IL-5 promotes induction of antigen-specific CD4+CD25+ T regulatory cells that suppress autoimmunity

Immune responses to foreign and self-Ags can be controlled by regulatory T cells (Tregs) expressing CD4 and IL-2Rα chain (CD25). Defects in Tregs lead to autoimmunity, whereas induction of Ag-specific CD4+CD25+ Tregs restores tolerance. Ag-specific CD4+CD25+ FOXP3+Tregs activated by the T helper type 2 (Th2) cytokine, IL-4, and specific alloantigen promote allograft tolerance. These Tregs expressed the specific IL-5Rα and in the presence of IL-5 proliferate to specific but not third-party Ag. These findings suggest that recombinant IL-5 (rIL-5) therapy may promote Ag-specific Tregs to mediate tolerance. This study showed normal CD4+CD25+ Tregs cultured with IL-4 and an autoantigen expressed Il-5rα. Treatment of experimental autoimmune neuritis with rIL-5 markedly reduced clinical paralysis, weight loss, demyelination, and infiltration of CD4+ (Th1 and Th17) CD8+ T cells and macrophages in nerves. Clinical improvement was associated with expansion of CD4+CD25+FOXP3+ Tregs that expressed Il-5rα and proliferated only to specific autoantigen that was enhanced by rIL-5. Depletion of CD25+ Tregs or blocking of IL-4 abolished the benefits of rIL-5. Thus, rIL-5 promoted Ag-specific Tregs, activated by autoantigen and IL-4, to control autoimmunity. These findings may explain how Th2 responses, especially to parasitic infestation, induce immune tolerance. rIL-5 therapy may be able to induce Ag-specific tolerance in autoimmunity.     

TNF downmodulates the function of human CD4+CD25hi T-regulatory cells

CD4+CD25+ T-regulatory cells (Tregs) play an essential role in maintaining immunologic homeostasis and preventing autoimmunity. However, little is known about the exogenous factors that regulate their differentiation and function. Here, we report that TNF inhibits the suppressive function of both naturally occurring CD4+CD25+ Tregs and TGFbeta1-induced CD4+CD25+ T-regulatory cells. The mechanism of this inhibition involves signaling through TNFRII that is constitutively expressed selectively on unstimulated Tregs and that is up-regulated by TNF. TNF-mediated inhibition of suppressive function is related to a decrease in FoxP3 mRNA and protein expression by the Tregs. Notably, CD4+CD25hi Tregs isolated from patients with active rheumatoid arthritis (RA) expressed reduced levels of FoxP3 mRNA and protein and poorly suppressed the proliferation and cytokine secretion of CD4+ effector T cells in vitro. Treatment with anti-TNF antibody (infliximab) increased FOXP3 mRNA and protein expression by CD4+CD25hi Tregs and restored their suppressive function. Thus, TNF has a novel action in modulating autoimmunity, by inhibiting CD4+CD25+ Treg activity.     

Functional role of CD4+CD25+ regulatory T cells and transforming growth factor-beta1 in childhood immune thrombocytopenic purpura

CD4+CD25+ regulatory T cells (Tregs) are critical in maintaining self tolerance and preventing organ specific autoimmune diseases. Their role in childhood immune thrombocytopenic purpura (ITP), an immune disorder in which the production of platelet autoantibodies might be caused by cytokine network dysregulation, has not been clearly defined. Transforming Growth Factor-beta1 (TGF-beta1) has been suggested to mediate Tregs suppressive function. The aim of this study was to assess cell populations of CD4+CD25+ T regulatory cells as well as mRNA expression of TGF-beta1 level in peripheral blood of children with ITP and evaluate their possible role in prediction of disease severity and response to therapy. Thirty-three children with ITP (13 acute and 20 chronic cases) and 10 healthy controls were studied. CD4+CD25+ regulatory T cells were assessed in peripheral blood by flowcytometry. Expression of TGF-beta1 mRNA was examined by real-time quantitative polymerase chain reaction assay. The frequency of CD4+CD25+ Tregs was significantly decreased in acute and chronic ITP patients as compared to controls. Acute ITP patients had significantly lower percentage of Tregs compared with chronic ITP patients. Higher frequency of CD4+CD25+ Tregs was detected in chronic ITP patients with platelet count >100 x 10(9)/L compared with patients with platelet count <100 x 10(9)/L and in steroid responsive patients compared with steroid resistant patients. The expression of mRNA TGF-beta1 level was significantly lower in acute and chronic ITP patients compared with controls and in chronic ITP patients with pltc <100 x 10(9)/L in comparison with patients with pltc > 100 x l0(9)/L. A significant positive correlation was found between percentage of CD4+CD25+ Tregs and mRNA expression of TGF-beta1 in chronic ITP patients. In conclusion, immune regulation of TGF-beta1 by Tregs may play a fundamental role in the pathogenesis of childhood immune thrombocytopenic purpura. This might form a base for future specific immunomodulatory therapies for ITP.     

Low percentages of regulatory T cells in common variable immunodeficiency (CVID) patients with autoimmune diseases and its association with increased numbers of CD4+CD45RO+ T and CD21low B cells

BackgroundCommon variable immunodeficiency (CVID) is a heterogeneous group of primary antibody deficiencies defined by marked reductions in serum IgG, IgA and/or IgM levels and recurrent bacterial infections. Some patients are associated with defects in T cells and regulatory T cells (Tregs), resulting in recurrent viral infections and early-onset autoimmune disease.MethodsWe analyzed whether there is an association between Tregs cells (CD4+CD25+CD127^low and CD4+CD25+FoxP3+); memory T cells (CD4+CD45RO+); memory B cells (CD19+CD27-IgD-); and CD21^low B cells (CD19+CD38^lowCD21^low); as well as autoimmune manifestations in 36 patients with CVID (25 women and 11 men, mean age 24 years), all by flow cytometry.ResultsFourteen patients presented with autoimmune diseases (AI) (39%), including 11 with autoimmune thrombocytopenia (ITP) (31%); two with vitiligo (6%); one with systemic lupus erythematosus (LES) (3%); and one with multiple sclerosis (MS) (3%). CVID patients with AI had a reduced proportion of Tregs (both CD4+CD25+CD127^low and FoxP3+ cells) compared with healthy controls. CVID patients with AI had expanded CD21^low B cell populations compared with patients who did not have AI. A correlation between increased CD4+CD45RO T cell populations and reduced Tregs was also observed.ConclusionsOur results showed that 39% of patients with CVID had AI and reduced Tregs populations. Research in this area might provide noteworthy data to better understand immune dysfunction and dysregulation related to CVID.     

Anti-CD3 prevents factor VIII inhibitor development in hemophilia A mice by a regulatory CD4+CD25+-dependent mechanism and by shifting cytokine production to favor a Th1 response

Non-Fc-receptor binding anti-CD3 Ab therapy, in the setting of several different autoimmune disorders, can induce antigen-specific and long-lasting immunologic tolerance. Because factor VIII (FVIII) inhibitor formation is the most serious treatment-related complication for hemophilia A patients, we tested the efficacy of anti-CD3 to prevent FVIII inhibitor formation in hemophilia A BALB/c and C57BL/6 mice. A short course of low-dose anti-CD3 significantly increased expression of CD25 and the proportion of CD4+CD25+ regulatory T cells in the spleen and potently prevented the production of inhibitory and non-neutralizing anti-FVIII antibodies in both strains of mouse. Depleting the CD4+CD25+ cells during anti-CD3 therapy completely ablated tolerance to FVIII. Further phenotypic characterization of regulatory cells in tolerant mice showed a consistently higher number of CD4+GITR+ and CD4+FoxP3+ cells in both strains of mice. In addition, in tolerant C57BL/6 mice we observed an increase in CD4+CD25+ CTLA-4+ and CD4+CD25+mTGF-beta1+ cells. Finally, in vitro cytokine profiling demonstrated that splenocytes from tolerant BALB/c and C57BL/6 were polarized toward a Th1-immune response. Taken together, these findings indicate that anti-CD3 induces tolerance to FVIII and that the mechanism(s) regulating this response almost certainly occurs through the generation of several distinct regulatory T-cell lineages and by influencing cytokine production and profile.     

Naturally occurring CD4+ CD25+ cells in modulating immune response to administered coagulation factor VIII in factor VIII‐deficient mice

Summary. CD4+ CD25+ T regulatory (Treg) cells are critical mediators of peripheral self‐tolerance and immune homeostasis. In this study, we characterized the ability of naturally occurring CD4+ CD25+ cells from the wild‐type mice to modulate the immune response to administered coagulation factor VIII (FVIII) in FVIII‐deficient mice. For the cell therapy, CD4+ CD25+ cells and CD4+ CD25− cells were purified from the spleens of wild‐type normal mice and administered to FVIII‐deficient mice prior to four injections of recombinant FVIII (rFVIII). The titre of FVIII antibodies and antibodies with inhibitory activity against FVIII was lower in the mice treated with natural CD4+ CD25+ cells or CD4+ CD25− cells compared with the mice treated only with rFVIII. We also demonstrated that CD4+ CD25− cells could differentiate to acquire the Treg phenotype expressing CD25 and FoxP3 if stimulated in vitro. These observations provide evidence that Treg cells can be used for designing cell therapy for controlling the immune response to FVIII.     

结核性胸膜炎患者胸液中CD4~+ CD25~+ FoxP3~+调节T细胞抑制抗结核免疫

目的研究结核性胸膜炎患者胸液中CD4+CD25+FoxP3+调节T细胞是否增多,这些调节T细胞是否抑制结核的特异细胞免疫反应。方法使用细胞分离、流式细胞分析及体外细胞培养作细胞增殖及增殖抑制等实验方法,对15例结核性胸膜炎患者及17例健康正常人群胸液及外周血白细胞中CD4+CD25+FoxP3+调节T细胞的量及特征作研究。结果结核性胸膜炎患者胸液中CD4+CD25+FoxP3+调节T细胞明显高于患者及健康人群外周血。在体外,结核性胸膜炎患者胸液中单核细胞对BCG刺激产生γ-干扰素(IFN-γ)的能力明显强于患者及健康人群外周血中单核细胞;把这些调节T细胞从胸液单核细胞中清除,增强了结核患者胸液单核细胞对BCG刺激产生IFN-γ;从结核患者胸液分离的这些调节T细胞能抑制结核患者Th1细胞产生IFN-γ。结论结核性胸膜炎患者胸液CD4+CD25+FoxP3+调节T细胞增多,抑制结核性胸膜炎患者Th1细胞免疫反应,从而参与了结核性胸膜炎的发病。     

CD4+ CD25+ 调节性T细胞与自身免疫性甲状腺疾病

CD4+ CD25+ 调节性T细胞为新近发现的一群功能成熟的T细胞亚群.其特征性表达叉头盒蛋白3(Foxp3)分子,专职免疫无能和免疫抑制,在维持外周免疫耐受,防止自身免疫性疾病发病中起着极为关键的作用.CD4+ CD25+ 调节性T细胞在自身免疫性甲状腺疾病(AITD)发病中的作用引起了人们的关注.动物实验发现CD4+ CD25+ 调节性T细胞存在与否决定了实验动物是否发生实验性自身免疫性甲状腺炎(EAT)和Graves病.人体研究发现CD4+ CD25+ 调节性T细胞数目和功能异常与人AITD发生密切相关.这些研究结果提示,CD4+ CD25+ 调节性T细胞可能在AITD发病中起重要作用.     

CD147 (Basigin/Emmprin) identifies FoxP3+CD45RO+CTLA4+-activated human regulatory T cells

Human CD4(+)FoxP3(+) T cells are functionally and phenotypically heterogeneous providing plasticity to immune activation and regulation. To better understand the functional dynamics within this subset, we first used a combined strategy of subcellular fractionation and proteomics to describe differences at the protein level between highly purified human CD4(+)CD25(+) and CD4(+)CD25(-) T-cell populations. This identified a set of membrane proteins highly expressed on the cell surface of human regulatory T cells (Tregs), including CD71, CD95, CD147, and CD148. CD147 (Basigin or Emmprin) divided CD4(+)CD25(+) cells into distinct subsets. Furthermore, CD147, CD25, FoxP3, and in particular CTLA-4 expression correlated. Phenotypical and functional analyses suggested that CD147 marks the switch between resting (CD45RA(+)) and activated (CD45RO(+)) subsets within the FoxP3(+) T-cell population. Sorting of regulatory T cells into CD147(-) and CD147(+) populations demonstrated that CD147 identifies an activated and highly suppressive CD45RO(+) Treg subset. When analyzing CD4(+) T cells for their cytokine producing potential, CD147 levels grouped the FoxP3(+) subset into 3 categories with different ability to produce IL-2, TNF-α, IFN-γ, and IL-17. Together, this suggests that CD147 is a direct marker for activated Tregs within the CD4(+)FoxP3(+) subset and may provide means to manipulate cells important for immune homeostasis.     

Expansion of CD4+CD25+FoxP3+ Regulatory T Cells in Chronic Hepatitis C Virus Infection

Background: Regulatory T cells (Tregs) have been involved in impaired immunity and may have a pivotal role in persistence of viral infections. Objective: To develop a simple and reliable in-house three color flow cytometery of peripheral blood to understand the role of HCV infection in the increase of Tregs. Methods: The level of naturally occurring CD4+CD25+FoxP3+ regulatory T cells (nTregs) in 20 chronically infected with hepatitis C virus (HCV) patients was compared to those of 15 healthy individuals by flowcytometry. In a different approach we performed permeabilization and intracellular staining before surface staining which allows the preservation of the surface molecules in the combined detection process and results in the normal frequency of nTregs in blood. Results: Using the optimized method, it was shown that a significantly higher proportion of nTregs in the total CD4+ T cell population was seen in the peripheral blood of chronic HCV patients (0.83 ± 0.21%, p=0.05) as compared to controls (0.26 ± 0.1, p=0.05). Conclusions: In accordance with other studies, we showed that HCV infection induces a dramatic increase in Tregs, which might contribute to the immune response failure during HCV infection.     

Diversity of regulatory CD4+T cells controlling distinct organ-specific autoimmune diseases

Depletion of selected regulatory CD4+ T cell subsets induces the spontaneous onset of various immune or autoimmune disorders. It is not clear, however, whether a given subset, notably CD4+CD25+ regulatory T cells, protects from a wide spectrum of immune disorders, or whether specialized subsets of regulatory T cells control each given disease or group of diseases. We report here, using diabetes prone nonobese diabetic (NOD) mice, that depending on the regulatory T cells that are depleted, i.e., CD25+, CD62L+, or CD45RB(low), distinct immune diseases appear after transfer into NOD severe combined immunodeficiency (SCID) recipients. Thus, reconstitution of NOD SCID mice with CD25- T cells induces major gastritis and late-onset diabetes, but no or mild colitis. Reconstitution with CD62L- T cells induces fulminant diabetes with no colitis or gastritis. Reconstitution with CD45RB(high) T cells induces major colitis with wasting disease and no or very moderate gastritis and diabetes. Major differences among the three regulatory T cell subsets are also seen in vitro. The bulk of suppressor cells inhibiting the proliferation of CD4+CD25- T cells in coculture is concentrated within the CD25+ but not the CD62L+ or CD45RB(low) T cell subsets. Similarly, cytokine production patterns are significantly different for each regulatory T cell subset. Collectively, these data point to the diversity and organ selectivity of regulatory T cells controlling distinct autoimmune diseases whatever the underlying mechanisms.     

Dysregulation of CD4+CD25+CD127lowFOXP3+ regulatory T cells in HIV-infected pregnant women

Pregnancy represents a major challenge to immunologic tolerance. How the fetal "semiallograft" evades maternal immune attack is unknown. Pregnancy success may involve alteration of both central (thymic) and peripheral tolerance mechanisms. HIV infection is characterized by CD4(+) T-cell depletion, chronic immune activation, and altered lymphocyte subsets. We studied immunologic consequences of pregnancy in 20 HIV-infected women receiving highly active antiretroviral therapy (HAART), and for comparison in 16 HIV-negative women. Lymphocyte subsets, thymic output, and cytokine profiles were measured prospectively during pregnancy and postpartum. A significant expansion of CD4(+)CD25(+)CD127(low)FoxP3(+) regulatory T cells indicating alteration of peripheral tolerance was seen during second trimester, but only in HIV-negative women. HIV-infected women had lower CD4 counts, lower thymic output and Th-2 cytokines, and more immune activation at all time points compared with controls. Immune activation was decreased in HIV-infected patients during pregnancy. In contrast, CD4 counts were increased in both groups. In conclusion, the study does not indicate that pregnancy adversely affects the immunologic course of HIV infection. However, despite HAART during pregnancy, HIV-infected women display different immunologic profiles from HIV-negative women, which may have importance for the induction of fetal-maternal tolerance and in part explain the increased risk of abortion in HIV-infected women.     

CTLA4 blockade expands FoxP3+ regulatory and activated effector CD4+ T cells in a dose-dependent fashion

Cytotoxic T lymphocyte-associated antigen 4 (CTLA4) delivers inhibitory signals to activated T cells. CTLA4 is constitutively expressed on regulatory CD4(+) T cells (Tregs), but its role in these cells remains unclear. CTLA4 blockade has been shown to induce antitumor immunity. In this study, we examined the effects of anti-CTLA4 antibody on the endogenous CD4(+) T cells in cancer patients. We show that CTLA4 blockade induces an increase not only in the number of activated effector CD4(+) T cells, but also in the number of CD4(+) FoxP3(+) Tregs. Although the effects were dose-dependent, CD4(+) FoxP3(+) regulatory T cells could be expanded at lower antibody doses. In contrast, expansion of effector T cells was seen only at the highest dose level studied. Moreover, these expanded CD4(+) FoxP3(+) regulatory T cells are induced to proliferate with treatment and possess suppressor function. Our results demonstrate that treatment with anti-CTLA4 antibody does not deplete human CD4(+) FoxP3(+) Tregs in vivo, but rather may mediate its effects through the activation of effector T cells. Our results also suggest that CTLA4 may inhibit Treg proliferation similar to its role on effector T cells. This study is registered at http://www.clinicaltrials.gov/ct2/show/NCT00064129, registry number NCT00064129.     

The immunosuppressive drug FK778 induces regulatory activity in stimulated human CD4+ CD25- T cells

The induction of transplantation tolerance involves a T-cell-mediated process of immune regulation. In clinical transplantation, the use of immunosuppressive drugs that promote or facilitate this process would be highly desirable. Here, we investigated the tolerance-promoting potential of the immunosuppressive drug FK778, currently under development for clinical therapy. Using a human allogeneic in vitro model we showed that, upon T-cell receptor (TCR) triggering, FK778 induced a regulatory phenotype in CD4+ CD25- T cells. Purified CD4+ CD25- T cells primed in the presence of FK778 showed hyporesponsiveness upon restimulation with alloantigen in the absence of the drug. This anergic state was reversible by exogenous interleukin-2 (IL-2) and was induced independent of naturally occurring CD4+ CD25+ regulatory T cells. Pyrimidine restriction was a crucial requirement for the de novo induction of regulatory activity by FK778. The FK778-induced anergic cells showed suppressor activity in a cell-cell contact-dependent manner; were CD25(high), CD45RO+, CD27-, and CD62L-; and expressed cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4), glucocorticoid-induced tumor necrosis factor receptor (GITR), and FoxP3. The cells revealed delayed p27(kip1) degradation and enhanced phosphorylation of STAT3. In conclusion, the new drug FK778 shows tolerizing potential through the induction of a regulatory T-cell subset in CD4+ CD25- T cells.     

IL-10-dependent infectious tolerance after the treatment of experimental allergic encephalomyelitis with redirected CD4+CD25+ T lymphocytes

How small numbers of CD4+CD25+ regulatory T cells suppress autoimmune responses in vivo is unclear. In this report we analyze the immunomodulatory activity of CD4+CD25+ T cells that are antigen-specifically redirected against myelin basic protein (MBP)89-101-specific autoreactive T cells by a MBP89-101-IA(s)-zeta chimeric receptor. We have previously shown that these redirected regulatory T cells are highly potent in treating a model autoimmune disease, experimental allergic encephalomyelitis. We show here that they have only limited effect in vivo on autoreactive T cell proliferation and therefore do not act by deleting or suppressing the expansion of pathologic effector cells. Rather, the redirected CD4+CD25+ T cells divert the pathologic T helper 1 self-specific T cell response to one characterized by high IL-10 and lower IL-4 production. Significantly, when isolated from the inducing CD4+CD25+ regulatory T cells, these self-specific T cells can independently suppress the autoreactive T cell response and experimental allergic encephalomyelitis development in an IL-10-dependent manner. These results provide evidence that CD4+CD25+ regulatory T cells can manipulate the adaptive immune response in vivo through the infectious induction of tolerance, specifically by promoting the formation of antigen-specific, IL-10-secreting regulatory T cells.     

CD4+CD25high Foxp3+ regulatory T cells in myelodysplastic syndrome (MDS)

Foxp3+ regulatory T cells (Tregs) play a central role in maintaining immune tolerance. A reduction in the function of Tregs is a key feature of autoimmune diseases, whereas their expansion in malignant diseases leads to the suppression of host antitumor responses. We analyzed the absolute number of CD4+ and CD8(+) Tregs in the peripheral blood of 52 patients with myelodysplastic syndrome (MDS) and show a significant correlation between increased number of CD4+ Tregs and MDS subgroups with 5% or more bone marrow blasts (P < .001), high International Prognostic Scoring System (IPSS) score (P < .001), and disease progression (P < .001), whereas no correlation between CD8+ Tregs and prognostic variables was observed. The CD4+ Tregs showed a polyclonal spectratype, and the percentage of the naive subset was significantly higher in the high-risk patients compared with low-risk or healthy age-matched donors (P = .032). Our data suggest that CD4+ Treg expansion is a feature of high-risk MDS and progression to aggressive subtypes of the disease.     

CD8+CD122+ T cells, a newly identified regulatory T subset, negatively regulate Graves' hyperthyroidism in a murine model

Graves' disease is a thyroid-specific autoimmune disease mediated by stimulatory autoantibodies against the TSH receptor (TSHR). We have previously shown in our mouse model with adenovirus expressing the TSHR that antibody mediated depletion of CD4(+)CD25(+) regulatory T cells (Tregs) enhances incidence and severity of hyperthyroidism in resistant and susceptible mouse strains, respectively. These data indicate that balance between effector T cells and Tregs is critical for disease development. This study was designed to evaluate the role played by another recently identified type of Treg, CD8(+)CD122(+) T cells, in our mouse model to delineate the significance of different types of Tregs in Graves' disease. Flow cytometry analysis showed that CD4(+)CD25(+) and CD8(+)CD122(+) T cells are distinct cell types, and anti-CD122 antibody effectively and selectively depleted CD8(+)CD122(+) T cells. As for CD4(+)CD25(+) Treg, CD8(+)CD122(+) T cell depletion increased the incidence of hyperthyroidism both in resistant and susceptible mice. Of interest, intrathyroidal lymphocytic infiltration was observed in some CD8(+)CD122(+) T cell-depleted, hyperthyroid resistant mice. These results indicate that in addition to CD4(+)CD25(+) T cells, CD8(+)CD122(+) T cells also play a crucial role in disease susceptibility in mouse Graves' disease. Thus, different types of Tregs appear to be involved in tolerance to a self-antigen, the TSHR.     

CD4~+ CD25~+调节性T细胞与恶性腹水的相关研究进展

CD4+ CD25+调节性T细胞(CD4+ CD25+Treg细胞)是具有独特免疫调节功能的T细胞亚群,抑制免疫反应,在机体免疫稳态维持、肿瘤免疫及移植耐受等方面发挥重要的作用。近年来,调节性T细胞在肿瘤免疫及治疗的研究中受到越来越广泛的关注。现就调节性T细胞在恶性腹水方面的研究做一简要综述。