Tet调控Th17/Treg细胞分化参与自身免疫性疾病发病的研究进展

CD4⁺ T细胞是人体免疫重要的一部分,其中辅助性T细胞17（T helper cell 17,Th17）和调节性T细胞（regulatory T cells,Treg）两个新近发现的CD4⁺ T细胞亚群在自身免疫疾病的发生发展中也起关键作用。Th17细胞能够引起自身免疫和炎性反应,而Treg细胞抑制疾病发生并维持免疫稳态;10～11易位蛋白（ten-eleven translocation,Tet）作为DNA去甲基化的关键因子,当其缺失时能够影响Th17/Treg免疫平衡。本文针对Tet调控Th17/Treg细胞分化参与自身免疫疾病发生发展的研究进展作一综述。     

Notch信号通路调控Treg/Th17细胞机制的研究进展

摘要： Notch家族是多细胞生物发育过程中一类高度保守的、 十分重要的跨膜信号蛋白, 通过与相邻细胞之间的相互作用, 在细胞增殖、 分化、 凋亡中发挥着关键作用。调节性T细胞 （Treg） 及辅助性T细胞17 （Th17） 是目前发现的一类新型CD4+ T细胞亚群, 在生理状态下, 两者可通过分泌多种细胞因子来调节机体免疫的平衡。近年来, 越来越多的研究发现Notch信号通路通过调控Treg、 Th17细胞参与机体多种疾病的发生。本文就Notch信号通路在血液系统疾病、 自身免疫系统疾病等疾病中对Treg/Th17细胞调控机制的研究进展作一简要综述。     

Targeting IL-17 in autoimmunity and inflammation

The discovery of two distinct subsets of helper T cells, IFN-γ-producing Th1 cells and IL-4-producing Th2 cells, about three decades ago enabled us to understand the immunopathology of cell-mediated and allergic inflammatory diseases in humans. The observation that T cell-mediated experimental autoimmune diseases can be induced in mice lacking Th1 and Th2 cell responses prompted many immunologists to hypothesize that there might be additional subsets in helper T cell population which mediate autoimmunity in the absence of Th1 and Th2 cells. Consequently, multiple independent research groups identified IL-17-expressing RORγt⁺CD4⁺ T cell population as a distinct subset of helper T cells which promotes autoimmune tissue inflammation. Subsequent studies have revealed that innate immune cells, including γδ T cells, NKT cells and innate lymphoid cells, also produce type 17 cytokines and contribute to tissue inflammation. In this review, we discuss our current understanding on the biology of IL-17 and the therapeutic potential of targeting IL-17 for the treatment of immune disorders in humans.     

Th17/Treg平衡在类风湿关节炎中作用的研究进展

类风湿性关节炎(rheumatoid arthritis,RA)是一种以慢性多关节滑膜炎为主要特征的自身免疫性疾病。虽然目前对于RA的确切发病机制尚不明确,但一般认为和T细胞相关。最近研究发现调节性T细胞(regulatory T cell,Treg)和Th17细胞在RA的发生发展中发挥重要作用。Th17细胞能够分泌促炎症因子IL-17,通过诱导基质金属蛋白酶(ma-trix metallo proteinases,MMPs)和破骨细胞生成,促进骨滑膜炎症、骨和关节损伤;而Treg则通过释放抑制性细胞因子IL-10和TGF-β发挥免疫效应,调控RA中的炎症性免疫应答过程。单独TGF-β作用下诱导初始T细胞分化为Treg,而在TGF-β和IL-6共同作用下诱导初始T细胞分化为Th17细胞,因此,Th17和Treg细胞在特定的细胞因子微环境下可以相互转化。调节Th17/Treg之间的平衡可能成为治疗RA的新方法。该文将对Th17/Treg平衡在RA发生发展中的调节作用作一综述。     

Generation of human regulatory T cells de novo with suppressive function prevent xenogeneic graft versus host disease

Treatment with rapamycin (RAPA) favorably affects regulatory T cells (Treg) in vivo, and RAPA induces the de novo expression of FOXP3 in murine alloantigen-specific T cells. Whether RAPA acts independently or with transforming growth factor beta (TGF-β) to produce ex vivo-induced Treg generation is unknown. Na?ve CD4(+) T cells isolated from peripheral blood mononuclear cells were stimulated with anti-CD3/CD28 coated beads in the presence of IL-2 for 5 to 7 days. Ten ng/ml of TGF-β (1 to 100 ng/mL RAPA) was added to some of the cultures. The phenotypes were analyzed with flow cytometry. The conditioned cells were cocultured with CFSE-labeled T cells in different ratios for 5 days. CFSE dilution indicating T response cell proliferation was analyzed by flow cytometry. Xenogeneic graft-versus-host disease (x-GVHD) was induced by transplanting human peripheral blood mononuclear cells into RAG2(-/-) γc(-/-) mice exposed to total body irradiation, and various factors in the subjects were subsequently compared. CD4 cells induced by rapamycin and TGF-β (CD4(RAPA/TGF-β)) expressed the natural Treg phenotypes and trafficking receptors, and no significant cytotoxicity was observed. CD4(RAPA/TGF-β) was anergic and demonstrated potent suppressive activity in vitro. Although the transfer of human peripheral blood mononuclear cells into RAG2(-/-) γc(-/-) mice caused x-GVHD, the cotransfer of CD4(RAPA/TGF-β) decreased human cell engraftment and extended survival in mice. RAPA plus TGF-β induces human na?ve T cells to become suppressor cells, a novel strategy for treating human autoimmune diseases and preventing allograft rejection.     

Molecular control of pathogenic Th17 cells in autoimmune diseases

IL-17A-producing CD4⁺ T helper cells (Th17) are crucial for the development of inflammatory and autoimmune diseases and thus are exploited for clinical immunotherapies. Emerging evidence suggests Th17 cells are heterogeneous and able to adopt both pathogenic and non-pathogenic phenotypes which are shaped by environmental and genetic factors. On one hand, IL-6 in concert with TGFβ1 can induce non-pathogenic Th17 cells (non-pTh17), which are not effective in inducing tissue inflammation. On the other hand, IL-6, IL-1β with IL-23 induce pathogenic Th17 cells (pTh17) to induce immune pathologies in various tissues. Th17 cells could be both pathogenic and non-pathogenic in a content-dependent manner in vivo. Understanding how the generation and pathogenicity of pTh17 cells are regulated will aid us to devise more effective immunotherapy. In this review, we summarize recent advances in the differentiation and regulation of Th17 cells especially pTh17 cells in vitro and in vivo. The emerging results revealing the specific molecular control of pTh17 cells are highlighted.     

Therapeutic benefits of regulating inflammation in autoimmunity

Autoimmunity results from the dysregulation of the immune system leading to tissue damage. Th1 and Th17 cells are known to be cellular mediators of inflammation in autoimmune diseases. The specific cytokine milieu within the site of inflammation or within secondary lymphatic tissues is important during the priming and effector phases of T cell response. In this review, we will address the nature of the inflammatory response in the context of autoimmune disease, specifically we will discuss the role of dendritic cells following stimulation of their innate pathogen recognition receptors in directing the development of T cell responses. We will focus on how dendritic cell subsets change the balance between major players in autoimmunity, namely Th1, Th17 and regulatory T cells. Th17 cells, once thought to only act as pathogenic effectors through production of IL-17, have been shown to have regulatory properties as well with co-production of the anti-inflammatory cytokine IL-10 by a subset now referred to as regulatory Th17 cells. IL-17 is important in the induction of autoimmune diseases such as experimental autoimmune encephalomyelitis (EAE) and inflammatory bowel disease (IBD). Study of the inflammatory process following encounter with agents that stimulate the innate immune responses such as adjuvants opens a new horizon for the discovery of therapeutic agents including those derived from microorganisms. Microbial products such as adjuvants that function as TLR ligands may stimulate the immune system by interacting with Toll-like receptors (TLR) on antigen-presenting cells. Microbial agents such as Bacille Calmette-Guérin (BCG) or Freund's adjuvant (CFA) that induce a Th17 response are protective in models of autoimmune diseases particularly EAE and type 1 diabetes (T1D). The induction of innate immunity by these microbial products alters the balance in the cytokine microenvironment and may be responsible for modulation of the inflammation and protection from autoimmunity.     

The JAK2 inhibitor AG490 regulates the Treg/Th17 balance and alleviates DSS-induced intestinal damage in IBD rats

The pathogenesis of inflammatory bowel disease (IBD) remains unclear, and it is currently believed that an imbalance in regulatory T (Treg) cells/T helper 17 cells (Th17 cells) is related to the occurrence and development of IBD. Recently, the JAK2 inhibitor AG490 has been used in animal models such as rheumatoid arthritis and bronchial asthma models and shown to exert immunoregulatory functions that improve disorder in the Treg/Th17 cell balance. This study aimed to evaluate the effect of AG490 on the intestinal inflammatory process in an IBD rat model. A dextran sulfate sodium (DSS)-induced IBD rat model was established, and disease activity index (DAI) scores were calculated. The histopathological damage score was determined by haematoxylin-eosin (H&E) staining. Treg/Th17 cells in the spleen were detected by flow cytometry. The levels of interleukin (IL)-10, IL-6 and IL-17A were detected by enzyme-linked immunosorbent assay (ELISA). AG490 attenuated DSS-induced IBD injury by regulating the Treg/Th17 balance and related cytokine secretion to reduce the DAI and colonic tissue damage. Thus, AG490 may be a new method for effective treatment of IBD.     

Dual effect of T helper cell 17 (Th17) and regulatory T cell (Treg) in liver pathological process: From occurrence to end stage of disease

Liver disease is a complicated pathological status with acute or chronic progressions, causing a series of damages to liver and massive burden to public health and society. Th17 and Treg, two subsets of CD4⁺ T helper cells, seem to keep a subtle balance in the maintenance of organic immune homeostasis including liver. The dysfunction of Th17/Treg balance in liver has been proved associated with hepatic injury and disease. Herein, we summarized the research advance of Th17 and Treg cells in different phenotypes of liver diseases in the past decade. It is known to all that hepatic diseases start from stimulations or infections like virus, autoimmune, alcohol and so on in the early stage, which would cause inflammation. With the disease consistently existed, severe outcomes like cirrhosis and hepatocellular carcinoma appear finally. In conclusion, it is found that Th17 and Treg cells serve as an important role in the immune response imbalance of liver diseases from the beginning to the end stage. However, the effect of these two subsets of CD4⁺ T helper cells is not a stereotype. Pathological role which exacerbates the disease and protective character which inhibits damage to liver are co-existed in the effect of Th17 and Treg cells. Still, more studies should be carried out to enrich the understandings of liver disease and Th17/Treg immune balance in the future.     

Thymol as a reciprocal regulator of T cell differentiation: Promotion of regulatory T cells and suppression of Th1/Th17 cells

Regulatory T cells (Tregs) are critical for maintaining immune response and enhancing their differentiation has therapeutic implications for autoimmune diseases. In this study, we investigated the effects of thymol a well-known monoterpene from Thyme on differentiation and function of Tregs. In vitro generation of Tregs from purified naïve CD4⁺CD25⁻ T cells in the presence of thymol was carried out. Suppressor activity of generated Tregs was examined by changes in the proliferation of CFSE-labeled conventional T cells. Thymol promotes differentiation of naïve CD4⁺CD25⁻ T cells to CD4⁺CD25⁺Foxp3⁺ Tregs [66.9–71.8% vs. control (47%)] and increased intensity of Foxp3 expression on Tregs (p < 0.01). In functional assay, an increased immune suppression by thymol-induced Tregs (≈2.5 times of untreated Tregs) was detected. For in vivo study, thymol was intraperitoneally administered to ovalbumin (Ova)-immunized mice. Flow cytometry assessment of spleens from thymol-treated Ova-immunized mice showed increased number of CD4⁺ Foxp3⁺ Tregs (>8%, p < 0.01(and decreased levels of CD4⁺T-bet⁺ Th1 and CD4⁺RORγt⁺ Th17 cells resulted in significant decreased Th1/Treg and Th17/Treg ratios. In ex vivo Ova challenge of splenocytes from thymol-treated Ova-immunized mice, similarly higher levels of CD4⁺ Foxp3⁺ Tregs, and also elevated TGF-β expression in CD4⁺Foxp3⁺ population (48.1% vs. 18.9% in untreated Ova-immunized group) and reduced IFN-γ-producing CD4⁺T-bet⁺ T cells and IL-17-producing CD4⁺RORγt⁺ T cells were detected. This led to marked decreased ratios of IFNγ/TGF-β and IL-17/TGF-β expressions. In conclusion, this study revealed thymol as a compound with enhancing effects on Treg differentiation and function, which may have potential benefits in treatment of immune-mediated diseases with Th1/Th17 over-activation.     

Th17-relevant cytokines vary with sera of different ANA staining patterns

Antinuclear antibodies (ANA) react with components located in the cell nucleus and cytoplasm. Differing ANA staining patterns may reflect the specificity of autoantibodies in sera and indicate some autoimmune diseases specifically, to some extent. Th17-relevant cytokines have been shown to be involved in a variety of autoimmune diseases, but not consistently. In this study, we investigated whether differences in Th17-relevant cytokines exist between different ANA pattern sera. Sera of 64 ANA-positive patients (12 homogeneous, 13 speckled particle, 11 nucleolar, 15 centromere, 6 peripheral nuclear) and 16 healthy donors were analyzed for IL-17, IL-6, IL-21, IL-22, IL-23 (p19), and TGF-β, and subsequently correlations between IL-17 and IL-6, IL-21, IL-22, IL-23, and TGF-β were analyzed. Results showed that these Th17-relevant cytokines varied with different ANA-positive sera compared with healthy donors, except TGF-β. Among them, IL-21 and IL-22 were higher with all ANA-positive sera and IL-17, IL-6, and IL-23 were higher with three or more ANA staining sera. No significant difference in these cytokines was seen between the different ANA staining sera except IL-17 levels in sera of peripheral nuclear staining positive subjects were higher than nucleolar. Additionally, in ANA-positive sera, IL-17 correlated with IL-6, IL-21, IL-22, and IL-23, but not with TGF-β. Thus, we demonstrated that Th17-relevant cytokines varied with different ANA staining pattern sera, suggesting that Th17-relevant cytokines play differing roles in autoimmune diseases.     

Butyrate inhibit collagen-induced arthritis via Treg/IL-10/Th17 axis

Rheumatoid arthritis (RA) is a chronic autoimmune disorder demanding the development of novel therapeutic strategy. Butyrate is a functional short-chain fatty acid produced by the anaerobic intestinal microbiota. This study aimed to investigate the attenuation of butyrate on RA. The collagen-induced arthritis (CIA) mouse model was established and butyrate was administered in drinking water along with the collagen immunization. The histopathological features, clinical score, paw swelling, as well as the production of pro-inflammatory cytokines including interleukin (IL)-1β, IL-6 and IL-17A were measured to determine the amelioration of butyrate on arthritis. The differentiation of Treg cells and Th17 cells in the splenic cells was assessed by flow cytometry. The expression of Foxp3, IL-10, Rorγt and IL-17A were detected by RT-PCR and FACS immunostaining. Anti-IL10R antibody was used in the CIA and CD4⁺ cell cultures to mediate the effects of butyrate. Butyrate significantly inhibited expressions of IL-1β, IL-6 and IL-17A, but promoted the expression of IL-10. Butyrate also increased systematical Treg cells and reduced Th17 cells. Mechanism study revealed that butyrate directly enhanced the polarization of Treg cells but not Th17 cells. All effects of butyrate on RA were inversed by the co-administered anti-IL10R antibody. This study showed that butyrate administration inhibited arthritis in CIA mice model, suppressed the expression of inflammatory cytokines. The modulation may be mediated the differentiation of CD4 T cells towards Treg cells, which produce anti-inflammatory cytokine IL-10, and thus influenced the function of Th17 cells.     

CNI induced Th17/Treg imbalance and susceptibility to renal dysfunction in renal transplantation

Calcineurin inhibitors (CNI) prevent graft rejection by blocking interleukin-2 (IL-2), which was required for development and function of Foxp3⁺CD4⁺CD25⁺ regulatory T cells (Treg). Recently, IL-2 was reported to play a part in the inhibition of Th17 cells. The renal transplantation recipient who used CNI regularly might have Th17/Treg imbalance with increased Th17 cells and decreased Treg cells, which would cause renal dysfunction even rejection. To assess the effect of CNI on Th17 cells and Treg cells, we included 123 renal transplantation recipients (101 in a stable stage and 22 with renal dysfunction) and 27 healthy volunteers. Among all the recipients, 103 recipients used CNI and 20 recipients used sirolimus without CNI. The recipients who used CNI were further classified into four groups according to the blood levels of CNI: Of all these subjects, Th17 and Treg frequencies in the peripheral blood were analyzed by flow cytometry (FCM). Serums IL-17, IL-23, IL-6, IFN-r, and TGF-β were analyzed by ELISA. The results demonstrated that the transplantation recipient treated by CNI revealed an obvious increase in peripheral Th17 frequencies and a significant decrease in Treg frequencies when compared with the sirolimus group and healthy people (P < 0.05). Even more, the transplantation recipient with renal dysfunction had the highest level of Th17 cells (P < 0.05) while the lowest Treg cells compared with stable recipient and healthy control, with increased serums IL-6 and IL-17. Our results indicated that CNI was associated with Th17/Treg imbalance in peripheral blood, which supported the followed generation of renal dysfunction after transplantation.     

Pre-differentiated Th1 and Th17 effector T cells in autoimmune gastritis: Ag-specific regulatory T cells are more potent suppressors than polyclonal regulatory T cells

Naïve antigen-specific CD4⁺ T cells (TxA23) induce autoimmune gastritis when transferred into BALB/c nu/nu mice. Transfer of in vitro pre-differentiated Th1 or Th17 TxA23 effector T cells into BALB/c nu/nu recipients induces distinct histological patterns of disease. We have previously shown that co-transfer of polyclonal naturally occurring Treg (nTreg) suppressed development of Th1-, but not Th17-mediated disease. Therefore, we analysed the suppressive capacity of different types of Treg to suppress Th1- and Th17-mediated autoimmune gastritis. We compared nTreg with polyclonal TGFβ-induced WT Treg (iTreg) or TGFβ-induced antigen-specific TxA23 iTreg in co-transfer experiments with Th1 or Th17 TxA23 effector T cells. 6 weeks after transfer in vitro pre-differentiated TxA23 Th1 and Th17 effector cells induced destructive gastritis. Th1-mediated disease was prevented by co-transfer of nTreg and also antigen-specific iTreg, whereas WT iTreg did not show an effect. However, Th17-mediated disease was only suppressed by antigen-specific iTreg. Pre-activation of nTreg in vitro prior to transfer did not increase their suppressive activity in Th17-mediated gastritis. Thus, antigen-specific iTreg are potent suppressors of autoimmune gastritis induced by both, fully differentiated Th1 and Th17 effector cells.