Impaired balance of Th17/Treg in patients with nasal polyposis

Nasal polyposis (NP) is a chronic inflammatory disease of the nasal cavity and sinuses that is regulated by T lymphocyte subsets. Imbalance of Th17/Treg has been considered critical in the development of inflammation and atopic reactions. To assess whether the balance of Th17/Treg is disrupted in patients with NP, we evaluated the distribution of Th17 and Treg cells among peripheral blood mononuclear cells (PBMCs) in atopic patients with NP, non-atopic patients with NP and controls. We then determined mRNA levels of RORc and Foxp3 and protein levels of IL-17, TGF-β and IL-10 in polyp tissue among the three groups. Finally, we investigated the correlation between Th17-, Treg- and Th1-, Th2-related cytokines (INF-γ, IL-4, IL-5). The results demonstrated that both atopic and non-atopic patients with NP revealed significantly increased Th17 proportion and decreased Treg proportion in PBMCs, as well as significantly increased RORc and IL-17 levels and decreased Foxp3 and TGF-β levels in polyp tissue. Furthermore, these differences were significant between atopic and non-atopic groups. The frequency of Treg in PBMCs was found to be negatively correlated with Th1 and Th2 cytokines in polyps. These results indicated that an impaired balance of Th17/Treg existed in patients with NP and was more severe in atopic patients, suggesting that the imbalance of Treg/Th17 may play an important role in the development of NP and that atopy may aggravate NP by promoting the imbalance of Th17/Treg.     

小檗碱调节睡眠剥夺大鼠的肠道菌群结构以及Th17/Treg细胞平衡

目的研究小檗碱对睡眠剥夺大鼠肠道菌群以及辅助性T细胞17(Th17)和调解性T细胞(Treg)细胞的影响。方法将大鼠随机分为对照组、模型组、低和高剂量小檗碱组(BBR1和BBR2,100和200 mg/kg,灌胃10 d干预)。小站台水环境法建立睡眠剥夺模型。检测大鼠直肠内容物中细菌数量,流式细胞计量技术分析大鼠肠道Th17/Treg细胞比值,并检测肠道白介素17(IL-17)、RAR相关孤儿受体(ROR)C和叉头框蛋白P3(Foxp3)表达。结果模型组大鼠肠道内产气荚膜梭菌数量增加(P0.05),而其他检测菌群数量降低(P0.05);Th17/Treg细胞比值升高(P0.05);IL-17和RORC表达升高(P0.05),Foxp3表达降低(P0.05)。小檗碱处理降低产气荚膜梭菌数量(P0.05),增加其他检测菌群的数量(P0.05);降低Th17/Treg细胞比值(P0.05);下调IL-17和RORC表达(P0.05),上调Foxp3表达(P0.05)。结论小檗碱能够拮抗睡眠剥夺诱导的大鼠肠道菌群结构和Th17/Treg细胞失衡。     

Oridonin promotes CD4+/CD25+ Treg differentiation, modulates Th1/Th2 balance and induces HO-1 in rat splenic lymphocytes

Objective and design: Oridonin is an ent-kaurene diterpenoid extracted from Isodon Serra, and we have previously demonstrated its immunosuppressive effect. Our goal was to study how Oridonin impacts CD4⁺/CD25⁺ regulatory T cells (Tregs) and Th1/Th2 balance, as well as its effect on the anti-inflammatory target HO-1. Material: Splenic lymphocytes were prepared from male 6–8-week-old SD rats. Treatment: Cells were cultured in four groups as Oridonin-L (Oridonin 12.5 μmol/l), Oridonin-H (Oridonin 25 μmol/l), Cobalt protoporphyrin (Copp 50 μmol/l) and control (DMSO) with stimulation of ConA (5 μg/ml) for 48 h or with no stimulation for 12 h. Method: We set up a model of Th1 polarization in vitro using ConA stimulation; ratios of CD4⁺/CD25⁺ Tregs (confirmed by the expression of Foxp3) were measured by flow cytometry, and levels of IL-2, IFN-γ, TGF-β and IL-10 were measured by ELISA. In addition, HO-1 expression was measured without stimulation with ConA by RT-PCR and Western blotting, and HO-1 level in vitro was then measured by enzyme activity assay. p<0.05 (t-test) was taken as the level of statistical significance. Result: Oridonin promoted differentiation towards CD4⁺/CD25⁺ Tregs, inhibited IL-2 and IFN-γ but induced TGF-β and IL-10, thus rectifed the Th1 polarization. Moreover, Oridonin induced the expression of HO-1 mRNA and protein, and HO-1 activity in vitro was enhanced accordingly. Conclusion: The results suggest that Oridonin has a distinct effect on promoting CD4⁺/CD25⁺ Treg differentiation and modulating Th1/Th2 balance, and this effect may be achieved via inducing the anti-inflammatory target HO-1. Received 16 October 2007; accepted by G. Wallace 26 November 2007     

Treg/Th17平衡在巨细胞病毒感染发病机制中的作用

目的:研究调节性T细胞(Treg/Th17)细胞失衡在巨细胞病毒感染(CMV)免疫调节机制中的作用。方法:将CMV感染患儿按诊断标准分为激活感染组与潜伏感染组,同时设立正常对照组,采用流式细胞术(FCM)分析各组外周血Treg、Th17的百分比,并计算Treg/Th17比值;同时采用ELISA和RT-PCR法检测Treg主要相关因子(IL-10、Foxp3)和Th17主要相关因子(TGF-β、IL-17、IL-6、IL-23及ROR-γt)表达水平。结果:与对照组比较,CMV感染后Treg细胞百分率降低,Th17细胞升高,致Treg/Th17比值下降(P<0.05);CMV感染后两组间比较,激活感染组Treg/Th17比值和Treg主要相关因子表达水平下降更明显,而Th17主要相关因子表达水平显著上调,差异均有统计学意义(P<0.05)。结论:Treg/Th17平衡参与了CMV感染发病免疫机制,并可能与病毒潜伏-激活状态相关。     

CD11b regulates the Treg/Th17 balance in murine arthritis via IL‐6

Th17 cells are often associated with autoimmunity and been shown to be increased in CD11b^?/? mice. Here, we examined the role of CD11b in murine collagen‐induced arthritis (CIA). C57BL/6 and CD11b^?/? resistant mice were immunized with type II collagen. CD11b^?/? mice developed arthritis with early onset, high incidence, and sustained severity compared with C57BL/6 mice. We observed a marked leukocyte infiltration, and histological examinations of the arthritic paws from CD11b^?/? mice revealed that the cartilage was destroyed in association with strong lymphocytic infiltration. The CD11b deficiency led to enhanced Th17‐cell differentiation. CD11b^?/? dendritic cells (DCs) induced much stronger IL‐6 production and hence Th17‐cell differentiation than wild‐type DCs. Treatment of CD11b^?/? mice after establishment of the Treg/Th17 balance with an anti‐IL‐6 receptor mAb significantly suppressed the induction of Th17 cells and reduced arthritis severity. Finally, the severe phenotype of arthritis in CD11b^?/? mice was rescued by adoptive transfer of CD11b⁺ DCs. Taken together, our results indicate that the resistance to CIA in C57BL/6 mice is regulated by CD11b via suppression of IL‐6 production leading to reduced Th17‐cell differentiation. Therefore, CD11b may represent a susceptibility factor for autoimmunity and could be a target for future therapy.     

Invariant NKT cells regulate experimental autoimmune uveitis through inhibition of Th17 differentiation

Although NKT cells have been implicated in diverse immunomodulatory responses, the effector mechanisms underlying the NKT cell-mediated regulation of pathogenic T helper cells are not well understood. Here, we show that invariant NKT cells inhibited the differentiation of CD4(+) T cells into Th17 cells both in vitro and in vivo. The number of IL-17-producing CD4(+) T cells was reduced following co-culture with purified NK1.1(+) TCR(+) cells from WT, but not from CD1d(-/-) or Jα18(-/-) , mice. Co-cultured NKT cells from either cytokine-deficient (IL-4(-/-) , IL-10(-/-) , or IFN-γ(-/-) ) or WT mice efficiently inhibited Th17 differentiation. The contact-dependent mechanisms of NKT cell-mediated regulation of Th17 differentiation were confirmed using transwell co-culture experiments. On the contrary, the suppression of Th1 differentiation was dependent on IL-4 derived from the NKT cells. The in vivo regulatory capacity of NKT cells on Th17 cells was confirmed using an experimental autoimmune uveitis model induced with human IRBP(1-20) (IRBP, interphotoreceptor retinoid-binding protein) peptide. NKT cell-deficient mice (CD1d(-/-) or Jα18(-/-) ) demonstrated an increased disease severity, which was reversed by the transfer of WT or cytokine-deficient (IL-4(-/-) , IL-10(-/-) , or IFN-γ(-/-) ) NKT cells. Our results indicate that invariant NKT cells inhibited autoimmune uveitis predominantly through the cytokine-independent inhibition of Th17 differentiation.     

川芎嗪对小鼠哮喘模型Th17细胞的调控作用以及对Th17、Treg细胞平衡的影响

目的：研究川芎嗪对小鼠哮喘模型外周血中Th17、Treg细胞比例以及特征性细胞因子IL-17、IL-10水平的影响。方法：雄性BALB/c小鼠随机分成四组：正常对照组、哮喘模型组、川芎嗪治疗组以及激素治疗组。 OVA诱导激发哮喘,治疗组小鼠在第0、7、14天以及每次雾化吸入前30 min腹腔注射川芎嗪或地塞米松注射液,正常对照组用生理盐水替代OVA进行腹腔注射以及雾化吸入。小鼠的肺组织HE染色,分离外周血淋巴细胞做流式细胞学检测并ELISA法检测血清中IL-17、IL-10的水平。结果：哮喘模型组造模成功,川芎嗪治疗组及激素治疗组哮喘表现较轻微。 HE染色显示哮喘模型组小鼠肺组织在支气管以及小血管周围发现大量的嗜酸性粒细胞、中性粒细胞、巨噬细胞等炎性细胞浸润;而川芎嗪治疗组和激素治疗组小鼠的肺组织切片中仅发现少量的炎性细胞。流式细胞仪检测显示哮喘组小鼠的Treg细胞较正常组小鼠比例明显降低,而Th17细胞占CD4＋T细胞显著升高;川芎嗪治疗组小鼠以及激素治疗组小鼠的变化趋势一致,Treg细胞和Th17细胞的比例趋于正常。 ELISA结果显示哮喘组小鼠的IL-17的水平显著高于正常组,IL-10的水平较正常组显著降低,川芎嗪治疗组小鼠的IL-17水平较哮喘模型组明显降低,而IL-10的水平显著升高,激素治疗组小鼠的变化趋势与川芎嗪治疗组一致。结论：在OVA诱导小鼠哮喘模型中,川芎嗪可以通过增强Treg细胞的功能,增加Treg细胞的数量,进而抑制Th17细胞的数量以及功能,减轻Th17细胞的应答,降低IL-17细胞因子的分泌,从而起到预防/控制哮喘发作的作用。     

IL‐6: Regulator of Treg/Th17 balance

IL‐6 is a pleiotropic cytokine involved in the physiology of virtually every organ system. Recent studies have demonstrated that IL‐6 has a very important role in regulating the balance between IL‐17‐producing Th17 cells and regulatory T cells (Treg). The two T‐cell subsets play prominent roles in immune functions: Th17 cell is a key player in the pathogenesis of autoimmune diseases and protection against bacterial infections, while Treg functions to restrain excessive effector T‐cell responses. IL‐6 induces the development of Th17 cells from naïve T cells together with TGF‐β; in contrast, IL‐6 inhibits TGF‐β‐induced Treg differentiation. Dysregulation or overproduction of IL‐6 leads to autoimmune diseases such as multiple sclerosis (MS) and rheumatoid arthritis (RA), in which Th17 cells are considered to be the primary cause of pathology. Given the critical role of IL‐6 in altering the balance between Treg and Th17 cells, controlling IL‐6 activities is potentially an effective approach in the treatment of various autoimmune and inflammatory diseases. Here, we review the role of IL‐6 in regulating Th17/Treg balance and describe the critical functions of IL‐6 and Th17 in immunity and immune‐pathology.     

Persistence of IL-2 expressing Th17 cells in healthy humans and experimental autoimmune uveitis

Compared with other T-helper subsets, Th17 cell numbers are very low in human blood but become elevated in chronic inflammatory diseases. In this study, we investigated mechanisms that may explain the frequent involvement of Th17 cells in autoimmune diseases such as uveitis. We compared Th17 and Th1 subsets and found that Th17 cells expressed lower IL-2 levels during Ag-priming and this correlated with their decreased susceptibility to activation-induced cell death (AICD). However, complete depletion of IL-2 with IL-2 neutralizing antibodies rendered Th17 cells as susceptible to apoptosis as Th1 cells, suggesting that the low levels of IL-2 produced by Th17 cells conferred survival advantages to this subset. We describe here a Th17 subtype that constitutively produces very low levels of IL-2 (Th17-DP). The Th17-DP population increased dramatically in the blood and retina of mice during experimental autoimmune uveitis, indicating their potential involvement in the etiology of uveitis. We further show that the majority of the memory Th17 cells in human blood are Th17-DP and are targets of daclizumab, an IL-2R antibody used in treating recalcitrant uveitis. Thus, Th17 cells may persist in tissues and contribute to chronic inflammation by limiting IL-2 production to levels that cannot provoke IL-2-induced AICD yet are sufficient to promote Th17 homeostatic expansion.     

TCR and Notch signaling in CD4 and CD8 T-cell development

Summary:  The generation of CD4 and CD8 αβ T-cell lineages from CD4⁺CD8⁺ double-positive (DP) thymocyte precursors is a complex process initiated by engagement of major histocompatibility complex (MHC) by T-cell receptor (TCR) and coreceptor. Quantitative differences in TCR signaling induced by this interaction impose an instructional bias on CD4/CD8 lineage commitment that must be reinforced by MHC recognition and TCR signaling over subsequent selection steps in order for the thymocyte to progress and mature in the adopted lineage. Our studies show that the transmembrane receptor Notch plays a role in this process by modifying TCR signal transduction in DP thymocytes. In this review, we consider the functional relationship of TCR and Notch signaling pathways in the selection and specification of CD4 and CD8 T-cell lineages.     

佐剂关节炎大鼠肺功能与Treg、Notch信号通路的关系

目的:观察佐剂关节炎(Adjuvant arthritis,AA)大鼠肺功能、外周血调节性T细胞(Treg)及肺组织Notch通路变化。方法:将20只SD大鼠随机分为正常对照组(NC)和模型对照组(MC),每组10只;向MC组大鼠右后足跖皮内注射弗氏完全佐剂0.1 ml致炎,复制成AA模型;致炎18天后,观察两组大鼠足跖肿胀度(E)、关节炎指数(AI)、肺功能、肺组织形态学变化,采用流式细胞术检测外周血Treg,PT-PCR法检测肺组织Notch受体及配体表达。结果:AA大鼠E、AI、1秒内平均呼气流量(FEV1/FVC%)、肺组织Notch3、Notch4及Delta1的表达明显升高;75%肺活量的最大呼气流量(FEF75)、用力最大呼气流量(PEF)、肺动态顺应性(Cldyn)降低,外周血CD4+CD25+Treg、肺组织Notch1、Jagged1、Jagged2的表达水平显著降低(P<0.01或P<0.05)。相关分析显示,肺功能参数FVC、FEF75与CD4+CD25+Treg呈正相关,FEF50、MMF与Jagged1、Notch1呈正相关;FEF25、FEF50、PEF与Delta1、Notch3、Notch4呈负相关(P<0.01或P<0.05)。结论:AA大鼠发生关节炎症同时,出现肺功能、Treg降低及Notch通路的变化;肺功能与Treg、Notch受体/配体呈高度相关性,提示Treg和Notch通路可能参与肺功能降低的过程。     

MOG诱导的实验性自身免疫性脑脊髓炎小鼠中枢及外周神经系统CD4~+ T及CD8~+ T调节性细胞变化

目的:观察实验性自身免疫性脑脊髓炎小鼠中枢及外周CD4+ CD25+调节性T细胞(CD4+ CD25+ Treg),CD8+ CD28-调节性T细胞(CD8+ CD28- Treg)表达的变化情况,并探讨相关的细胞免疫学机制。方法:雌性C57BL/6小鼠随机分为未使用髓鞘少突胶质细胞糖蛋白35-55(MEVGWYR-SPFSRVVHLYRNGK)(MOG35-55)免疫的对照组和使用MOG35-55免疫诱导的EAE小鼠模型组,采用临床症状评分记录小鼠行为学变化、HE染色观察CNS炎症细胞浸润及病理改变,使用流式细胞术(FCM)检测小鼠中枢及外周CD8+ CD28- Treg,CD4+ CD25+ Treg细胞表达水平。结果:MOG35-55诱导的EAE模型组动物出现典型的EAE临床行为学及病理学表现,FCM检测EAE模型组小鼠脾细胞CD4+ CD25+ Treg较对照组升高但无统计学差异,CD8+ CD28- Treg表达水平明显低于对照组(P0.01),EAE模型组中枢有CD4+ CD25+ Treg,CD8+CD28-Treg淋巴细胞的浸润,且CD4+ CD25+ Treg,CD8+ CD28- Treg在中枢的表达均高于外周,对照组中枢神经系统未检测到淋巴细胞浸润。结论:CD4+ CD25+ Treg,CD8+ CD28- Treg均参与调控EAE的病理过程,CD4+ CD25+ Treg,CD8+ CD28- Treg在EAE小鼠中枢及外周分布及变化的不同,提示其进入中枢神经系统(CNS)并参与调节中枢局部炎症。     

Notch1 信号通路调控银屑病模型小鼠Th17 细胞分化和功能

目的:探讨Notch1 信号通路对银屑病模型小鼠Th17 细胞分化和功能的调控作用。方法:以5% 咪喹莫特外涂联合 2b 干扰素腹腔注射的方法制备20 只银屑病模型小鼠,免疫磁珠分离小鼠脾脏CD4+ T 淋巴细胞,流式细胞术检测 Th17 细胞比例,实时荧光定量RT-PCR 检测Th17 细胞特异性转录因子ROR-t、效应性细胞因子IL-17A、Notch1 信号分子及其靶基因Hes-1 的mRNA 表达水平,并与10 只对照组小鼠相比较。将银屑病模型小鼠CD4+ T 淋巴细胞分为未干预对照组和Notch1 抑制剂组( 分泌酶抑制剂DAPT),检测DAPT 阻断Notch1 信号对银屑病模型小鼠Notch1 信号分子及Hes-1、Th17 细胞比例、ROR-t 及IL-17A 表达水平的影响。结果:银屑病模型小鼠CD4+ T 淋巴细胞中Th17 细胞比例,ROR-t、IL-17A、Notch1及Hes-1 的mRNA 表达水平均显著高于对照小鼠[分别为(2.97±0.86)% 比(0.65±0.11)%,t =15.083;(5.75±0.61)比(1.57±0.43),t =21.630;(7.83±0.97)比(1.63±0.31),t =25.348;(7.10±1.37)比(1.47±0.34),t = 17.386;(7.30±1.15)比(1.67±0.48),t = 18.840,P 均<0.01];与未干预对照组相比,银屑病模型小鼠CD4+ T 淋巴细胞各DAPT 处理组中Notch1、Hes-1mRNA 表达水平,Th17 细胞比例、ROR-t 与IL-17A mRNA 表达水平及培养上清液中IL-17A 含量均明显下降,组间比较差异具有统计学意义(F 值分别为74.368、89.719、126.572、94.558、124.323 和123.231,P 均<0.01),且随DAPT 浓度的增加呈剂量依赖性降低。结论:Notch1 信号通路能够调控银屑病模型小鼠Th17 细胞的分化和功能,对银屑病的免疫靶向治疗有潜在价值。     

茯苓多糖对系统性红斑狼疮患者Th17/Treg平衡的影响

目的:研究茯苓多糖对系统性红斑狼疮(SLE)患者外周血辅助性T细胞17(Th17)/调节性T细胞(Treg)平衡的免疫调节作用。方法:选取45例SLE患者和35例健康对照者,应用磁珠分选法分离外周血CD4~+ T细胞,流式细胞术检测CD4~+ T细胞中Th17和Treg细胞的比例。用茯苓多糖分别处理健康对照者及患者的CD4~+ T细胞,MTT法检测细胞活力以测定茯苓多糖毒性,ELISA检测细胞中白细胞介素17(IL-17)、IL-6、IL-10及转化生长因子β(TGF-β)的含量,RT-q PCR和Western blot法分别测定维甲酸相关孤儿受体γt(RORγt)与叉头框蛋白P3(Foxp3)的mRNA和蛋白表达水平。结果:与健康对照组相比,SLE患者的Th17细胞比例显著升高,Treg细胞比例明显降低(P0.05)。用100μg/L的茯苓多糖处理SLE患者CD4~+ T细胞,与空白对照组相比,IL-17和IL-6的含量显著降低,IL-10和TGF-β的含量明显上升(P0.05);RORγt的mRNA和蛋白表达显著下降,同时Foxp3的表达在mRNA和蛋白水平上明显增加(P0.05);并且Th17/Treg的比值降低(P0.05)。结论:茯苓多糖可以通过升高Treg并降低Th17细胞的比例,对SLE起到一定的治疗作用。