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IL-17+Foxp3+ T cells: An intermediate differentiation stage between Th17 cells and regulatory T cells
Abstract
Foxp3(+) Tregs have been known as a major regulator of immune homeostasis through their immunosuppressive function. Th17 lineage is a CD4(+) T cell subset that exerts its function by secreting proinflammatory cytokines and protecting host against microbial infections. The altered ratio between Foxp3(+) Tregs and Th17 cells plays an important role in the pathogenesis of immune-related diseases. Recent mice and human studies have demonstrated that Tregs can be reprogrammed into a novel population, IL-17(+)Foxp3(+) T cells, phenotypically and functionally resembling Th17 cells under the complicated cytokine stimulation. The identification of IL-17(+)Foxp3(+) T cells may provide a new understanding of therapy targeting Tregs and Th17 cells in autoimmune diseases and cancer. Here, we highlight significant data regarding the phenotype profile, origination, differentiation, and the pleiotropic functions of IL-17(+)Foxp3(+) T cells and the reciprocal relationships of these cells to Tregs and Th17 cells. Furthermore, the role of IL-17(+)Foxp3(+) T cells in tumorigenesis and clinical implications in cancer therapy are discussed in this review.
... Hence, the balance between FOXP3+ Treg cells and Th17 cells is considered an important factor for treatment of autoimmune diseases [145] and cancers [146]. Indeed, Maruyama et al. [54] reported that the infiltration of Th17 cells gradually decreased compared to increased Treg cells in gastric cancer progression. ...
... Notably, CD4 + CD25 + FOXP3 + (GFP + ) T cells can differentiate into Th17 cells in the presence of IL-6 [57] and T helper 17 (Th17) cells, one of the CD4 + T cells, can produce IL-17 to protect against microbial infection [214], while excessive activation of Treg cells suppresses antipathogenic or anticancer immunity, leading to chronic infection and tumor progression [144]. The balance between FOXP3 + Treg cells and Th17 cells is considered an important target for treatment of autoimmune diseases [145] and cancers [146]. Thus, next generation DC immunotherapy is required for more effective cancer therapy. ...
... In the same line, combination therapy of DCs with checkpoint inhibitors including ipilimumab [216] or other immune cells or oncolytic virus [216] is considered attractive in cancer therapy [216,217]. In this regard, we suggest the cocktail of specialized DC vaccines and Th17 cells by reprogramming Treg cells into Th17 cells [146] or ex vivo expansion of Th17 cells from human PBMCs [218] is suggested as a next generation cellular cancer immunotherapy, which should be further investigated in vivo and clinically. ...
Though Forkhead box P (FOXP) transcription factors comprising of FOXP1, FOXP2, FOXP3 and FOXP4 are involved in the embryonic development, immune disorders and cancer progression, the underlying function of FOXP3 targeting CD4 + CD25+ regulatory T (Treg) cells and the dual roles of FOXP proteins as an oncogene or a tumor suppressor are unclear and controversial in cancers to date. Thus, the present review highlighted research history, dual roles of FOXP proteins as a tumor suppressor or an oncogene, their molecular networks with other proteins and noncoding RNAs, cellular immunotherapy targeting FOXP3, and clinical implications in cancer progression.
... In addition, JNJ-61803534 did not affect nTreg suppressive activity on Teff cells, as evaluated by Teff proliferation and IFNγ production. It has been reported that Tregs can convert into IL-17-producing FOXP3 + RORγt + CD4 + T cells under certain microenvironments 48 . This IL-17-producing FOXP3 + Treg population, considered as an intermediate differentiation stage that can transform to Th17 cells, has been demonstrated both in mouse and human 48 . ...
... It has been reported that Tregs can convert into IL-17-producing FOXP3 + RORγt + CD4 + T cells under certain microenvironments 48 . This IL-17-producing FOXP3 + Treg population, considered as an intermediate differentiation stage that can transform to Th17 cells, has been demonstrated both in mouse and human 48 . While the cell populations in the current study were not examined by co-staining FOXP3 and RORγt, treatment of activated CCR6 + T cells from human peripheral blood with RORγt inhibitor JNJ-54271074 in a previous study, slightly increased IL-10 production while greatly reducing IL-17A production 22 , suggesting skewing towards Tregs by inhibition of RORγt. ...
The nuclear receptor retinoid-related orphan receptor gamma t (RORγt) plays a critical role in driving Th17 cell differentiation and expansion, as well as IL-17 production in innate and adaptive immune cells. The IL-23/IL-17 axis is implicated in several autoimmune and inflammatory diseases, and biologics targeting IL-23 and IL-17 have shown significant clinical efficacy in treating psoriasis and psoriatic arthritis. JNJ-61803534 is a potent RORγt inverse agonist, selectively inhibiting RORγt-driven transcription versus closely-related family members, RORα and RORβ. JNJ-61803534 inhibited IL-17A production in human CD4 ⁺ T cells under Th17 differentiation conditions, but did not inhibit IFNγ production under Th1 differentiation conditions, and had no impact on in vitro differentiation of regulatory T cells (Treg), nor on the suppressive activity of natural Tregs. In the mouse collagen-induced arthritis model, JNJ-61803534 dose-dependently attenuated inflammation, achieving ~ 90% maximum inhibition of clinical score. JNJ-61803534 significantly inhibited disease score in the imiquimod-induced mouse skin inflammation model, and dose-dependently inhibited the expression of RORγt-regulated genes, including IL-17A, IL-17F, IL-22 and IL-23R. Preclinical 1-month toxicity studies in rats and dogs identified doses that were well tolerated supporting progression into first-in-human studies. An oral formulation of JNJ-61803534 was studied in a phase 1 randomized double-blind study in healthy human volunteers to assess safety, pharmacokinetics, and pharmacodynamics. The compound was well tolerated in single ascending doses (SAD) up to 200 mg, and exhibited dose-dependent increases in exposure upon oral dosing, with a plasma half-life of 164 to 170 h. In addition, dose-dependent inhibition of ex vivo stimulated IL-17A production in whole blood was observed, demonstrating in vivo target engagement. In conclusion, JNJ-61803534 is a potent and selective RORγt inhibitor that exhibited acceptable preclinical safety and efficacy, as well as an acceptable safety profile in a healthy volunteer SAD study, with clear evidence of a pharmacodynamic effect in humans.
... This lineage also performs a crucial role in the pathogenesis of immune-related diseases. The altered ratio between regulatory T (Treg) cells generally suppresses or downregulates the induction and proliferation of effector T cells (9). Identification of Treg cells was originally based on CD25 expression; however, CD25 is also expressed by activated effector T cells. ...
... Similar to this finding, Qin et al reported that decreased regulatory T cells (12) are commonly seen in critically ill patients, suggesting dysregulated immune responses. Data from experimental study show IL-6 that arise in ARDS, may have contextual protective or exacerbating roles including severity of infection, survival and tissue remodeling (5)(6)(7)(8)(9)(10)(11)(12)(13)(14). ...
Aim:
Evaluating the expression level of CD4+ FoxP3+ CD25+ T cells and IL-6 in peripheral blood samples of hospitalized COVID-19 patients.
Background:
COVID-19 is an emerging disease with worldwide distribution. However, there is a little data about the correlation between the disease and the host immune responses.
Methods:
Whole blood samples of 30 COVID-19 patients and eight healthy people were collected during March to June 2020. Total RNA was extracted from the samples, cDNA synthesis was performed, and the expression level of targeted genes was evaluated using quantitative real-time PCR.
Results:
The expression level of CD4, CD25, and Foxp3 was significantly downregulated 5-, 2-, and 3-fold, respectively, among COVID-19 patients in comparison to healthy controls (P-value < 0.0001). The expression level of IL-6 represented almost 18-fold increase in COVID-19 patients compared to healthy controls.
Conclusion:
Our findings indicated the expression profile analysis of CD4+ FoxP3+ CD25+ T cells could be a potential marker for the assessment of severity of COVID-19 patients.
... inflammatory Th17 cells (Bovenschen et al., 2011;Remedios et al., 2018). FoxP3 + cells from individuals with autoimmune diseases can easily differentiate into IL-17A producing cells as FoxP3 expression is progressively lost ("Treg instability": Du et al., 2014). These IL-17A + FoxP3 + T-cells may also retain the expression of other Treg-related proteins like CTLA-4 (Du et al., 2014). ...
... FoxP3 + cells from individuals with autoimmune diseases can easily differentiate into IL-17A producing cells as FoxP3 expression is progressively lost ("Treg instability": Du et al., 2014). These IL-17A + FoxP3 + T-cells may also retain the expression of other Treg-related proteins like CTLA-4 (Du et al., 2014). We found higher Il-17a splenic expression in F344/LEW-BM rats coincident with high levels of Il-6 but less Il-23 and Il-1b mRNA (the expression of which varies together with Tgf-β1 and is lower in F344/LEW-BM rats). ...
One of the main challenges to understand drug addiction is defining the biological mechanisms that underlie individual differences in recidivism. Studies of these mechanisms have mainly focused on the brain, yet we demonstrate here a significant influence of the peripheral immune system on this phenomenon. Lewis (LEW) and Fischer 344 (F344) rats have different immunological profiles and they display a distinct vulnerability to the reinforcing effects of cocaine, with F344 more resistant to reinstate cocaine-seeking behavior. Bone marrow from male LEW and F344 rats was transferred to male F344 rats (F344/LEW-BM and F344/F344-BM), and these rats were trained to self-administer cocaine over 21 days. Following extinction, these animals received a sub-threshold primer dose of cocaine to evaluate reinstatement. F344/LEW-BM but not F344/F344-BM rats reinstated cocaine-seeking behavior, in conjunction with changes in their peripheral immune cell populations to a profile that corresponded to that of the LEW donors. Cocaine treatment increased the CD4⁺ T-cells in F344/LEW-BM rats, and the splenic expression of Il-17a, Tgf-β, Tlr-2, Tlr-4 and Il-1β was altered in both groups. We propose that peripheral T-cells respond to cocaine, with CD4⁺ T-cells in particular undergoing Th17 polarization and generating long-term memory, these cells releasing mediators that trigger central mechanisms to induce reinstatement after a second encounter. This immune response may explain the high rates of recidivism observed despite long periods of detoxification, shedding light on the mechanisms underlying the vulnerability and resilience of specific individuals, and opening new perspectives for personalized medicine in the treatment of relapse.
... Nevertheless, whether Th17 cells certainly convert toward Treg phenotype in vivo in a tumor microenvironment is not known yet. Additionally, though Th17/Treg (IL-17 þ Foxp3 þ ) hybrid cells have been reported in human tumors, they mainly originate from real Treg cells [44]. Those immunosuppressive IL17 þ Foxp3 þ T cells have been shown in human colorectal and esophageal cancers, but not in ovarian cancer, melanoma, or renal cell carcinoma [12,44,45]. ...
... Additionally, though Th17/Treg (IL-17 þ Foxp3 þ ) hybrid cells have been reported in human tumors, they mainly originate from real Treg cells [44]. Those immunosuppressive IL17 þ Foxp3 þ T cells have been shown in human colorectal and esophageal cancers, but not in ovarian cancer, melanoma, or renal cell carcinoma [12,44,45]. ...
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.
... En contraste, la expresión del factor transcripcional Th2 (GATA3) se encuentra disminuida en las mismas biopsias de Pt, lo que puede deberse al efecto inhibitorio de T-bet sobre la expresión de GATA3 [16][17][18][19] . El aumento de la expresión de IL-17 y Foxp3 en muestras de Pt sugiere la presencia de linfocitos Th IL-17 + Foxp3 + , un subconjunto de la población Th recientemente descrito con la capacidad de suprimir la proliferación de células T y de promover la progresión tumoral 20 . En estudios realizados previamente en pacientes con Pt primario y recidivante, se encontró que ambos mostraban un nivel elevado de IL-17 en la película lagrimal, por lo que se sugiere que podría estar asociado con su causa y recurrencia 21 . ...
... Las células Th IL-17 + Foxp3 + han sido identificadas en diversas enfermedades humanas, como la enfermedad inflamatoria intestinal 22 , tumores de colon 23 , periodontitis 24 , psoriasis 25 y artritis reumatoide 26 . Característicamente estas células conservan la actividad inmunosupresora de los linfocitos Treg, al tiempo que adquieren capacidad efectora por la producción de IL-17 20 . Es conocido el hecho de que el microambiente en el que se encuentren los linfocitos Th precursores (naive) determina el fenotipo del linfocito Th diferenciado; sin embargo, es importante también considerar que los linfocitos Th efectores pueden modificar su fenotipo en respuesta a los estímulos circundantes. ...
... En contraste, la expresión del factor transcripcional Th2 (GATA3) se encuentra disminuida en las mismas biopsias de Pt, lo que puede deberse al efecto inhibitorio de T-bet sobre la expresión de GATA3 [16][17][18][19] . El aumento de la expresión de IL-17 y Foxp3 en muestras de Pt sugiere la presencia de linfocitos Th IL-17 + Foxp3 + , un subconjunto de la población Th recientemente descrito con la capacidad de suprimir la proliferación de células T y de promover la progresión tumoral 20 . En estudios realizados previamente en pacientes con Pt primario y recidivante, se encontró que ambos mostraban un nivel elevado de IL-17 en la película lagrimal, por lo que se sugiere que podría estar asociado con su causa y recurrencia 21 . ...
... Las células Th IL-17 + Foxp3 + han sido identificadas en diversas enfermedades humanas, como la enfermedad inflamatoria intestinal 22 , tumores de colon 23 , periodontitis 24 , psoriasis 25 y artritis reumatoide 26 . Característicamente estas células conservan la actividad inmunosupresora de los linfocitos Treg, al tiempo que adquieren capacidad efectora por la producción de IL-17 20 . Es conocido el hecho de que el microambiente en el que se encuentren los linfocitos Th precursores (naive) determina el fenotipo del linfocito Th diferenciado; sin embargo, es importante también considerar que los linfocitos Th efectores pueden modificar su fenotipo en respuesta a los estímulos circundantes. ...
Introduction: Various pro-inflammatory cytokines and growth factors have been implicated in the pathogenesis of pterygium (Pt); however, the effect of cytokines produced by helper T lymphocytes (Th) has been poorly studied. To date, only interleukin 4 (IL-4) from Th lymphocytes has been linked to Pt recurrence. Therefore, this study aimed to evaluate the expression of Th cytokines and Th transcription factors in primary Pt. Methods: Pt biopsies were obtained from 28 eyes of 28 Mexican patients undergoing primary excision of Pt with conjunctival autograft. Conjunctival biopsies of 8 patients undergoing cataract surgery were used as the control group. Gene expression of Th cytokines - interferon gamma (IFN-γ), IL-13, IL-17, IL-10-, as well as gene expression of Th transcription factors T-bet, GATA3, Foxp3 and RORγt, was analyzed by real-time RT-PCR. results: We found decreased expression of markers T-bet, GATA3, RORγt, IFN- γ, IL-13 and IL-10 in Pt. We found a sixfold increase in the expression of IL-17 and Foxp3 in Pt compared to healthy conjunctiva (p < 0.0001); this increase was dependent on Pt size. Conclusions: The increased expression of IL-17 and Foxp3 in Pt samples suggests the presence of IL-17+ Foxp3⁺ Th lymphocytes, a subset of the Th population with the ability to suppress T cell proliferation and promote tumor progression. Therefore, IL17⁺ Foxp3⁺ Th cells may be involved in the occurrence and growth of Pt.
... FOXP3 protein plays a crucial role in regulating immune function. 35,36 Our study's genomic analysis highlighted a strong association between FOXP3 and alterations in numerous immune checkpoints in invasive BRCA. Notably, this association was concurrent but not mutually exclusive. ...
Background
Breast cancer (BC) continues to pose a substantial challenge to global health, necessitating an enhanced understanding of its fundamental mechanisms. Among its various pathological classifications, breast invasive carcinoma (BRCA) is the most prevalent. The role of the transcription factor forkhead box P3 (FOXP3), associated with regulatory T cells, in BRCA’s diagnosis and prognosis remains insufficiently explored, despite its recognized importance.
Methods
We examined the mRNA expression profile of FOXP3 in BRCA patients, assessing its correlation with disease detection, patient survival, immune checkpoint alterations, and response to anticancer drugs.
Results
Our analysis revealed significantly elevated FOXP3 mRNA levels in BRCA patients, with a 95.7% accuracy for BRCA detection based on the area under the curve. High FOXP3 mRNA levels were positively correlated with overall survival and showed significant associations with CTLA4, CD274, PDCD1, TMB, and immune cell infiltration status. Furthermore, FOXP3 mRNA expression was linked to the efficacy of anticancer drugs and the tumor inflammation signature.
Discussion
These findings suggest that FOXP3 serves as a promising biomarker for BRCA, offering valuable insights into its diagnosis and prognosis. The correlation between FOXP3 expression and immune checkpoint alterations, along with its predictive value for treatment response, underscores its potential in guiding therapeutic strategies.
Conclusion
FOXP3 stands out as an influential factor in BRCA, highlighting its diagnostic accuracy and prognostic value. Its association with immune responses and treatment efficacy opens new avenues for research and clinical applications, positioning FOXP3 as a vital target for further investigation in BRCA management.
... 8 Under the regulation of Hp, gastric cancer stem cells can act as regulator of Th17/Treg ratio, Treg IL-17 − lymphocytes can be transferred to IL-17 + phenotype while Th17 Foxp3 − lymphocytes can become to Foxp3 + Tregs. 15,16 In brief, Hp infection causes local inflammation and leads to immunosuppressive environment, which result from the negative immune regulation described above (as shown in Figure 1). Followed by superficial gastritis (SG), chronic atrophic gastritis (CAG), intestinal metaplasia (IM), dysplasia and leading to GC. ...
Introduction
Despite advancements in the methods for prevention and early diagnosis of gastric cancer (GC), GC continues to be the fifth in incidence among major cancers and the third most common cause of cancer‐related death. The therapeutic effects of surgery and drug treatment are still unsatisfied and show notable differences according to the tumor microenvironment (TME) of GC.
Methods
Through screening Pubmed, Embase, and Web of Science, we identified and summarized the content of recent studies that focus on the investigation of Helicobacter pylori (Hp) infection, regulatory T cells (Tregs), and tumor‐associated macrophages (TAMs) in the TME of GC. Furthermore, we searched and outlined the clinical research progress of various targeted drugs in GC treatment including CTLA‐4, PD‐1\PD‐L1, and VEGF/VEGFR.
Results
In this review, the findings indicate that Hp infection causes local inflammation and leads to immunosuppressive environment. High Tregs infiltration in the TME of GC is associated with increased induction and recruitment; the exact function of infiltrated Tregs in GC was also affected by phenotypes and immunosuppressive molecules. TAMs promote the development and metastasis of tumors, the induction, recruitment, and function of TAMs in the TME of gastric cancer are also regulated by various factors.
Conclusion
Discussing the distinct tumor immune microenvironment (TIME) of GC can deepen our understanding on the mechanism of cancer immune evasion, invasion, and metastasis, help us to reduce the incidence of GC, and guide the innovation of new therapeutic targets for GC eventually.
... Nevertheless, there is evidence suggestive of transforming the IL17 − Treg into IL-17 + Treg with gaining proinflammatory functions. On the other hand, FOXP3 − Th17 has been shown to turn into FOXP3 + Th17 with the functions like regulatory T lymphocytes [84,85]. Lymphocytes with both IL-17 and FOXP3 expression in colorectal cancer provoke the CSCs development and expansion in hypoxic niche through activating the MAPK and AKT kinases signaling pathways [86]. ...
Cancer stem cells (CSCs) defined as a small fraction of cells within malignancies have been isolated from tumors with different histological origins with stem related characteristics such as self-replicating potential, tumorigenesis, and therapy resistance. The dynamic communication between CSCs and tumor microenvironment particularly immune cells orchestrates their fate and plasticity as well as the patient outcome. According to recent evidence, it has been reported that they harness different immunological pathways to escape immunosurveillance and express aberrantly immunomodulatory agents or decreased levels of factors forming antigen presenting machinery (APM), subsequently followed by impaired antigen presentation and suppressed immune detection. As effective therapies are expected to be able to eradicate CSCs, mechanistic understanding of such interactions can provide insights into causes of therapy failure particularly in immunotherapy. Also, it can contribute to enhance the practical interventions against CSCs and their immunomodulatory features resulting in CSCs eradication and improving patient clinical outcome. The aim of this review is to explain the present knowledge regarding the immunobiology of CSCs and the immunoevasion mechanisms they use.
... Murine and human studies have demonstrated that Treg cells can be reprogrammed to a distinct population, Foxp3 + /IL17 + T cells, phenotypically and functionally resembling Th17 cells. 38 Particularly in CRC, the presence of Foxp3 + /IL17 + T cells has been reported to be increased in the mucosa and peripheral blood of patients with chronic colitis as well as in colorectal tumours. 39 Mutations in the gene encoding APC are the most frequent driver mutations leading to sporadic CRC, together with mutations in TP53 and KRAS. ...
Objective:
Helicobacter pylori infection is the most prevalent bacterial infection worldwide. Besides being the most important risk factor for gastric cancer development, epidemiological data show that infected individuals harbour a nearly twofold increased risk to develop colorectal cancer (CRC). However, a direct causal and functional connection between H. pylori infection and colon cancer is lacking.
Design:
We infected two Apc-mutant mouse models and C57BL/6 mice with H. pylori and conducted a comprehensive analysis of H. pylori-induced changes in intestinal immune responses and epithelial signatures via flow cytometry, chip cytometry, immunohistochemistry and single cell RNA sequencing. Microbial signatures were characterised and evaluated in germ-free mice and via stool transfer experiments.
Results:
H. pylori infection accelerated tumour development in Apc-mutant mice. We identified a unique H. pylori-driven immune alteration signature characterised by a reduction in regulatory T cells and pro-inflammatory T cells. Furthermore, in the intestinal and colonic epithelium, H. pylori induced pro-carcinogenic STAT3 signalling and a loss of goblet cells, changes that have been shown to contribute-in combination with pro-inflammatory and mucus degrading microbial signatures-to tumour development. Similar immune and epithelial alterations were found in human colon biopsies from H. pylori-infected patients. Housing of Apc-mutant mice under germ-free conditions ameliorated, and early antibiotic eradication of H. pylori infection normalised the tumour incidence to the level of uninfected controls.
Conclusions:
Our studies provide evidence that H. pylori infection is a strong causal promoter of colorectal carcinogenesis. Therefore, implementation of H. pylori status into preventive measures of CRC should be considered.
... Additionally, specific chemokine receptors and adhesion molecules are expressed, especially in tissue-directed and tissue-resident Tregs (14). For example, CCR4 is important for migration to skin and CCR9 is needed for homing to intestine (15,16). Furthermore, memory Tregs can be divided into Th1, Th17 and Th2 Tregs, and these subpopulations emerge for two main reasons (17)(18)(19). ...
Regulatory T cells (Tregs) are the master regulators of immunity and they have been implicated in different disease states such as infection, autoimmunity and cancer. Since their discovery, many studies have focused on understanding Treg development, differentiation, and function. While there are many players in the generation and function of truly suppressive Tregs, the role of checkpoint pathways in these processes have been studied extensively. In this paper, we systematically review the role of different checkpoint pathways in Treg homeostasis and function. We describe how co-stimulatory and co-inhibitory pathways modulate Treg homeostasis and function and highlight data from mouse and human studies. Multiple checkpoint pathways are being targeted in cancer and autoimmunity; therefore, we share insights from the clinic and discuss the effect of experimental and approved therapeutics on Treg biology.
... Pathogenesis of RA is still unknown but may be due to some genetic causes and environmental factors that affect immune system and attack joints accordingly (National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIH) 2014; Majithia and Geraci 2007 Forkhead box P3 (FOXP3) is a master regulator of regulatory T cells (Treg cells) development and function. CD4 + Foxp3 + Treg cells suppress other immune cells but can also produce pro-inflammatory cytokines like IL17A (Du et al. 2014;Jung et al. 2017). T helper 17 (Th17) cells are CD4 + T cells functions by proinflammatory cytokines secretion and protection against microbial infections. ...
Background
Rheumatoid arthritis (RA) is a known debilitating autoimmune disease. Immune-suppressants that are used for disease treatment have serious side effects, therefore, trivalent chromium (Cr (III)); which has shown evidence of its influences on some inflammatory pathways and cytokines; was used in this study for the first time to be assessed for its therapeutic effect in RA rat model and was compared to prednisolone in a trial to find a treatment with lesser side effects.
Methods
Adult male albino rats were randomly divided into four groups: normal, untreated RA, prednisolone treated RA (1.25 mg/kg/day) and Cr (III) treated RA groups (80 μg/kg/day), induction of RA was done by subcutaneous complete Freund adjuvant injection. Study duration was 4 weeks throughout which arthritis scoring and weight measurement were pursued. Histopathological examination and immunohistochemical FOXP3 assessment were done for joint biopsies. Serum inflammatory markers (interleukin 17, interleukin 10, CRP) and synovial erosive arthritis marker (Cathepsin G) were measured. HDL and non-HDL cholesterol were estimated as well.
Results
Cr (III) treatment showed marked clinical and histopathological improvement, also astonishing anti-inflammatory effects (increase in FOXP3 expression and interleukin 10, with decrease in interleukin 17, CRP and synovial Cathepsin G) to the extent that Cr (III) effects on inflammation abolishment were comparable to that of prednisolone and even better at some aspects. Moreover, Cr (III) was protective from side effects, i.e., weight gain and dyslipidemia that were seen with prednisolone treatment.
Conclusions
Cr (III) is promising in treating RA and it lacks some side effects of accustomed immune-modulatory agents including prednisolone. Further experimental studies and clinical trials should be held to see the efficacy of Cr (III) in different doses and to assess its long term side effects when used for rheumatoid arthritis and other autoimmune diseases treatment.
... Accumulating evidence demonstrated that FOXPs proteins play important roles in the regulation of immune function 50,51 . In this work, genomic analysis revealed that FOXPs was closely related to the alteration of extensive immune checkpoints in breast invasive carcinoma. ...
Forkhead-box-P family include FOXP1/2/3/4 and its clinical significance still remains unclear in breast cancer (BRCA). We analysed the expressions of FOXPs in BRCA patients to determine diagnostic and prognostic values. Our results indicated that the transcriptional levels of FOXP3/4 were up-regulated in BRCA patients, but FOXP2 were down-regulated. No statistically significant correlation were found between the expression levels of FOXPs in Pathologic stage. FOXP2/3 had a significantly high AUC value in the detection of breast cancer, with 96.8% or 95.7% in accuracy respectively. Our study also suggested that BRCA patients with high transcription levels of FOXP1/2/4 were significantly associated with longer Overall Survival (OS). In contrast, BRCA patients with high transcription level of FOXP3 was not statistically related with OS. Our work revealed that FOXPs were closely related to the alteration of extensive immune checkpoints in breast invasive carcinoma. Additionally, FOXP3 has a significant positive correlation with PDCD1, CD274, CTLA4 and TMB in breast cancer, and FOXP3 expression showed a statistically significant correlation with infiltration of immune cells. Finally, we found that FOXP3 expression predicted the breast cancer cells response to anticancer drugs. Altogether, our work strongly suggested that FOXPs could serve as a biomarker for tumor detection, therapeutic design and prognosis.
... It is of particular interest that the latter cell cluster has an immunosuppressive capacity and represents an intermediate differentiation stage between Th17 cells and regulatory T cells. The cHL TME may influence Tregs to produce IL17 and to acquire a more immunosuppressive phenotype by releasing TGF-β or other molecules, which are known to promote IL17 + Tregs [41]. Even the HRS cells themselves produce TGF-β [21], which is consistent with the hypothesis. ...
Simple Summary
Classic Hodgkin lymphoma (cHL) is one of the most enigmatic study models of the tumor microenvironment (TME), in which the Hodgkin–Reed Sternberg (HRS) cells are distributed throughout an abundant but ineffective immune ecosystem. The hyperactivation of HRS cells due to somatic mutations leads to complex interactions with the different subsets of immune cell populations, modeling the pathophysiology of the disease. There are remaining loose ends regarding the identification of the immune functional states in the cHL microenvironment and their influence on tumor cell survival. Here, we review the most relevant immune populations identified in the cHL context, focusing on integrative functional signatures.
Abstract
The classic Hodgkin lymphoma (cHL) tumor microenvironment (TME) is by far the most abundant component of tumors and is responsible for most of their biological and clinical characteristics. Recent advances in our knowledge of these networks in cellular interactions allow us to understand that the neoplastic Hodgkin and Reed Sternberg (HRS) cells, although they are in the minority, are the main architects of this dysregulated immune milieu. Here, we review the major changes that have happened in recent years: from TME as a helpless bystander, reflecting an ineffective immune response, to a dynamic tumor-promoting and immunosuppressive element. The HRS cells promote survival through interconnected intrinsic and extrinsic alterations, boosting pro-tumoral signaling pathways through genetic aberrations and autocrine growth signals, in parallel with abnormal cytokine secretion for the recruitment and selection of the best cell partners for this immunosuppressive TME. In turn, cHL is already proving to be the perfect model with which to address an immune checkpoint blockade. Preliminary data demonstrate the utility of druggable key signaling pathways in this ensemble, such as JAK-STAT, NF-κB, and others. In addition, myriad biomarkers predicting a response await validation by new in situ multiplex analytical methods, single-cell gene expression, and other techniques. Together, these components will define the functional phenotypes with which we will elucidate the molecular pathogenesis of the disease and improve the survival of patients who are refractory to conventional therapies.
... Accumulating evidence demonstrated that FOXPs proteins play important roles in the regulation of immune function 50,51 . In this work, genomic analysis revealed that FOXPs was closely related to the alteration of extensive immune checkpoints in breast invasive carcinoma. ...
Forkhead-box-P family include FOXP1/2/3/4 and its clinical significance still remains unclear in breast cancer(BRCA). We analysed the expressions of FOXPs in BRCA patients to determine diagnostic and prognostic values. Our results indicated that the transcriptional levels of FOXP3/4 were up-regulated in BRCA patients, but FOXP2 were down-regulated. No statistically significant correlation were found between the expression levels of FOXPs in Pathologic stage. FOXP2/3 had a significantly high AUC value in the detection of breast cancer, with 96.8% or 95.7% in accuracy respectively. Our study also suggested that BRCA patients with high transcription levels of FOXP1/2/4 were significantly associated with longer OS. In contrast, BRCA patients with high transcription levels of FOXP3 was not statistically related with OS. Our work revealed that FOXPs was closely related to the alteration of extensive immune checkpoints in breast invasive carcinoma. Additionally, FOXP3 has a significant positive correlation with PDCD1, CD274, CTLA4 and TMB in breast cancer, and FOXP3 expression showed a statistically significant correlation with infiltration of immune cells. Finally, we found that FOXP3 expression predicted the breast cancer cells response to anticancer drugs. Altogether, our work strongly suggests that FOXPs could serve as a biomarker for tumor detection, therapeutic design and prognosis.
... Indeed, several studies demonstrate the negative influence of lymphopenia on the Treg population associated with the lack of FoxP3 expression and decreased suppression activity of ex-FoxP3-cells [3,9]. The lack of FoxP3 expression in Treg is known to be associated with differentiation to Th17-lymphocytes due to the induced RORγt expression [2]. The reconstitution of the T-cell pool by adoptive transfer prevents the negative influence of lymphopenia by increasing IL-2 production. ...
Cytokines IL-7 and IL-15 are the most important humoral factors providing T-conventional cell pool reconstitution during homeostatic proliferation caused by lymphopenia. However, whether these cytokines can provide homeostatic maintenance and proliferation of T-regulatory (Treg) cells is largely unknown. Considering the association between homeostatic proliferation and the development of autoimmunity, we decided to investigate the ability of these factors to cause differentiation of Treg-cells into Th17-lymphocytes. Therefore, the purpose of this study was to investigate the influence of humoral factors of homeostatic proliferation (IL-7 and IL-15) on Treg-cells in vitro . The study used peripheral blood sampled from 22 healthy donors. PBMC fraction was isolated by Ficoll density gradient centrifugation. Proliferation was induced by IL-7, IL-15, and by a combination of IL-2 with anti-CD3-antibodies. The proliferation intensity of Tregs was evaluated by flow cytometry using CFSE in PBMC cultures by phenotype CD3 ⁺ CD4 ⁺ CD25 ⁺ FoxP3 ⁺ and in the previously purified population of CD3 ⁺ CD4 ⁺ CD25 ⁺ CD127lo-cells. In this case Treg-cells were obtained by immunomagnetic separation from PBMCs using a MACS Treg Isolation Kit. Also, the RORyt expression in CD3 ⁺ CD4 ⁺ CD25 ⁺ FoxP3 ⁺ -cells was evaluated during cultivation. Here, we have shown that IL-7 and IL-15 could support Treg-cells by number and phenotype. Also, we revealed that these factors provide FoxP3 expression in Treg-cells; meanwhile, stimulation with IL-2 + anti-CD3 can also cause induction of FoxP3 expression de novo in conventional CD4+ cells. Also, we have shown that IL-7 and IL-15 can cause lower-intensity proliferation of Treg-cells in comparison with IL-2 + anti-CD3. Herewith homeostatic cytokines didn’t have the ability to induce RORyt expression in both T-regulatory cells and CD4+ conventional T-lymphocytes. Thus, it has been shown that IL-7 and IL-15 can potentially participate in maintaining the total pool of Treg-cells during lymphopenia, when IL-2 deficiency occurs, without causing the induction of RORyt expression. However, how homeostatic cytokines affect the functional activity of Treg-cells remains unclear and requires further investigation.
... Although Th17 lymphocytes are usually viewed as a pro-inflammatory and totally opposite to anti-inflammatory regulatory T (T reg ) cells, there is evidence that mixed phenotypes can exist between these cells, and these are associated with CSC induction. For example, it is known that T reg IL-17 − lymphocytes can become IL-17 + , transforming into a pro-inflammatory phenotype [55], while Th17 lymphocytes without transcription factor, FOXP3, expression can become to FOXP3 + , functioning as T reg lymphocytes [56]. These IL-17 + FOXP3 + lymphocytes have been found in colorectal tumor tissue and have been shown to promote generation of CSCs in hypoxic environments by inducing activation of MAPK and AKT kinases in cancer cells (Fig. 2). ...
Cross talk between cancer cells and the immune system is determinant for cancer progression. Emerging evidence demonstrates that GC characteristics such as metastasis, treatment resistance, and disease recurrence are associated with a tumor subpopulation called gastric cancer stem cells (GCSCs). However, the specific interaction between GCSCs and the immune microenvironment is still under investigation. Although immune evasion has been well described for cancer stem cells (CSCs), recent studies show that GCSCs can also regulate the immune system and even benefit from it. This review will provide an overview of bidirectional interactions between CSCs and immune cells in GC, compiling relevant data about how CSCs can induce leukocyte reprogramming, resulting in pro-tumoral immune cells that orchestrate promotion of metastasis, chemoresistance, tumorigenicity, and even increase in number of cancer cells with stem properties. Some immune cells studied are tumor-associated macrophages (TAMs), neutrophils, Th17 and T regulatory (T reg ) cells, mesenchymal stem cells (MSCs), and cancer-associated fibroblasts (CAFs), as well as the signaling pathways involved in these pro-tumoral activities. Conversely, although there are cytotoxic leukocytes that can potentially eliminate GCSCs, we describe mechanisms for immune evasion in GCSCs and their clinical implications. Furthermore, we describe current available immunotherapy targeting GCSC-related markers as possible treatment for GC, discussing how the CSC-modified immune microenvironment can mitigate or inactivate these immunotherapies, limiting their effectiveness. Finally, we summarize key concepts and relevant evidence to understand the cross talk between GCSCs and the immune microenvironment as an important process for effective design of therapies against GCSCs that improve the outcome of patients with GC.
... Here we found that Tregs acquired Th17-like features, an adaptation that has been reported to result in reduced suppressive potential under inflammatory conditions (42,47). Thus far, it is not clear whether Th17-like Tregs are a stable cell type or represent an intermediate state in a conversion process (50,51). Nonetheless, our finding is important, as it demonstrates that AIH pathogenesis does not require a generalized Treg defect, as had been proposed before (26). ...
The liver is an immune-privileged organ that can deactivate autoreactive T cells. Yet in autoimmune hepatitis (AIH), autoreactive T cells can defy hepatic control and attack the liver. To elucidate how tolerance to self-antigens is lost during AIH pathogenesis, we generated a spontaneous mouse model of AIH, based on recognition of an MHC class II-restricted model peptide in hepatocytes by autoreactive CD4 T cells. We find that the hepatic peptide was not expressed in the thymus leading to deficient thymic deletion and resulting in peripheral abundance of autoreactive CD4 T cells. In the liver, autoreactive CD4 effector T cells accumulated within portal ectopic lymphoid structures and maturated towards pathogenic IFNγ and TNF co-producing cells. Expansion and pathogenic maturation of autoreactive effector T cells was enabled by a selective increase of plasticity and instability of autoantigen-specific Tregs, but not of non-specific Tregs. Indeed, antigen-specific Tregs were reduced in frequency and manifested increased IL-17 production, reduced epigenetic demethylation and reduced expression of Foxp3. As a consequence, autoantigen-specific Tregs had a reduced suppressive capacity, as compared to non-specific Tregs. In conclusion, loss of tolerance and the pathogenesis of AIH were enabled by combined failure of thymic deletion and peripheral regulation.
... Interestingly, this Treg reprogramming does not represent a classical Th17 conversion as these IL17-producing Tregs did not lose their suppressive function (Blatner et al. 2010). It has been suggested that these Tregs might be transitional and have reserved the ability to differentiate into FoxP3 + Tregs or TH17 cells depending on the local milieu of tumor microenvironment (Du et al. 2014). In a mouse model of CRC, TGFβ and prostaglandin E2 mediated the differentiation of Th17 cells into suppressive IL17 + and IL17 À FoxP3 + Tregs (Downs-Canner et al. 2017). ...
Mucosal surfaces are distinctive sites exposed to environmental, dietary, and microbial antigens. Particularly in the gut, the host continuously actively adapts via complex interactions between the microbiota and dietary compounds and immune and other tissue cells. Regulatory T cells (Tregs) are critical for tuning the intestinal immune response to self- and non-self-antigens in the intestine. Its importance in intestinal homeostasis is illustrated by the onset of overt inflammation caused by deficiency in Treg generation, function, or stability in the gut. A substantial imbalance in Tregs has been observed in intestinal tissue during pathogenic conditions, when a tightly regulated and equilibrated system becomes dysregulated and leads to unimpeded and chronic immune responses. In this chapter, we compile and critically discuss the current knowledge on the key factors that promote Treg-mediated tolerance in the gut, such as those involved in intestinal Treg differentiation, specificity and suppressive function, and their immunophenotype during health and disease. We also discuss the current state of knowledge on Treg dysregulation in human intestine during pathological states such as inflammatory bowel disease (IBD), necrotizing enterocolitis (NEC), graft-versus-host disease (GVHD), and colorectal cancer (CRC), and how that knowledge is guiding development of Treg-targeted therapies to treat or prevent intestinal disorders.
... Interestingly, the increment of Treg cells has been observed in patients with BC receiving IL-2 29,30 because it can induce the overexpression of the miR-182-183 cluster, which impairs FOXO1 expression. 12,[31][32][33] Although miR-182-183 expression is induced by IL-2, bioinformatics studies have shown that miR-182-183 potentially targets the IL-2/IL-2RA pathway and NFATs as IL-2 inducers (http://www.targe tscan.org). ...
T cells are polarized toward regulatory T cells (Tregs) in tumor microenvironment by shuttling of microRNAs that target T cell‐activating signaling pathways. We evaluated the expression of miR‐182 cluster (miR‐96, 182, and 183) in peripheral blood mononuclear cells (PBMCs) of patients with breast cancer (BC), and T cell polarization by the expression of FOXO1, NFATs, ITK, TCR/CD3 complex, and IL‐2/IL‐2RA. Twenty‐six microRNAs overexpressed in tumor tissues and sera of these patients were extracted by a meta‐analysis. Then the expression of miR‐182 cluster was investigated in PBMCs and sera of these patients and correlated with their targets in PBMCs. Finally, miR‐182 was cloned into Jurkat cells to evaluate its effects on T cell polarization. FOXO1, CD3d, ITK, NFATc3, NFATc4, and IL‐2RA were targeted by miR‐182, which their expression decreased in PBMCs of patients. Although IL‐6, IL‐17, and TGF‐β increased after miR‐182 transduction, IL‐2 dramatically decreased. We revealed CD4+FOXP3+ T cell differentiation in the miR‐182‐transduced group. Although miR‐182 has Inhibitory effects on T cells by inhibition of FOXO1, TCR/CD3 complex, NFATs, and IL‐2/IL‐2RA signaling pathways, it increases FOXP3, TGF‐β, and IL‐17 expression to possibly drive T cell deviation toward the transitional state of IL‐17‐producing Tregs and Treg formation in the end.
... Moreover, antisense oligonucleotide-mediated splicing of FOXP3 exon 7 in naive CD4 + T cells promotes the differentiation of proinflammatory Th17 cells (55). Mostly described for inflamed tissues, phenotypically mixed CD4 + FOXP3 + IL-17A + /ROR-γt + T cells have been identified [reviewed in Du et al. (64)]. However, the extent of FOXP3 exon 7 splicing and the threshold that sufficiently drives Treg-cell plasticity in these cells is currently unknown. ...
The transcription factor FOXP3 controls the immunosuppressive program in CD4+ T cells that is crucial for systemic immune regulation. Mutations of the single X-chromosomal FOXP3 gene in male individuals cause the inherited autoimmune disease immune dysregulation, polyendocrinopathy, enteropathy, and X-linked (IPEX) syndrome. Insufficient gene expression and impaired function of mutant FOXP3 protein prevent the generation of anti-inflammatory regulatory T (Treg) cells and fail to inhibit autoreactive T cell responses. Diversification of FOXP3 functional properties is achieved through alternative splicing that leads to isoforms lacking exon 2 (FOXP3Δ2), exon 7 (FOXP3Δ7), or both (FOXP3Δ2Δ7) specifically in human CD4+ T cells. Several IPEX mutations targeting these exons or promoting their alternative splicing revealed that those truncated isoforms cannot compensate for the loss of the full-length isoform (FOXP3fl). In this review, IPEX mutations that change the FOXP3 isoform profile and the resulting consequences for the CD4+ T-cell phenotype are discussed.
... Another possibility is the activation of destructive effector cells as well as protective Tregs during the active stage of LN, which therefore results in activation-induced Foxp3 mRNA (21) . Moreover, recent mice and human studies have demonstrated that Treg cells can be reprogrammed into a novel population, IL-17+Foxp3+T cells, phenotypically and functionally resembling Th17 cells under the complicated cytokine stimulation which could be one of the explanation for increased level of Foxp3 expression in active LN (22) . In summary, our result is in line with the above observations. ...
... Ces cellules TH17/Treg sont très stables et expriment à la fois FoxP3 et RORgt (Ye et al., 2013). Cependant, une étude montre que ces cellules TH17/Treg dériveraient plutôt de LTreg pouvant exprimer les facteurs spécifiques des LTH17 (Du et al., 2014b). En plus de cette conversion, les LTH17 peuvent exercer leur rôle immuno-suppresseur grâce à l'expression d'ectonucleotidases à leur surface. ...
Les cellules T auxiliaires TH17 sont caractérisées par l’expression de la cytokine IL-17A, ainsi que le facteur de transcription RORɣt. Elles sont connues pour jouer un rôle clé dans la pathogenèse de la sclérose en plaques. Ces cellules existent sous deux formes : les cellules régulatrices, immunomodulatrices, et les cellules pathogènes qui sont critiques pour l'inflammation. Il est donc important de comprendre le mécanisme qui sous-tend la différenciation des cellules TCD4+ naïves en ces deux types cellulaires. J'ai trouvé que le facteur de transcription Ikaros est un répresseur indirect de la transcription des gènes pathogéniques (Il3, Csf2, Ifng, Stat4…) dans les cellules TCD4+ naïves murines, cultivées pour induire une polarisation vers le phénotype TH17 régulateur. De plus, en absence d’Ikaros et en conditions de culture régulatrice, l’ajout d’IL-6 seul augmente l’expression de GM-CSF, facteur clé dans l’induction des maladies auto-immunes, suggérant un rôle d’Ikaros dans la régulation de cette voie. En conclusion, nos résultats suggèrent que Ikaros est nécessaire pour polariser correctement les cellules TCD4+ naïves dans le programme TH17.
... These findings suggest that Tregs cannot suppress the production of pro-inflammatory cytokines under conditions that strongly activate effector T-cells. In pro-inflammatory microenvironments, Tregs can be induced to secrete the cytokine IL-17 [29,30]. IL-17 + FoxP3 + Tregs, isolated from human peripheral blood, are suppressive in the presence of low TCR stimulation [28,29]; however, IL-17 + FoxP3 + Tregs concomitantly lose their immunosuppressive ability and gain the capacity to secrete IL-17 when they are strongly activated in the presence of pro-inflammatory cytokines IL-1β and IL-6 [29]. ...
Background
Tumor-infiltrating lymphocytes include tumor-reactive lymphocytes and regulatory T-cells. However, the prognostic value of tumor-infiltrating lymphocytes in oral squamous cell carcinoma (OSCC) remains unclear.
Methods
We used immunohistochemistry to evaluate the presence of tumor-infiltrating FoxP3⁺ T-cells and CTLA-4⁺ cells in four distinct histological compartments (tumor parenchyma and stroma at the tumor center, and parenchyma and stroma at the invasive front) and assessed the association between the prevalence of these cells and the histopathological status of 137 patients with OSCC.
Results
Five-year overall survival, disease-specific survival, and recurrence-free survival were favorable in patients with high numbers of FoxP3⁺ T-cells in the parenchyma of the invasive front. Recurrence-free survival and metastasis-free survival were decreased in patients with high numbers of CTLA-4⁺ cells in the parenchyma of the invasive front.
Conclusions
The presence of FoxP3⁺ T-cells in the parenchyma of the invasive front may be a useful prognostic factor. Our results indicate that FoxP3⁺ T-cells may exert site-specific anti-tumor effects but may not play an immunosuppressive role in OSCC. In addition, our results suggest that CTLA-4⁺ cells suppress the function of FoxP3⁺ T-cells and promote anti‐tumor immunity in OSCC.
... Conventional Tregs perform an immunosuppressive function via the production of anti-in ammatory cytokines like TGF-β, IL-35, and IL-10. But there have been studies directed towards a type of Tregs with the property of secreting proin ammatory cytokines [109][110][111][112]. IL-17Aproducing FOXP3 + Tregs originated from the induction of naïve CD25 neg Tregs either through the ectopic action of FOXP3 or TGF-β signaling [113]. ...
T cells are conventionally categorized into two basic types, viz., CD4⁺ helper and CD8⁺ cytotoxic T cells. CD4⁺ T cells were known to “help” in the activation and differentiation of various immune cells such as NK cells, macrophages, and dendritic cells, whereas CD8⁺ T cells were known to kill foreign antigens. In 1970s, it was reported that functions exhibited by T cells were not merely restricted to augmenting an immune response but also to dampen it [1]. These T supressor cells were famously named as regulatory T cells or Tregs. Suppression caused by Tregs on various T cells was believed to mediate immunological tolerance by discriminating between self- and non-self-antigen [2, 3]. Tregs are believed to play an important role in maintaining homeostasis of the immune system by restricting the enormity of effector responses and permitting the initiation of immunological tolerance [4–6]. Treg populations are majorly divided into two major types: nTregs (natural Tregs) originating from the thymus and iTregs (induced Tregs) arising extrathymically, i.e., from secondary lymphoid organs or inflamed tissues [7]. Tregs are further differentiated into five subtypes based upon their origin, phenotypes, and expression of markers.
... Activation by stimulation of T cell antigen receptor (TCR) shows the characteristics of inhibition of the proliferation of CD4 + and CD8 + T cells. 2 Transforming growth factor beta (TGF-β) can induce T cells to differentiate into Treg cells and subsets of Treg produce interleukin-10 (IL-10) and TGF-β. 3 IL-10 is an important immune mediator, with multiple effects on anti-inflammatory and immune regulatory functions. ...
Objective:
To investigate potential differences in circulating levels of T regulatory (Treg)/T helper 17 (Th17) cells, related inflammatory cytokines and specific transcription factors in healthy individuals and patients with psoriasis conforming to one of three Traditional Chinese Medicine (TCM) syndromes: blood-heat syndrome (BHS), blood-stasis syndrome (BSS) and blood-dryness syndrome (BDS).
Methods:
Sixty-seven patients with psoriasis were recruited and assigned to one of three corresponding TCM syndrome groups: BHS (n = 40), BSS (n = 14) and BDS (n = 13 patients). The control group comprised 21 healthy individuals. The circulating levels of Treg/Th17 cells in peripheral blood were assessed using flow cytometry; the levels of inflammatory cytokines interleukin (IL)-10 and tumor necrosis factor (TNF)-α by enzyme-linked immunosorbent assay; and the mRNA expression of T cell-specific transcription factors retinoic acid-related orphan receptor γt (RORγt) and forkhead box P3 (Foxp3) by quantitative real-time PCR.
Results:
The ratio of Th17 cells and the levels of TNF-α and RORγt were all significantly higher in the BHS and BSS groups than the control group (P < 0.05), while the ratio of Treg cells and the levels of IL-10 and Foxp3 mRNA in the BHS group were significantly lower compared with the control group (P < 0.05). No significant differences were seen between the BSS group and the control group. The ratio of Th17 cells and the levels of TNF-α and RORγt in the BDS group were not significantly different from those of the control group; however, the ratio of Treg cells and the levels of IL-10 and Foxp3 were all lower than those in the healthy controls (P < 0.05).
Conclusion:
Compared with healthy individuals, the ratio of Th17 cells and the levels of related cytokines were higher, while the ratio of Treg cells and the levels of related cytokines were lower, in the peripheral blood of psoriasis/BHS patients; corresponding results for the BSS and BDS groups also showed differences. We propose that patterns of differentiation of immunological cells in psoriasis patients are reflected in corresponding TCM blood syndromes.
... Knock-down of Runx1 expression significantly reduced the expression of Rorγc and Foxp3, suggesting that Runx1 is involved in the production of IL-17 in Tregs. Higher levels of Runx3 protein were also found in CD4+ CD25-Tregs producing IL-17 compared to IL-17-Tregs [85,86]. Interestingly, phenotypic and functional characterization of Th17 TILs from melanoma, ovarian, breast and colon cancers by FACS analysis revealed the possibility of the conversion of Th17 cells into Foxp3 + T cells (Figure 2e) [87]. ...
Th17 cells represent a subset of CD4+ T cells characterized by the master transcription factor RORγt and the production of IL-17. Epigenetic modifications such as post-translational histone modifications and DNA methylation play a key role in Th17 cell differentiation and high plasticity. Th17 cells are highly recruited in many types of cancer and can be associated with good or bad prognosis. Here, we will review the remodeling of the epigenome induced by the tumor microenvironment, which may explain Th17 cell predominance. We will also discuss the promising treatment perspectives of molecules targeting epigenetic enzymes to remodel a Th17-enriched tumor microenvironment.
... These data are supporting the idea that conversion of Th17 cells from Tregs is developed by stable CD25 hi Tregs. During inflammation, Foxp3 + IL-17 + CD4 + T cells are generated as an intermediate stage between Tregs and Th17 cells in the periphery [39,40]. Although Foxp3 + IL-17 + CD4 + T cells were not observed in this study, it is clear that IL-17-producing cells were developed by Tregs, not naïve T cells, because CD25 lo Tregs were slightly changed to Th17 cells in the presence of IL-33-matDCs. ...
Interleukin (IL)-33 is an alarmin factor that is highly secreted in a variety of autoimmune diseases, induces maturation of dendritic cells (DCs) and differentiation of T helper 17 (Th17) cells. As the balance between Th17 cells and regulatory T cells (Tregs) is important to maintain immune homeostasis, in this study, we investigated the effects of IL-33 on Treg cell response. We observed that direct treatment with IL-33 had no effect on Treg differentiation, whereas IL-33-matured DCs (IL33-matDCs) inhibited the differentiation of CD4+ T cells to Tregs by decreasing the expression of Foxp3. Furthermore, co-culture with IL-33-matDCs changed stable Tregs (CD25hiCD4+ Tregs) to IL-17-producing cells, whereas IL-33-matDCs had little effects on unstable Tregs (CD25loCD4+ Tregs). The stable Tregs were demonstrated to express high levels of IL-6 receptors. Blocking of IL-6 secreted from IL-33-matDCs suppressed the conversion of Tregs to Th17 cells, indicating the greater propensity to convert stable Tregs to Th17 cells is due to IL-6 signaling. Taken together, these results demonstrate that IL-33 inhibits Treg differentiation and the conversion of stable Tregs to Th17 cells via DCs.
... Th17-like Tregs are preferentially generated from Tregs. It has been reported that Th17-like Tregs can be generated from either Foxp3 + Tregs or Foxp3 − Th17 cells depending on the inflammatory context 25 . To determine if IL-23 driven Th17-like Tregs could be generated from Treg or conventional T (Tconv) cells, GFP − (Tconv) and GFP + (Treg) CD4 + T cells were sorted from Foxp3-GFP reporter mice. ...
Foxp3+ regulatory T cells (Tregs) represent a major fraction of skin resident T cells. Although normally protective, Tregs have been shown to produce pro-inflammatory cytokines in human diseases, including psoriasis. A significant hurdle in the Treg field has been the identification, or development, of model systems to study this Treg plasticity. To overcome this gap, we analyzed skin resident Tregs in a mouse model of IL-23 mediated psoriasiform dermatitis. Our results demonstrate that IL-23 drove the accumulation of Tregs; including a subpopulation that co-expressed RORγt and produced IL-17A. Genesis of this population was attenuated by a RORγt inverse agonist compound and clinically relevant therapeutics. In vitro, IL-23 drove the generation of CD4+Foxp3+RORγt+IL-17A+ cells from Treg cells. Collectively, our data shows that IL-23 drives Treg plasticity by inducing a population of CD4+Foxp3+RORγt+IL-17A+ cells that could play a role in the disease pathogenesis. Through this work, we define an in vitro system and a pre-clinical in vivo mouse model that can be used to further study Treg homeostasis and plasticity in the context of psoriasis.
... CD4 + IL-17 + Foxp3 + cells were found in human tumors, and could inhibit tumor-specific CD8 + T effector cells. 22,23 Herein, we found that this population was significantly increased in NSCLC patients (Figure S1A-C), suggesting that CD4 + IL-17 + Foxp3 + cells may play a promoting role in the pathogenesis of NSCLC. ...
The presence of IL‐17‐ producing T cells has recently been reported in non‐small cell lung cancer (NSCLC) patients. However, the long‐term prognostic significance of these populations in NSCLC patients remains unknown. In this study, we collected peripheral blood from 82 NSCLC patients and 22 normal healthy donors (NCs). The percentages of IL‐17‐producing CD4⁺T (Th17), CD8⁺T (Tc17) and γδT cells (γδT17) were measured to determine their association with clinical outcomes and overall survival (OS) in NSCLC. All NSCLC patients were followed up until July 2018. The median follow‐up time was 13.5 months (range 1‐87 months). The 3‐year and 5‐year survival rate was 27% and 19.6%, respectively. We found that Th17 cells and γδT17 cells were significantly increased, whereas the Tc17 cells were markedly decreased in patients with NSCLC compared with those in NCs. In addition, Th17 cells were significantly positively associated with Th1, whereas γδT17 cells were significantly negatively associated with γδT ⁺IFN‐γ⁺ cells. High percentages of peripheral Tc17 cells were significantly associated with favorable 5‐year OS (P = 0.025), especially in patients with early TNM stage (P = 0.016). Furthermore, high percentages of peripheral Th17 cells were positively associated with favorable 5‐year OS in patients with late TNM stage (P = 0.002). However, no significant association was observed between γδT17 cells and OS, regardless of the TNM stage. In conclusion, our findings suggested that enhanced Th17 and reduced Tc17 cells, in the peripheral blood could be a significant predictor of a favorable prognosis for NSCLC patients.
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... They represent partially Th17 (inflammatory) and Treg (inhibitory) cells. Moreover, IL-17 + Foxp3 + T cells, as proinflammatory Treg cells produce IL-17 and moderate levels of IL-2, IFN-γ, and TNF-α resulting in the aggravated inflammatory response (56). Identification of FoxP3 hi and FoxP3 lo -IL-17 producing Tregs is evidence for de novo FoxP3 expression in IL-17 producing T cells in human (57). ...
Gastric cancer stem cells (GCSCs), a small population among tumor cells, are responsible for tumor initiation, development, metastasis, and recurrence. They play a crucial role in immune evasion, immunomodulation, and impairment of effector immunity and believed to be emerged to change the balance of the immune system, importantly CD4+ T cells in the chronic inflamed tumor site. However, different subtypes of innate and adaptive immune cells are involved in the formation of the immune system in the tumor microenvironment, we would look at T cells in this study. Tumor microenvironment induces differentiation of CD4+ T cells into different subsets of T cells, mainly suppressive regulatory T cells (Treg), and T helper 17 (Th17) cells, although their exact role in tumor immunity is still under debate depending on tumor types and stages. Counterbalance between Th17 and Treg cells in the gastrointestinal system result in the homeostasis and normal function of the immune system, particularly mucosal immunity. Recent data demonstrated a high infiltration of Th17 and Treg cells into the gastric tumor site and proved that tumor microenvironment might disturb the balance between Th17 and Treg. It is possible to assume an association between activation of CSCs which contribute to metastasis in late stages, and the imbalanced Th17/Treg cells observed in advanced gastric cancer patients. This review intends to clarify the importance of gastric tumor microenvironment specifically CSCs in relation to Th17/Tregs balance firstly and to highlight the relevance of imbalanced Th17/Treg subsets in determining the stages and behavior of the tumor secondly. Finally, the present study suggests a clinical approach looking at the plasticity of T cells with a focus on Th17 as a promising dedicated arm in cancer immunotherapy.
... Several reports have demonstrated that TGFβ-induced iTregs can redifferentiate into FoxP3-negative conventional T cells upon restimulation in the absence of TGFβ, which suppresses Th1 and Th2 differentiation [31]. Moreover, differentiation into IL-17-producing Th17 cells is not inhibited by the presence of TGFβ and intermediate differentiation stage IL17 + FoxP3 + T cells have been described [32]. We therefore aimed to determine whether the presence of CTLA4Ig affects the cytokine profile, especially the regulatory cytokine IL-10 and the inflammatory cytokine IL-17 (Figure 4(a) and 4(b)). ...
Blockade of the CD28:CD80/86 costimulatory pathway has been shown to be potent in blocking T cell activation in vitro and in vivo . The costimulation blocker CTLA4Ig has been approved for the treatment of autoimmune diseases and transplant rejection. The therapeutic application of regulatory T cells (Tregs) has recently gained much attention for its potential of improving allograft survival. However, neither costimulation blockade with CTLA4Ig nor Treg therapy induces robust tolerance on its own. Combining CTLA4Ig with Treg therapy would be an attractive approach for minimizing immunosuppression or for possibly achieving tolerance. However, since the CD28 pathway is more complex than initially thought, the question arose whether blocking CD80/86 would inadvertently impact immunological tolerance by interfering with Treg generation and function. We therefore wanted to investigate the compatibility of CTLA4Ig with regulatory T cells by evaluating direct effects of CTLA4Ig on murine Treg generation and function in vitro . For generation of polyclonal-induced Tregs, we utilized an APC-free in vitro system and added titrated doses of CTLA4Ig at different time points. Phenotypical characterization by flow cytometry and functional characterization in suppressor assays did not reveal negative effects by CTLA4Ig. The costimulation blocker CTLA4Ig does not impair but rather improves murine iTreg generation and suppressor function in vitro .
... 25,26 Phenotypic and functional plasticity in both populations can even allow differentiated cells to redifferentiate. 25,41 Increased T helper 17 cell/regulatory T-cell ratio and hair follicle homeostasis Historically, keratotic occlusion of the terminal HFs has been regarded as the initial cause of HS. 2,5 Instabilities of the HF infundibulum have been the recent focus of HS pathogenesis. 3,4 Hyperplasia of follicular epithelium was evident in 77% of HS specimens and pronounced perifolliculitis was seen in 68%, with rupture of the follicle structure in 28% of HS specimens. ...
Background
Disintegration of the infundibula of terminal hair follicles (HF) in intertriginous skin areas exhibits the histological hallmark of hidradenitis suppurativa (HS)/acne inversa, featuring a dissecting terminal hair folliculitis. Elevated serum levels of interleukin 17 and local increase in the ratio of proinflammatory Th17 cells and antiinflammatory regulatory T cells (Tregs) have been reported. Perifollicular Tregs play a key role for HF stem cell homeostasis and infundibular integrity.
Objectives
In this review, we evaluate the Th17/Treg ratio in HS, its aggravating conditions and associated comorbidities. Furthermore, we intended to clarify whether drugs with reported beneficial effects in HS readjust the deviated Th17/Treg axis.
Methods
PubMed‐listed, peer‐reviewed original research articles characterizing Th17/Treg regulation in hidradentitis suppurativa/acne inversa and associated comorbidities have been selected.
Results
This review presents HS as a disease exhibiting an increased Th17/Treg ratio. Perifollicular deficiencies in Treg numbers or function may disturb HF stem cell homeostasis initiating infundibular dissection of terminal HFs and perifollicular inflammation. The Th17/Treg imbalance is aggravated by obesity, smoking as well as decreased Notch signalling. In addition, HS‐associated autoimmune diseases exhibit a disturbed Th17/Treg axis resulting in a Th17‐dominant state. All drugs that have beneficial effects in the treatment of HS normalise the Th17/Treg ratio.
Conclusions
HS immunopathogenesis is closely related to deviations of the Th17/ Treg balance, which may negatively affect Treg‐controlled HF stem cell homeostasis and infundibular integrity. Pharmacological intervention should not only attenuate Th17/IL‐17 signalling but should improve Treg function to stabilize HF stem cell homeostasis and infundibular integrity
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Background
Food allergy is a leading cause of anaphylaxis worldwide. Allergen‐specific immunotherapy is the only treatment shown to modify the natural history of allergic disease, but application to food allergy has been hindered by risk of severe allergic reactions and short‐lived efficacy. Allergen‐derived peptides could provide a solution. PVX108 comprises seven short peptides representing immunodominant T‐cell epitopes of major peanut allergens for treatment of peanut allergy.
Methods
Pre‐clinical safety of PVX108 was assessed using ex vivo basophil activation tests ( n = 185). Clinical safety and tolerability of single and repeat PVX108 doses were evaluated in a first‐in‐human, randomized, double‐blind, placebo‐controlled trial in peanut‐allergic adults (46 active, 21 placebo). The repeat‐dose cohort received six doses over 16 weeks with safety monitored to 21 weeks. Exploratory immunological analyses were performed at pre‐dose, Week 21 and Month 18 after treatment.
Results
PVX108 induced negligible activation of peanut‐sensitised basophils. PVX108 was safe and well tolerated in peanut‐allergic adults. There were no treatment‐related hypersensitivity events or AEs of clinical concern. The only events occurring more frequently in active than placebo were mild injection site reactions. Exploratory immunological analyses revealed a decrease in the ratio of ST2 ⁺ Th2A:CCR6 ⁺ Th17‐like cells within the peanut‐reactive Th pool which strengthened following treatment.
Conclusion
This study supports the concept that PVX108 could provide a safe alternative to whole peanut immunotherapies and provides evidence of durable peanut‐specific T‐cell modulation. Translation of these findings to clinical efficacy in ongoing Phase 2 trials would provide important proof‐of‐concept for using peptides to treat food allergy.
The helper CD4⁺ T cell-type 17 (Th17) cells and regulatory CD4⁺ T cells (Tregs) are balanced through numerous molecular regulators, particularly metabolic factors, and their alteration causes immune dysregulation. Herein, we report that peroxisome proliferator of activated receptor-alpha (Pparα), a lipid metabolism regulator, suppresses Th17 differentiation. We demonstrated that Pparα ablation improves Th17 and pro-Th17 factor HIF-1α by enhancing the expression and nuclear localization of NFκB-activator IκB kinase-alpha (IKKα). Unexpectedly, we found that IKKα directly interacts with RORγt and enhances the expression of Il17a gene. Meanwhile, IKKα also interacts with Foxp3, leading to the post-translational regulation of Foxp3 by elevating its proteasomal degradation, and influencing Th17 development. Pparα deficiency leads to enhanced Th17 development in vivo and is associated with enhanced pathology in a murine experimental autoimmune encephalomyelitis (EAE) model. Overall, our data indicate that Pparα may serve as a potential therapeutic target for autoimmune and inflammatory diseases.
Cancer stem cells (CSCs) defined as a small fraction of cells within malignancies have been isolated from tumors with different histological origins with stem related characteristics such as self-replicating potential, tumorigenesis, and therapy resistance. The dynamic communication between CSCs and tumor microenvironment particularly immune cells orchestrates their fate and plasticity as well as the patient outcome. According to recent evidence, it has been reported that they harness different immunological pathways to escape immunosurveillance and express aberrantly immunomodulatory agents or decreased levels of factors forming antigen presenting machinery (APM), subsequently followed by impaired antigen presentation and suppressed immune detection. As effective therapies are expected to be able to eradicate CSCs, mechanistic understanding of such interactions can provide insights into causes of therapy failure particularly in immunotherapy. Also, it can contribute to enhance the practical interventions against CSCs and their immunomodulatory features resulting in CSCs eradication and improving patient clinical outcome. The aim of this review is to explain the present knowledge regarding the immunobiology of CSCs and the immunoevasion mechanisms they use.
Die Rheumatoide Arthritis ist eine chronische Autoimmunerkrankung, deren Pathogenese noch unvollständig verstanden ist. Eine Dysbalance von pro- und antiinflammatorischen T-Helferzellen ist an der Entzündungsreaktion wesentlich beteiligt. Die Rolle von proinflammatorischen Th17-Zellen und regulatorischen Tregs ist bereits recht gut erforscht und bietet bereits erfolgreiche Therapieansätze. Umstritten bleibt die Rolle der sogenannten Th9-Zellen, welche pro- und antiinflammatorische Wirkungen zu vereinen scheinen. Die vorliegende Untersuchung umfasst die Daten von 7 RA-Patientinnen und -Patienten und 10 gesunden Kontrollen. Nach Isolation von naiven CD4+ T-Zellen erfolgte eine durchflusszytometrische Phänotypisierung der T-Zellen. Diese Phänotypisierung zeigte keinen relevanten Unterschied zwischen Pat und HD. Im Anschluss an eine 3-tägige Zellkultur von naiven und nicht naiven CD4+ T-Zellen unter Th9- und Th17-Stimulation erfolgte eine Auswertung mittels Durchflusszytometrie, RT-PCR und ELISA. Hierbei wurde eine sehr ähnliche Wirkung beider Stimuli auf die T-Zelldifferenzierung, die Zytokinproduktion und die Expression von Oberflächenmarkern beobachtet. Insbesondere die IL-9-Produktion wurde durch beide Stimuli signifikant induziert. Es scheint, dass IL-9-produzierende Zellen durch unterschiedliche Differenzierungswege induziert werden. Dies unterstützt die kontroverse Hypothese, dass Th9-Zellen keinen eigenständigen T-Helferzellsubtyp darstellen. Im Vergleich zu den Pat in Remission war die induzierte Zytokinproduktion (IL-9, IL-17, IFNγ) bei den HD betont. Dies liegt vermutlich an unspezifischen Effekten durch die immunsuppressive Therapie. Bei den beiden Patientinnen und Patienten mit der höchsten Krankheitsaktivität jedoch wurde die höchste IL-9-Konzentration in den Überständen nach Stimulation festgestellt. Ein IL-9-Exzess bzw. eine IL-9-Blockade hatte keinen signifikanten Einfluss auf die T-Zelldifferenzierung, die Zytokinproduktion und die Expression der Oberflächenmarker und des Transkriptionsfaktors. Das Vorliegen eines relevanten IL-9-regulierenden, autokrinen Mechanismus ist daher unwahrscheinlich. Zusammenfassend ergeben sich Hinweise, dass es sich bei IL-9-produzierenden Zellen aufgrund der diversen Differenzierungswege nicht um einen eigenständige T-Helferzellsubtyp handelt. Es benötigt einen sehr gut ausbalancierten Zytokincocktail und vermutlich eine längere Kulturzeit, um tatsächlich Th9-Zellen zu polarisieren. Ob diese in vivo vorkommen und eine Rolle in der Pathogenese der RA spielen ist zum jetzigen Zeitpunkt ebenso unklar wie die Rolle des IL-9 in der Pathogenese der RA. Daher kommt IL-9 als therapeutisches Ziel nach jetziger Kenntnis nicht in Betracht.
Background:
Circulating CCR9+ IL-17+ regulatory T (Treg) cells and intestinal barrier biomarkers, such as trefoil factor 3 (TFF3), intestinal-fatty acid binding protein (I-FABP), and Zonulin, are associated with gastrointestinal inflammatory diseases. So far, it is still difficult for clinicians to predict early necrotizing enterocolitis (NEC).
Study design:
This study included 13 patients with stage I NEC-like presentation (early NEC), 24 patients with one stage of II or III NEC (confirmed NEC), and 80 non-NEC and nonsepsis preterm infants (control group). Another 16 patients experienced at least two stages. We used flow cytometry to measure the frequency of CCR9+ IL-17+ Treg cells and enzyme-linked immunosorbent assay to measure TFF3, I-FABP, and Zonulin levels in the peripheral blood.
Results:
The demographic and clinical characteristics of patients were comparable. Compared with controls, CCR9+ IL-17+ Treg cells were markedly increased in early NEC and slightly increased in confirmed NEC; in contrast, plasma TFF3, I-FABP, and Zonulin concentrations were notably elevated in confirmed NEC and slightly elevated or not elevated in early NEC. Moreover, for patients who experienced at least two stages, dynamic monitoring of the above indicators also verified this.
Conclusions:
This study suggested that an elevated frequency of circulating CCR9+ IL-17+ Treg cells could be a predictor of intestinal inflammation in patients with early NEC.
Cancer stem cells are a population of cells enable to reproduce the original phenotype of the tumor and capable to self-renewal, which is crucial for tumor proliferation, differentiation, recurrence, and metastasis, as well as chemoresistance. Therefore, the cancer stem cells (CSCs) have become one of the main targets for anticancer therapy and many ongoing clinical trials test anti-CSCs efficacy of plenty of drugs. This chapter describes CSCs starting from general description of this cell population, through CSCs markers, signaling pathways, genetic and epigenetic regulation, role of epithelial-mesenchymal transition (EMT) transition and autophagy, cooperation with microenvironment (CSCs niche), and finally role of CSCs in escaping host immunosurveillance against cancer.
Aim
Lupus nephritis (LN) is one of the most serious complications of SLE. Tregs (Regulatory T lymphocytes) are thought to play a part in the pathogenesis of SLE. According to recent research, Foxp3, a Treg identification marker, plays a significant role in the pathogenesis of SLE. This study aimed to compare the urinary Foxp3 mRNA levels of patients with active and inactive forms of LN and healthy control subjects to see whether it played a role in disease activity.
Methods
We measured FOXP3 messenger RNA (mRNA) expression in the urine of 50 people with active LN, 50 people with inactive lupus, and 50 healthy people.
Results
We found that the expression level of FOXP3 was significantly higher in urine from patients with active LN than from subjects with inactive lupus and healthy controls (22.93 ± 4.13 vs 5.66 ± 0.47 vs 0.57 ± 0.15copy; P < 0.001). Urinary FOXP3 mRNA level significantly correlated with SLEDAI (0.000057) In the active group, urinary FOXP3 mRNA level also significantly correlated with histological activity index (< 0.00001).
Conclusion
We concluded that urinary FOXP3 mRNA is elevated in patients with active LN and that it is linked to the SLEDAI and the severity of the disease. FOXP3 mRNA in urine sediment may be used as a non-invasive biomarker for evaluating the severity of LN and risk stratification.
Derived from a common precursor cell, the balance between Th17 and Treg cells must be maintained within immune system to prevent autoimmune diseases. IL-1β-mediated IL-1 receptor (IL-1R) signaling is essential for Th17-cell biology. Fine-tuning of IL-1R signaling is controlled by two receptors, IL-1RI and IL-RII, IL-1R accessory protein, and IL-1R antagonist. We demonstrate that the decoy receptor, IL-1RII, is important for regulating IL-17 responses in TCR-stimulated CD4 ⁺ T cells expressing functional IL-1RI via limiting IL-1β responsiveness. IL-1RII expression is regulated by NFAT via its interaction with Foxp3. The NFAT/FOXP3 complex binds to the IL-1RII promoter and is critical for its transcription. Additionally, IL-1RII expression is dysregulated in CD4 ⁺ T cells from patients with rheumatoid arthritis. Thus, differential expression of IL-1Rs on activated CD4 ⁺ T cells defines unique immunological features and a novel molecular mechanism underlies IL-1RII expression. These findings shed light on the modulatory effects of IL-1RII on Th17 responses.
Immune checkpoint proteins including programmed cell death protein 1 (PD-1), its ligand PD-L1 and cytotoxic T lymphocyte-associated antigen-4 (CTLA‐4) are involved in proliferation, angiogenesis, metastasis, chemoresistance via immune escape and immune tolerance by disturbing cytotoxic T cell activation. Though many clinical trials have been completed in several cancers by using immune checkpoint inhibitors alone or in combination with other agents to date, recently multi-target therapy is considered more attractive than monotherapy, since immune checkpoint proteins work with other components such as surrounding blood vessels, dendritic cells, fibroblasts, macrophages, platelets and extracellular matrix within tumor microenvironment. Thus, in the current review, we look back on research history of immune checkpoint proteins and discuss their associations with platelets or tumor cell induced platelet aggregation (TCIPA) and FOXP3+ regulatory T cells (Tregs) related molecules involved in immune evasion and tumor progression, clinical implications of completed trial results and signaling networks by phytochemicals for combination therapy with immune checkpoint inhibitors and suggest future research perspectives.
Background
Recently beneficial roles of ginsenoside F2 (GF2), a minor constituent of Panax ginseng, have been demonstrated in diverse inflammatory diseases. However, its roles in alcoholic liver inflammation and injury have not been clearly understood. Here, we investigated the underlying mechanism by which GF2 ameliorated alcoholic liver injury
Methods
To induce alcoholic liver injury, C57BL/6J wild type (WT) or interleukin (IL)-10 knockout (KO) mice were orally administered with ethanol (3 g/kg) or ethanol containing GF2 (50 mg/kg) for 2 weeks. Liver injury and infiltration of macrophages and neutrophils were evaluated by serum biochemistry and immunohistochemistry, respectively. The changes of hepatic immune cells were assessed by flow cytometry and PCR analysis. In vitro differentiation of naïve T cells were performed
Results
GF2 treatment significantly attenuated alcoholic liver injury, in which infiltrations of inflammatory macrophages and neutrophils were decreased. Moreover, the frequencies of Foxp3⁺ regulatory T cells (Tregs) increased but interleukin (IL)-17-producing T (Th17) cells decreased in GF2-treated mice compared to controls. Furthermore, the mRNA expression of IL-10 and Foxp3 was significantly increased, whereas IL-17 mRNA expression was suppressed in GF-treated mice. However, these beneficial roles of GF2 were not observed in GF2-treated IL-10 KO mice, suggesting a critical role of IL-10. Similarly, GF2 treatment suppressed differentiation of naïve T cells into Th17 cells by inhibiting RORγt expression and stimulating Foxp3 expression.
Conclusion
The present study suggests that GF2 treatment attenuates alcoholic liver injury by increasing IL-10 expression and Tregs and decreasing IL-17 expression and Th17 cells.
Background:
Hemodynamic aberrations after severe burns are treated with aggressive intravenous (IV) fluid resuscitation however, oral resuscitation has been proposed in resource poor scenarios. Previously we have shown that animals receiving oral fluid following burns were able to recover kidney function. However, immune function such as circulating and splenic immune cell populations after oral or intravenous fluid administration was not examined. Herein, we perform a follow up analysis of splenic tissue and plasma from the previous animal study to examine the splenic response following these resuscitation strategies after burn injury.
Methods:
Eighteen anesthetized Yorkshire swine receiving 40%TBSA contact burns were randomized to receive either: (1) no fluids (Fluid Restricted; negative control), (2) 70 mL/kg/d Oral Rehydration Salt solution (Oral), or (3) 2 mL/kg/%TBSA/d of lactated Ringer's solution IV. Blood was drawn for blood cell analysis, and CT scans were performed before and 48 h post-burn, at which point spleens were harvested for histological, Western blot, and RT-PCR analyses.
Results:
Splenic artery diameter decreased by -0.97 ± 0.14 mm in fluid-restricted animals, while IV led to an increase of 0.68 ± 0.30 mm. No significant differences were detected in white and red pulp. IV fluids reduced the population of splenic monocytes (CD163; P = 0.001) and neutrophils (MPO protein; P = 0.13), as well as cytokines IL-8 (P = 0.003), IFN-γ (P = 0.11) and TNFα (P = 0.05). Additionally, withholding IV fluids consistently decreased the expression of FoxP3, CCR6, and IL17β in spleen, suggesting a shift in T-cell phenotype with IV resuscitation.
Conclusions:
The route of fluid administration has a minor influence on the changes in circulating and splenic leukocytes post-burn in the acute phase. Further research is needed to help guide resuscitation approaches using immunologic markers of splenic function following burns.
In classical Hodgkin lymphoma (cHL), the significance of the interplay between Hodgkin and Reed–Sternberg cells (HRS) and reactive T cells remains poorly defined. By immunohistochemistry on bioptic cHL specimens, we found that HRS and surrounding T lymphocytes stained positive for IL-17 in 40% of cases. IL-17 was detectable in a similar proportion of patients’ sera and correlated with disease burden. Supernatants of KM-H2 and HDLM-2 cHL cell lines guided preferential chemotaxis of CCR6⁺ T lymphocytes. Coculture of cHL cell lines with PBMC promoted the enrichment of Th17 lymphocytes and Foxp3⁺/IL-17⁺ cells, whereas T regulatory cells slightly decreased. Soluble CD30 downmodulated membrane CD30 expression on T cells and contributed to their polarization shift by stimulating IL-17 production and reducing IFN-γ synthesis. Thus, HRS and a number of reactive CD4⁺ T cells, attracted by tumor-secreted chemokines, produce an IL-17 tumor-shaped inflammatory milieu in a cHL subset.
L’échappement des tumeurs à la surveillance du système immunitaire est une des raisons pour lesquelles le cancer parvient à se développer. Un des objectifs de notre équipe consiste à étudier les lymphocytes T CD4+ et leurs rôles dans un contexte de cancer. Mon travail de thèse a eu pour objectif de déterminer si les résultats murins obtenus au laboratoire étaient transposables à l’Homme, et surtout dans le cancer du sein. Nous avons tout d’abord mis en évidence que les lymphocytes Th17 infiltrant les cancers du sein inhibent les fonctions effectrices des lymphocytes T cytotoxiques de manière dépendante des ectonucléotidases. Enfin, nous avons montré qu’une infiltration tumorale riche en cellules Th17 est associée à un moins bon pronostic clinique des patientes atteintes de cancer du sein. Nous avons ensuite eu pour objectif de tenter de limiter la différenciation de ces cellules Th17. Nous avons démontré que l’activation de SIRT1 diminue l’acétylation de STAT3 ce qui perturbe le programme de différenciation de ces cellules. L’activation de SIRT1 in vivo limite l’expansion des Th17 et conduit à un ralentissement de la croissance tumorale. Ce concept fut validé chez l’Homme et ouvre donc la possibilité d’association d’agonistes de SIRT1 avec la chimiothérapie. Le dernier projet porte sur les lymphocytes Th9 et leur rôle pronostique dans le cancer du sein. Nous avons mis en évidence que les propriétés effectrices des lymphocytes Th9 humains pouvaient être augmentées par l’IFNα via l’activation du facteur de transcription IRF1. Nous avons enfin démontré que l’infiltration tumorale de lymphocytes Th9 était associée au meilleur pronostic des patientes.
Regulatory T lymphocytes (Tregs) exist as natural ideal immunosuppressors in the immune system. Autologous or allogeneic Treg transfusion therapy has been carried out in animal models and humans as a new strategy for treating autoimmune disease. Recent studies have shown that Tregs can be engineered with chimeric antigen receptors to be antigen-specific, which are more effective than polyclonal Tregs with fewer target limitations and a lack of major histocompatibility complex restriction. This review describes the potential for applying chimeric antigen receptor–engineered regulatory T cells in autoimmune diseases.
CD4⁺CD25⁺FoxP3⁺ regulatory T (Treg) cells play major roles in the maintenance of immune homeostasis. In this review, we comprehensively describe the relationship between tumor necrosis factor (TNF) and Treg cells, focusing on the effects of TNF on Treg cells and on TNF-producing Treg cells. Contradictory results have been reported for the effect of TNF on the suppressive activity of Treg cells. In patients with rheumatoid arthritis, TNF has been shown to reduce the suppressive activity of Treg cells. Meanwhile, however, TNF has also been reported to maintain the suppressive activity of Treg cells via a TNFR2-mediated mechanism. In addition, Treg cells have been found to acquire the ability to produce TNF under inflammatory conditions, such as acute viral hepatitis. These TNF-producing Treg cells exhibit T helper 17-like features and hold significance in various human diseases.
The cytokines Tumor necrosis factor α (TNFα) and Interleukin 1 β (IL-1β) are both strong NF-κB activators, and some of the first cytokines to be released in an inflammatory process. TNFα and IL-1β are present in many autoimmune diseases, such as rheumatoid arthritis (RA). TNFα and IL-1β-blocking therapies are quite successful and established in the treatment of RA, but may also be promising in other diseases. For the treatment of recurring autoimmune diseases, strong controlled sensor-effector cells inhibiting TNFα or IL-1β, appear highly predestined. Such cells detect a disease biomarker and autonomously react with the dose-dependent production of therapeutic proteins. Hence, we aim to harness and assemble the interactions of TNFα, IL-1β and NF-κB, which are an ideal match for synthetic biology-based circuits to rewire the transmission to approved TNFα- or IL-1β-blocking biologicals. Considering the high impact of environmental influences on the dynamics of cell-based systems, we established closed-loop controllable cytokine neutralizer cells, monitoring cytokine levels and autonomously delivering powerful biologicals. This real-time processing system may provide dose-dependent drug delivery, which may be tailored for prospective cell and gene therapies against RA, and may offer a more personalized medicine than calculated drug dosing based on body weight.
Interleukin 17 (IL-17)-producing T helper cells (T(H)-17 cells) have been characterized in mice as a distinct subset of effector cells, but their identity and properties in humans remain elusive. We report here that expression of CCR6 and CCR4 together identified human memory CD4+ T cells selectively producing IL-17 and expressing mRNA encoding the human ortholog of mouse RORgammat, a transcription factor, whereas CCR6 and CXCR3 identified T(H)1 cells producing interferon-gamma and T helper cells producing both interferon-gamma and IL-17. Memory T cells specific for Candida albicans were present mainly in the CCR6+CCR4+ T(H)-17 subset, whereas memory T cells specific for Mycobacterium tuberculosis were present in CCR6+CXCR3+ T helper type 1 subset. The elicitation of IL-17 responses correlated with the capacity of C. albicans hyphae to stimulate antigen-presenting cells for the priming of T(H)-17 responses in vitro and for the production of IL-23 but not IL-12. Our results demonstrate that human T(H)-17 cells have distinct migratory capacity and antigenic specificities and establish a link between microbial products, T helper cell differentiation and homing in response to fungal antigens.
The nuclear hormone receptor retinoic acid receptor–related orphan receptor t (RORt) is required for the generation of T helper 17 cells expressing the proinflammatory cytokine interleukin (IL)-17. In vivo, however, less than half of RORt+ T cells express IL-17. We report here that RORt+ T cells include Foxp3+ cells that coexist with IL-17–producing RORt+ T cells in all tissues examined. The Foxp3+ RORt+ T express IL-10 and CCL20, and function as regulatory T cells. Furthermore, the ratio of Foxp3+ to IL-17– producing RORt+ T cells remains remarkably constant in mice enduring infection and inflammation. This equilibrium is tuned in favor of IL-10 production by Foxp3 and CCL20, and in favor of IL-17 production by IL-6 and IL-23. In the lung and skin, the largest popu- lation of RORt+ T cells express the T cell receptor and produce the highest levels of IL- 17 independently of IL-6. Thus, potentially antagonistic proinflammatory IL-17–producing and regulatory Foxp3+ RORt+ T cells coexist and are tightly controlled, suggesting that a perturbed equilibrium in RORt+ T cells might lead to decreased immunoreactivity or, in contrast, to pathological inflammation.
Objectives:
Investigations of FOXP3 protein expression in cervical oesophageal cancer cells and the number of FOXP3 + lymphocytes infiltrating tumour tissue were undertaken.
Methods:
FOXP3 protein expression and FOXP3 + tumour-infiltrating lymphocytes were studied immunohistochemically, in cervical oesophageal cancer tissue samples from 42 cases and paracancerous tissue samples from 30 of these cases.
Results:
The percentage of parenchymal cells expressing FOXP3 protein was significantly higher in cancer tissue (42.9%, 18/42) than in paracancerous tissue 6.67% (two of 30). FOXP3 + lymphocyte infiltration was significantly more frequent in cancer (38.1%, 16/42) than in paracancerous (13.33%, four of 30) tissue. FOXP3 protein expression in cancer parenchymal cells in patients with lymph node metastasis was significantly greater than expression in those without lymph node metastasis. FOXP3 protein expression was significantly higher in cancer tissue samples from clinical stage III or IV than those from stage I or II disease. FOXP3 + lymphocyte infiltration of tumours was significantly greater in patients with lymph node metastasis than in those without metastasis.
Conclusions:
Abnormal FOXP3 expression in cervical oesophageal cancer parenchyma and FOXP3 + lymphocyte infiltration might be closely related to metastasis of this cancer by promoting immune escape of the tumour. FOXP3 might be a potential marker for the assessment of postoperative metastasis in cervical oesophageal cancer.
Treg cells are critical for the prevention of autoimmune diseases and are thus prime candidates for cell-based clinical therapy. However, human Treg cells are 'plastic', and able to produce IL-17 under inflammatory conditions. Here, we identify and characterize the human Treg sub-population that can be induced to produce IL-17 and identify its mechanisms. We confirm that a sub-population of human Treg cells produces IL-17 in vitro when activated in the presence of IL-1β, but not IL-6. "IL-17 potential" is restricted to population III (CD4(+) CD25(hi) CD127(lo) CD45RA(-) ) Treg cells expressing the natural killer cell marker CD161. We show that these cells are functionally as suppressive and have similar phenotypic/molecular characteristics to other sub-populations of Treg cells and retain their suppressive function following IL-17 induction. Importantly, we find that IL-17 production is STAT3-dependent, with Treg cells from patients with STAT3 mutations unable to make IL-17. Finally, we show that CD161(+) population III Treg cells accumulate in inflamed joints of patients with inflammatory arthritis and are the predominant IL-17 producing Treg-cell population at these sites. As IL-17 production from this Treg-cell sub-population is not accompanied by a loss of regulatory function, in the context of cell therapy, exclusion of these cells from the cell product may not be necessary. This article is protected by copyright. All rights reserved.
Whereas Th17 cells are associated with aggravated inflammation, regulatory T cells (Tregs) provide an environment to control overt responses. Nevertheless, Tregs display a certain degree of plasticity demonstrating that T cell differentiation processes are not absolute. Previously, we showed that human Treg clones induced B cells to produce IgG4. Here we focus on the actions of freshly isolated CD4(+)CD25(+)Foxp3(+)CD127(dim) Tregs on Ig production by B cells and the consequences of prior TLR activation of B cells. In the absence of TLR stimuli, Tregs, but not conventional T cells, dampened B cell proliferation, plasma cell formation and, with the exception of IgG4, all other Ig production. Although IgG4 levels were unchanged in total B cell:Treg co-cultures, levels were increased in Treg co-cultures of naive, but not memory, B cells. Triggering TLR on B cells skewed both Ig and cytokine secretion patterns and, surprisingly, Tregs within TLR4- and TLR9- but not TLR2-triggered B cell co-cultures up-regulated retinoic acid related orphan receptor (RORC) and produced IL-17. These data indicate that under conditions like bacterial or viral infections, B cells can escape Treg control, and provides an explanation as to why patients suffering from allergy or helminth infections display polar immunopathological symptoms despite being exposed to the same agent.
Defective development and function of CD4CD25Foxp3 regulatory T cells (Tregs) contribute to the pathogenesis of psoriasis and other autoimmune diseases. Little is known about the influence of adhesions molecules on the differentiation of Foxp3 Tregs into proinflammatory Th17 cells occurring in lesional skin and blood of psoriasis patients. In the CD18 PL/J mouse model of psoriasis, reduced expression of CD18/β integrin to 2-16% of wild-type levels is associated with progressive loss of Tregs, impaired cell-cell contact between Tregs and dendritic cells (DCs), as well as Treg dysfunction as reported earlier. In the present investigation, Tregs derived from CD18 PL/J mice were analyzed for their propensity to differentiate into IL-17-producing Th17 cells in vivo and in in vitro Treg-DC cocultures. Adoptively transferred CD18 PL/J Tregs were more inclined toward conversion into IL-17-producing Th17 cells in vivo in an inflammatory as well as noninflammatory environment compared with CD18 PL/J Tregs. Addition of neutralizing Ab against CD18 to Treg-DC cocultures in vitro promoted conversion of CD18 PL/J Tregs to Th17 cells in a dose-dependent manner similar to conversion rates of CD18 PL/J Tregs. Reduced thymic output of naturally occurring Tregs and peripheral conversion of Tregs into Th17 cells therefore both contribute to the loss of Tregs and the psoriasiform dermatitis observed in CD18 PL/J mice. Our data overall indicate that CD18 expression levels impact Treg development as well as Treg plasticity and that differentiation of Tregs into IL-17-producing Th17 cells is distinctly facilitated by a subtotal deficiency of CD18.
The role of regulatory T cells (T(regs)) in human colon cancer (CC) remains controversial: high densities of tumor-infiltrating T(regs) can correlate with better or worse clinical outcomes depending on the study. In mouse models of cancer, T(regs) have been reported to suppress inflammation and protect the host, suppress T cells and protect the tumor, or even have direct cancer-promoting attributes. These different effects may result from the presence of different T(reg) subsets. We report the preferential expansion of a T(reg) subset in human CC with potent T cell-suppressive, but compromised anti-inflammatory, properties; these cells are distinguished from T(regs) present in healthy donors by their coexpression of Foxp3 and RORγt. T(regs) with similar attributes were found to be expanded in mouse models of hereditary polyposis. Indeed, ablation of the RORγt gene in Foxp3(+) cells in polyp-prone mice stabilized T(reg) anti-inflammatory functions, suppressed inflammation, improved polyp-specific immune surveillance, and severely attenuated polyposis. Ablation of interleukin-6 (IL-6), IL-23, IL-17, or tumor necrosis factor-α in polyp-prone mice reduced polyp number but not to the same extent as loss of RORγt. Surprisingly, loss of IL-17A had a dual effect: IL-17A-deficient mice had fewer polyps but continued to have RORγt(+) T(regs) and developed invasive cancer. Thus, we conclude that RORγt has a central role in determining the balance between protective and pathogenic T(regs) in CC and that T(reg) subtype regulates inflammation, potency of immune surveillance, and severity of disease outcome.
CD4(+)Foxp3(+) T regulatory cells (Tregs) display phenotypic and functional plasticity that is regulated by cytokines and other immune cells. Previously, we determined that during co-culture with CD4(+)CD25(-) T cells and antigen presenting cells, Tregs produced IL-17. Here, we investigated the mechanisms underlying the differentiation of IL-17-producing Treg (Tr17) cells and their molecular and functional properties. We determined that during stimulation via TCR/CD3 and CD28, the combination of IL-1β and IL-2 was necessary and sufficient for the generation of Tr17 cells. Tr17 cells expressed Runx1 transcription factor, which was required for sustained expression of Foxp3 and RORγt and for production of IL-17. Surprisingly, Tr17 cells also expressed Runx3, which regulated transcription of perforin and granzyme B thereby mediating cytotoxic activity. Our studies indicate that Tr17 cells concomitantly express Foxp3, RORγt, Runx1 and Runx3 and are capable of producing IL-17 while mediating potent suppressive and cytotoxic function.
While natural CD4(+)Foxp3(+) regulatory T (nT(REG)) cells have long been viewed as a stable and distinct lineage that is committed to suppressive functions in vivo, recent evidence supporting this notion remains highly controversial. We sought to determine whether Foxp3 expression and the nT(REG) cell phenotype are stable in vivo and modulated by the inflammatory microenvironment. Here, we show that Foxp3(+) nT(REG) cells from thymic or peripheral lymphoid organs reveal extensive functional plasticity in vivo. We show that nT(REG) cells readily lose Foxp3 expression, destabilizing their phenotype, in turn, enabling them to reprogram into Th1 and Th17 effector cells. nT(REG) cell reprogramming is a characteristic of the entire Foxp3(+) nT(REG) population and the stable Foxp3(NEG) T(REG) cell phenotype is associated with a methylated foxp3 promoter. The extent of nT(REG) cell reprogramming is modulated by the presence of effector T cell-mediated signals, and occurs independently of variation in IL-2 production in vivo. Moreover, the gut microenvironment or parasitic infection favours the reprogramming of Foxp3(+) T(REG) cells into effector T cells and promotes host immunity. IL-17 is predominantly produced by reprogrammed Foxp3(+) nT(REG) cells, and precedes Foxp3 down-regulation, a process accentuated in mesenteric sites. Lastly, mTOR inhibition with the immunosuppressive drug, rapamycin, stabilizes Foxp3 expression in T(REG) cells and strongly inhibits IL-17 but not RORγt expression in reprogrammed Foxp3(-) T(REG) cells. Overall, inflammatory signals modulate mTOR signalling and influence the stability of the Foxp3(+) nT(REG) cell phenotype.
Interleukin-17F (IL-17F), produced by Th17 cells and other immune cells, is a member of IL-17 cytokine family with highest homology to IL-17A. IL-17F has been shown to have multiple functions in inflammatory responses. While IL-17A plays important roles in cancer development, the function of IL-17F in tumorigenesis has not yet been elucidated. In the current study, we found that IL-17F is expressed in normal human colonic epithelial cells, but this expression is greatly decreased in colon cancer tissues. To examine the roles of IL-17F in colon cancer, we have used IL-17F over-expressing colon cancer cell lines and IL-17F-deficient mice. Our data showed decreased tumor growth of IL-17F-transfected HCT116 cells comparing to mock transfectants when transplanted in nude mice. Conversely, there were increased colonic tumor numbers and tumor areas in Il-17f(-/-) mice than those from wild-type controls after colon cancer induction. These results indicate that IL-17F plays an inhibitory role in colon tumorigenesis in vivo. In IL-17F over-expressing tumors, there was no significant change in leukocyte infiltration; instead, we found decreased VEGF levels and CD31(+) cells. While the VEGF levels were increased in the colon tissues of Il-17f(-/-) mice with colon cancer. Together, our findings demonstrate a protective role for IL-17F in colon cancer development, possibly via inhibiting tumor angiogenesis.
The maintenance of immune homeostasis requires regulatory T cells (Treg cells). Here we found that Treg cell–specific ablation of Ubc13, a Lys63 (K63)-specific ubiquitin-conjugating enzyme, caused aberrant T cell activation and autoimmunity. Although Ubc13 deficiency did not affect the survival of Treg cells or expression of the transcription factor Foxp3, it impaired the in vivo suppressive function of Treg cells and rendered them sensitive to the acquisition of T helper type 1 (TH1) cell– and interleukin 17 (IL-17)-producing helper T (TH17) cell–like effector phenotypes. This function of Ubc13 involved its downstream target, the kinase IKK. The Ubc13-IKK signaling axis controlled the expression of specific Treg cell effector molecules, including IL-10 and SOCS1. Collectively, our findings suggest that the Ubc13-IKK signaling axis regulates the molecular program that maintains Treg cell function and prevents Treg cells from acquiring inflammatory phenotypes.
Helios, an Ikaros family transcription factor, is preferentially expressed at the mRNA and protein level in regulatory T cells. Helios expression previously appeared to be restricted to thymic-derived Treg. Consistent with recent data, we show here that Helios expression is inducible in vitro under certain conditions. To understand phenotypic and functional differences between Helios(+) and Helios(-) Treg, we profiled cell-surface markers of FoxP3(+) Treg using unmanipulated splenocytes. We found that CD103 and GITR are expressed at high levels on a subset of Helios(+) Treg and that a Helios(+) Treg population could be significantly enriched by FACS sorting using these two markers. Quantitative real-time PCR (qPCR) analysis revealed increased TGF-β message in Helios(+) Treg, consistent with the possibility that this population possesses enhanced regulatory potential. In tumor-bearing mice, we found that Helios(+) Treg were relatively over-represented in the tumor-mass, and BrdU studies showed that, in vivo, Helios(+) Treg proliferated more than Helios(-) Treg. We hypothesized that Helios-enriched Treg might exert increased suppressive effects. Using in vitro suppression assays, we show that Treg function correlates with the absolute number of Helios(+) cells in culture. Taken together, these data show that Helios(+) Treg represent a functional subset with associated CD103 and GITR expression.
Immunosuppressive factors such as regulatory T cells (Tregs) limit the efficacy of immunotherapies. Histone deacetylase (HDAC) inhibitors have been reported to have antitumor activity in different malignancies and immunomodulatory effects. Herein, we report the Tregs-targeting and immune-promoting effect of a class I specific HDAC inhibitor, entinostat, in combination with either IL-2 in a murine renal cell carcinoma (RENCA) model or a survivin-based vaccine therapy (SurVaxM) in a castration resistant prostate cancer (CR Myc-CaP) model.
RENCA or CR Myc-CaP tumors were implanted orthotopically or subcutaneously, respectively. Inoculated mice were randomized into four treatment groups: vehicle, entinostat, cytokine or vaccine, and combination. Tregs in the blood were assessed by FACS analysis. Real time quantitative PCR and Western blot analysis of isolated T cell subpopulations from spleen were performed to determine Foxp3 gene and protein expression. The suppressive function of Tregs was tested by T cell proliferation assay. Low dose (5 mg/kg) entinostat reduced Foxp3 levels in Tregs and this was associated with enhanced tumor growth inhibition in combination with either IL-2 or a SurVaxM vaccine. Entinostat down-regulated Foxp3 expression transcriptionally and blocked Tregs suppressive function without affecting T effector cells (Teffs). In vitro low dose entinostat (0.5 µM) induced STAT3 acetylation and a specific inhibitor of STAT3 partially rescued entinostat-induced down-regulation of Foxp3, suggesting that STAT3 signaling is involved in Foxp3 down-regulation by entinostat.
These results demonstrate a novel immunomodulatory effect of class I HDAC inhibition and provide a rationale for the clinical testing of entinostat to enhance cancer immunotherapy.
Regulatory T cells (T(regs)) can suppress a wide variety of cell types, in diverse organ sites and inflammatory conditions. Whereas T(regs) possess multiple suppressive mechanisms, the number required for maximal function is unclear. Furthermore, whether any interrelationship or cross-regulatory mechanisms exist to orchestrate and control their utilization is unknown. In this study, we assessed the functional capacity of T(regs) lacking the ability to secrete both IL-10 and IL-35, which individually are required for maximal T(reg) activity. Surprisingly, IL-10/IL-35 double-deficient T(regs) were fully functional in vitro and in vivo. Loss of IL-10 and IL-35 was compensated for by a concurrent increase in cathepsin E (Ctse) expression, enhanced TRAIL (Tnfsf10) expression, and soluble TRAIL release, rendering IL-10/IL-35 double-deficient T(regs) functionally dependent on TRAIL in vitro and in vivo. Lastly, whereas C57BL/6 T(regs) are normally IL-10/IL-35 dependent, BALB/c T(regs), which express high levels of cathepsin E and enhanced TRAIL expression, are partially TRAIL dependent by default. These data reveal that cross-regulatory pathways exist that control the utilization of suppressive mechanisms, thereby providing T(reg) functional plasticity.
IL-17-producing CD4(+) Th (Th17) cells are a unique subset of proinflammatory cells expressing the retinoic acid-related orphan receptor γt and associated with different forms of inflammatory autoimmune pathologies. The development of Th17 cells, mediated by TGF-β and IL-1, is closely related to that of FOXP3(+) suppressor/regulatory T cells (Treg). In this study, we report that ex vivo expression of IL-1RI in human circulating CD4(+) T cells identifies a subpopulation of FOXP3(+) Treg that coexpress retinoic acid-related orphan receptor γt, secrete IL-17, and are highly enriched among CCR7(+) central memory cells. Consistent with the concept that IL-1RI expression in Treg identifies a subpopulation at an early stage of differentiation, we show that, in Th17 populations differentiated in vitro from natural naive FOXP3(+) Treg, IL-1RI(+) IL-17-secreting cells are central memory cells, whereas IL-1RI(-) cells secreting IL-17 are effector memory cells. Together with the absence of detectable IL-1RI and IL-17 expression in resting naive CD4(+) T cells, these data identify circulating CCR7(+) Treg expressing IL-1RI ex vivo as early intermediates along an IL-1-controlled differentiation pathway leading from naive FOXP3(+) Treg to Th17 effectors. We further show that, whereas IL-1RI(+) central memory Treg respond to stimulation in the presence of IL-1 by generating IL-17-secreting effectors, a significant fraction of them maintain FOXP3 expression, consistent with an important role of this population in maintaining the Treg/Th17 memory pool in vivo.
Regulatory T cells (T(reg) cells) maintain immune homeostasis by limiting inflammatory responses. SOCS1 (suppressor of cytokine signaling 1), a negative regulator of cytokine signaling, is necessary for the suppressor functions of T(reg) cells in vivo, yet detailed mechanisms remain to be clarified. We found that Socs1(-/-) T(reg) cells produced high levels of IFN-γ and rapidly lost Foxp3 when transferred into Rag2(-/-) mice or cultured in vitro, even though the CNS2 (conserved noncoding DNA sequence 2) in the Foxp3 enhancer region was fully demethylated. Socs1(-/-) T(reg) cells showed hyperactivation of STAT1 and STAT3. Because Foxp3 expression was stable and STAT1 activation was at normal levels in Ifnγ(-/-)Socs1(-/-) T(reg) cells, the restriction of IFN-γ-STAT1 signaling by SOCS1 is suggested to be necessary for stable Foxp3 expression. However, Ifnγ(-/-)Socs1(-/-) T(reg) cells had hyperactivated STAT3 and higher IL-17A (IL-17) production compared with Ifnγ(-/-)Socs1(+/+) T(reg) cells and could not suppress colitis induced by naive T cells in Rag2(-/-) mice. In vitro experiments suggested that cytokines produced by Socs1(-/-) T(reg) cells and Ifnγ(-/-)Socs1(-/-) T(reg) cells modulated antigen-presenting cells for preferential Th1 and Th17 induction, respectively. We propose that SOCS1 plays important roles in T(reg) cell integrity and function by maintaining Foxp3 expression and by suppressing IFN-γ and IL-17 production driven by STAT1 and STAT3, respectively.
T cell differentiation into distinct functional effector and inhibitory subsets is regulated, in part, by the cytokine environment present at the time of antigen recognition. Here, we show that hypoxia-inducible factor 1 (HIF-1), a key metabolic sensor, regulates the balance between regulatory T cell (T(reg)) and T(H)17 differentiation. HIF-1 enhances T(H)17 development through direct transcriptional activation of RORγt and via tertiary complex formation with RORγt and p300 recruitment to the IL-17 promoter, thereby regulating T(H)17 signature genes. Concurrently, HIF-1 attenuates T(reg) development by binding Foxp3 and targeting it for proteasomal degradation. Importantly, this regulation occurs under both normoxic and hypoxic conditions. Mice with HIF-1α-deficient T cells are resistant to induction of T(H)17-dependent experimental autoimmune encephalitis associated with diminished T(H)17 and increased T(reg) cells. These findings highlight the importance of metabolic cues in T cell fate determination and suggest that metabolic modulation could ameliorate certain T cell-based immune pathologies.
Naturally occurring CD4(+)CD25(+)FOXP3(+) regulatory T cells suppress the activity of pathogenic T cells and prevent development of autoimmune responses. There is growing evidence that TLRs are involved in modulating regulatory T cell (Treg) functions both directly and indirectly. Specifically, TLR2 stimulation has been shown to reduce the suppressive function of Tregs by mechanisms that are incompletely understood. The developmental pathways of Tregs and Th17 cells are considered divergent and mutually inhibitory, and IL-17 secretion has been reported to be associated with reduced Treg function. We hypothesized that TLR2 stimulation may reduce the suppressive function of Tregs by regulating the balance between Treg and Th17 phenotype and function. We examined the effect of different TLR2 ligands on the suppressive functions of Tregs and found that activation of TLR1/2 heterodimers reduces the suppressive activity of CD4(+)CD25(hi)FOXP3(low)CD45RA(+) (naive) and CD4(+)CD25(hi)FOXP3(hi)CD45RA(-) (memory or effector) Treg subpopulations on CD4(+)CD25(-)FOXP3(-)CD45RA(+) responder T cell proliferation while at the same time enhancing the secretion of IL-6 and IL-17, increasing RORC, and decreasing FOXP3 expression. Neutralization of IL-6 or IL-17 abrogated Pam3Cys-mediated reduction of Treg suppressive function. We also found that, in agreement with recent observations in mouse T cells, TLR2 stimulation can promote Th17 differentiation of human T helper precursors. We conclude that TLR2 stimulation, in combination with TCR activation and costimulation, promotes the differentiation of distinct subsets of human naive and memory/effector Tregs into a Th17-like phenotype and their expansion. Such TLR-induced mechanism of regulation of Treg function could enhance microbial clearance and increase the risk of autoimmune reactions.
Psoriasis is an autoimmune-related chronic inflammatory skin disease that is strongly associated with IL-23 and T helper-17 (Th17) effector cytokines. In addition, CD4+CD25(high) regulatory T-cell (Treg) function appeared to be impaired in psoriasis. CD4+CD25(high)Foxp3+ Tregs are typically considered inhibitors of autoimmune responses. However, under proinflammatory conditions, Tregs can differentiate into inflammation-associated Th17 cells--a paradigm shift, with as yet largely unknown consequences for human disease initiation or progression. Th17 cells are highly proinflammatory T cells that are characterized by IL-17A and IL-22 production and expression of the transcription factor retinoic acid-related orphan receptor γt (RORγt). We here show that Tregs of patients with severe psoriasis, as compared with those of healthy controls, have an enhanced propensity to differentiate into IL-17A-producing cells on ex vivo stimulation. This enhanced Treg differentiation was linked to unexpectedly high RORγt levels and enhanced loss of Foxp3. Notably, IL-23 boosted this Treg differentiation process particularly in patients with psoriasis but less so in controls. IL-23 further reduced Foxp3 expression while leaving the high RORγt levels unaffected. The histone/protein deacetylase inhibitor, Trichostatin-A, prevented Th17 differentiation of Tregs in psoriasis patients. Importantly, IL-17A+/Foxp3+/CD4+ triple-positive cells were present in skin lesions of patients with severe psoriasis. These data stress the clinical relevance of Treg differentiation for the perpetuation of chronic inflammatory disease and may pave novel ways for immunotherapy.
Although polyclonal regulatory T cells (Tregs) that once expressed Foxp3 (ex-Tregs) derived from Foxp3(+) Tregs have been described in homeostatic and autoimmune settings, little is known regarding the influence of the tumor environment on ex-Treg development. After adoptive transfer of HY-specific green Tregs (peripheral or thymic) to Rag2(-/-) B6 female mice bearing syngeneic HY-expressing MB49 tumors, a significant fraction rapidly lost expression of Foxp3. On the second transfer to a Rag2(-/-) B6 male environment, these ex-Tregs expanded strongly, whereas Tregs that maintained expression of Foxp3 expression did not. Both FACS and quantitative real-time-PCR analysis revealed that ex-Tregs upregulated genes characteristic of a Th1 effector-memory phenotype including IFN-γ and downregulated a panel of Treg-specific genes. Peripheral HY-specific green Tregs were adoptively transferred to Rag2(-/-) B6 male mice, to dissect the factors regulating ex-Treg differentiation. Development of ex-Tregs was more efficient in the mesenteric lymph node (mLN) than peripheral lymph node environment, correlating with a much greater level of IL-6 mRNA in mLN. In addition, the preferential development of ex-Tregs in mLN was significantly impaired by cotransfer of HY-specific naive CD4 T cells. Collectively, our study not only demonstrates the plasticity of Ag-specific Tregs in the context of the tumor environment, but also defines key molecular and cellular events that modulate ex-Treg differentiation.
Accumulating evidence suggests that Th17 cells and Tregs may exhibit development plasticity and that CD4(+) Tregs can differentiate into IL-17-producing T cells; however, whether Th17 cells can reciprocally convert into Tregs has not been described. In this study, we generated Th17 clones from tumor-infiltrating T lymphocytes (TILs). We showed that Th17 clones generated from TILs can differentiate into IFN-γ-producing and FOXP3(+) cells after in vitro stimulation with OKT3 and allogeneic peripheral blood mononuclear cells. We further demonstrated that T-cell receptor (TCR) engagement was responsible for this conversion, and that this differentiation was due to the epigenetic modification and reprogramming of gene expression profiles, including lineage-specific transcriptional factor and cytokine genes. In addition to expressing IFN-γ and FOXP3, we showed that these differentiated Th17 clones mediated potent suppressive function after repetitive stimulation with OKT3, suggesting that these Th17 clones had differentiated into functional Tregs. We further demonstrated that the Th17-derived Tregs, unlike naturally occurring CD4(+) CD25(+) Tregs, did not reconvert back into Th17 cells even under Th17-biasing cytokine conditions. These results provide the critical evidence that human tumor-infiltrating Th17 cells can differentiate into Tregs and indicate a substantial developmental plasticity of Th17 cells.
Foxp3(+)CD4(+) regulatory T (Treg) cells inhibit immune responses and temper inflammation. IL-17(+)CD4(+) T (Th17) cells mediate inflammation of autoimmune diseases. A small population of IL-17(+)Foxp3(+)CD4(+) T cells has been observed in peripheral blood in healthy human beings. However, the biology of IL-17(+)Foxp3(+)CD4(+) T cells remains poorly understood in humans. We investigated their phenotype, cytokine profile, generation, and pathological relevance in patients with ulcerative colitis. We observed that high levels of IL-17(+)Foxp3(+)CD4(+) T cells were selectively accumulated in the colitic microenvironment and associated colon carcinoma. The phenotype and cytokine profile of IL-17(+)Foxp3(+)CD4(+) T cells was overlapping with Th17 and Treg cells. Myeloid APCs, IL-2, and TGF-β are essential for their induction from memory CCR6(+) T cells or Treg cells. IL-17(+)Foxp3(+)CD4(+) T cells functionally suppressed T cell activation and stimulated inflammatory cytokine production in the colitic tissues. Our data indicate that IL-17(+)Foxp3(+) cells may be "inflammatory" Treg cells in the pathological microenvironments. These cells may contribute to the pathogenesis of ulcerative colitis through inducing inflammatory cytokines and inhibiting local T cell immunity, and in turn may mechanistically link human chronic inflammation to tumor development. Our data therefore challenge commonly held beliefs of the anti-inflammatory role of Treg cells and suggest a more complex Treg cell biology, at least in the context of human chronic inflammation and associated carcinoma.
Interest in the use of regulatory T cells (Tregs) as cellular therapeutics has been tempered by reports of naturally occurring Tregs losing Foxp3 expression and producing IL-17, raising concerns over a switch to pathogenic function under inflammatory conditions in vivo. TGF-β-induced Tregs (inducible Tregs [iTregs]), generated in large numbers in response to disease-relevant Ags, represent the most amenable source of therapeutic Tregs. Using Foxp3-reporter T cells recognizing myelin basic protein (MBP), we investigated the capacity of iTregs to produce effector-associated cytokines under proinflammatory cytokine conditions in vitro and whether this translated into proinflammatory function in vivo. In contrast with naturally occurring Tregs, iTregs resisted conversion to an IL-17-producing phenotype but were able to express T-bet and to produce IFN-γ. iTregs initiated their T-bet expression during their in vitro induction, and this was dependent on exposure to IFN-γ. IL-12 reignited iTreg expression of T-bet and further promoted iTreg production of IFN-γ upon secondary stimulation. Despite losing Foxp3 expression and expressing both T-bet and IFN-γ, MBP-responsive IL-12-conditioned iTregs induced only mild CNS inflammation and only when given in high numbers. Furthermore, iTregs retained an ability to suppress naive T cell clonal expansion in vivo and protected against the development of experimental autoimmune encephalomyelitis. Therefore, despite bearing predictive hallmarks of pathogenic effector function, previously Foxp3(+) iTregs have much lower proinflammatory potential than that of MBP-responsive Th1 cells. Our results demonstrate that autoprotective versus autoaggressive functions in iTregs are not simply a binary relationship to be determined by their relative expression of Foxp3 versus T-bet and IFN-γ.
RORγt(+) T(H)17 cells are a proinflammatory CD4(+) T-cell population associated with autoimmune tissue injury. In mice, priming of T(H)17 requires TGF-β, which alone directs the priming of FOXP3(+) regulatory T cells (Treg), in association with inflammatory cytokines. Priming of human T(H)17 cells from conventional naive CD4(+) T cells under similar conditions, however, has proved difficult to achieve. Here, we report that differentiation of human T(H)17 cells preferentially occurs from FOXP3(+) naive Treg (NTreg) in the presence of IL-2 and IL-1β and is increased by IL-23 and TGF-β. IL-1β-mediated differentiation correlated with IL-1RI expression in stimulated NTreg and was accompanied by induction of RORγt along with down-regulation of FOXP3. IL-17-secreting cells in NTreg cultures cosecreted TNF-α and IL-2 and contained distinct subpopulations cosecreting or not cosecreting IFN-γ and other T(H)17-associated cytokines. Polarized NTreg contained significant subpopulations of CCR6-expressing cells that were highly enriched in IL-17-secreting cells. Finally, analysis of CCR6 expression with respect to that of IL-1RI identified distinct IL-17-secreting subpopulations that had maintained or lost their suppressive functions. Together our results support the concept that priming of human T(H)17 from naive CD4(+) T cells preferentially takes place from FOXP3(+) Treg precursors in the presence of lineage-specific polarizing factors.
Recent studies have suggested a close relationship between CD4+FOXP3+ regulatory T cells (Treg) and pro-inflammatory IL-17-producing T helper cells (TH17) expressing the lineage-specific transcription factor RORγt. We report here the unexpected finding that human memory Treg secrete IL-17 ex-vivo and constitutively express RORγt. IL-17 secreting Treg share some phenotypic and functional features with conventional TH17 cells, expressing high levels of CCR4 and CCR6 and low levels of CXCR3. However, at variance with conventional TH17 cells, they express low levels of CD161 and mostly fail to co-secrete IL-22 and TNF-α ex-vivo. Ex-vivo secretion of IL-17 and constitutive expression of RORγt by human memory Treg suggest that, beside their well known suppressive functions, these cells likely play additional yet un-described pro-inflammatory functions.
Approximately 10% of peripheral CD4+ cells and less than 1% of CD8+ cells in normal unimmunized adult mice express the IL-2 receptor a-chain (CD25++) molecules. When CD4+ cell suspensions prepared from BALB/c nu/+ mice lymph nodes and spleens were depleted of CD25++ cells by specific mAb and C, and then inoculated into BALB/c athymic nude (nu/nu) mice, all recipients spontaneously developed histologically and serologically evi dent autoimmune diseases (such as thyroiditis, gastritis, insulitis, sialoadenitis, adrenalitis, oophoritis, glomerulo nephritis, and polyarthritis); some mice also developed graft-vs-host-like wasting disease. Reconstitution of CD4+ cells within a limited period after transfer of CD4+ cells prevented these autoimmune de velopments in a dose-dependent fashion, whereas the reconstitution several days later, or inoculation of an equivalent dose of CD8+ cells, was far less efficient for the prevention. When nu/nu mice were transplanted with allogeneic skins or immunized with xenogeneic proteins at the time of CD25++ cell inoculation, they showed significantly heightened immune responses to the skins or proteins, and reconstitution of CD4+ cells normalized the responses. Taken together, these results indicate that CD4+ cells contribute to maintaining self-tolerance by down-regulating immune response to self and non-self Ags in an Ag-nonspecific manner, pre sumably at the T cell activation stage; elimination/reduction of CD4+ cells relieves this general suppres sion, thereby not only enhancing immune responses to non-self Ags, but also eliciting autoimmune responses to certain self-Ags. Abnormality of this T cell-mediated mechanism of peripheral tolerance can be a possible cause of various autoimmune diseases.
Autoimmune diseases often result from an imbalance between regulatory T (Treg) cells and interleukin-17 (IL-17)-producing T helper (TH17) cells; the origin of the latter cells remains largely unknown. Foxp3 is indispensable for the suppressive function of Treg cells, but the stability of Foxp3 has been under debate. Here we show that TH17 cells originating from Foxp3(+) T cells have a key role in the pathogenesis of autoimmune arthritis. Under arthritic conditions, CD25(lo)Foxp3(+)CD4(+) T cells lose Foxp3 expression (herein called exFoxp3 cells) and undergo transdifferentiation into TH17 cells. Fate mapping analysis showed that IL-17-expressing exFoxp3 T (exFoxp3 TH17) cells accumulated in inflamed joints. The conversion of Foxp3(+)CD4(+) T cells to TH17 cells was mediated by synovial fibroblast-derived IL-6. These exFoxp3 TH17 cells were more potent osteoclastogenic T cells than were naive CD4(+) T cell-derived TH17 cells. Notably, exFoxp3 TH17 cells were characterized by the expression of Sox4, chemokine (C-C motif) receptor 6 (CCR6), chemokine (C-C motif) ligand 20 (CCL20), IL-23 receptor (IL-23R) and receptor activator of NF-κB ligand (RANKL, also called TNFSF11). Adoptive transfer of autoreactive, antigen-experienced CD25(lo)Foxp3(+)CD4(+) T cells into mice followed by secondary immunization with collagen accelerated the onset and increased the severity of arthritis and was associated with the loss of Foxp3 expression in the majority of transferred T cells. We observed IL-17(+)Foxp3(+) T cells in the synovium of subjects with active rheumatoid arthritis (RA), which suggests that plastic Foxp3(+) T cells contribute to the pathogenesis of RA. These findings establish the pathological importance of Foxp3 instability in the generation of pathogenic TH17 cells in autoimmunity.
CD4(+) T cells stimulate immune responses through distinct patterns of cytokine produced by Th1, Th2 or Th17 cells, or inhibit immune responses through Foxp3-expressing regulatory T cells (Tregs). Paradoxically, effector T cells were recently shown to activate Tregs, however, it remains unclear which Th subset is responsible for this effect. In this study, we found that Th17 cells expressed the highest levels of TNF among in vitro generated Th subsets, and most potently promoted expansion and stabilized Foxp3 expression by Tregs when co-transferred into Rag1(-/-) mice. Both TNF and IL-2 produced by Th17 cells contributed to this effect. The stimulatory effect of Th17 cells on Tregs was largely abolished when co-transferred with TNFR2-deficient Tregs. Furthermore, Tregs deficient in TNFR2 also supported a much lower production of IL-17A and TNF expression by co-transferred Th17 cells. Thus, our data indicate that the TNF-TNFR2 pathway plays a crucial role in the reciprocal stimulatory effect of Th17 cells and Tregs. This bidirectional interaction should be taken into account when designing therapy targeting Th17 cells, Tregs, TNF and TNFR2.
Lethal autoimmunity associated with IL-2Rβ-deficient mice is prevented after thymic transgenic expression of wild-type IL-2Rβ in IL-2Rβ−/− mice (Tg −/− mice). Here, we show that CD4+CD25+ regulatory T cells were not readily detected in IL-2Rβ−/− mice, but the production of functional CD4+CD25+ T cells was reconstituted in Tg −/− mice. Adoptive transfer of normal CD4+CD25+ T cells into neonatal IL-2Rβ-deficient mice prevented this lethal autoimmune syndrome. The CD4+CD25+ T cells in disease-free adult IL-2Rβ-deficient recipient mice were present at a near normal frequency, were solely donor-derived, and depended on IL-2 for expansion. These observations indicate that the essential function of the IL-2/IL-2R system primarily lies at the level of the production of CD4+CD25+ regulatory T cells.
T helper cells that produce interleukin 17 (IL-17; 'T-H-17 cells') are a distinct subset of proinflammatory cells whose in vivo function requires IL-23 but whose in vitro differentiation requires only IL-6 and transforming growth factor-beta (TGF-beta). We demonstrate here that IL-6 induced expression of IL-21 that amplified an autocrine loop to induce more IL-21 and IL-23 receptor in naive CD4(+) T cells. Both IL-21 and IL-23, along with TGF-beta, induced IL-17 expression independently of IL-6. The effects of IL-6 and IL-21 depended on STAT3, a transcription factor required for the differentiation of T-H-17 cells in vivo. IL-21 and IL-23 induced the orphan nuclear receptor ROR gamma t, which in synergy with STAT3 promoted IL-17 expression. IL-6 therefore orchestrates a series of 'downstream' cytokine-dependent signaling pathways that, in concert with TGF-beta, amplify ROR gamma t-dependent differentiation of TH-17 cells.
Purpose:
Regulatory T cells (Tregs) are potent tools to prevent graft-versus-host disease (GVHD) induced after allogeneic stem cell transplantation or donor lymphocyte infusions. Toward clinical application of Tregs for GVHD treatment, we investigated the impact of Tregs on the therapeutic graft-versus-tumor (GVT) effect against human multiple myeloma tumors with various immunogenicities, progression rates, and localizations in a humanized murine model.
Experimental design:
Immunodeficient Rag2(-/-)γc(-/-) mice, bearing various human multiple myeloma tumors, were treated with human peripheral blood mononuclear cell (PBMC) alone or together with autologous ex vivo cultured Tregs. Mice were analyzed for the in vivo engraftment, homing of T-cell subsets, development of GVHD and GVT. In additional in vitro assays, Tregs that were cultured together with bone marrow stromal cells were analyzed for phenotype and functions.
Results:
Treatment with PBMC alone induced variable degrees of antitumor response, depending on the immunogenicity and the growth rate of the tumor. Coinfusion of Tregs did not impair the antitumor response against tumors residing within the bone marrow, irrespective of their immunogenicity or growth rates. In contrast, Tregs readily inhibited the antitumor effect against tumors growing outside the bone marrow. Exploring this remarkable phenomenon, we discovered that bone marrow stroma neutralizes the suppressive activity of Tregs in part via production of interleukin (IL)-1β/IL-6. We furthermore found in vitro and in vivo evidence of conversion of Tregs into IL-17-producing T cells in the bone marrow environment.
Conclusions:
These results provide new insights into the Treg immunobiology and indicate the conditional benefits of future Treg-based therapies.
A Shift in Cancer's Inflammatory Balance
One of the many factors that contribute to the initiation and progression of cancer is inflammation. Inflammation can support tumor development, both directly and indirectly, and tumors can promote a chronic inflammatory environment that results in immunosuppression, which benefits the tumor. Coussens et al. (p. 286 ) review the components of the immune system that contribute to the chronic inflammation seen in tumors. Potential therapies might shift this inflammatory environment toward one more characteristic of an acute, resolving inflammation, similar to what is observed during a pathogenic infection. Such a shift would relieve immunosuppression and drive antitumor immunity that, when combined with other therapies, may ultimately result in tumor cell clearance.
Several lines of evidence indicate the instability of CD4(+)Foxp3(+) regulatory T cells (Tregs). We have therefore investigated means of promoting the stability of Tregs. In this study, we found that the proportion of Tregs in mouse strains deficient in TNFR2 or its ligands was reduced in the thymus and peripheral lymphoid tissues, suggesting a potential role of TNFR2 in promoting the sustained expression of Foxp3. We observed that upon in vitro activation with plate-bound anti-CD3 Ab and soluble anti-CD28 Ab, Foxp3 expression by highly purified mouse Tregs was markedly downregulated. Importantly, TNF partially abrogated this effect of TCR stimulation and stabilized Foxp3 expression. This effect of TNF was blocked by anti-TNFR2 Ab, but not by anti-TNFR1 Ab. Furthermore, TNF was not able to maintain Foxp3 expression by TNFR2-deficient Tregs. In a mouse colitis model induced by transfer of naive CD4 cells into Rag1(-/-) mice, the disease could be inhibited by cotransfer of wild-type Tregs, but not by cotransfer of TNFR2-deficient Tregs. Furthermore, in the lamina propria of the colitis model, most wild-type Tregs maintained Foxp3 expression. In contrast, an increased number of TNFR2-deficient Tregs lost Foxp3 expression. Thus, our data clearly show that TNFR2 is critical for the phenotypic and functional stability of Tregs in the inflammatory environment. This effect of TNF should be taken into account when designing future therapy of autoimmunity and graft-versus-host disease by using TNF inhibitors.
Solid tumors consist of neoplastic cells, non-malignant stromal cells, and migratory hematopoietic cells. Complex interactions between the cell types in this microenvironment regulate tumor growth, progression, metastasis, and angiogenesis. The cells and mediators of inflammation form a major part of the epithelial tumor microenvironment. In some cancers, inflammatory conditions precede development of malignancy; in others, oncogenic change drives a tumor-promoting inflammatory milieu. Whatever its origin, this "smoldering" inflammation aids proliferation and survival of malignant cells, stimulates angiogenesis and metastasis, subverts adaptive immunity, and alters response to hormones and chemotherapy. Cytokines are major mediators of communication between cells in the inflammatory tumor microenvironment. It is known that neoplastic cells often over-express proinflammatory mediators including proteases, eicosanoids, cytokines, and chemokines. Several cytokines such as macrophage migratory inhibitory factor (MIF), TNF-α, IL-6, IL-17, IL-12, IL-23, IL-10, and TGF-β have been linked with both experimental and human cancers and can either promote or inhibit tumor development. MIF is a major cytokine in many cancers and there is evidence that the cytokine is produced by both malignant cells and infiltrating leukocytes. In this article we will discuss the role of cancer-associated inflammation and the particular role of MIF in malignant disease.
The transcription factor Foxp3 is essential for the development of regulatory T (Treg) cells, yet its expression is insufficient for establishing the Treg cell lineage. Here we showed that Treg cell development was achieved by the combination of two independent processes, i.e., the expression of Foxp3 and the establishment of Treg cell-specific CpG hypomethylation pattern. Both events were induced by T cell receptor stimulation. The Treg cell-type CpG hypomethylation began in the thymus and continued to proceed in the periphery and could be fully established without Foxp3. The hypomethylation was required for Foxp3(+) T cells to acquire Treg cell-type gene expression, lineage stability, and full suppressive activity. Thus, those T cells in which the two events have concurrently occurred are developmentally set into the Treg cell lineage. This model explains how Treg cell fate and plasticity is controlled and can be exploited to generate functionally stable Treg cells.
The transcription factor Foxp3 is essential for the development of regulatory T (Treg) cells, yet its expression is insufficient for establishing the Treg cell lineage. Here we showed that Treg cell development was achieved by the combination of two independent processes, i.e., the expression of Foxp3 and the establishment of Treg cell-specific CpG hypomethylation pattern. Both events were induced by T cell receptor stimulation. The Treg cell-type CpG hypomethylation began in the thymus and continued to proceed in the periphery and could be fully established without Foxp3. The hypomethylation was required for Foxp3(+) T cells to acquire Treg cell-type gene expression, lineage stability, and full suppressive activity. Thus, those T cells in which the two events have concurrently occurred are developmentally set into the Treg cell lineage. This model explains how Treg cell fate and plasticity is controlled and can be exploited to generate functionally stable Treg cells.
Foxp3(+) T regulatory (Treg) cells can be induced to produce interleukin (IL)-17 by in vitro exposure to proinflammatory cytokines, drawing into question their functional stability at sites of inflammation. Unlike their splenic counterparts, Treg cells from the inflamed central nervous system (CNS-Treg cells) during EAE resisted conversion to IL-17 production when exposed to IL-6. We show that the highly activated phenotype of CNS-Treg cells includes elevated expression of the Th1-associated molecules CXCR3 and T-bet, but reduced expression of the IL-6 receptor α chain (CD126) and the signaling chain gp130. We found a lack of IL-6 receptor on all CNS CD4(+) T cells, which was reflected by an absence of both classical and trans-IL-6 signaling in CNS CD4(+) cells, compared with their splenic counterparts. We propose that extinguished responsiveness to IL-6 (via down-regulation of CD126 and gp130) stabilizes the regulatory phenotype of activated Treg cells at sites of autoimmune inflammation.
Increasing evidence suggests that distinct inflammatory cytokines convert forkhead box protein P3 (FOXP3(+)) regulatory T-cells (Tregs) into IL-17-producing cells (Th17 cells) in vitro. However, this functional plasticity has not been examined in the pathogenesis of periodontal disease. In this study, we analyzed the IL-17A(+)FOXP3(+) cells present in periodontitis lesions to determine the association between Treg conversion and the pathogenesis of periodontitis. The immunohistochemical analysis of gingival tissues demonstrated that the numbers of Th17 cells (IL-17A(+)FOXP3(-)) and Tregs (IL-17A(-)FOXP3(+)) were greater in periodontitis lesions than in gingivitis lesions. We further identified a small number of IL-17A(+)FOXP3(+) cells in periodontitis lesions but not in gingivitis lesions. The flow cytometry analysis of CD4(+) T-cell lines established from gingival tissues and the peripheral blood of periodontitis patients showed that the proportion of Tregs was reduced and the proportion of IL-17A(+)FOXP3(+) cells among all FOXP3(+) cells was elevated in gingival tissue T-cell lines relative to the proportions in peripheral blood T-cell lines. Our findings indicate that Treg-Th17 conversion may occur in periodontitis lesions. Further studies addressing the role of Treg conversion during inflammatory responses against periodontopathic bacteria are needed.
Patients with autoimmune hepatitis (AIH) have reduced numbers and function of CD4+CD25(high)FOXP3+ T regulatory cells (Tregs). Tregs can be generated from CD25⁻ (ngTreg) cells, which suppress the immune response less efficiently than Tregs. We investigated whether their differentiation into T-helper (Th)17 cells, an effector subset that has the same CD4+ progenitors as Tregs, accounts for the reduced suppressive functions of ngTregs. We investigated whether blocking interleukin (IL)-17 increased the immunosuppressive activity of Tregs.
ngTregs were generated from 36 patients with AIH and 23 healthy subjects (controls). During Treg differentiation, expression of IL-17 was inhibited by physical removal of IL-17-secreting cells, exposure to recombinant transforming growth factor β or neutralizing antibodies against IL-6 and IL-1β (to promote differentiation of ngTregs vs Th17 cells), small inhibitory RNAs specific for the Th17 transcription factor RORC, or a combination of all these approaches.
ngTregs from patients with AIH contained greater proportions of IL-17+ and RORC+ cells than Tregs from controls. All approaches to inhibit IL-17 increased expression of FOXP3 by ngTregs and their suppressive functions. Inhibition of IL-17 led to development of ngTregs that were phenotypically stable and did not acquire proinflammatory properties after exposure to IL-6 and IL-1β.
Blocking Th17 allows ngTregs to differentiate into functionally stable immune inhibitory cells; this approach might be developed for therapy of patients with AIH.
The emerging notion of environment-induced reprogramming of Foxp3(+) regulatory T (Treg) cells into helper T (Th) cells remains controversial. By genetic fate mapping or adoptive transfers, we have identified a minor population of nonregulatory Foxp3(+) T cells exhibiting promiscuous and transient Foxp3 expression, which gave rise to Foxp3(-) ("exFoxp3") Th cells and selectively accumulated in inflammatory cytokine milieus or in lymphopenic environments including those in early ontogeny. In contrast, Treg cells did not undergo reprogramming under those conditions irrespective of their thymic or peripheral origins. Moreover, although a few Treg cells transiently lose Foxp3 expression, such "latent" Treg cells retained their memory and robustly re-expressed Foxp3 and suppressive function upon activation. This study establishes that Treg cells constitute a stable cell lineage, whose committed state in a changing environment is ensured by DNA demethylation of the Foxp3 locus irrespectively of ongoing Foxp3 expression.
There has been a long controversy as to whether interleukin-17 (IL17) has an impact on tumor growth. In order to assess whether IL17 may affect tumor growth, it would be convenient to achieve high levels of this pro-inflammatory cytokine at the tumor neo-vasculature, since IL17 is known to promote angiogenesis. Here, we have generated and tested in vivo a fusion protein, consisting of the F8 antibody (specific to the alternatively spliced EDA domain of fibronectin, a marker of angiogenesis) and of murine IL17 (mIL17). The resulting immunocytokine (termed F8-mIL17) was shown to selectively localize at the tumor neo-vasculature and to vigorously promote tumor angiogenesis, without however reducing or enhancing tumor growth rate both in immunocompetent and in immunodeficient mice.
IL-17 is a newly described, T cell-derived cytokine with ill-defined physiologic properties. As such, we examined the release of proinflammatory mediators by human macrophages in response to recombinant human (rh) IL-17. IL-1beta and TNF-alpha expression and synthesis were up-regulated by rhIL-17 in a dose (ED50 was 50 +/- 9 ng/ml)- and time-dependent fashion, with cytokine accumulation reaching a zenith after 9 h. Release of IL-6, PGE2, IL-10, IL-12, IL-1R antagonist, and stromelysin was also stimulated by rhIL-17. IL-1beta and TNF-alpha mRNA expression levels were controlled by rhIL-17 in a complex manner with an initial 30-min inhibitory phase, and then up-regulation beginning at 1 h and reaching a plateau at about 3 h. The latter expression pattern closely mirrored the nuclear accumulation of the transcription factor nuclear factor-kappaB. cAMP mimetics isobutyl-1-methylxanthine (IBMX), forskolin, PGE2, and cholera toxin reversed rhIL-17-induced release of TNF-alpha, but had no consistent effect on induced IL-1beta synthesis. Induced release of TNF-alpha was also inhibited by serine/threonine protein kinase inhibitors KT-5720 (protein kinase A) and Calphostin C (protein kinase C), mitogen-activated protein kinase kinase inhibitor PD098059, and a nonspecific tyrosine kinase inhibitor, genistein. Calphostin C alone abrogated the rhIL-17-induced release of IL-1beta. The antiinflammatory cytokines IL-4 (p < 0.01) and IL-10 (p < 0.02) completely reversed rhIL-17-stimulated IL-1beta release, while IL-13 and TGF-beta2 were partially effective (59 and 43% diminution, respectively). IL-10 exerted a significant suppressive effect on IL-17-induced TNF-alpha release (99%, p < 0.02), while the inhibitory effects of IL-4, IL-13, and TGF-beta2 on TNF-alpha secretion were partial (48, 10, and 23%, respectively). The data suggest a pivotal role for IL-17 in initiating and/or sustaining an inflammatory response.
The adoptive transfer of in vitro expanded Treg is a promising treatment option for autoimmune as well as alloantigen-induced diseases. Yet, concerns about the phenotypic and functional stability of Tregs upon in vitro culture command both careful selection of the starting population and thorough characterization of the final cell product. Recently, a high degree of developmental plasticity has been described for murine Treg and Th17 cells. Similarly, IL-17-producing FOXP3(+) cells have been detected among the CD45RA(-) memory-type subpopulation of human Tregs ex vivo. This prompted us to investigate the predisposition of human naïve and memory Tregs to develop into Th17 cells during polyclonal in vitro expansion. Here, we show that stimulation-induced DNA demethylation of RORC, which encodes the lineage-defining transcription factor for Th17 cells, occurs selectively in CD45RA(-) memory-type Tregs, irrespective of their FOXP3 expression level. On the contrary, naïve CD45RA(+) Tregs retain stable CpG methylation across the RORC locus even upon prolonged ex vivo expansion and in consequence show only a marginal tendency to express RORC and develop into IL-17-producing cells. These findings are highly relevant for the generation of therapeutic Treg products.
The pathogenesis of cancer is remained to be further understood. This study aims to investigate the role of tumour-derived Foxhead box P3 (Foxp3)(+) interleukin (IL)-17(+) T cells on suppressing tumour-specific CD8(+) T cells. Colorectal cancer (CRC) tissue was collected from surgically removed cancer tissue of 22 patients with CRC. Foxp3(+) IL-17(+) T cells in cancer tissue were examined by flow cytometry. A set of cell markers and cytokines expressed by Foxp3(+) IL-17(+) T cells were determined by immune staining. By coculture with isolated peripheral CD8(+) T cells, the immune regulatory capacity of Foxp3(+) IL-17(+) T cells was examined. The results showed that a number of Foxp3(+) IL-17(+) T cells were found in CRC tissue (22.8 ± 2.6 cells/mm(2) tissue) that was significantly more than in non-cancer colonic mucosa (5.6 ± 1.04 cells/mm(2) tissue). The Foxp3(+) IL-17(+) cells also CD4(+), CCR6(+), transforming growth factor (TGF)-beta(+) and IL-6(+) . The CD8(+) T cells proliferated markedly after exposure to tumour protein in culture that was suppressed in the presence of CRC-derived Foxp3(+) IL-17(+) T cells; the suppression was attenuated by pretreatment with anti-IL-17 antibody. We conclude that CRC-derived Foxp3(+) IL-17(+) T cells have the ability to suppress tumour-specific CD8(+) T cells. This subset of T cells may be a novel therapeutic target in the treatment of CRC.
The etiology of cancer is unclear. Recent studies indicate that some cytokines, such as interleukin (IL)-17, and regulatory T cells are involved in the development of cancer. This study aims to detect a subset of T cell, IL17+Foxp3+ T cell, in the pathogenesis of esophageal cancer (Eca). Twelve patients with squamous Eca were recruited in this study. The surgically removed Eca tissue was collected. Cells isolated from Eca tissue were analyzed by flow cytometry. The results showed that 2-10% Eca tissue-derived CD4(+) T cells expressed Foxp3; only 0.2-0.8% non-ca tissue-derived CD4(+) T cells expressed Foxp3. Further analysis showed that 3-15% Eca-isolated CD4(+) T cells were also IL-17 positive whereas only 0.4-1.5% non-ca tissue-isolated CD4(+) T cells were IL-17 positive. We also found that about 4.8-11.2% Foxp3(+) IL-17(+) T cells in isolated CD4(+) T cells from Eca tissue that were significantly less than in non-ca tissue derived CD4(+) T cells. Less than 1% Foxp3(+) IL-17(+) T cells in isolated CD4(+) T cells in both Eca patients and healthy controls. Treatment with hypoxia markedly increased the expression of IL-6 in peripheral CD68+ cells. Coculturing CD68+ cells and Foxp3+ T cells under hypoxic environment resulted in abundant expression of IL-17 in Foxp3+ T cells that could be blocked by pretreatment with either anti-IL-17 or anti-transforming growth factor beta antibodies. We conclude that IL-17+Foxp3+ T cells may contribute to the development of Eca.
The transcription factor IRF4 is involved in several T-cell-dependent chronic inflammatory diseases. To elucidate the mechanisms for pathological cytokine production in colitis, we addressed the role of the IRF transcription factors in human inflammatory bowel disease (IBD) and experimental colitis.
IRF levels and cytokine production in IBD patients were studied as well as the effects of IRF4 deficiency in experimental colitis.
In contrast to IRF1, IRF5, and IRF8, IRF4 expression in IBD was augmented in the presence of active inflammation. Furthermore, IRF4 levels significantly correlated with IL-6 and IL-17 mRNA expression and to a lesser extent with IL-22 mRNA expression in IBD. To further explore the role of IRF4 under in vivo conditions, we studied IRF4-deficient and wildtype mice in experimental colitis. In contrast to DSS colitis, IRF4 deficiency was protective in T-cell-dependent transfer colitis associated with reduced RORα/γt levels and impaired IL-6, IL-17a, and IL-22 production, suggesting that IRF4 acts as a master regulator of mucosal Th17 cell differentiation. Subsequent mechanistic studies using database analysis, chromatin immunoprecipitation, and electrophoretic mobility shift assays identified a novel IRF4 binding site in the IL-17 gene promoter. Overexpression of IRF4 using retroviral infection induced IL-17 production and IL-17 together with IL-6 induced RORγt expression.
IRF4 can directly bind to the IL-17 promotor and induces mucosal RORγt levels and IL-17 gene expression thereby controlling Th17-dependent colitis. Targeting of this molecular mechanism may lead to novel therapeutic approaches in human IBD.
Interleukin (IL)-17-producing CD4(+) helper T cells (Th17) mediate mucosal immunity and are involved in the pathogenesis of inflammatory bowel disease (IBD). They are believed to arise from the same precursor population as regulatory T (Treg) cells, but little is known about how these T-cell subsets interact under chronic inflammatory conditions. We studied Th17 and Treg cells isolated from intestinal lamina propria of patients with IBD to investigate their role in pathogenesis.
FoxP3 expression (a marker of Treg cells) and IL-17 production were assessed in CD4(+) lamina propria lymphocytes isolated from IBD patients and healthy subjects. IL-17(+)FoxP3(+) and IL-17(+) CD4(+) T-cell clones were generated by limiting dilution. An in vitro suppression assay was performed to assess the functional capacity of derived T-cell clones.
IL-17(+)FoxP3(+) T cells were identified in inflamed intestinal mucosa of patients with Crohn disease (CD), but not in patients with ulcerative colitis (UC) or healthy controls. These cells shared phenotypic characteristics of Th17 and Treg cells, and showed potent suppressor activity in vitro. Transforming growth factor-β was necessary and sufficient to induce development of an IL-17(+) FoxP3(+) cell population in CD4(+) lamina propria lymphocytes derived from patients with UC.
The inflammatory environment in the intestinal mucosa of patients with CD contributes to the generation of a distinct population of Treg cells that are FoxP3(+) and produce IL-17. These cells are likely to arise during differentiation of Th17 and Treg cells. Specific microenvironmental cues from tissues are likely to determine their commitment to either lineage and affect the balance between regulation and inflammation in the intestine.
The pathogenesis of CRC remains to be further understood. This study was designed to elucidate the role of Foxp3+IL-17+ T cells in the pathogenesis of CRC. Surgically removed CRC tissue was collected from 12 patients with CRC. The frequency and cytokine profile of Foxp3+IL-17+ T cells in CRC were examined by flow cytometry. Chemokine CXCL11 was examined in CRC tissue by Western blotting. Treg chemotaxis was examined in a transwell system. The effect of Foxp3+IL-17+ T cells on induction of cancer-initiating cells was examined; the latter's Akt and MAPK activities and colony formation were examined afterward. Abundant Foxp3+IL-17+ T cells were detected in CRC tissue that expresses high levels of TGF-β, CXCR3, CCR6, and RORγt. High levels of CXCL11 were detected in CRC tissue-derived CD68+ cells, which had a strong chemotactic effect on Foxp3+ Tregs. Hypoxia induced the expression of IL-17 in Foxp3+ Tregs; Foxp3+IL-17+ T cells were capable of inducing CRC-associated cell markers in BMMo and drove the cells to be cancer-initiating cells. High levels of phosphorylated Akt and MAPK were detected in the induced cancer-initiation cells; the latter has the capability to form a colony. CRC tissue-derived Foxp3+IL-17+ cells have the capacity to induce cancer-initiating cells.