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.     

Indoles mitigate the development of experimental autoimmune encephalomyelitis by induction of reciprocal differentiation of regulatory T cells and Th17 cells

Background and Purpose

Dietary indole derivatives, indole-3-carbinol (I3C) and diindolylmethane (DIM), possess anti-cancer properties and exhibit the characteristics of aryl hydrocarbon receptor (AhR) ligands. Because AhR activation has recently been shown to regulate T cell differentiation, we tested the hypothesis that I3C and DIM may mediate anti-inflammatory properties by promoting regulatory T cell (T-regs) differentiation while inhibiting Th17 cells.

Experimental Approach

We investigated the therapeutic efficacy of I3C and DIM against experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS). The efficacy was evaluated based on clinical scores of paralysis, histopathology, serum cytokines and infiltration of T cells in the CNS. We next studied the mechanism of induction of T cells against myelin oligodendrocyte glycoprotein (MOG_35–55) peptide, both in vivo and in vitro, specifically investigating the differentiation of T-regs and Th17 cells, and determined if indoles were acting through AhR.

Key Results

Pretreatment of EAE mice with I3C or DIM completely prevented the clinical symptoms and cellular infiltration into the CNS. Also, post-treatment of EAE with I3C or DIM proved highly effective in curtailing the overall severity of the disease. In addition, I3C or DIM promoted the generation of T-regs, while down-regulating the induction of MOG-specific Th17 cells. The regulation of FoxP3 induction and suppression of Th17 cells by indoles in vivo and in vitro were found to be AhR-dependent.

Conclusions and Implications

Together, our studies demonstrate for the first time that I3C and DIM may serve as novel therapeutics to suppress neuroinflammation seen during MS through activation of AhR.     

自身免疫性甲状腺炎小鼠发病中调节性T细胞、Th17细胞的变化

目的观察高碘诱发自身免疫性甲状腺炎(AIT)动物模型中CD4+CD25+调节性T细胞、Th17细胞的变化。方法选取NOD.H-2h4雌鼠饮0.005%碘化钠水,HE染色观察淋巴细胞浸润情况并进行甲状腺炎症程度评分;测定血清甲状腺球蛋白抗体(TgA b)水平;免疫荧光染色流式细胞仪分析CD4+CD25+Foxp3+调节性T细胞、Th17细胞比例的变化;实时定量RT-PCR检测Foxp3 mRNA、IL-17 mRNA、RORγt mRNA表达水平。结果 NOD.H-2h4小鼠高碘饮水后,甲状腺炎的发生率明显高于对照组,甲状腺组织有不同程度的淋巴细胞浸润,甲状腺相对重量及血清TgA b水平均较对照组小鼠明显升高(P<0.05)。脾细胞中CD4+CD25+Foxp3+调节性T细胞所占比例和Foxp3 mRNA表达量均较对照组明显降低(P<0.01);脾细胞中Th17细胞所占比例和IL-17 mRNA表达量、RORγt mRNA表达水平均较对照组明显升高(P<0.05)。结论 NOD.H-2h4小鼠在高碘诱导发生甲状腺炎时,脾脏CD4+CD25+调节性T细胞比例明显降低,Th17细胞比例明显升高。     

Patent Evaluation: Polyamide bile acid sequestrants are more potent than cholestyramine

Summary

Novelty: Amide polymers, potentially useful in the treatment of hypercholesterolaemia, are claimed. The polymers form ionic and hydrogen bonds with bile acids, preventing reabsorption and facilitating their excretion. The compositions have greater efficacy than the known therapeutic agents, cholestyramine and cholestipol, which have a similar mode of action.

Biology: A six month in vivo canine study evaluating the ability of the compounds to lower plasma cholesterol levels is described (animals were fed on a semi-synthetic, low cholesterol diet with and without a bile acid sequestrant). The results indicate that the compounds are effective for the control of blood cholesterol levels and show superior activity to cholestyramine.

Chemistry: Dimethylaminopropylmethacrylamide (DMAPMA) is one of thirty-four polymers specifically claimed. Preparative details using standard techniques are presented.     

Th17 and regulatory T cell subsets in diseases and clinical application

In the past decade it has been established that regulatory T cells (Tregs) control all immune responses. As the induction and effector mechanisms used by Tregs are being unraveled, it is emerging that a reciprocal population of CD4(+) T lymphocytes exists in the immune system that produces inflammatory cytokine IL-17, and coined "Th17 cells". Th17 cells have been implicated in the pathogenesis of many forms of human disease. The development, function, mechanism of action, and homeostasis of Tregs and Th17 cells, and the reciprocal control between Tregs and Th17 cells were presented at the Second International Conference on Regulatory T Cells and Th17 Cells and Clinical Application in Human Diseases in Shanghai on 17-20 July 2010 (China Tregs/Th17 2010). In this Special Issue of International Immunopharmacology, several paper submitted to the conference will highlight the biology of Tregs and Th17 cells, and their clinical application in human disease.     

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.     

Rapamycin inhibits differentiation of Th17 cells and promotes generation of FoxP3+ T regulatory cells

Reciprocal differentiation of immunosuppressive CD4⁺CD25⁺FoxP3⁺ T regulatory cells (Tregs) and proinflammatory IL-17-producing cells (Th17) from naïve CD4 cells is contingent upon the cytokine environment. Using MACS-purified CD4 cells, we found that rapamycin and cyclosporine A (CsA) potently inhibited the TGFβ and IL-6-induced generation of IL-17-producing cells. Intriguingly, rapamycin promoted, while CsA markedly inhibited, TGFβ-mediated generation of Tregs. The aforementioned effects of rapamycin and CsA were also observed for Flow-sorted CD4⁺CD25⁻ T cells, indicating that the effect of these two immunosuppressive agents was based on their action on de novo generation of Tregs and Th17 cells from naïve CD4 cells. Our observation suggests a distinct mode of immunosuppressive action and tolerance induction by rapamycin and CsA. The capacity of rapamycin to generate immunosuppressive Tregs and to suppress differentiation of pathogenic Th17 cells furthers our understanding of the basis for the therapeutic immunosuppressive effects of rapamycin in patients with autoimmune diseases and allo-transplantation reactions.     

Detection of Th1, Th2 and Th17 immune responses in rats with experimental autoimmune encephalomyelitis

目的 探讨不同类型免疫反应在实验性自身免疫性脑脊髓炎(EAE)发病中的作用.方法 24只SD大鼠随机均分为EAE组和对照组,通过行为观测和大脑微观形态学确认免疫诱导的EAE模型,用ELISA法检测淋巴结和脾细胞培养上清液中IL-4、IFN-γ和血清中IgG水平,流式细胞术检测淋巴结和脾细胞中IL-17及Foxp3细胞频数.结果 与对照组相比,EAE组IgG、IFN-γ、IL-17及IL-4水平均明显增高(P＜0.05或P＜0.01).结论 Th1、Th2和Th17免疫细胞在EAE的发病均起着重要的作用.     

实验性自身免疫性脑脊髓炎大鼠Th1、Th2和Th17免疫反应检测

目的探讨不同类型免疫反应在实验性自身免疫性脑脊髓炎(EAE)发病中的作用。方法 24只SD大鼠随机均分为EAE组和对照组,通过行为观测和大脑微观形态学确认免疫诱导的EAE模型,用ELISA法检测淋巴结和脾细胞培养上清液中IL-4、IFN-γ和血清中IgG水平,流式细胞术检测淋巴结和脾细胞中IL-17及Foxp3细胞频数。结果与对照组相比,EAE组IgG、IFN-γ、IL-17及IL-4水平均明显增高(P<0.05或P<0.01)。结论 Th1、Th2和Th17免疫细胞在EAE的发病均起着重要的作用。     

Immunoregulatory Cordyceps sinensis increases regulatory T cells to Th17 cell ratio and delays diabetes in NOD mice

Cordyceps sinensis (CS) is a parasitic fungus, and it has been used widely in traditional Chinese medicines (TCM) for centuries. Many studies have shown that CS has immunoregulatory activity in many disease models, but the underlying mechanism remains elusive. We studied whether CS could suppress the onset of diabetes by altering T lymphocyte subsets in non-obese diabetic (NOD) mice. We found that the onset of type1 diabetes in NOD mice was associated with an imbalance of CD4⁺CD25⁺FoxP3⁺ regulatory T (Treg) cells and IL-17 producing Th17 cells. Oral administration of CS resulted in reduction in the overall incidence of diabetes, and this was due to an increase in the ratio of Treg cells to Th17 in the spleen and pancreatic lymph nodes (PLNs). Taken together, these data imply that CS is able to modulate Treg to Th17 cell ratio in vivo, thus contributing to the inhibition of diabetes.     

C-Phycocyanin ameliorates experimental autoimmune encephalomyelitis and induces regulatory T cells

For decades Experimental Autoimmune Encephalitis (EAE) has remained as an unsurpassed multiple sclerosis (MS) animal model. C-Phycocyanin (C-Pc) has been reported to exhibit pharmacological properties that may be expected to symptomatically improve EAE and MS. However, in this paper we reveal a basic underlying mechanism that may provide a new approach to the rationale of the overall beneficial effect of this natural antioxidant. We demonstrate that C-Pc is able to trigger mechanisms preventing or downgrading EAE expression and induces a regulatory T cell (Treg) response, in peripheral blood mononuclear cells (PBMC) from MS patients. These results agree with reports suggesting that Treg limit acute MS attacks and that C-Pc may act as a neuroprotector and thereby reverts the organic and functional damage in neurodegenerative disorders of the central nervous system (CNS). Moreover, evidence is provided on the antioxidant activity of C-Pc within the CNS, intended to improve the myelin and axonal damage of EAE induced Lewis rats. Our results indicate that specific Treg activation may represent a central and essential mechanism in supporting the therapeutic potential of C-Pc for MS and may lead to new and more effective therapies; this property would then complement and enhance other proven active principles such as interferons (IFN), giving rise to combined therapies.     

系统性红斑狼疮Th17细胞分子调控机制研究进展

系统性红斑狼疮(SLE)是一种典型的多系统炎症性自身免疫性疾病,发病机制复杂。Th17细胞是一种以分泌IL-17为主的CD4⁺T细胞亚群,与肿瘤、感染、过敏以及自身免疫性疾病等相关。Th17细胞生物学特性及其相关的细胞因子与其致病机制密切相关,研究发现Th17细胞在SLE的发生发展中发挥重要作用。归纳近年来Th17细胞的研究进展,探讨其在SLE可能的发病机制,为治疗提供新的依据。     

类风湿关节炎患者外周血Th1、Th2和Th17细胞亚群表达比例变化的特点

目的研究类风湿关节炎（rheumatoidarthritis,RA）患者外周血中Th1、Th2和Th17细胞亚群表达比例变化,分析RA免疫学发病机制。方法运用流式细胞术检测RA患者及对照组外周血单个核细胞（PBMCs）中Th1、Th2、Th17细胞表达比例。并用酶联免疫吸附法检测RA患者及对照组血清中IFN-γ、IL-4、IL-17的浓度。结果RA患者CD4＋T细胞明显高于健康对照组（P〈0．05）;活动期RA患者Th1、Th17及其分泌的细胞因子含量明显高于健康对照组（P〈0．05）;非活动期RA患者Th17也明显高于健康对照组（P〈0．05）。结论类风湿关节炎Th1、Th2、Th17平衡改变与其疾病的活动度有关,处于活动期Th1、Th17反应增强,非活动期Th1、Th2、Th17趋于平衡。通过调节Th1、Th2、Th17平衡,有可能为RA的治疗提供一条新思路。     

类风湿关节炎患者Th1、Th17细胞的表达

目的 分析类风湿关节炎(RA)患者外周血和关节滑液中Th极化细胞(Th1、Th17细胞)的表达.方法 流式细胞技术测定RA患者外周血、关节滑液中CD4T细胞上 IFN-γ~+ IL-17~- (Th1)、IL-17~1 IFN-γ(Th17)的表达,并与健康人比较,分析疾病活动指数(DAS28)和Th极化细胞表达量、Th1/Th17比值之间的关系.结果 RA患者关节滑液Th1、Th17细胞的平均百分比与RA外周血、健康人外周血比较,差异有统计学意义(P＜0.01).RA患者关节滑液中Th1细胞、Th1/Th17细胞比值与DAS28指数呈正相关(P＜0.01).结论 在RA疾病部位关节液中是Th1细胞占优势,IL-Th1细胞的表达与疾病活动性密切相关.     

CD4+CD25+调节性T细胞及Th17细胞与Graves病机制研究

目的：探讨CD4+CD25+调节性T细胞以及Th17细胞与Graves病的关系。方法：检测GD患者和对照组外周血单个核细胞CD4+CD25+Tregs、Th17细胞的数量,PBMC中TGF-β、IL-10和FoxP3 mRNA的表达水平以及血清TGF-β、IL-10和IL-17水平。结果：GD组外周血Th17数量高于对照组,血清中IL-17水平也高于对照组（P＜0.05）;而CD4+CD25+Tregs的数量低于对照组,而且TGF-β、FoxP3表达水平低于对照组（P＜0.05）;GD组血清中TGF-β、IL-10水平高于对照组（P＜0.05）。结论：CD4+CD25+Tregs数量的减少或功能障碍以及Th17细胞数量的升高,可能参与GD的发病过程。     

The relationship between TIGIT+ regulatory T cells and autoimmune disease

The role of regulatory T cells (Treg cell) in controlling autoimmune disease is an area of intense study. As such, the characterization and understanding the function of Treg markers has the potential to provide a considerable impact in developing treatments and understanding the pathogenesis of autoimmune diseases. One such inhibitory Treg cell marker that has been recently discovered is T cell immunoglobulin and ITIM domain (TIGIT). In this review, we discuss what is known about the expression and function of TIGIT on Treg cells, and we discuss the relationship between TIGIT expressing Treg cells and different autoimmune diseases such as atopic dermatitis, autoimmune thyroiditis, type 1 diabetes, autoimmune uveitis, aplastic anemia, multiple sclerosis, systemic lupus erythematosus, arthritis, and colitis.