram autologous antigen-specific T-cells in vitro, in vivo in a murine model of ovarian cancer, and ex vivo using human cells from patients.ResultsWe show that dendritic cells (DCs) treated with a p38 MAPK inhibitor and transduced with a recombinant adenovirus associated vector (AAV) expressing Sperm protein (Sp) 17 are highly effective in generating antigen-specific T-cell cytotoxic response against ovarian cancer cells. Additionally, these DCs enhanced the differentiation of effector T-cells while reducing the frequency of Foxp3+ T-reg cells in vitro.ConclusionsThis work provides a rationale for translation of pharmacologically reprogrammed DCs into clinical trials for prevention of tumor recurrence and progression in high-risk ovarian cancer patients.

Ectonucleotidase CD39 and Checkpoint Signalling Receptor Programmed Death 1 are Highly Elevated in Intratumoral Immune Cells in Non–small-cell Lung Cancer

Lung cancer is the leading cause of cancer death in both sexes worldwide and has a predicted 5-year survival rate of <20%. Immunotherapy targeting immune checkpoints such as the programmed death 1 (PD-1) signaling pathway has led to a shift of paradigm in the treatment of advanced non–small-cell lung cancer (NSCLC) but remains without effect in ∼80% of patients. Accumulating evidence suggests that several immunosuppressive mechanisms may work together in NSCLC. The contribution and cooperation between different immunosuppressive mechanisms in NSCLC remain unknown. Recently, the CD39-adenosine pathway has gained increasing attention as a crucial immunosuppressive mechanism and possible target for immunotherapy. Immune cells were extracted from lung and tumor tissue after lung resection in 12 patients by combined enzymatic and mechanical tissue disaggregation. A multiparameter flow cytometry panel was established to investigate the expression and coexpression of CD39 and PD-1 on key lymphocyte subtypes. Frequencies of CD39+, PD-1+, and CD39+/PD-1+cells were higher among both CD4+ and CD8+ T cells isolated from NSCLC tumor tissue than in T cells from normal lung tissue. Similarly, the frequency of FoxP3+ CD4+ T cells (Tregs) was highly significantly elevated in tumor tissue compared to adjacent lung tissue. The consistent upregulation of CD39 on immune cells in tumor microenvironment indicates that the CD39 signaling pathway may, in addition to the PD-1 pathway, represent another important mechanism for tumor-induced immunosuppression in NSCLC. In addition, the present study indicates that a comprehensive immune response profiling with flow cytometry may be both feasible and clinically relevant.

Foxp3+Helios+ regulatory T cells are associated with monocyte subsets and their PD-1 expression during acute HIV-1 infection

BackgroundHelios has been reported to stabilize regulatory T (Treg) suppressive function. Programmed cell death protein 1 (PD-1) expression in three human monocyte subsets modulates immune responses. Recently, our team reported that three monocyte subsets are associated with T helper cell differentiation in HIV-1-infected patients. Until now, the effects of monocyte subsets and their PD-1 expression on Foxp3+Helios+ Treg cells have not been fully characterized, especially during acute HIV-1 infection.ResultsThe frequency of Foxp3+Helios+CD45RA+ Treg cells is significantly higher in patients with acute HIV-1 infection than those of healthy controls and chronic HIV-1-infected patients undergoing combined antiretroviral therapy. The frequency of Foxp3+Helios+CD45RA+ Treg cells is inversely correlated with CD4 T-cell counts and the CD4/CD8 ratio in chronic HIV-1-infected patients. During acute HIV-1 infection, the frequency of Foxp3+Helios+CD45RA+ Treg cells is inversely correlated with the frequency of the intermediate CD14++CD16+ monocyte subset, but positively correlated with PD-1 expression in both intermediate CD14++CD16+ and non-classical CD14+CD16++ monocyte subsets.ConclusionsIn this study, the perturbations of Foxp3+Helios+ Treg cells were characterized, and the association between monocyte subsets and their PD-1 expression and Foxp3+Helios+ Treg cells was evaluated during HIV-1 infection. Our observations provide new evidence of the roles for Foxp3+Helios+ Treg cells and PD-1 expression on monocyte subsets in HIV pathogenesis.

Short-chain fatty acids regulate regulatory T cells and intestinal pathology during oral mucosal infection

Abstract Complex interactions between the microbial flora and the host exert sophisticated means of immune tolerance and regulation mechanisms. One mechanism is by inducing the accumulation of regulatory T (Treg) cells. Here we show that the depletion of resident bacteria using antibiotics (Abx) causes oral and gut immunopathology during Oropharyngeal Candidiasis (OPC) infection. Abx treatment causes decrease in the frequency of Foxp3+ regulatory cells (Tregs) and IL-17A producing T cells, with a concomitant increase in oral tissue pathology. Although oral C. albicans (CA) is commonly controlled in the oral cavity, Abx treatment led to CA dependent oral and gut inflammation. The combination of short chain fatty acids (SCFA) partially controlled the pathology in Abx treated mice, correlating to an increase in the frequency of Foxp3+, IL-17A+, and Foxp3+IL-17A+ double positive (Treg17) cells in tongue and oral draining lymph nodes. SCFA enabled the restoration of Th17 cells and Treg cells and oral infection clearance, but did not reverse weight loss. Because SCFA treatment did not fully reverse the gut inflammation, it is evident that resident microbiota have SCFA independent homeostatic mechanisms in gut mucosa. We also found that SCFA potently induce Foxp3 and IL-17A expression in CD4+ T cells, depending on the cytokine milieu in vitro. Taken together, our data reveal that SCFA derived from resident bacteria play a critical role in controlling gut immunopathology by regulating T cell cytokines during oral mucosal infections.

RNA-binding protein PCBP1 shapes tolerance and immune response by constraining the formation of regulatory T cells

Abstract Maintaining adequate regulatory T cell (Treg) and effector T cell balance is critical for efficient regulation of tolerance and immune response, respectively. There is increasing evidence that in addition to regulating RNA fate, RNA-binding proteins (RBPs) may also control gene expression and determine the fate of immune cells. Here by studying mice in which the RBP Pcbp1 gene was disrupted in T cells, we define PCBP1 as a global regulator of immune activity that stabilizes immune responses, while repressing the signature programs of Tregs. PCBP1 expression was upregulated in response to T cell activation, but functionally inhibited by TGF-β-mediated phosphorylation. Loss of PCBP1 expression in T cells was associated with dysregulated peripheral homeostasis and impaired effector functions in knockout mice. Indeed, PCBP1-deficient T cells produced less IFN-γ and TNF-α upon stimulation with PMA, plus ionomycin, compared to wild-type littermates. Strikingly, lack of PCBP1 constrained the pathogenicity of effector T cells by conferring resistance to acute graft-vs-host disease in mice. Mechanistically, assessment of the genome-wide function of PCBP1 revealed that it limits the expression of genes associated with commitment to the Treg lineage. Consequently, its absence resulted in a marked increase in the frequency of Foxp3+Treg cells. Together, our findings identify PCBP1 as a distinct regulator of effector T cell functions and the formation of Treg cells in order to control the balance between tolerance and immunity.

Polarization of M2 Macrophages to Promote Tumor Immune Escape via Il-10 in SCLC but Not NSCLC Patients

Background and Aims: Because of significant differences in pathology, etiology, clinical features, and treatment options between small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC), it is necessary to investigate differences in macrophage polarization and function in SCLC vs. NSCLC. Macrophage polarization and function are important factors in the growth and metastasis of lung cancer. Methods: Peripheral blood and bronchoalveolar lavage fluid (BALF) were collected from 32 SCLC patients, 35 NSCLC patients, and 14 control subjects (CS). Levels of various subtypes of macrophages, monocytes, and regulatory T cells (Tregs) were determined by flow cytometry. Levels of IL-10 in serum and BALF were evaluated using cytometric bead array (CBA). Lung cancer was diagnosed based on the results of hematoxylin-eosin (HE) staining and histochemistry. Results: Total macrophages (CD14+) and frequency of M2 macrophages (CD206+CD14+ or IL-10+CD206+CD14+) were significantly increased in SCLC patients, compared to NSCLC patients. These changes were accompanied by increased levels of IL-10 in BALF of SCLC patients, compared to NSCLC patients. Increased frequency of M2 macrophages and elevated IL-10 concentration in BALF were positively correlated with advanced tumor stage in SCLC. In SCLC patients, levels of M2 macrophages were positively correlated with IL-10 levels but negatively correlated with survival time. In the same group of patients, IL-10 levels in BALF and frequency of IL-10+CD206+CD14+ M2 macrophages were positively correlated with frequency of Foxp3+CD25+CD4+ Tregs. Finally, levels of circulating M2 monocytes were significantly increased after chemotherapy in SCLC patients, compared to NSCLC patients. Conclusions: Compared to NSCLC, SCLC is characterized by strong polarization of M2 macrophages, which promotes tumor immune escape via IL-10. Funding: This work was supported by the following sponsors: the National Natural Science Foundation of China (No. 30972610, 81273240, 91742107, 81570002, and 81700027), the National Key Research and Development Program (No. 2017YFC0910000 and 2017YFD0501300), the Jilin Province Science and Technology Agency (No. 20160101037JC, 20170622009JC, 2017C021, 2017J039, SXGJXX2017-8, JJKH20180197KJ, DBXM154-2018, and 2018SCZWSZX-015), the Norman Bethune Program of Jilin University (2012206), and the fund of the State Key Laboratory of Kidney Diseases at PLA General Hospital. Declaration of Interest: The authors declare no potential conflicts of interest. Ethical Approval: This study was approved by the Ethical Committee of the First Hospital of Jilin University.

Immunological Changes in Monocyte Subsets and Their Association With Foxp3+ Regulatory T Cells in HIV-1-Infected Individuals With Syphilis: A Brief Research Report

The incidence of syphilis has increased dramatically in men who have sex with men (MSM), especially those with HIV-1 infection. Treponema pallidum and HIV-1 are bidirectionally synergistic, accelerating disease progression reciprocally in co-infected individuals. We have shown that monocytes have different effects on T helper cells at different stages of HIV-1 infection. However, the immunological changes in the three monocyte subsets and in regulatory T cells (Tregs), and the associations between these cell types during syphilis infection among HIV-1-infected MSM remain unclear. Herein, we used cell staining methods to explore changes in monocyte subsets and Tregs and any associations between these cells. We found that the frequency of classical monocytes was higher in the rapid plasma reagin (RPR+) group than in the healthy controls (HCs) and the chronic HIV-1 infection (CHI) plus RPR+ (CHI&RPR+) group. The frequencies of Foxp3+CD25+CD45RA+ and Foxp3+Helios+CD45RA+ Tregs were significantly higher in the RPR+, CHI, and CHI&RPR+ groups than in HCs, whereas the frequency of CD45RA+ Tregs was lower in the CHI&RPR+ group than in CHI group. The frequencies of Foxp3+CD25+CD45RO+ and Foxp3+Helios+CD45RO+ Tregs were lower in the RPR+, CHI, and CHI&RPR+ groups than in HCs. The frequency of intermediate monocytes was inversely correlated with the frequency of CD45RA+ Tregs and positively correlated with the frequency of CD45RO+ Tregs. These results demonstrate for the first time that intermediate monocytes control the differentiation of Treg subsets in Treponema pallidum/HIV-1 co-infections. These findings provide new insights into an immunological mechanism involving monocytes/Tregs in HIV-infected individuals with syphilis.

The significance of cytokine-producing B cells in breast tumor-draining lymph nodes.

The role of cytokine-producing B cells in antitumor immunity is mostly overlooked. In the present study, we investigated changes in B cell cytokine profiles in breast tumor-draining lymph nodes (TDLNs) during disease progression, and associations of these changes with prognostic indicators. Flow cytometry was used to measure the expression of TNF-α, IL-10, TGF-β, IL-2 and IFN-γ in B cells from 42 axillary lymph nodes. The frequencies of IL-10+ and FoxP3+ T regulatory cells (Tregs) were also determined. No significant changes in B cell cytokine profiles were observed during breast cancer progression from stage I to III, but the percentage of B cells with high TNF-α expression (TNFhi) showed a negative relationship with lymph node involvement and Her2 expression (p < 0.05). The percentage of IL-10+ B cells was found to be significantly higher in non-metastatic lymph nodes in node-negative compared to node-positive patients (p = 0.001). The frequencies of IL-10+ and TNFhi B cells were found to be negatively correlated with the number of involved lymph nodes. The frequency of TNFhi B cells showed an inverse correlation with the frequency of FoxP3+ Tregs, which in turn was associated with indicators of a poor prognosis. Our data indicate that the cytokine profiles of B cells in TDLNs of patients with breast cancer show associations with various disease parameters. TNFhi and IL-10+ B cells correlated positively with indicators of a good prognosis. Further functional studies are required to elucidate the role of cytokine production by B cells in immunity against breast cancer.

Dendritic Cells Exposed to Triiodothyronine Deliver Pro-Inflammatory Signals and Amplify IL-17-Driven Immune Responses.

Although a cross-talk between immune and endocrine systems has been well established, the precise pathways by which these signals co-regulate pro- and antiinflammatory responses on antigen-presenting cells remain poorly understood. In this work we investigated the mechanisms by which triiodothyronine (T3) controls T cell activity via dendritic cell (DC) modulation. DCs from wild-type (WT) and IL-6-deficient mice were pulsed with T3. Cytokine production and programmed death protein ligands (PD-L) 1 and 2 expression were assayed by flow cytometry and ELISA. Interferon-regulatory factor-4 (IRF4) expression was evaluated by RT-qPCR and flow cytometry. The ability of DCs to stimulate allogenic splenocytes was assessed in a mixed lymphocyte reaction and the different profile markers were analyzed by flow cytometry and ELISA. For in vivo experiments, DCs treated with ovalbumin and T3 were injected into OTII mice. Proliferation, cytokine production, frequency of FoxP3+ regulatory T (Treg) cells and PD-1+ cells were determined by MTT assay, ELISA and flow cytometry, respectively. T3 endows DCs with pro-inflammatory potential capable of generating IL-17-dominant responses and down-modulating expression of PD-L1 and 2. T3-stimulated WT-DCs increased the proportion of IL-17-producing splenocytes, an effect which was eliminated when splenocytes were incubated with T3-treated DCs derived from IL-6-deficient mice. Enhanced IL-17 expression was recorded in both, CD4- and CD4+ populations and involved the IRF-4 pathway. Particularly, γδ-T cells but not natural killer (NK), NKT, B lymphocytes nor CD8+ T cells were the major source of IL-17-production from CD4- cells. Moreover, T3-conditioned DCs promoted a decrease of the FoxP3+ Treg population. Furthermore, T3 down-modulated PD-1 expression on CD4- cells thereby limiting inhibitory signals driven by this co-inhibitory pathway. Thus, T3 acts at the DC level to drive proinflammatory responses in vitro. Accordingly, we found that T3 induces IL-17 and IFNγ-dominant antigen-specific responses in vivo. These results emphasize the relevance of T3 as an additional immune-endocrine checkpoint and a novel therapeutic target to modulate IL-17-mediated pro-inflammatory responses.

Study of FoxP3+ CD4+ CD25+ in systemic lupus erythematosus and rheumatoid arthritis

BackgroundSystemic lupus erythematosus (SLE) and rheumatoid arthritis (RA) as autoimmune diseases arise owing to failure of immunological self-tolerance. One of the mechanisms employed to control these potentially damaging cells are regulatory T cells (Tregs). The importance of Tregs is underscored by the overwhelming inflammation and autoimmunity that result from their absence. Forkhead box p3 (FoxP3) is an important regulator of Treg function, and the expression of FoxP3 correlates with the expression of other Treg-associated markers such as CD25 and CTLA-4.AimTo investigate the frequency of FoxP3+ CD4+ CD25+high cells (Tregs) in peripheral blood from patients with SLE and those with RA.Patients and methodsA total of 25 patients with SLE (15 patients with active SLE and 10 patients with inactive SLE), 25 patients with RA (15 patients with active RA and 10 patients with inactive RA), and 10 age-matched and sex-matched healthy controls were enrolled in the study. Patients underwent clinical and laboratory assessment. The frequency of Tregs was determined by flow cytometry.ResultsThe distribution of FoxP3+ CD4+ CD25+high cells (Tregs) revealed a highly significant decrease in the frequency of Treg in patients with SLE compared with healthy controls. Moreover, patients with active SLE showed significantly lower Tregs percent when compared with inactive group.Moreover, the distribution of FoxP3+ CD4+ CD25+high cells (Tregs) revealed a high significantly decrease in the frequency of Treg in patients with RA compared with healthy controls.ConclusionCD4+ CD25+ FoxP3 Tregs (as a percent of total CD4 cells) were significantly lower in patients with SLE and those with RA when compared with healthy controls.

Modulation of B Cells and Homing Marker on NK Cells Through Extracorporeal Photopheresis in Patients With Steroid-Refractory/Resistant Graft-Vs.-Host Disease Without Hampering Anti-viral/Anti-leukemic Effects

Graft-vs.-host disease (GvHD), a severe complication of allogeneic hematopoietic stem cell transplantation, significantly affects the post-transplant morbidity and mortality. Systemic steroids remain the gold standard for the initial management of GvHD. However, up to 60% of patients will not sufficiently respond to steroids. Extracorporeal photopheresis (ECP), a cell-based immunotherapy, has shown good clinical results in such steroid-refractory/resistant GvHD patients. Given its immunomodulatory, but not global immunosuppressive and steroid-sparing capacity, ECP constitutes an attractive option. In the case of GvHD, the balance of immune cells is destroyed: effector cells are not any longer efficiently controlled by regulatory cells. ECP therapy may restore this balance. However, the precise mechanism and the impact of ECP on anti-viral/anti-leukemic function remain unclear. In this study, 839 ECP treatments were performed on patients with acute GvHD (aGvHD) and chronic GvHD (cGvHD). A comprehensive analysis of effector and regulatory cells in patients under ECP therapy included multi-parametric flow cytometry and tetramer staining, LuminexTM-based cytokine, interferon-γ enzyme-linked immunospot, and chromium-51 release assays. Gene profiling of myeloid-derived suppressor cells (MDSCs) was performed by microarray analysis. Immunologically, modulations of effector and regulatory cells as well as proinflammatory cytokines were observed under ECP treatment: (1) GvHD-relevant cell subsets like CD62L+ NK cells and newly defined CD19hiCD20hi B cells were modulated, but (2) quantity and quality of anti-viral/anti-leukemic effector cells were preserved. (3) The development of MDSCs was promoted and switched from an inactivated subset (CD33−CD11b+) to an activated subset (CD33+CD11b+). (4) The frequency of Foxp3+CD4+ regulatory T cells (Tregs) and CD24+CD38hi regulatory B cells was considerably increased in aGvHD patients, and Foxp3+CD8+ Tregs in cGvHD patients. (5) Proinflammatory cytokines like IL-1β, IL-6, IL-8, and TNF-α were significantly reduced. In summary, ECP constitutes an effective immunomodulatory therapy for patients with steroid-refractory/resistant GvHD without impairment of anti-viral/leukemia effects.

Patients with intracranial aneurysms presented defects in regulatory T cells, which were associated with impairment in Tim-3 upregulation

Pathogenic inflammation contributes to aneurysm formation by mediating the destruction of the endothelium and the extracellular matrix and promoting pathogenic proliferation of smooth muscle cells. In mouse models, tolerance-inducing T regulatory (Treg) cells could significantly reduce the incidence and severity of aneurysms. Hence, it should be investigated why in human intracranial aneurysm (IA) patients, Treg cells failed to provide protection against aneurysm formation. In this study, the frequency and function of Treg cells in IA patients were examined. The frequency of Foxp3+ Treg cells was significantly lower in IA patients than in healthy controls. This downregulation was only specific to the Treg subset of CD4+ T cells, as the frequency of total CD4+ T cell was increased in IA patients. Subsequently, we found that the expressions of Treg-associated molecules, including Foxp3, CTLA-4, TGF-β, and IL-10, were significantly lower in Foxp3+ Treg cells from IA patients than in Foxp3+ Treg cells from healthy controls. In both healthy controls and IA patients, Foxp3+ Treg cells were distinguished into a more potent Tim-3+ subset and a less potent Tim-3− subset. The Tim-3+ subset of Foxp3+ Treg cells was significantly reduced in IA patients. Signaling via IL-2, IL-7, IL-15 and IL-21 was shown to promote Tim-3 upregulation in CD4+ and CD8+ T cells. Interestingly, we found that Tim-3 could be upregulated in Treg cells via the same mechanism, but compared to the Treg cells from healthy controls, the Treg cells from IA patients presented defects in Tim-3 upregulation upon cytokine stimulation. Together, our results demonstrated that Foxp3+ Treg cells in IA patients presented reduced function, which was associated with a defect in Tim-3 upregulation.

Role of Short Chain Fatty Acids in Controlling Tregs and Immunopathology During Mucosal Infection.

Interactions between mucosal tissues and commensal microbes control appropriate host immune responses and inflammation, but very little is known about these interactions. Here we show that the depletion of resident bacteria using antibiotics (Abx) causes oral and gut immunopathology during oropharyngeal candidiasis (OPC) infection. Antibiotic treatment causes reduction in the frequency of Foxp3+ regulatory cells (Tregs) and IL-17A producers, with a concomitant increase in oral tissue pathology. While C. albicans (CA) is usually controlled in the oral cavity, antibiotic treatment led to CA dependent oral and gut inflammation. A combination of short chain fatty acids (SCFA) controlled the pathology in Abx treated mice, correlating to an increase in the frequency of Foxp3+, IL-17A+, and Foxp3+IL-17A+ double positive (Treg17) cells in tongue and oral draining lymph nodes. However, SCFA treatment did not fully reverse the gut inflammation suggesting that resident microbiota have SCFA independent homeostatic mechanisms in gut mucosa. We also found that SCFA potently induce Foxp3 and IL-17A expression in CD4+ T cells, depending on the cytokine milieu in vitro. Depletion of Tregs alone in FDTR mice recapitulated oral inflammation in CA infected mice, showing that Abx mediated reduction of Tregs was involved in infection induced pathology. SCFA did not control inflammation in Treg depleted mice in CA infected FDTR mice, showing that Foxp3+ T cell induction was required for the protective effect mediated by SCFA. Taken together, our data reveal that SCFA derived from resident bacteria play a critical role in controlling immunopathology by regulating T cell cytokines during mucosal infections. This study has broader implications on protective effects of resident microbiota in regulating pathological infections.

Celastrol mediates Th17 and Treg cell generation via metabolic signaling

A T helper 17 (Th17) cell/regulatory T (Treg) cell imbalance is involved in many immune disorders and diseases. Celastrol, a Chinese herbal compound that has anti-inflammatory and immunosuppressive properties, has been indicated to suppress T cell proliferation and Th17 cell induction, while facilitating Forkhead box P3 (Foxp3) expression and Treg cell generation. In this study, we explored the impact and mechanism of celastrol on Th17 cell/induced Treg (iTreg) cell induction. CD4+CD25− T cells were purified, stimulated with anti-CD3 and anti-CD28 antibodies, and polarized in vitro to generate Th17 or iTreg cells in the presence or absence of celastrol. Initially, we determined that Interleukin (IL)-17 expression by celastrol-treated Th17 was significantly decreased compared with untreated cells; however, the frequency of Foxp3+ cells was increased in celastrol-treated cells. We verified that celastrol inhibited phospho-STAT3 expression in cultured Th17 cells and up-regulated phospho-STAT5 expression in iTreg cells. Furthermore, T cells treated with celastrol were more likely to participate in FAO metabolism instead of glycolysis. Celastrol suppressed the expression of glucose transporter, Glut1, and the rate-limiting enzyme, HK2, in addition to mTOR, HIF-1α, c-Myc and Akt expression in Th17 cells. Conversely, celastrol promoted FAO of lipids by up-regulating CPT1A and AMPKα expression in iTreg cells. Our results suggest that celastrol suppresses Th17 cell induction, while promoting the generation of iTreg cells. We found that celastrol inhibits glycolysis in Th17 cells and promotes FAO by iTreg cells, suggesting that celastrol could mediate the metabolism of Th17 and iTreg cells.

The Balance of Th1/Th2 and LAP+Tregs/Th17 Cells Is Crucial for Graft Survival in Allogeneic Corneal Transplantation.

Purpose CD4+LAP+ T cells are newly discovered regulatory T cells (Tregs). The aim of this study is to investigate the balance of Th1/Th2 and LAP+Tregs/Th17 in mice after allogeneic corneal transplantation. Methods A total of 65 mice received orthotopic penetrating transplantation. According to the survival scores of the grafts, the mice were divided into the rejection group and the survival group 3 weeks after transplantation. Th1, Th2, Th17, and regulatory T cells in the ipsilateral drainage lymph nodes and spleens were measured with flow cytometry. The related cytokines in aqueous humor were also analyzed. Results The frequencies of Foxp3+Tregs, GARP+Tregs, and LAP+Tregs in the survival group were significantly higher than those in the rejection group. And the expression trend of CD4+LAP+ T cells and CD4+GARP+ T cells was consistent. The level of IFN-γ, TNF, IL-6, and IL-17A markedly increased in aqueous humor during corneal allograft rejection. The ratio of Th1/Th2 and Th17/LAP+Tregs significantly increased in the rejection group at the 3rd week after corneal transplantation. Conclusion LAP+Tregs might be regarded as substitute for Foxp3+Tregs. The balance of Th1/Th2 and LAP+Tregs/Th17 is crucial for corneal allograft survival.

Regulatory T-cells in acute dengue viral infection.

Although regulatory T-cells (Tregs ) have been shown to be expanded in acute dengue, their role in pathogenesis and their relationship to clinical disease severity and extent of viraemia have not been fully evaluated. The frequency of Tregs was assessed in 56 adult patients with acute dengue by determining the proportion of forkhead box protein 3 (FoxP3) expressing CD4+ CD25+ T-cells (FoxP3+ cells). Dengue virus (DENV) viral loads were measured by quantitative real-time polymerase chain reaction (PCR) and DENV-specific T-cell responses were measured by ex-vivo interferon (IFN)-γ enzyme-linked immunospot (ELISPOT) assays to overlapping peptide pools of DENV-NS3, NS1 and NS5. CD45RA and CCR4 were used to phenotype different subsets of T-cells and their suppressive potential was assessed by their expression of cytotoxic T lymphocyte-antigen 4 (CTLA-4) and Fas. While the frequency of FoxP3+ cells in patients was significantly higher (P<0·0001) when compared to healthy individuals, they did not show any relationship with clinical disease severity or the degree of viraemia. The frequency of FoxP3+ cells did not correlate with either ex-vivo IFN-γ DENV-NS3-, NS5- or NS1-specific T-cell responses. FoxP3+ cells of patients with acute dengue were predominantly CD45RA+ FoxP3low , followed by CD45RA-FoxP3low , with only a small proportion of FoxP3+ cells being of the highly suppressive effector Treg subtype. Expression of CCR4 was also low in the majority of T-cells, with only CCR4 only being expressed at high levels in the effector Treg population. Therefore, although FoxP3+ cells are expanded in acute dengue, they predominantly consist of naive Tregs , with poor suppressive capacity.

Macrophages in keloid are potent at promoting the differentiation and function of regulatory T cells

The mechanistic details of keloid formation are still not understood. Given that the immune system is engaged in skin lesion repair, we examined the CD14+ macrophages and CD3+ T cells in keloid tissues and in the normal skin. Compared to the normal skin, keloid tissues presented significantly elevated infiltration by CD14+ macrophages. Moreover, the transcription and protein expression of iNOS, IL-12, IL-10, and TGF-β were significantly higher in keloid macrophages than in normal skin macrophages, in which the expression of M2-associated genes were further elevated compared to M1-associated genes in keloid. We also observed that keloid tissues presented higher infiltration by CD3+ T cells, of which the majority was CD4+ T cells. Notably, the frequency of Foxp3+ regulatory T cells (Tregs) in keloid tissues was significantly higher compared to that in the peripheral blood. Furthermore, macrophages from keloid tissues possessed potent capacity to induce Foxp3 expression in circulating CD3+ T cells. Together, this study suggested that macrophages in keloid tissues presented high activation status and were polarized toward the M2 subtype; moreover, these macrophages could promote Treg differentiation by upregulating Foxp3 expression.

CCR6 signaling inhibits suppressor function of induced-Treg during gut inflammation

CCR6 is a G protein-coupled receptor (GPCR) that binds to a specific chemokine, CCL20. The role of CCR6-CCL20 is very well studied in the migration of immune cells, but the non-chemotaxis functions of CCR6 signaling were not known. Here, we show that during gut inflammation, the frequency of Foxp3+CD4+ T cells (Tregs) reduced in the secondary lymphoid tissues and CCR6+ Tregs enhanced the expression of RORγt. The peripheral blood mononuclear cells (PBMCs) of ulcerative colitis (UC) patients showed lower percentages of Foxp3+CD4+ T cells, as compared to healthy individuals, with CCR6+ Tregs showing higher RORγt expression as compared to CCR6−Tregs. CCL20 inhibited the TGF-β1-induced Treg (iTreg) differentiation and directed them towards the pathogenic Th17-lineage in a CCR6-dependent manner. The iTreg that differentiated in the presence of CCL20 showed lower surface expression of suppressor molecules such as CD39, CD73 and FasL, and had impaired suppressive function. Furthermore, CCR6 signaling induced phosphorylation of Akt, mTOR, and STAT3 molecules in T cells. In conclusion, we have identified a new role of CCR6 signaling in the differentiation of iTregs during inflammation and gut autoimmunity.

Cells with Treg-specific FOXP3 demethylation but low CD25 are prevalent in autoimmunity

Identification of alterations in the cellular composition of the human immune system is key to understanding the autoimmune process. Recently, a subset of FOXP3+ cells with low CD25 expression was found to be increased in peripheral blood from systemic lupus erythematosus (SLE) patients, although its functional significance remains controversial. Here we find in comparisons with healthy donors that the frequency of FOXP3+ cells within CD127lowCD25low CD4+ T cells (here defined as CD25lowFOXP3+ T cells) is increased in patients affected by autoimmune disease of varying severity, from combined immunodeficiency with active autoimmunity, SLE to type 1 diabetes. We show that CD25lowFOXP3+ T cells share phenotypic features resembling conventional CD127lowCD25highFOXP3+ Tregs, including demethylation of the Treg-specific epigenetic control region in FOXP3, HELIOS expression, and lack of IL-2 production. As compared to conventional Tregs, more CD25lowFOXP3+HELIOS+ T cells are in cell cycle (33.0% vs 20.7% Ki-67+; P = 1.3 × 10−9) and express the late-stage inhibitory receptor PD-1 (67.2% vs 35.5%; P = 4.0 × 10−18), while having reduced expression of the early-stage inhibitory receptor CTLA-4, as well as other Treg markers, such as FOXP3 and CD15s. The number of CD25lowFOXP3+ T cells is correlated (P = 3.1 × 10−7) with the proportion of CD25highFOXP3+ T cells in cell cycle (Ki-67+). These findings suggest that CD25lowFOXP3+ T cells represent a subset of Tregs that are derived from CD25highFOXP3+ T cells, and are a peripheral marker of recent Treg expansion in response to an autoimmune reaction in tissues.