Intestine-derived Clostridium leptum induces murine tolerogenic dendritic cells and regulatory T cells in vitro

Patients with autoimmune and allergic diseases frequently present with reduced numbers and functionally impaired regulatory T cells (Tregs) and/or tolerogenic dendritic cells (tDCs). tDC-mediated regulation of Treg proliferation (numbers) and activation is crucial to establishing and maintaining an appropriate level of immune tolerance. Colonic colonization of Clostridium spp. is associated with accumulation of Tregs, which inhibits development of inflammatory lesions. To investigate whether infection with the Clostridium leptum sp. can specifically induce Tregs and/or tDCs bone marrow-derived dendritic cells were cultured in the presence or absence of C. leptum then co-cultured with CD4⁺CD25⁻ T cells or not. Changes in tDC numbers, Treg numbers, percentages of T cell subsets, and expression of cytokines related to Tregs (IL-10 and transforming growth factor-beta (TGF-β1)), DCs (IL-12p40 and IL-6) and effector T cells (IFN-γ, IL-4, IL-5, IL-13, and IL-17A) were measured. In the co-culture system, C. leptum-stimulated tDCs were able to increase the percentage and total number of Tregs attenuate activation of T helper cells (Th1, Th2, and Th17), and decrease the amount of secreted IL-4, IL-5, IL-13, IFN-γ and IL-17A. Thus, C. leptum exposure can induce the tDC-mediated stimulation of Tregs while disrupting the immune inflammatory response mediated by Th1, Th2 and Th17 cells.     

Changes in T helper cell-related factors in patients with type 2 diabetes mellitus after empagliflozin therapy

The immune factors related to T helper (Th) 1 (T-bet, STAT1, and IFN-γ), Th17 (ROR-γt, STAT3, and IL-17), and Treg (FOXP3, STAT5, and IL-10) cells, and SOCS1/3 and the proliferation of Th cells were investigated in type 2 diabetes mellitus patients before (baseline) and after empagliflozin therapy.A total of 56 patients on metformin and gliclazide were separated into two groups: Group 1 did not receive empagliflozin (EMPA^?) and the Group 2 received 10 mg/day of empagliflozin for 6 months (EMPA⁺). The expressions of T-bet, ROR-γt, FOXP3, STAT1/3/5 and SOCS1/3 were evaluated in CD4⁺ T cells with real-time PCR. The production of IFN-γ, IL-17, and IL-10 from CD4⁺ T cells was measured using ELISA. The proliferation of Th cells was assessed with flow cytometry.Six months of empagliflozin therapy significantly reduced the expression of ROR-γt and increased FOXP3 and STAT5 expression, compared to baseline. Production of IL-17 decreased after empagliflozin treatment, while IL-10 was enhanced in the EMPA⁺ group. Oral administration of empagliflozin or the addition of empagliflozin to the cell cultures diminished the proliferation of Th cells.Empagliflozin showed anti-inflammatory effects on Th cells by decreasing Th17-related factors, reducing proliferation capacity, and increasing Treg cell properties.     

Liver X receptor regulates Th17 and RORγt+ Treg cells by distinct mechanisms

The gastrointestinal microenvironment, dominated by dietary compounds and the commensal bacteria, is a major driver of intestinal CD4⁺ T helper (Th) cell differentiation. Dietary compounds can be sensed by nuclear receptors (NRs) that consequently exert pleiotropic effects including immune modulation. Here, we found that under homeostatic conditions the NR Liver X receptor (LXR), a sensor of cholesterol metabolites, regulates RORγt⁺ CD4 T cells in the intestine draining mesenteric lymph node (MLN). While LXR activation led to a decrease, LXR-deficiency resulted in an increase in MLN Th17 and RORγt⁺ Tregs. Mechanistically, LXR signaling in CD11c⁺ myeloid cells was required to control RORγt⁺ Treg. By contrast, modulation of MLN Th17 was independent of LXR signaling in either immune or epithelial cells. Of note, horizontal transfer of microbiota between LXRα^−/− and WT mice was sufficient to only partially increase MLN Th17 in WT mice. Despite LXRα deficiency resulted in an increased abundance of Ruminococcaceae and Lachnospiraceae bacterial families compared to littermate controls, microbiota ablation (including SFB) was not sufficient to dampen LXRα-mediated expansion of MLN Th17. Altogether, our results suggest that LXR modulates RORγt⁺ Treg and Th17 cells in the MLN through distinct mechanisms.     

Imbalance of Th17 cells and regulatory T cells in tuberculous pleural effusion

Both T helper interleukin 17 (IL-17)-producing cells (Th17 cells) and regulatory T cells (Tregs) have been found to be increased in human tuberculous pleural effusion (TPE); however, the possible interaction between Th17 cells and Tregs in TPE remains to be elucidated. The objective of the present study was to investigate the distribution of Th17 cells in relation to Tregs, as well as the mechanism of Tregs in regulating generation and differentiation of Th17 cells in TPE. In the present study, the numbers of Th17 cells and Tregs in TPE and blood were determined by flow cytometry. The regulation and mechanism of CD39⁺ Tregs on generation and differentiation of Th17 cells were explored. Our data demonstrated that the numbers of Th17 cells and CD39⁺ Tregs were both increased in TPE compared with blood. Th17 cell numbers were correlated negatively with Tregs in TPE but not in blood. When naïve CD4⁺ T cells were cultured with CD39⁺ Tregs, Th17 cell numbers decreased as CD39⁺ Treg numbers increased, and the addition of the anti-latency-associated peptide monoclonal antibody to the coculture reversed the inhibitory effect exerted by CD39⁺ Tregs. This study shows that Th17/Treg imbalance exists in TPE and that pleural CD39⁺ Tregs inhibit generation and differentiation of Th17 cells via a latency-associated peptide-dependent mechanism.     

The role of T helper 17 and regulatory T cells in tumor microenvironment

T helper 17 (Th17) cells were first described as a novel T helper cell lineage independent from Th1 and Th2 subsets. Th17 cells play vital roles in inflammation and tumor immunity. It causes the dissipation of antitumor immunity and contribution to the survival of tumor cells, worsening tumor growth and metastasis. Tumor-infiltrating Th17 cells were seen innumerous cancers in mice and humans. There has been an association between intratumoral Th17 cell infiltration and both good and bad prognoses. Besides the protumoral roles defined for IL-17 andTh17 cells, several reports have shown that Th17 cells also drive antitumoral immunity. Various mechanisms by which Th17 cells control tumor growth are as following: recruitment of several immune cells including DCs, CD4⁺ T cells, and CD8⁺ T cells within tumors, activation of CD8⁺ T cells, and probably plasticity toward Th1 phenotype, related to IFN-γ and TNF-α production. Regulatory T cells (Tregs) have been exhibited to infiltrate human tumors and are believed to restrict antitumor immunity. The effect of Treg cells has been more controversial. Whereas some studies have proposed that a high density of Treg cells within the tumor associated with a poor clinical prognosis, other studies have presented a positive clinical prognosis, underlining the importance of elucidating the clinical significance of Treg cells further. Treg and Th17 cells play both positive and negative roles in regulating antitumor immune responses. In spite of the presence of these cells, yet some tumors develop and grow. These T cells by themselves are not adequate to efficiently mount antitumor immune responses.     

Mesenchymal Stem Cells Attenuate the Adverse Effects of Immunosuppressive Drugs on Distinct T Cell Subopulations

Immunosuppressive drugs are widely used to treat undesirable immune reaction, however their clinical use is often limited by harmful side effects. The combined application of immunosuppressive agents with mesenchymal stem cells (MSCs) offers a promising alternative approach that enables the reduction of immunosuppressive agent doses and simultaneously maintains or improves the outcome of therapy. The present study aimed to determinate the effects of immunosuppressants on individual T cell subpopulations and to investigate the efficacy of MSC-based treatment combined with immunosuppressive drugs. We tested the effect of five widely used immunosuppressants with different action mechanisms: cyclosporine A, mycophenolate mofetil, rapamycin, and two glucocorticoids - prednisone and dexamethasone in combination with MSCs on mouse CD4⁺ and CD8⁺ lymphocyte viability and activation, Th17 (RORγt⁺), Th1 (T-bet⁺), Th2 (GATA-3⁺) and Treg (Foxp3⁺) cell proportion and on the production of corresponding key cytokines (IL-17, IFNγ, IL-4 and IL-10). We showed that MSCs modulate the actions of immunosuppressants and in combination with immunosuppressive drugs display distinct effect on cell activation and balance among different T lymphocytes subpopulations and exert a suppressive effect on proinflammatory T cell subsets while promoting the functions of anti-inflammatory Treg lymphocytes. The results indicated that MSC-based therapy could be a powerful strategy to attenuate the negative effects of immunosuppressive drugs on the immune system.     

T cell-specific BLIMP-1 deficiency exacerbates experimental autoimmune encephalomyelitis in nonobese diabetic mice by increasing Th1 and Th17 cells

Recently, we demonstrated that B lymphocyte-induced maturation protein 1 (BLIMP-1) has a role in regulating the differentiation and effector function of Th1 and Th17 cells. As these cells play critical roles in the induction and pathogenesis of experimental autoimmune encephalomyelitis (EAE), we investigated the potential role of T cell BLIMP-1 in modulating MOG_35–55-induced EAE. We established T cell-specific BLIMP-1 conditional knockout (CKO) NOD mice to dissect the role of BLIMP-1 in EAE using loss-of-function model. Our results indicate that EAE severity is dramatically exacerbated in CKO mice. The numbers of CNS-infiltrating Th1, Th17, IFN-γ⁺IL-17A⁺, and IL-21⁺IL-17A⁺ CD4⁺ T cells are remarkably increased in brain and spinal cord of CKO mice. Moreover, the ratio of Tregs/effectors and IL-10 production of Tregs are significantly downregulated in CNS of CKO mice. We conclude that BLIMP-1 suppresses autoimmune encephalomyelitis via downregulating Th1 and Th17 cells and impairing Treg cells.     

Infiltration of IL-17-Producing T Cells and Treg Cells in a Mouse Model of Smoke-Induced Emphysema

Chronic obstructive pulmonary disease (COPD) is a progressive and irreversible chronic inflammatory disease associated with the accumulation of activated T cells. To date, there is little information concerning the intrinsic association among Th17, Tc17, and regulatory T (Treg) cells in COPD. The objective of this study was to investigate the variation of lungs CD4⁺Foxp3⁺ Treg cells and IL-17-producing CD4 and CD8 (Th17 and Tc17) lymphocytes in mice with cigarette-induced emphysema. Groups of mice were exposed to cigarette smoke or room air. At weeks 12 and 24, mice were sacrificed to observe histological changes by HE stain. The frequencies of Th17 (CD4⁺IL-17⁺T), Tc17 (CD8⁺IL-17⁺T), and Treg (CD4⁺Foxp3⁺T) cells in lungs from these mice were analyzed by flow cytometry. The mRNA levels of orphan nuclear receptor ROR γt and Foxp3 were performed by real-time quantitative polymerase chain reaction. The protein levels of interleukin-17 (IL-17), IL-6, IL-10, and transforming growth factor-beta (TGF-β1) were measured by enzyme-linked immunosorbent assay. Cigarette smoke caused substantial enlargement of the air spaces accompanied by the destruction of the normal alveolar architecture and led to emphysema. The frequencies of Th17 and Tc17 cells, as well as the expressions of IL-6, IL-17, TGF-β1, and ROR γt were greater in the lungs of cigarette smoke (CS)-exposed mice, particularly in the 24-week CS-exposed mice. The frequencies of Treg cells and the expressions of IL-10 and Foxp3 were lower in CS-exposed mice compared to control group. More important, the frequencies of Tregs were negatively correlated with Th17 cells and with Tc17 cells. Interestingly, a significant portion of the cells that infiltrate the lungs was skewed towards a Tc17 phenotype. Our findings suggest the contribution of Th17, Tc17, and Treg cells in the pathogenesis of COPD. Rebalance of these cells will be helpful for developing and refining the new immunological therapies for COPD.     

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

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

Regulatory T cells and T helper 17 cells in viral infection

CD4⁺ T cells are the central element of the adaptive immune responses and protect the body from a variety of pathogens. Starting from naive cells, CD4⁺ T cells can differentiate into various effector cell subsets with specialized functions including T helper (Th) 1, Th2, Th17, regulatory T (Treg) and T follicular helper (Tfh) cells. Among them, Tregs and Th17 cells show a strong plasticity allowing the functional adaptation to various physiological and pathological environments during immune responses. Although they are derived from the same precursor cells and their differentiation pathways are interrelated, the terminally differentiated cells have totally opposite functions. Studies have shown that Tregs and Th17 cells have rather complex interplays in viral infection: Th17 cells may contribute to immune activation and disease progression while Tregs may inhibit this process and play a key role in the maintenance of immune homoeostasis, possibly at the cost of compromised viral control. In this review, we take respiratory syncytial virus (RSV), hepatitis B virus (HBV)/hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections as examples to discuss these interplays and their impacts on disease progression in viral infection.     

Kidney GATA3+ regulatory T cells play roles in the convalescence stage after antibody-mediated renal injury

FoxP3⁺ regulatory T cells (Tregs) play crucial roles in peripheral immune tolerance. In addition, Tregs that reside or accumulate in nonlymphoid tissues, called tissue Tregs, exhibit tissue-specific functions and contribute to the maintenance of tissue homeostasis and repair. In an experimental mouse model of crescentic glomerulonephritis induced by an anti-glomerular basement membrane antibody, Tregs started to accumulate in the kidney on day 10 of disease onset and remained at high levels (~30–35% of CD4⁺ T cells) during the late stage (days 21–90), which correlated with stable disease control. Treg depletion on day 21 resulted in the relapse of renal dysfunction and an increase in Th1 cells, suggesting that Tregs are essential for disease control during the convalescence stage. The Tregs that accumulated in the kidney showed tissue Treg phenotypes, including high expression of GATA3, ST2 (the IL33 receptor subunit), amphiregulin (Areg), and PPARγ. Although T-bet⁺ Tregs and RORγt⁺ Tregs were observed in the kidney, GATA3⁺ Tregs were predominant during the convalescence stage, and a PPARγ agonist enhanced the accumulation of GATA3⁺ Tregs in the kidney. To understand the function of specific genes in kidney Tregs, we developed a novel T cell transfer system to T cell-deficient mice. This experiment demonstrates that ST2, Areg, and CCR4 in Tregs play important roles in the accumulation of GATA3⁺ Tregs in the kidney and in the amelioration of renal injury. Our data suggest that GATA3 is important for the recruitment of Tregs into the kidney, which is necessary for convalescence after renal tissue destruction.     

Distinct different sensitivity of Treg and Th17 cells to Fas-mediated apoptosis signaling in patients with acute coronary syndrome

Objective: An imbalance in CD4⁺CD25⁺ regulatory T (Treg) cells and Th17 cells has been found to correlate to occurrence of acute coronary syndrome [ACS, including unstable angina (UA) and acute myocardial infarction (AMI)]. However, the mechanisms of Th17/Treg imbalance in ACS patients are still unclear. The purpose of this study is to investigate the possibility of differences in sensitivity of Th17 and Tregs to Fas-mediated apoptosis which could lead to Th17/Treg imbalance in ACS patients. Methods: We examined the apoptosis of Th17 and Treg cells, apoptosis-related Fas/Fas ligand(FasL) pathway, and inflammatory markers in patients with AMI, UA, stable angina (SA) and controls by Flow cytometry and ELISA. Then we analysed the correlation of inflammatory markers and sFasL to Treg apoptosis, and the effect of anti-FasL antibody on Treg apoptois in vitro. Results: Our study demonstrated that apoptotic Tregs, Fas and FasL expression, Caspase-3 activity of Tregs were significantly higher in ACS patients than those in NCA and SA patients (all P < 0.05). The percentage of apoptotic Tregs is positively correlated with the levels of inflammatory markers and sFasL. In vitro incubation of peripheral blood mononuclear cells from ACS patients with anti-FasL antibody resulted in a markedly reduction of apoptotic Treg cells. However, there were no significant differences in apoptotic Th17 cells and in Fas and FasL expression for Th17 cells between the four groups (all P >0.05). Conclusions: Tregs, but not Th17 cells, become apoptotic through Fas/FasL pathway, which contributed to reduction of Tregs leading to an imbalance between Th17 and Treg cells. This could be the mechanism underlying Th17/Treg imbalance and occurrence of ACS.     

T cell‐derived leptin contributes to increased frequency of T helper type 17 cells in female patients with Hashimoto's thyroiditis

Leptin modulates T cell function and plays an important role in autoimmune diseases. Our study aimed to explore the role of leptin and T helper type 17 (Th17) cells in Hashimoto's thyroiditis patients. Twenty‐seven patients with Hashimoto's thyroiditis (HT) and 20 healthy controls were enrolled into the current study. A modest increase of plasma leptin in HT patients and the CD4⁺ T cell‐derived leptin from HT patients was stronger than that from healthy controls. In HT patients, there are no statistically significant correlations between plasma leptin concentrations and the percentage of Th17 cells or the level of retinoic acid‐related orphan receptor γt (RORγt), but strong positive correlations were observed between CD4⁺ T cell‐derived leptin and the percentage of Th17 cells or the level of RORγt mRNA, and additionally significantly up‐regulated leptin, interleukin (IL)17 and RORγt mRNA levels in the thyroid tissue. Furthermore, neutralization of leptin decreases the frequency of Th17 cells in vitro. Current study has revealed an increased leptin involvment in Hashimoto's thyroiditis associated with an increased number of Th17 cells.     

PDCD5 negatively regulates autoimmunity by upregulating FOXP3+ regulatory T cells and suppressing Th17 and Th1 responses

Maintenance of FOXP3 protein expression is crucial for differentiation and maturation of regulatory T (Treg) cells, which play important roles in immune homeostasis and immune tolerance. We demonstrate here that PDCD5 interacts with FOXP3, increases acetylation of FOXP3 in synergy with Tip60 and enhances the repressive function of FOXP3. In PDCD5 transgenic (PDCD5tg) mice, overexpression of PDCD5 enhanced the level of FOXP3 protein and percentage of CD4⁺CD25⁺FOXP3⁺ cells. Naïve CD4⁺ T cells from PDCD5tg mice were more sensitive to TGF-β-induced Treg polarization and expansion. These induced Tregs retained normal suppressive function in vitro. Severity of experimentally-induced autoimmune encephalomyelitis (EAE) in PDCD5tg mice was significantly reduced relative to that of wild-type mice. The beneficial effect of PDCD5 likely resulted from increases of Treg cell frequency, accompanied by a reduction of the predominant pathogenic Th17/Th1 response. Activation-induced cell death enhanced by PDCD5 was also linked to this process. This is the first report revealing that PDCD5 activity in T cells suppresses autoimmunity by modulating Tregs. This study suggests that PDCD5 serves as a guardian of immunological functions and that the PDCD5-FOXP3-Treg axis may be a therapeutic target for autoimmunity.     

Lactobacillus plantarum C29 alleviates NF-κB activation and Th17/Treg imbalance in mice with TNBS-induced colitis

In this study, we examined whether Lactobacillus plantarum C29 could restore 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced T helper 17 (Th17)/regulatory T cells (Tregs) imbalance in mice. Treatment with C29 inhibited the differentiation of splenic T cells into Th17 cells and the expression of retinoic acid receptor-related orphan receptor gamma t (RORγt) and IL-17 in vitro, whereas promoting the differentiation into Tregs. Oral administration of Lactobacillus plantarum C29 in mice attenuated TNBS-induced colon shortening, myeloperoxidase (MPO) activity, inducible Nitric oxide (NO) synthase, and cyclooxygenase-2 expression, and activation of NF-κB in the colon of mice. C29 treatment downregulated TNF-α, IL-17, and IL-1β expression, while increasing IL-10 expression. C29 treatment suppressed TNBS-induced Th17 cell differentiation and reduced IL-17 and RORγt expression, while promoting the TNBS-suppressed Tregs differentiation and IL-10 and forkhead box P3 expression. These findings suggest that C29 can alleviate colitis by modulating NF-κB activation as well as Th17/Treg balance.