Function Of Treg Cells Research Articles

A combination therapy of ingredients from TCM for the prevention and treatment of gastric cancer by targeting HER2/ PD-L1

Gastric cancer (GC) is a globally important disease. Targeted therapies licensed to treat gastric cancer include trastuzumab, which targets HER2, and nivolumab or pembrolizumab, which block PD-1/PD-L1. The main challenge of tumor molecule-targeted drugs is resistance. Based on the multicomponent and multitarget characteristics of Traditional Chinese Medicine (TCM) and our previous research on Guiqi Baizhu Prescription (GQBZP) in targeting HER2/PD-L1, we used different cell models to verify the effect of quercetin targeting of HER2 and formononetin targeting of PD-L1. The results show that quercetin can inhibit MKN-45 cell proliferation by targeting HER2 to inhibit the PI3K-AKT pathway, and formononetin can regulate T-cell function by inhibiting the PD-1/PD-L1 pathway. We also used GC MKN-45 cells in a T lymphocyte co-culture model to show that quercetin and formononetin co-treatment can regulate T cell function and inhibit MKN-45 cell proliferation. More importantly, quercetin with formononetin treatment had a greater effect than treatment with either quercetin or formononetin alone. We believe that quercetin and formononetin may be useful to target HER2 and inhibit PD-1/PD-L1 in GC.

Assessment of the Impact of Cytokines on TFH, TREG, and TFR Cell Populations After Influenza Infection.

Within the last several years, great strides have been made in understanding the molecular and cellular mechanisms that control the generation of T follicular helper (TFH), T regulatory (TREG), and T follicular regulatory (TFR) cells. As a result, it is now clear that cytokines play a critical role in regulating the development and function of these CD4+ T cell subsets. One of the critical limitations when studying the effect of individual cytokines in these populations is differentiating between the intrinsic and extrinsic effects of these cytokines in vivo. Here we describe how to utilize mixed bone marrow chimeras in combination with MHC class II tetramers to characterize the direct role played by cytokines on controlling the development, function, and maintenance of TFH, TREG, and TFR cells in vivo.

Th17/Treg cells regulated by interleukin 6 in the pathogenesis of chronic rhinosinusitis with nasal polyps.

To be investigated whether Th17/Treg cells regulated by Interleukin-6 in the pathogenesis of chronic rhinosinusitis with nasal polyps. The distributions of Th17 and Treg cells in peripheral blood mononuclear cells (PBMCs) and tissues got from 46 CRSwNP patients and 14 controls were evaluated. Th17 and Treg cells and cells-related cytokines in serum were assessed in means of cytometric bead array (CBA) multiplex assays and enzyme-linked immunosorbent assays (ELISAs). Spleen cells were isolated from spleen of 20 normal BALB/c mice (male), isolated and purified with CD4 antibody immunomagnetic bead kit. CD4+ cells were divided into three groups, including TGF-β1, TGF-β1+ IL-6 and control (PBS). Treg and Th17 cells and cells related cytokines were detected by flow cytometry (FCM) after collecting spleen cells. The level of IL-10 and IL-17 in supernatant was measured by ELISA. Th17/Treg ratio and the level of IL-6 in both ECRSwNP (P < 0.05) and non-ECRSwNP (P < 0.05) were significantly increased when compared with control group, these were consistent with the previous findings. Experiments in vitro suggested that the level of Th17 cells in IL-6+ TGF-β1 group was significantly increased than TGF-β1 group and control group (P < 0.05). The ratio of cells expressed RoRγt in IL-6+ TGF-β1 group was much higher than TGF-β1 group (P < 0.05). IL-6 might regulate the function of Th17 and Treg cells and the Th17/Treg ratio and have a role in the pathogenesis of CRSwNP.

MiR-146a enhances regulatory T-cell differentiation and function in allergic rhinitis by targeting STAT5b.

MicroRNA (miR)-146a, as an important immune regulatory factor with an anti-inflammatory effect, plays a crucial role in regulatory T-cell (Tregs) differentiation and function in allergic rhinitis (AR). The present study aimed to investigate the regulatory mechanism employed by miR-146a to control Treg differentiation and function in AR. Expression of miR-146a and STAT5b in peripheral blood mononuclear cells (PBMCs) and nasal mucosa from patients with AR was detected by qPCR and Western blotting. Tregs were quantified by flow cytometry in miR-146a knockdown or STAT5b knockdown PBMCs. FOXP3, IL-10, and TGF-β levels were detected by Western blotting or ELISA in miR-146a knockdown or STAT5b overexpressing PBMCs, as well as in STAT5b knockdown PBMCs overexpressing miR-146a. The effect of miR-146a on STAT5b was observed by luciferase assay and knockdown experiments. Levels of miR146a and STAT5b in the nasal mucosa or PBMCs were significantly lower in the AR group than in the control group. There were significantly fewer Tregs in miR-146a knockdown or STAT5b knockdown PBMCs compared to control PBMCs. Expression of FOXP3, IL-10, and TGF-β was decreased in the miR-146a knockdown group but increased in the STAT5b overexpression group. In contrast, miR-146a overexpression increased the levels of these factors, but knockdown of STAT5b significantly inhibited this effect. Luciferase assay and knockdown experiments showed that miR-146a bound directly to STAT5b. miR-146a enhances Treg differentiation and function in AR by positively targeting STAT5b.

Role of FoxP3-positive regulatory T-cells in regressive and progressive cervical dysplasia.

Forkhead Box Protein 3 (FoxP3) is known as a key mediator in the immunosuppressive function of regulatory T-cells (Tregs). The aim of our study was to investigate whether FoxP3-positive Tregs have the potential to act as an independent predictor in progression as well as in regression of cervical intraepithelial neoplasia, especially in patients with intermediate cervical intraepithelial neoplasia (CIN II). Nuclear FoxP3 expression was immunohistochemically analysed in 169 patient samples (CIN I, CIN II with regressive course, CIN II with progressive course, CIN III). The median numbers were calculated for each slide and correlated with the histological CIN grade. Statistical analysis was performed by SPSS 26 (Mann-Whitney U test, Spearman's rank correlation). An increased FoxP3 expression in CIN II with progression could be detected in comparison to CIN II with regression (p = 0.003). Total FoxP3 expression (epithelium and dysplasia-connected stroma) was higher in more advanced CIN grades (p < 0.001 for CIN I vs. CIN II; p = 0.227 for CIN II vs. CIN III). A positive correlation could be detected between FoxP3-positive cells in epithelium and total FoxP3 expression (Spearman's Rho: 0,565; p < 0.01). Expression of FoxP3 could be a helpful predictive factor to assess the risks of CIN II progression. As a prognosticator for regression and progression in cervical intraepithelial lesions it might thereby help in the decision process regarding surgical treatment vs. watchful waiting strategy to prevent conisation-associated risks for patients in child-bearing age. In addition, the findings support the potential of Tregs as a target for immune therapy in cervical cancer patients.

The role of regulatory T cells on the activation of astrocytes in the brain of high-fat diet mice following lead exposure

Lead (Pb) exposure can cause damage to the central nervous system (CNS)*. Pb can accumulate in the hippocampus, leading to learning and memory impairments. Recent studies have shown that high-fat diet (HFD) is also associated with cognitive impairment. However, there are few reports on CNS damage due to HFD and Pb exposure. We aimed to investigate the effect of Pb on cognitive functions of HFD-fed mice, focusing on the role of regulatory T (Treg) cells in astrocyte activation. C57BL/6J mice were randomly divided into control, HFD, Pb, and HFD + Pb groups. TGF-β and IL-10 secreted by Treg cells and the intracellular transcription factor Foxp3 were evaluated as a measure of Treg cell function; astrocyte activation was assessed by evaluating glial fibrillary acidic protein (GFAP) expression. The learning and memory ability was significantly lower in the HFD + Pb group than in other groups. The brain Treg cell ratio was significantly decreased and the protein levels of TGF-β, IL-10, and Foxp3 were significantly lower, whereas the protein level of GFAP was higher in the HFD + Pb group. The hippocampus of the HFD + Pb group mice showed significantly higher levels of neurotoxic reactive astrocyte markers and astrogliosis was also much higher compared to HFD and Pb groups. Furthermore, all-trans retinoic acid treatment increased the brain Treg cell ratio, reversed cognitive decline, and suppressed astrocyte activation in the HFD + Pb group mice. We concluded that HFD along with Pb exposure could aggravate the activation of astrocytes in the brain, and the brain Treg cells may be involved in inhibiting astrocyte activation in HFD-fed mice exposed to Pb.

The Effect of Orally Administering Forkhead Box P3 Recombinant Lactobacillus plantarum on Regulatory T-cell Functionality in a Humanized Mouse Model for Rheumatoid Arthritis: A Research Protocol

Introduction: Inflammation and joint stiffening are common symptoms of rheumatoid arthritis (RA), an autoimmune inflammatory disease. Previous treatments of RA have focused on decreasing symptomatic effects but have limited effects on disease progression. In RA, an influx of pro-inflammatory cytokines occurs at the synovium, which is the soft tissue surrounding the joints. The production of pro-inflammatory cytokines is controlled by regulatory T-cells, which have a deficit in function in RA patients. Regulatory T-cell development and function is regulated by the forkhead box P3 (FOXP3). The FOXP3 gene is a viable therapeutic target to restore regulatory T-cell functionality because FOXP3 is underexpressed in RA patients. Therefore, this study ventures to treat RA regulatory T-cell functionality by increasing FOXP3 gene expression through FOXP3 recombinant Lactobacillus plantarum bactofection. We hypothesize that bactofection will lead to a decrease in RA progression by restoring normal function in regulatory T-cells, thus decreasing inflammation. Methods: We propose a study using severe combined immunodeficient mouse models engrafted with human RA synovium. The mice will be given either no treatment (control group) or a 2×109 CFU/g dose of recombinant Lactobacillus plantarum strain. The mice will be sacrificed after 0 days, 10 days, 20 days, and 30 days (control group and treatment groups respectively). Synovial tissue samples will be obtained from the hip joints. Through immunofluorescence and western blotting, the prevalence of FOXP3, regulatory T-cells and pro-inflammatory cytokines such as tumor necrosis factor-alpha, Interleukin-1 and Interleukin-6 will be compared between the control and treatment groups. For statistical analysis, a one-way MANOVA test, Levene’s test, and a Shapiro-Wilk test will be performed using GraphPad Prism. Results: As a result of bactofection, there will be an increase in FOXP3 and regulatory T-cells, resulting in a decrease of pro-inflammatory cytokines. Discussion: Analysis of mice treated with recombinant Lactobacillus plantarum compared to mice with no treatment will set a correlation between FOXP3, regulatory T-cells, and pro-inflammatory cytokines prevalence and RA progression after treatment. Conclusion: The findings of this study will provide evidence that bactofection is a viable treatment for RA, and may be more effective than conventional treatments.

Activation of AMPKα1 is essential for regulatory T cell function and autoimmune liver disease prevention.

Regulatory T cells (Treg cells) are crucial for maintaining immune tolerance. Compromising the regulatory function of Treg cells can lead to autoimmune liver disease. However, how Treg cell function is regulated has not been fully clarified. Here, we report that mice with AMP-activated protein kinase alpha 1 (AMPKα1) globally knocked out spontaneously develop immune-mediated liver injury, with massive lymphocyte infiltration in the liver, elevated serum alanine aminotransferase levels, and greater production of autoantibodies. Both transplantation of wild-type bone marrow and adoptive transfer of wild-type Treg cells can prevent liver injury in AMPKα1-KO mice. In addition, Treg cell-specific AMPKα1-KO mice display histological features similar to those associated with autoimmune liver disease, greater production of autoantibodies, and hyperactivation of CD4+ T cells. AMPKα1 deficiency significantly impairs Treg cell suppressive function but does not affect Treg cell differentiation or proliferation. Furthermore, AMPK is activated upon T cell receptor (TCR) stimulation, which triggers Foxp3 phosphorylation, suppressing Foxp3 ubiquitination and proteasomal degradation. Importantly, the frequency of Treg cells and the phosphorylation levels of AMPK at T172 in circulating blood are significantly lower in patients with autoimmune liver diseases. Conclusion: Our data suggest that AMPK maintains the immunosuppressive function of Treg cells and confers protection against autoimmune liver disease.

Interleukin-2 and regulatory T cells in rheumatic diseases.

Failure of regulatory T (Treg) cells to properly control immune responses leads invariably to autoimmunity and organ damage. Decreased numbers or impaired function of Treg cells, especially in the context of inflammation, has been documented in many human autoimmune diseases. Restoration of Treg cell fitness and/or expansion of their numbers using low-dose natural IL-2, the main cytokine driving Treg cell survival and function, has demonstrated clinical efficacy in early clinical trials. Genetically modified IL-2 with an extended half-life and increased selectivity for Treg cells is now in clinical development. Administration of IL-2 combined with therapies targeting other pathways involved in the expression of autoimmune diseases should further enhance its therapeutic potential. Ongoing clinical efforts that capitalize on the early clinical success of IL-2 treatment should bring the use of this cytokine to the forefront of biological treatments for autoimmune diseases.

DNAM-1 versus TIGIT: competitive roles in tumor immunity and inflammatory responses.

The co-stimulatory and co-inhibitory immunoreceptors, DNAX accessory molecule-1 (DNAM-1) and T-cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibitory motif (ITIM) domain (TIGIT), are paired activating and inhibitory receptors on T cells and natural killer (NK) cells. They share the ligands poliovirus receptor (PVR, CD155) and its family member nectin-2 (CD112), which are highly expressed on antigen-presenting cells (APCs), tumors and virus-infected cells. Upon ligation with the ligands, DNAM-1 and TIGIT show reciprocal functions; whereas DNAM-1 promotes activation, proliferation, cytokine production and cytotoxic activity in effector lymphocytes, including CD4+ T-helper cells, CD8+ cytotoxic T lymphocytes and NK cells, TIGIT inhibits these DNAM-1 functions. On the other hand, DNAM-1 competes with TIGIT on regulatory T (Treg) cells in binding to CD155 and therefore regulates TIGIT signaling to down-regulate Treg cell function. Thus, whereas DNAM-1 enhances anti-tumor immunity and inflammatory responses by augmenting effector lymphocyte function and suppressing Treg cell function, TIGIT reciprocally suppresses these immune responses by suppressing effector lymphocyte function and augmenting Treg cell function. Thus, blockade of DNAM-1 and TIGIT function would be potential therapeutic approaches for patients with inflammatory diseases and those with cancers and virus infection, respectively.

Synergistic and additive interactions between receptor signaling networks drive the regulatory T cell versus T helper 17 cell fate choice

CD4+ T cells differentiate into subsets that promote immunity or minimize damage to the host. T helper 17 cells (Th17) are effector cells that function in inflammatory responses. T regulatory cells (Tregs) maintain tolerance and prevent autoimmunity by secreting immunosuppressive cytokines and expressing check point receptors. While the functions of Th17 and Treg cells are different, both cell fate trajectories require T cell receptor (TCR) and TGF-β receptor (TGF-βR) signals, and Th17 polarization requires an additional IL-6 receptor (IL-6R) signal. Utilizing high-resolution phosphoproteomics, we identified that both synergistic and additive interactions between TCR, TGF-βR, and IL-6R shape kinase signaling networks to differentially regulate key pathways during the early phase of Treg versus Th17 induction. Quantitative biochemical analysis revealed that CD4+ T cells integrate receptor signals via SMAD3, which is a mediator of TGF-βR signaling. Treg induction potentiates the formation of the canonical SMAD3/4 trimer to activate a negative feedback loop through kinases PKA and CSK to suppress TCR signaling, phosphatidylinositol metabolism, and mTOR signaling. IL-6R signaling activates STAT3 to bind SMAD3 and block formation of the SMAD3/4 trimer during the early phase of Th17 induction, which leads to elevated TCR and PI3K signaling. These data provide a biochemical mechanism by which CD4+ T cells integrate TCR, TGF-β, and IL-6 signals via generation of alternate SMAD3 complexes that control the development of early signaling networks to potentiate the choice of Treg versus Th17 cell fate.

Intranuclear Delivery of HIF-1α-TMD Alleviates EAE via Functional Conversion of TH17 Cells

T helper 17 (TH17) cells are involved in several autoimmune diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA). In addition to retinoic acid receptor-related orphan nuclear receptor gamma t (ROR-γt), hypoxia-inducible factor-1α (HIF-1α) is essential for the differentiation and inflammatory function of TH17 cells. To investigate the roles of HIF-1α in the functional regulation of TH17 cells under the normal physiological condition without genetic modification, the nucleus-transducible form of transcription modulation domain (TMD) of HIF-1α (ntHIF-1α-TMD) was generated by conjugating HIF-1α-TMD to Hph-1 protein transduction domain (PTD). ntHIF-1α-TMD was effectively delivered into the nucleus of T cells without cellular cytotoxicity. ntHIF-1α-TMD significantly blocked the differentiation of naïve T cells into TH17 cells in a dose-dependent manner via IL-17A and ROR-γt expression inhibition. However, T-cell activation events such as induction of CD69, CD25, and IL-2 and the differentiation potential of naïve T cells into TH1, TH2, or Treg cells were not affected by ntHIF-1α-TMD. Interestingly, TH17 cells differentiated from naïve T cells in the presence of ntHIF-1α-TMD showed a substantial level of suppressive activity toward the activated T cells, and the increase of Foxp3 and IL-10 expression was detected in these TH17 cells. When mRNA expression pattern was compared between TH17 cells and ntHIF-1α-TMD-treated TH17 cells, the expression of the genes involved in the differentiation and functions of TH17 cells was downregulated, and that of the genes necessary for immune-suppressive functions of Treg cells was upregulated. When the mice with experimental autoimmune encephalomyelitis (EAE) were treated with ntHIF-1α-TMD with anti-IL-17A mAb as a positive control, the therapeutic efficacy of ntHIF-1α-TMD in vivo was comparable with that of anti-IL-17A mAb, and ntHIF-1α-TMD-mediated therapeutic effect was contributed by the functional conversion of TH17 cells into immune-suppressive T cells. The results in this study demonstrate that ntHIF-1α-TMD can be a new therapeutic reagent for the treatment of various autoimmune diseases in which TH17 cells are dominant and pathogenic T cells.

Probiotics and prebiotics: potential prevention and therapeutic target for nutritional management of COVID-19?

Scientists are working to identify prevention/treatment methods and clinical outcomes of coronavirus disease 2019 (COVID-19). Nutritional status and diet have a major impact on the COVID-19 disease process, mainly because of the bidirectional interaction between gut microbiota and lung, that is, the gut-lung axis. Individuals with inadequate nutritional status have a pre-existing imbalance in the gut microbiota and immunity as seen in obesity, diabetes, hypertension and other chronic diseases. Communication between the gut microbiota and lungs or other organs and systems may trigger worse clinical outcomes in viral respiratory infections. Thus, this review addresses new insights into the use of probiotics and prebiotics as a preventive nutritional strategy in managing respiratory infections such as COVID-19 and highlighting their anti-inflammatory effects against the main signs and symptoms associated with COVID-19. Literature search was performed through PubMed, Cochrane Library, Scopus and Web of Science databases; relevant clinical articles were included. Significant randomised clinical trials suggest that specific probiotics and/or prebiotics reduce diarrhoea, abdominal pain, vomiting, headache, cough, sore throat, fever, and viral infection complications such as acute respiratory distress syndrome. These beneficial effects are linked with modulation of the microbiota, products of microbial metabolism with antiviral activity, and immune-regulatory properties of specific probiotics and prebiotics through Treg cell production and function. There is a need to conduct clinical and pre-clinical trials to assess the combined effect of consuming these components and undergoing current therapies for COVID-19.

Nanocurcumin supplementation ameliorates Behcet’s disease by modulating regulatory T cells: A randomized, double‐blind, placebo‐controlled trial

Current research was designed to assess the effects of nanocurcumin supplementation on regulatory T (Treg) cells frequency and function in Behçet's disease (BD). In this randomized double-masked, placebo-controlled trial, 36 BD subjects were randomly put into two groups to take one 80 mg nanocurcumin capsule or placebo daily for 8 weeks. Before and after trial, disease activity, Treg cells frequency and expression of related immunologic parameters including forkhead box protein P3 (Foxp3) transcription factor messenger RNA (mRNA) and microRNAs (miRNAs) such as miRNA-25 and miRNA-106b as well as cytokines including transforming growth factor (TGF)-β and interleukin (IL)-10 were studied. Thirty-two patients (17 in the nanocurcumin and 15 in the placebo groups) completed the trial. Treg cells frequency increased significantly in the nanocurcumin group compared with baseline (P < 0.001) and placebo group (P < 0.001). Moreover, FoxP3, TGF-β, IL‐10, miRNA-25, and miRNA-106b mRNA expression levels increased considerably in the nanocurcumin group compared to baseline (P < 0.001) and placebo group (P < 0.001, P < 0.001, P = 0.025, P = 0.011, and P < 0.001, respectively). Significant increases in serum TGF-β and IL-10 were seen in nanocurcumin group compared with baseline (P < 0.001) and placebo group (P = 0.001 and P < 0.001, respectively). Significant decrease in disease activity was found in nanocurcumin group compared with placebo group (P = 0.044). Our study provided a promising view for desirable effects of nanocurcumin supplementation in improving immunological parameters and disease activity in BD.

The mechanism of rapamycin in promoting asthmatic regulatory T cell differentiation and function.

To investigate the mechanism of rapamycin in promoting asthmatic regulatory T cell differentiation . Asthma model was prepared by sensitization and challenge of ovalbumin in mice. Spleen CD4CD25 T cells were sorted from the asthmatic mice and normal mice by ultrahigh speed flow cytometer, and divided into three groups. Transforming growth factor-β and interleukin-2, or combined with rapamycin (final concentration of 500 nmol/L) were given in the model group or the rapamycin group. The levels of Treg cells and CD4CD25 T cells were detected by flow cytometry. The phosphorylation level of downstream proteins of S6 and Akt in the mTORC1/2 signaling pathway were examined by Western blotting. Compared with the model group, the differentiation level of Treg cells in the rapamycin group was significantly increased, the proliferation level of CD4CD25 T cells was decreased, and the phosphorylations of the mTORC1/2 substrates, S6 protein and Akt were decreased （all <0.05）. Rapamycin can promote the differentiation and function of Treg cells via inhibition of the mTORC1/2 signaling pathway.

Prostaglandin E2 directly inhibits the conversion of inducible regulatory T cells through EP2 and EP4 receptors via antagonizing TGF-β signalling.

Regulatory T (Treg) cells are essential for control of inflammatory processes by suppressing effector T‐cell functions. The actions of PGE2 on the development and function of Treg cells, particularly under inflammatory conditions, are debated. In this study, we employed pharmacological and genetic approaches to examine whether PGE2 had a direct action on T cells to modulate de novo differentiation of Treg cells. We found that TGF‐β‐induced Foxp3 expression and iTreg cell differentiation in vitro is markedly inhibited by PGE2, which was mediated by the receptors EP2 and EP4. Mechanistically, PGE2‐EP2/EP4 signalling interrupts TGF‐β signalling during iTreg differentiation. Moreover, EP4 deficiency in T cells impaired iTreg cell differentiation in vivo. Thus, our results demonstrate that PGE2 negatively regulates iTreg cell differentiation through a direct action on T cells, highlighting the potential for selectively targeting the PGE2‐EP2/EP4 pathway to control T cell‐mediated inflammation.

Study on the preparation and function of regulatory T cells from human peripheral blood.

BackgroundRegulatory T cells (Tregs) are an important cell subgroup of CD4+ T cells. Treg cells are critically involved in inducing immune tolerance, maintaining immune environment homeostasis, and preventing the occurrence of autoimmune diseases. Under normal conditions, the number of Tregs in the body is very small. This research was designed to establish an effective method to expand human peripheral blood Tregs in vitro and to analyze phenotype, purity, and function of Treg cells post-expansion.MethodsPeripheral blood was obtained from healthy donors. CD4+CD25+CD127dim/− Treg cells were isolated from peripheral blood mononuclear cells (PBMCs) by magnetic-activated cell sorting (MACS), and an optimized culture system was used for amplification. The in vitro amplification ability of Treg cells was evaluated to determine the expression and purity of Treg cell-specific surface markers in different culture cycles. The suppressive function of Treg was determined by in vitro lymphocyte proliferation assay.ResultsTreg cells could be successfully isolated by magnetic activated cell sorting (MACS). After 21 days of in vitro culture, the mean expansion fold was 2,009±452.2 in ≤60 years, and there was a significant difference between the younger group and the older than 60 years group (1,238±330.0). Flow cytometry analysis revealed that the percentages of CD4+CD25+ cells and FOXP3+ cells were (93.25±3.05)% and (94.19±4.21)% on day 14, and (92.86±4.36)% and (91.55±5.62)% on day 21, respectively. In addition, the proportions of CD8+ T, CD19+ B, CD3−CD56+ natural killer cell (NK), and CD3+ CD56+ natural killer T cell (NKT) were extremely low. Lymphocyte proliferation assay demonstrated that Tregs could inhibit the proliferation of CD8+ T cells more effectively than that of CD4+ T cells. Furthermore, the suppressive capacity of Tregs was correlated with Treg-to-PBMCs ratios.ConclusionsWe successfully established a technical protocol for manufacturing a large quantity of Tregs with high efficiency in vitro. The expanded Tregs have a steady FOXP3 expression and exhibited a potent immune suppression, which might have great significance in adoptive Treg therapy for treating graft-versus-host disease and autoimmune diseases.

Macrophages Control the Bioavailability of Vitamin D and Vitamin D-Regulated T Cell Responses.

The active form of vitamin D3 (1,25(OH)2D3) has a great impact on T cell effector function. Thus, 1,25(OH)2D3 promotes T helper 2 (Th2) and regulatory T (Treg) cell function and concomitantly inhibits Th1 and Th17 cell function. Thus, it is believed that vitamin D exerts anti-inflammatory effects. However, vitamin D binding protein (DBP) strongly binds both 1,25(OH)2D3 and the precursor 25(OH)D3, leaving only a minor fraction of vitamin D in the free, bioavailable form. Accordingly, DBP in physiological concentrations would be expected to block the effect of vitamin D on T cells and dendritic cells. In the present study, we show that pro-inflammatory, monocyte-derived M1 macrophages express very high levels of the 25(OH)D-1α-hydroxylase CYP27B1 that enables them to convert 25(OH)D3 into 1,25(OH)2D3 even in the presence of physiological concentrations of DBP. Co-cultivation of M1 macrophages with T cells allows them to overcome the sequestering of 25(OH)D3 by DBP and to produce sufficient levels of 1,25(OH)2D3 to affect T cell effector function. This study suggests that in highly inflammatory conditions, M1 macrophages can produce sufficient levels of 1,25(OH)2D3 to modify T cell responses and thereby reduce T cell-mediated inflammation via a vitamin D-mediated negative feed-back loop.

The semaphorin 4A–neuropilin 1 axis alleviates kidney ischemia reperfusion injury by promoting the stability and function of regulatory T cells

Previous studies have suggested the role of CD4+Foxp3+ regulatory T cells (Tregs) in protection against kidney ischemia reperfusion injury via their immunosuppressive properties. Unfortunately, the associated mechanisms of Tregs in kidney ischemia reperfusion injury have not been fully elucidated. Semaphorin 4A (Sema4A) is essential for maintaining the immunosuppressive capacity of Tregs in tumors. However, whether Sema4A can alleviate kidney ischemia reperfusion injury through Tregs has not yet been demonstrated. Here, we investigated the effect and mechanism of Sema4A on the development of kidney ischemia reperfusion injury. Administration of recombinant human Sema4A-Fc chimera protein prior to ischemia reperfusion injury promoted the expansion and function of Tregs and decreased the accumulation of neutrophils and proinflammatory macrophages thereby attenuating functional and histological injury of the injured kidneys. Depletion of Tregs abrogated the protective effect of Sema4A on kidney ischemia reperfusion injury, suggesting Tregs as the main target cell type for Sema4A in the development of this injury. Mechanistically, Sema4A bound to neuropilin 1 (Nrp1), a cell surface receptor for Sema4A and other ligands and a key regulator of Tregs, which then promoted recruitment of phosphatase and tensin homologue and suppressed the Akt-mTOR pathway in Foxp3Cre mice but not in Nrp1f/fFoxp3Cre mice. Consistently, Treg-specific deletion of Nrp1 blocked the effect of Sema4A on the expansion and function of Treg cells. Thus, our results demonstrate that the Sema4A-Nrp1 axis alleviates the development of ischemia reperfusion injury by promoting the stability and function of Tregs in mouse kidneys.

Effects of α1-adrenoreceptor on Treg cell function in collagen-induced arthritis

Objective: An animal model of collagen-induced arthritis (CIA) was used to investigate the effects of norepinephrine (NE) and α1-adrenoreceptor (α1-AR) on Treg cells in CIA. Methods: Thirty-two male DBA/1 mice were randomly divided into control group and CIA model group. CIA was prepared by intradermal injection of collagen type II (CII, 100 μl) at the tail base of DBA/1 mice. On the 41th day following primary immunization, co-expression of CD4+T and α1-AR in mouse spleens was observed. Protein expressions of α1-AR in the ankle joints and the spleens of mice were measured by Western blot analysis. The CD4+ T cells were isolated from the mouse spleen tissues in CIA mice and treated with NE or α1-AR agonist phenylephrine. Percentage of Treg cells in mouse CD4+ T cells of CIA mice was determined by flow cytometry. Expressions of α1-AR, transforming growth factor-β (TGF-β) and IL-10 in CD4+T cells of CIA mice were assessed by Western blot. Results: Co-expression of CD4 and α1-AR was observed in spleens of both intact and CIA mice. Compared with intace mice, α1-AR expressions in the ankle joints and spleens were down-regulated in CIA mice. NE increased the function of Treg cells in CD4+ T cells of CIA mice compared with that of nothing-treated CD4+T cells of CIA mice. Moreover, the α1-AR agonist phenylephrine increased the Treg cell function in CD4+ T cells of CIA mice relative to that of nothing-treated CD4+T cells of CIA mice. Conclusion: Activating α1-AR on CD4+T cells of CIA mice enhances Treg cell function,facilitating a shift of CD4+T cells toward Treg polarization.