Increased Treg Frequency Research Articles

Nicotinamide enhances Treg differentiation by promoting Foxp3 acetylation in immune thrombocytopenia.

Imbalanced nicotinamide adenine dinucleotide (NAD+) homeostasis has been reported in multiple autoimmune diseases and supplementation with NAD+ precursors has consistently demonstrated positive therapeutic benefits for these conditions. Immune thrombocytopenia (ITP) is an acquired autoimmune disease, in which the decreased number and impaired function of regulatory T cells (Tregs) contribute to the main pathogenesis. Here we found NAD+ level was decreased in the plasma and CD4+ T cells of ITP patients. Supplementation with NAD+ precursor nicotinamide (NAM), but not nicotinamide mononucleotide (NMN), increased Treg frequency and ameliorated thrombocytopenia in an ITP murine model. Moreover, whilst both NAM and NMN restored cytosolic NAD+ level in the CD4+ T cells from ITP patients, only NAM promoted Treg differentiation. Mechanistically, Sirtuin1 (Sirt1), a major consumer of NAD+, was highly expressed in the CD4+ T cells of ITP patients, potentially contributing to the low level of NAD+. NAM, which could act as Sirt1 inhibitor, promoted Foxp3 acetylation and stability in induced Tregs derived from naïve CD4+ T cells of ITP patients. These findings suggest that NAM holds promise as a novel therapeutic strategy for restoring immune balance in ITP.

Cytokine and T cell responses in post-chikungunya viral arthritis: A cross-sectional study.

To define the relationship between chronic chikungunya post-viral arthritis disease severity, cytokine response and T cell subsets in order to identify potential targets for therapy. Participants with chikungunya arthritis were recruited from Colombia from 2019-2021. Arthritis disease severity was quantified using the Disease Activity Score-28 and an Arthritis-Flare Questionnaire adapted for chikungunya arthritis. Plasma cytokine concentrations (interleukin (IL)-1β, IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, interferon-γ and tumor necrosis factor (TNF)) were measured using a Meso Scale Diagnostics assay. Peripheral blood T cell subsets were measured using flow cytometry. Among participants with chikungunya arthritis (N = 158), IL-2 levels and frequency of regulatory T cells (Tregs) were low. Increased arthritis disease activity was associated with higher levels of inflammatory cytokines (IL-6, TNF and CRP) and immunoregulatory cytokine IL-10 (p<0.05). Increased arthritis flare activity was associated with higher Treg frequencies (p<0.05) without affecting T effector (Teff) frequencies, Treg/Teff ratios and Treg subsets. Finally, elevated levels of IL-2 were correlated with increased Treg frequency, percent Tregs out of CD4+ T cells, and Treg subsets expressing immunosuppressive markers, while also correlating with an increased percent Teff out of live lymphocytes (p<0.05). Chikungunya arthritis is characterized by increased inflammatory cytokines and deficient IL-2 and Treg responses. Greater levels of IL-2 were associated with improved Treg numbers and immunosuppressive markers. Future research may consider targeting these pathways for therapy.

Regulatory T cell homing and activation is a signature of neonatal sepsis.

Regulatory T cells (Treg) play a prominent role in utero tolerating non-inherited maternal antigens and in regulating immune responses against pathogens at birth. This study investigates Treg immunity in newborns in West Africa, where sepsis remains a major public health problem. Treg phenotypes on neonates subgroups with early-onset sepsis (EOS), presumed sepsis, and healthy newborn with and without prenatal risk factors were evaluated. Treg phenotypes varied according to prenatal conditions, with increase in Treg frequency and Foxp3 expression in healthy newborns with prenatal risk factors compared to those with none risk. Compared to healthy newborns with prenatal risk factors, EOS neonates had a significantly reduced frequency of Treg and Foxp3 expression. In the Treg pool, higher frequency of activated Treg was observed in EOS neonates, suggesting an in-utero activation upstream of the sepsis onset. Their migration to the infection site may explain the reduced frequency of circulating Integrin α4β1+ Treg suggestive of homing to the endothelial tissue. EOS neonates show increases expression of CTLA-4, PD-1 and CD39 on Treg, which negatively regulate the activation of effector T cells (Teff) corroborating by the lower frequency of Teff in EOS neonates. The higher frequency of CD39+ Treg and the lower frequency of integrinα4β1+ Treg in EOS non-survivor suggests that Treg exhaustement and endothelial homing are associated with outcome severity. Neonates developing EOS are born with an altered Treg phenotypic profile. Treg expression of CTLA-4, PD-1, CD39, and integrinα4β1 cell markers can be considered as early warning or diagnostic markers of EOS.

The impact of fingolimod on Treg function in brain ischaemia.

Fingolimod has generally shown neuroprotective effects in stroke models. Here, we tested the hypothesis that fingolimod modulates T-cell cytokine production towards a regulatory phenotype. Second, we investigated how fingolimod altered the Treg suppressive function and the sensitivity of effector T cells to regulation. Mice that had underwent the permanent electrocoagulation of the left middle cerebral artery received saline or fingolimod (0.5mg/kg) daily for 10-days post-ischaemia. Fingolimod improved neurobehavioural recovery compared to saline control and increased Treg frequency in the periphery and brain. Tregs from fingolimod-treated animals had a higher expression of CCR8. Fingolimod increased the frequencies of CD4+ IL-10+ , CD4+ IFN-γ+ and CD4+ IL-10+ IFN-γ+ cells in spleen and blood, and CD4+ IL-17+ cells in the spleen, with only minor effects on CD8+ T-cell cytokine production. Treg from post-ischaemic mice had reduced suppressive function compared toTreg from non-ischaemic mice. Fingolimod treatment rescued this function against saline-treated but not fingolimod-treated CD4+ effector T cells. In conclusion, fingolimod seems to improve the suppressive function of Treg post-stroke while also increasing the resistance of CD4+ effector cells to this suppression. Fingolimod's capacity to increase both effector and regulatory functions may explain the lack of consistent improvement in functional recovery in experimental brain ischaemia.

Immunomodulatory effect of different statin regimens on regulatory T-cells in patients with acute coronary syndrome: a systematic review and network meta-analysis of randomized clinical trials.

We conducted a network meta-analysis (NMA) to determine the effects of low-dose (20mg/day or less) conventional statin therapy (CST) and high-dose (40mg/day or more) intensive statin therapy (IST) on the frequency of Tregs and their associated cytokines (IFN-γ, IL-10, TGF-β) compared with control. PubMed, Cochrane Library, and EMBASE databases were searched for randomized clinical trials (RCTs) to identify relevant articles published until June 2021. We pooled data extracted from the included studies using the standardized mean difference (SMD). A random-effects model was used to conduct this NMA. Heterogeneity was evaluated using Cochran's Q test and the I2 test. The Grading of Recommendations, Assessment, Development and Evaluations (GRADE) was used to assess the quality of the study. Data analysis was conducted using R software.A total of 505 patients were enrolled in the five RCTs. NMA indicated a significant increase in Treg frequency in the CST group compared with the control group (SMD 1.77; 95% CI: 0.77-2.76; P=0.0005) and a larger increase in the Treg frequency associated with the IST group compared with the control group (SMD 2.12; 95% CI: 1.15-3.10; P-value<0.0001). However, there was significant heterogeneity and inconsistency among the included studies [τ2=0.6096; τ = 0.7808; I2=91.2% (80.5%; 96.0%)]. When compared with control, both CST and IST increased the levels of secreted IL-10 (SMD 2.69; 95% CI: 2.07-3.31; P-value<0.0001 and SMD 2.14; 95% CI: 1.76-2.52; P-value<0.0001). Compared with the control group, CST was associated with increased levels of TGF-β (SMD 3.83; 95% CI: 0.63-7.0; P-value=0.0189); this association was not seen in the IST group. IFN-γ levels decreased significantly in both the IST and CST groups (SMD -1.52; 95% CI: -1.94-1.10; P-value<0.0001 and SMD -2.34; 95% CI: -2.73-1.95; P-value<0.0001, respectively). The findings of our study indicated that both high- and low-dose statin groups increased Treg frequency compared with the control group. IST demonstrated greater benefits than CST. Furthermore, statin therapy increased IL-10 and TGF-β levels and decreased IFN-γ levels. Overall, these results have significant implications for patients with ACS who would benefit from Treg-induced immunomodulatory balance.

Development of an extended half-life GM-CSF fusion protein for Parkinson's disease.

Transformation of CD4+ T cell effector to regulatory (Teff to Treg) cells have been shown to attenuate disease progression by restoring immunological balance during the onset and progression of neurodegenerative diseases. In our prior studies, we defined a safe and effective pathway to restore this balance by restoring Treg numbers and function through the daily administration of the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF). These studies were conducted as a proof-of-concept testing in Parkinson's disease (PD) preclinical models and early phase I clinical investigations. In both instances, they served to ameliorate disease associated signs and symptoms. However, despite the recorded efficacy, the cytokine's short half-life, low bioavailability, and injection site reactions proved to be limitations for any broader use. To overcome these limitations, mRNA lipid nanoparticles encoding an extended half-life albumin-GM-CSF fusion protein were developed for both mouse (Msa-GM-CSF) and rat (Rsa-GM-CSF). These formulations were tested for immunomodulatory and neuroprotective efficacy using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and human wild-type alpha-synuclein (αSyn) overexpression preclinical models of PD. A single dose of the extended half-life mouse and rat mRNA lipid nanoparticles generated measurable GM-CSF plasma cytokine levels up to four days. Increased Treg frequency and function were associated with a resting microglial phenotype, nigrostriatal neuroprotection, and restoration of brain tissue immune homeostasis. These findings were substantively beyond the recorded efficacy of daily recombinant wild-type GM-CSF with a recorded half-life of six hours. Mechanistic evaluation of neuropathological transcriptional profiles performed in the disease-affected nigral brain region demonstrated an upregulation of neuroprotective CREB and synaptogenesis signaling and neurovascular coupling pathways. These findings highlight the mRNA-encoded albumin GM-CSF fusion protein modification linked to improvements in therapeutic efficacy. The improvements achieved were associated with the medicine's increased bioavailability. Taken together, the data demonstrate that mRNA LNP encoding the extended half-life albumin-GM-CSF fusion protein can serve as a benchmark for PD immune-based therapeutics. This is especially notable for improving adherence of drug regimens in a disease-affected patient population with known tremors and gait abnormalities.

Helminth infection modulates number and function of adipose tissue Tregs in high fat diet-induced obesity.

BackgroundEpidemiological and experimental studies have shown a protective effect of helminth infections in weight gain and against the development of metabolic dysfunctions in the host. However, the mechanisms Treg cells exert in the helminth-obesity interface has been poorly investigated. The present study aimed to verify the influence of Heligmosomoides polygyrus infection in early stages of high fat diet-induced obesity.Principal findingsThe presence of infection was able to prevent exacerbated weight gain in mice fed with high fat diet when compared to non-infected controls. In addition, infected animals displayed improved insulin sensitivity and decreased fat accumulation in the liver. Obesity-associated inflammation was reduced in the presence of infection, demonstrated by lower levels of leptin and resistin, lower infiltration of Th1 and Th17 cells in adipose tissue, higher expression of IL10 and adiponectin, increased infiltration of Th2 and eosinophils in adipose tissue of infected animals. Of note, the parasite infection was associated with increased Treg frequency in adipose tissue which showed higher expression of cell surface markers of function and activation, like LAP and CD134. The infection could also increase adipose Treg suppressor function in animals on high fat diet.ConclusionThese data suggest that H. polygyrus modulates adipose tissue Treg cells with implication for weight gain and metabolic syndrome.

Induction of Foxp3 and activation of Tregs by HSP gp96 for treatment of autoimmune diseases

SummaryUpregulation and stabilization of Foxp3 expression in Tregs are essential for regulating Treg function and immune homeostasis. In this study, gp96 immunization showed obvious therapeutic effects in a Lyn–/– mouse model of systemic lupus erythematosus. Moreover, gp96 alleviated the initiation and progression of MOG-induced experimental autoimmune encephalomyelitis. Immunization of gp96 increased Treg frequency, expansion, and suppressive function. Gene expression profiling identified the NF-κB family member p65 and c-Rel as the key transcription factors for enhanced Foxp3 expression in Treg by gp96. Mutant gp96 within its Toll-like receptor (TLR) binding domain, TLR2 knockout mice, and mice with cell-specific deletion of MyD88, were used to demonstrate that gp96 activated Tregs and induced Foxp3 expression via a TLR2-MyD88-mediated NF-κB signaling pathway. Taken together, these results show that gp96 immunization restricted antibody-induced and Th-induced autoimmune diseases by integrating Treg expansion and activation, indicating its potential clinical usefulness against autoimmune diseases.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Mechanism of Increased Treg Frequency Induced by Irradiated Esophageal Squamous Cell Carcinoma

<h3>Purpose/Objective(s)</h3> The infiltration of CD4+CD25+Foxp3+ regulatory T cells (Treg) in the tumor microenvironment is one of the main reasons for radiation resistance and tumor recurrence after radiotherapy. It has been established that Treg is more resistant to radiation than other T cells, but the proliferation of immune cells after radiotherapy is affected by other factors, including tumor cells. Treg frequency in the tumor microenvironment after radiotherapy has not been defined. We studied the effect and mechanism of increased Treg frequency induced by irradiated ESCC cell, TE-1. <h3>Materials/Methods</h3> After 2 Gy irradiation, TE-1 cells were co-cultured with normal peripheral blood lymphocytes for 48 hours. Flow cytometry was used to detect Treg/CD4+T cell frequency. The mRNA expression of TGF-β1 and TGF-β2 in TE-1 was detected by qPCR, and the protein content of TGF-β1 and TGF-β2 in the medium was detected by ELISA. <h3>Results</h3> Compared with non-irradiation group, the expression of TGF-β1 and TGF-β2 in TE-1 cells of irradiation group increased, and the protein content of TGF-β1 and TGF-β2 in culture medium increased, the difference was statistically significant (<i>P</i> < 0.001). Flow cytometry showed that CD4+CD25+/CD4+Tcell and CD4+CD25+Foxp3+/CD4+Tcell were increased in the radiotherapy group after co-culture, and the difference was statistically significant (<i>P</i> < 0.001). <h3>Conclusion</h3> The expression of TGF-β1 and TGF-β2 in esophageal squamous cell carcinoma cells increased after irradiation, and the frequency of Treg induced by co-culture increased, suggesting that esophageal squamous cell carcinoma cells after radiotherapy can induce the increase of Treg cells, which may be achieved mainly through the mechanism of increasing the secretion of TGF-β1 and TGF-β2.

Induction of Foxp3 and Activation of Regulatory T Cells by Heat Shock Protein Gp96 for Treatment of Autoimmune Diseases

Utilizing regulatory T cells (Treg) is a promising approach for treating autoimmune diseases, including systemic lupus erythematosus (SLE) and rheumatic diseases. Upregulation and stabilization of forkhead box P3 (Foxp3) expression in Tregs are essential for regulating Treg function and immune homeostasis. In this study, we examined the potential effect and mechanism of heat shock protein gp96 in regulating Foxp3 expression and Treg activation in vivo. High-dose gp96 immunization showed obvious therapeutic effects in a Lyn-/- mouse model of SLE, as evidenced by decreased auto-antibody titers, follicular helper T cells, and antibody-production plasma cells. Moreover, immunization of mice with gp96 alleviated the initiation and progression of myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis (EAE). Immunization of gp96 increased Treg frequency, expansion, and suppressive function with no obvious effector T cell activation. Gene expression profiling was used to identify the NF-κB family member p65 and c-Rel as the key transcription factors for enhanced Foxp3 expression in Treg by gp96. Mutant gp96 within its Toll-like receptor (TLR) binding domain, TLR2 knockout (KO) mice, and mice with cell-specific deletion of MyD88, were used to demonstrate that gp96 activated Tregs and induced Foxp3 expression via a TLR2-MyD88-mediated NF-κB signaling pathway. Taken together, these results show that gp96 immunization restricted antibody- and Th-induced autoimmune diseases by integrating Treg expansion and activation, indicating its potential clinical usefulness against autoimmune diseases.

Lymphocyte Immune Response and T Cell Differentiation in Fontan Patients with protein-losing enteropathy.

Background Protein-losing enteropathy (PLE) is a severe complication of the Fontan circulation. There is increasing discussion about whether lymphatic dysregulation is involved as pathomechanism of PLE. This investigation focuses on the interplay between alteration of lymphatic cells and immunologic pathway alterations. Methods Micro-ribonucleic acid (miRNA) expression profiling was performed in 49 patients ( n = 10 Fontan patients with PLE, n = 30 Fontan patients without PLE, and n = 9 patients with dextro-transposition of the great arteries (dTGA). miRNA pathway analysis was performed to identify significantly enriched pathways. To determine lymphocyte populations and subtypes multiparameter flow cytometry was used. Results miRNAs pathway analysis of Fontan patients with PLE revealed 20 significantly changed networks of which four of the ten largest were associated with immunologic processes. This finding is supported by significant T cell deficiency with decreased CD4+ count ( p = 0.0002), altered CD4 +/CD8+ ratio, and significantly modified CD4+ ( p < 0.0001) and CD8+ ( p = 0.0002) T cell differentiation toward effector and terminal differentiated T cells in Fontan patients with PLE. Analyses of CD4+ T cell subsets demonstrated significantly increased frequencies of CD4+ CD25+ CD127– regulatory T cells (Treg) in Fontan patients with PLE ( p = 0.0011). Conclusion PLE in Fontan patients is associated with severe lymphopenia, T cell deficiency, significant alterations of T cell differentiation, and increased Treg frequency reflecting an immune status of chronic inflammation and shortened protection against pathogens and autoimmunity. These cellular alterations seemed to be dysregulated by several miRNA controlled immunological pathways.

Interleukin-6 blockade with tocilizumab increases Tregs and reduces T effector cytokines in renal graft inflammation: A randomized controlled trial.

Interleukin-6 (IL-6) is a proinflammatory cytokine and key regulator of Treg: T effector cell (Teff) balance. We hypothesized that IL-6 blockade with tocilizumab, a monoclonal antibody to IL-6R, would increase Tregs, dampen Teff function, and control graft inflammation. We conducted a randomized controlled clinical trial (2014-2018) of clinically stable kidney transplant recipients on calcineurin inhibitor, mycophenolate mofetil, and prednisone, with subclinical graft inflammation noted on surveillance biopsies during the first year posttransplant. Subjects received tocilizumab (8mg/kg IV every 4weeks; 6 doses; n=16) or no treatment (controls; n=14) on top of usual maintenance immunosuppression. Kidney biopsies pre- and post-treatment were analyzed using Banff criteria. Blood was analyzed for serum cytokines, Treg frequencies, and T cell effector molecule expression (IFN-γ, IL-17, granzyme B) post-stimulation ex vivo. Tocilizumab-treated subjects were more likely to show improved Banff ti-score (62.5% vs. 21.4%, p=.03), increased Treg frequency (7.1% ± 5.55% vs. 3.6% ± 1.7%, p = .0168), and a blunted Teff cytokine response compared to controls. Changes in Banff i- and t-scores were not significantly different. The treatment was relatively well tolerated with no patient deaths or graft loss. Blockade of IL-6 is a novel and promising treatment option to regulate the T cell alloimmune response in kidney transplant recipients. NCT02108600.

InsB9-23 Gene Transfer to Hepatocyte-Based Combined Therapy Abrogates Recurrence of Type 1 Diabetes After Islet Transplantation.

The induction of antigen (Ag)-specific tolerance represents a therapeutic option for autoimmune diabetes. We demonstrated that administration of a lentiviral vector enabling expression of insulin B chain 9-23 (InsB9-23) (LV.InsB) in hepatocytes arrests β-cell destruction in prediabetic NOD mice by generating InsB9-23-specific FoxP3+ T regulatory cells (Tregs). LV.InsB in combination with a suboptimal dose of anti-CD3 monoclonal antibody (combined therapy [CT], 1 × 5 μg [CT5]) reverts diabetes and prevents recurrence of autoimmunity after islet transplantation in ∼50% of NOD mice. We investigated whether CT optimization could lead to abrogation of recurrence of autoimmunity. Therefore, alloislets were transplanted after optimized CT tolerogenic conditioning (1 × 25 μg [CT25]). Diabetic NOD mice conditioned with CT25 when glycemia was <500 mg/dL remained normoglycemic for 100 days after alloislet transplantation and displayed reduced insulitis, but independently from the graft. Accordingly, cured mice showed T-cell unresponsiveness to InsB9-23 stimulation and increased Treg frequency in islet infiltration and pancreatic lymph nodes. Additional studies revealed a complex mechanism of Ag-specific immune regulation driven by CT25, in which both Tregs and PDL1 costimulation cooperate to control diabetogenic cells, while transplanted islets play a crucial role, although transient, recruiting diabetogenic cells. Therefore, CT25 before alloislet transplantation represents an Ag-specific immunotherapy to resolve autoimmune diabetes in the presence of residual endogenous β-cell mass.

Increased Regulatory T Cells Precede the Development of Bronchopulmonary Dysplasia in Preterm Infants.

Regulatory T cells (Tregs) are important for the ontogenetic control of immune activation and tissue damage in preterm infants. However, the role of Tregs for the development of bronchopulmonary dysplasia (BPD) is yet unclear. The aim of our study was to characterize CD4+ CD25+ forkhead box protein 3 (FoxP3)+ Tregs in peripheral blood of well-phenotyped preterm infants (n = 382; 23 + 0 – 36 + 6 weeks of gestational age) with a focus on the first 28 days of life and the clinical endpoint BPD (supplemental oxygen for longer than 28 days of age). In a subgroup of preterm infants, we characterized the immunological phenotype of Tregs (n = 23). The suppressive function of Tregs on CD4+CD25- T cells was compared in preterm, term and adult blood. We observed that extreme prematurity was associated with increased Treg frequencies which peaked in the second week of life. Independent of gestational age, increased Treg frequencies were noted to precede the development of BPD. The phenotype of preterm infant Tregs largely differed from adult Tregs and displayed an overall naïve Treg population (CD45RA+/HLA-DR-/Helios+), especially in the first days of life. On day 7 of life, a more activated Treg phenotype pattern (CCR6+, HLA-DR+, and Ki-67+) was observed. Tregs of preterm neonates had a higher immunosuppressive capacity against CD4+CD25- T cells compared to the Treg compartment of term neonates and adults. In conclusion, our data suggest increased frequencies and functions of Tregs in preterm neonates which display a distinct phenotype with dynamic changes in the first weeks of life. Hence, the continued abundance of Tregs may contribute to sustained inflammation preceding the development of BPD. Functional analyses are needed in order to elucidate whether Tregs have potential as future target for diagnostics and therapeutics.

Overexpressed immunoglobulin-like transcript (ILT) 4 in lung adenocarcinoma is correlated with immunosuppressive T cell subset infiltration and poor patient outcomes

BackgroundThe poor response to current PD-1/PD-L1 inhibitors in lung cancer patients requires development of novel immunotargets. Immunoglobulin-like transcript (ILT)4 is an immunosuppressive molecule mainly expressed in myeloid innate cells. Recent studies showed that ILT4 was highly expressed in multiple malignant cells and regulated tumor biologies including proliferation, invasion and metastasis. However, the immunomodulatory role of tumor cell-derived ILT4 is unclear. Here we aimed to analyze the correlation of tumor cell ILT4 expression with T cell infiltration and subset distribution, illustrate ILT4-regulated immunosuppressive microenvironment, and raise tumor cell-derived ILT4 as a novel immunotherapeutic target and prognostic biomarker for lung adenocarcinoma (LUAD) patients.MethodsWe collected the tissue samples and corresponding clinicopathological data from 216 primary LUAD patients. Using immunohistochemical staining and public database analyses we investigated the relationship between ILT4 expression and different T cell subset density as well as patient outcomes.ResultsEnriched ILT4 expression in tumor cells of LUAD tissues indicated reduced T cell infiltration in the tumor microenvironment (TME), advanced diseases and poor patient overall survival (OS). Further T cell subset analyses revealed that ILT4 expression was correlated with decreased CD8+T cell and increased Treg frequency in both cancer nest and stroma, but not with altered CD4+T cell frequency. High ILT4 level combined with low CD8+T cell/high Treg density predicted markedly poorer clinical outcomes compared with any of these biomarkers alone.ConclusionsTumor cell-derived ILT4 is correlated with immunosuppressive T cell subset infiltration and poor clinical outcomes, and might be a potential immunotherapeutic target and prognostic biomarker for LUAD patients. Combined ILT4 expression and CD8+ T cell/Treg frequency in tumor infiltrating lymphocytes (TILs) are stronger predictors for patient outcomes.

The Effect of Statins on the Functionality of CD4+CD25+FOXP3+ Regulatory T-cells in Acute Coronary Syndrome: A Systematic Review and Meta-analysis of Randomised Controlled Trials in Asian Populations.

Acute coronary syndrome (ACS) is characterised by increased effector cells and decreased regulatory T-cells (Tregs). Statins have been shown to be clinically beneficial in ACS patients. This effect could be mediated via the induction of Tregs in ACS patients. The aim of this systemic review and meta-analysis was to evaluate whether statin therapy enhances the frequency of Tregs determined by CD4+CD25+FOXP3+ in this subset of patients. A comprehensive search of PubMed and Embase was performed. Studies were restricted to randomised controlled trials that quantified CD4+CD25+FOXP3+ cell frequency by flow cytometric analysis before and after statin treatment in adults diagnosed with ACS. A minimum of at least two of the conventional markers to identify Tregs was compulsory. Four randomised controlled trials studies (439 participants) were included, all with low-to-moderate risk of bias. Pooled data showed a significant increase in Treg frequency after statin therapy in ACS patients. A further meta-regression and subgroup analysis also showed a negative dose-related effect, and a statin type-related effect (rosuvastatin versus atorvastatin), respectively. The results confirmed that statins positively alter the frequency of Tregs, which may indicate a potential mechanism of their therapeutic effect. However, there was a risk of information bias due to the markers used to identify Tregs, which was not fully explored, therefore, further randomised controlled trials should utilise markers of Tregs, such as the FOXP3 locus (Treg-specific demethylated region), for identification.

Alemtuzumab in Multiple Sclerosis: Short- and Long-Term Effects of Immunodepletion on the Peripheral Treg Compartment.

Treatment with alemtuzumab is followed by an early increase in Treg frequencies. Whether naïve and memory subsets are differentially affected and how depletion influences dysfunctional MS-Treg is unclear. In this study, we analyzed the effect of alemtuzumab on regulatory T-cells (Treg) in patients with multiple sclerosis (MS). For this purpose 182 blood samples from 25 MS patients were taken shortly before treatment and serially for up to 24 months after two alemtuzumab cycles. We studied Treg by flow cytometry (quantitation, phenotypical characterization), real-time polymerase chain reaction (T-cell receptor (TCR) excision circles [TREC] content), CDR3-spectratyping (clonal distribution), and proliferation assays (suppressive function). CD52-mediated cytolysis of Treg and conventional T-cells was determined by a complement-dependent cytolysis assay. Our studies revealed that 1 week post-alemtuzumab, Treg were depicted at constant frequencies among CD4+ T-cells. In contrast, Treg frequencies were massively increased at month 1. Post-depletional Treg exhibited a CD45RO+ memory phenotype, a skewed TCR repertoire, and contained minimum TREC numbers. Naïve Treg, thymic markers, and TCR-variability commenced to rise after 6 months but did not attain baseline levels. In vitro, Treg exhibited higher susceptibility to lysis than Tcon. Treg suppressive function constantly increased within 1 year when co-cultured with syngeneic T-cells, but remained stable against allogeneic T-cells from normal donors. Our findings suggest that (1) Treg are not spared from alemtuzumab-mediated depletion and thymopoiesis does not considerably contribute to long-term recovery, (2) either homeostatic proliferation and/or conversion from residual Tcon contributes to Treg expansion during the early post-treatment phase (3) the enhanced inhibitory effect of Treg following alemtuzumab is due to altered composition and reactivity of post-depletional Tcon.

The co-inhibitory molecule PD-L1 contributes to regulatory T cell-mediated protection in murine crescentic glomerulonephritis

Immune-mediated glomerular diseases like crescentic glomerulonephritis (cGN) are driven by inappropriately regulated cellular and humoral immune responses subsequently leading to renal tissue injury. Recent studies demonstrated the crucial role for regulatory T cells (Tregs) in suppressing pathogenic T-cell responses during nephrotoxic nephritis (NTN), a murine model of cGN. However, mechanisms of immune regulation in cGN are less clear. Here, we aim at investigating the role of the co-inhibitory PD-1/PD-L1 pathway in Treg-mediated suppression of renal inflammation. We demonstrated that Foxp3+ Tregs expressing PD-L1 infiltrate the kidney during NTN. Inhibition of PD-L1 signalling by using PD-L1−/− mice or by blockage of PD-L1 in wildtype mice resulted in an increased Treg frequency in the inflamed kidney. However, mice lacking PD-L1 developed more severe NTN associated with an elevated pathogenic renal Th1 immune response, which was reversed by blockage of IFNγ in these mice. Interestingly, lack of PD-L1 altered the gene expression profile of Tregs in homeostasis and kidney inflammation. Functionally, Tregs from nephritic PD-L1−/− mice had impaired suppressive capacity in vitro and failed to protect from NTN in vivo. Thus, PD-L1 displays a protective role in NTN, which is related to Treg-mediated suppression of the Th1 immune response.

FOXP3+ Regulatory T Cell Compartment Is Altered in Children With Newly Diagnosed Type 1 Diabetes but Not in Autoantibody-Positive at-Risk Children.

The dysfunction of FOXP3-positive regulatory T cells (Tregs) plays a key role in the pathogenesis of autoimmune diseases, including type 1 diabetes (T1D). However, previous studies analyzing the peripheral blood Treg compartment in patients with T1D have yielded partially conflicting results. Moreover, the phenotypic complexity of peripheral blood Tregs during the development of human T1D has not been comprehensively analyzed. Here, we used multi-color flow cytometry to analyze the frequency of distinct Treg subsets in blood samples from a large cohort comprising of 74 children with newly diagnosed T1D, 76 autoantibody-positive children at-risk for T1D and 180 age- and HLA-matched control children. The frequency of CD4+CD25+CD127lowFOXP3+ Tregs was higher in children with T1D compared to control children, and this change was attributable to a higher proportion of naïve Tregs in these subjects. Further longitudinal analyses demonstrated that the increase in Treg frequency correlated with disease onset. The frequencies of the minor subsets of CD25+FOXP3low memory Tregs as well as CD25lowCD127lowFOXP3+ Tregs were also increased in children with T1D. Moreover, the ratio of CCR6-CXCR3+ and CCR6+CXCR3- memory Tregs was altered and the frequency of proliferating Ki67-positive and IFN-γ producing memory Tregs was decreased in children with T1D. The frequency of CXCR5+FOXP3+ circulating follicular T regulatory cells was not altered in children with T1D. Importantly, none of the alterations observed in children with T1D were observed in autoantibody-positive at-risk children. In conclusion, our study reveals multiple alterations in the peripheral blood Treg compartment at the diagnosis of T1D that appear not to be features of early islet autoimmunity.