Therapeutic potential of self-antigen-specific CD4+ CD25+ regulatory T cells selected in vitro from a polyclonal repertoire

CD4+ CD25+ regulatory T cells (Treg) play a major role in the prevention of autoimmune diseases. Converging evidence indicates that Treg specific for self-antigens expressed by target tissues have a greater therapeutic potential than polyclonal Treg. Therefore, the selective expansion of rare self-antigen-specific T(reg) naturally present in a polyclonal repertoire of Treg is of major importance. In this work, we investigated the potential of different dendritic cell (DC) subsets to expand antigen-specific Treg in mice. The in vitro selective expansion of rare islet-specific Treg from polyclonal Treg could only be achieved efficiently by stimulation with CD8+ splenic DC presenting islet antigens. These islet-specific Treg exerted potent bystander suppression on diabetogenic T cells and prevented type 1 diabetes. This approach opens new perspectives for cell therapy of autoimmune diseases.     

Flt3 ligand expands CD4+FoxP3+ regulatory T cells in human subjects

CD4⁺CD25⁺FoxP3⁺ naturally occurring regulatory T (Treg) cells play a crucial role in the maintenance of immune tolerance and in preventing autoimmune pathology. Interventions that expand Treg cells are highly desirable, as they may offer novel treatment options in a variety of autoimmune and transplantation settings. Paralleling previous preclinical studies, we demonstrate here that administration of the hematopoietic growth factor Flt3L to human subjects increases the frequency and absolute number of Treg cells, and reduces the ratio of CD8⁺ T cells to Treg cells in the peripheral blood. The increase in Treg cells was due to enhanced Treg‐cell proliferation rather than release of Treg cells from the thymus. Further studies revealed that Flt3L‐induced proliferation of Treg cells was an indirect effect that occurred via the interaction of Treg cells with the Flt3L‐expanded pool of CD1c⁺ myeloid dendritic cells. On the basis of these findings, Flt3L may represent a promising agent for promoting immune tolerance in a variety of clinical settings.     

Reduced suppressive effect of CD4+CD25high regulatory T cells on the T cell immune response against myelin oligodendrocyte glycoprotein in patients with multiple sclerosis

Immunoregulatory T cells of (CD4+)CD25+ phenotype suppress T cell function and protect rodents from organ-specific autoimmune disease. The human counterpart of this subset of T cells expresses high levels of CD25 and its role in human autoimmune disorders is currently under intense investigation. In multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system (CNS), the activation of circulating self-reactive T cells with specificity for myelin components is considered to be an important disease initiating event. Here, we investigated whether MS is associated with an altered ability of (CD4+)CD25high regulatory T cells (Treg) to confer suppression of myelin-specific immune responses. Whereas Treg frequencies were equally distributed in blood and cerebrospinal fluid of MS patients and did not differ compared to healthy controls, the suppressive potency of patient-derived (CD4+)CD25high T lymphocytes was impaired. Their inhibitory effect on antigen-specific T cell proliferation induced by human recombinant myelin oligodendrocyte protein as well as on immune responses elicited by polyclonal and allogeneic stimuli was significantly reduced compared to healthy individuals. The effect was persistent and not due to responder cell resistance or altered survival of Treg, suggesting that a defective immunoregulation of peripheral T cells mediated by (CD4+)CD25high T lymphocytes promotes CNS autoimmunity in MS.     

CD52 is a novel costimulatory molecule for induction of CD4+ regulatory T cells

We previously reported that 4C8 monoclonal antibody (mAb) provides a costimulatory signal to human CD4+ T cells and consequently induces regulatory T (Treg) cells, which are hypo-responsive and suppress the polyclonal response of bystander CD4+ cells in a contact-dependent manner. In this study, we identified the antigen of 4C8 mAb as CD52. Costimulation with Campath-1H, a humanized anti-CD52 mAb, also induced Treg cells. Anti-CD52-induced Treg cells suppressed the proliferation of both CD4+ and CD8+ T cells provided with polyclonal or allogeneic stimulation. When Treg cells were induced from Staphylococcal enterotoxin B (SEB) treated cells, they suppressed the response to SEB more efficiently than that to another superantigen, SEA. Furthermore, anti-CD52-induced Treg cells could be expanded by culture with IL-2 followed by CD52-costimulation, and co-injection of expanded Treg cells suppressed lethal xenogeneic graft versus host disease (GvHD) reactions in SCID mice caused by human peripheral blood mononuclear cells (PBMCs).     

CD4+CD25+T细胞的扩增方法与临床应用

CD4+CD25+T细胞是最重要的一类调节性T细胞(Tr).体内固有CD4+CD25+T细胞的自然扩增率极低,不能满足临床治疗的需要.通过采用FoxP3基因转染技术、阻断细胞活化信号、DC诱导、加入细胞因子等方法,对CD4+CD25+T细胞的数量和功能进行扩增,使其在器官移植、自身免疫性疾病和肿瘤免疫等领域具有广泛的临床应用前景.     

IDO expands human CD4+CD25high regulatory T cells by promoting maturation of LPS-treated dendritic cells

We have previously shown that human monocyte-derived dendritic cells (DC) express indoleamine 2,3-dioxygenase (IDO), as well as several other enzymes of the kynurenine pathway at the mRNA level upon maturation. The tolerogenic mechanisms of this pathway remain unclear. Here we show that LPS-treated DC metabolize tryptophan as far as quinolinate. We found that IDO contributes to LPS and TNF-alpha + poly(I:C)-induced DC maturation since IDO inhibition using two different inhibitors impairs DC maturation. IDO knock-down using short-hairpin RNA also led to diminished LPS-induced maturation. In line with these results, the tryptophan-derived catabolites 3-hydroxyanthranilic acid and 3-hydroxykynurenine increased maturation of LPS-treated DC. Concerning the molecular mechanisms of this effect, IDO acts as an intermediate pathway in LPS-induced production of reactive oxygen species and NF-kappaB activation, two processes that lead to DC maturation. Finally, we show that mature DC expand CD4(+)CD25(high) regulatory T cells in an IDO-dependent manner. In conclusion, we show that IDO constitutes an intermediate pathway in DC maturation leading to expansion of CD4(+)CD25(high) regulatory T cells.     

CD4+CD25+T调节细胞的研究进展

CD4 CD25 TH细胞通过抗原特异性方式或细胞接触的方式抑制自身反应性T细胞的活化,能有效地维持自身免疫耐受,是调节自身反应性T细胞和防止自身免疫病发生的重要调节细胞。     

CD4~+CD25~+调节性T细胞在同种异体移植免疫耐受中的研究进展

同种异体器官移植已经成为终末器官衰竭患者修复病损的组织和器官,并重建其功能必要的治疗方法。受者往往都需要长期接受免疫抑制剂来维持移植物在其体内的存活。由于免疫抑制剂的非特异性及毒副作用,长期应用该类药物会导致受者免疫力低下而出现易感染、癌症等副作用。因此,解决该问题的关键是诱导受者对供者移植抗原产生特异性地"移植免疫耐受"。大量研究结果显示,CD4+CD25+调节性T细胞能特异性地抑制免疫排斥反应,在调控免疫应答与维持外周免疫耐受中发挥着重要作用。本文对近年来CD4+CD25+调节性T细胞的作用机制及其在同种异体移植免疫耐受方面的研究进行了综述。     

Maintenance of immune tolerance by Foxp3+ regulatory T cells requires CD69 expression

Although FoxP3⁺ regulatory T cells are key players in the maintenance of immune tolerance and autoimmunity, the lack of specific markers constitute an obstacle to their use for immunotherapy protocols. In this study, we have investigated the role of the C-type lectin receptor CD69 in the suppressor function of Tregs and maintenance of immune tolerance towards harmless inhaled antigens. We identified a novel FoxP3⁺CD69⁺ Treg subset capable to maintain immune tolerance and protect to developing inflammation. Although CD69⁺ and CD69⁻FoxP3⁺ Tregs exist in homeostasis, only CD69-expressing Tregs express high levels of CTLA-4, ICOS, CD38 and GITR suppression-associated markers, secrete high amounts of TGFβ and have potent suppressor activity. This activity is regulated by STAT5 and ERK signaling pathways and is impaired by antibody-mediated down-regulation of CD69 expression. Moreover, immunotherapy with FoxP3⁺CD69⁺ Tregs restores the homeostasis in Cd69^−/− mice, that fail to induce tolerance, and is also highly proficient in the prevention of inflammation. The identification of the FoxP3⁺CD69⁺ Treg subset paves the way toward the development of new therapeutic strategies to control immune homeostasis and autoimmunity.     

Human regulatory T cells and autoimmunity

CD4+CD25+ regulatory T cells (Treg) appear to be critical in regulating immune responses to self-antigens. Treg deficiency is associated with several human autoimmune diseases. Although substantial progress has been made in the study of murine and human Treg, their fundamental mechanism of action remains unknown. In this review, we discuss the phenotype of human natural Treg, their functional mechanism, and their role in autoimmune disease.     

CD38+ CD45RBlow CD4+ T cells: a population of T cells with immune regulatory activities in vitro

An antibody reactive with CD38 revealed both phenotypic and functional heterogeneity amongst CD45RB^low cells. Functional analysis of the CD38⁺ and CD38⁻ fractions showed that the latter contained T cells which responded to recall antigens and produced high levels of cytokine in response to polyclonal stimulation. In contrast, the CD38⁺ population failed to proliferate or to produce detectable levels of cytokines. Despite appearing unresponsive, the CD38⁺ population significantly inhibited anti-CD3-induced proliferation and cytokine secretion by the reciprocal CD38⁻ population. Immune suppression required stimulation through the TCR and was dependent on a physical interaction between regulatory and responding CD4⁺ populations. It did not involve killing of the responding T cells or secretion of IL-10 or TGF-β. Despite some similarities there is no direct correlation between the in vitro suppression characteristic of the CD38⁺ CD45RB^low subset and in vivo suppression which has been shown to be mediated by unseparated CD45RB^low CD4⁺ T cells. However, these results demonstrate that two functionally distinct subsets of T cells reside within the antigen-exposed or CD45RB^low CD4⁺ T cell population and are thus generated in vivo: (1) conventional memory T cells which proliferate and secrete cytokines in response to activation and (2) a population of regulatory T cells which inhibit T cell activation in vitro. Antibodies reactive with CD38 may provide a useful tool with which to study the role of these T cell subsets in the induction and regulation of the immune response.     

Aging and human CD4 regulatory T cells

Alterations in immunity that occur with aging likely contribute to the development of infection, malignancy and inflammatory diseases. Naturally occurring CD4⁺ regulatory T cells (Treg) expressing high levels of CD25 and forkhead box P3 (FOXP3) are essential for regulating immune responses. Here we investigated the effect of aging on the number, phenotypes and function of CD4⁺ Treg in humans. The frequency and phenotypic characteristics of CD4⁺, FOXP3⁺ T cells as well as their capacity to suppress inflammatory cytokine production and proliferation of CD4⁺, CD25⁻ T cells (target cells) were comparable in young (age ≤40) and elderly (age ≥65) individuals. However, when CD4⁺, FOXP3⁺ Treg and CD4⁺, CD25⁻ T cells were co-cultured at a ratio of 1:1, the production of anti-inflammatory cytokine IL-10 from CD4⁺, CD25⁻ T cells was more potently suppressed in the elderly than in the young. This finding was not due to changes in CTLA-4 expression or apoptosis of CD4⁺, FOXP3⁺ Treg and CD4⁺, CD25⁻ T cells. Taken together, our observations suggest that aging may affect the capacity of CD4⁺, FOXP3⁺ T cells in regulating IL-10 production from target CD4⁺ T cells in humans although their other cellular characteristics remain unchanged.     

CD4+ CD25high regulatory T cells reduce T cell transendothelial migration in cancer patients

Cell-mediated immunity is thought to be the main mechanism of anti-tumour responses of the host, but it is not known if cancer disease affects T cell recruitment from blood to tissues. Therefore, we compared Heliobacter pylori-induced T cell transendothelial migration (TEM) in H. pylori-infected gastric carcinoma patients, colon and lung carcinoma patients and healthy volunteers. H. pylori induced significant T cell migration from all groups. However, there was a dramatic reduction of T cell TEM in gastric carcinoma patients (80%) compared to healthy individuals. A similarly reduced transmigration was also seen in colon and lung carcinoma patients. We found significantly increased frequencies of T(reg) cells in the blood of gastric carcinoma patients compared to healthy individuals, and depletion of T(reg) cells from the blood of these patients prior to TEM restored T cell migration. The effect of T(reg) cells was largely dependent on cell-cell contact, but not on IL-10 or TGF-beta. In addition, the presence of T(reg) cells led to reduced T cell attachment to endothelium and decreased production of T cell-recruiting chemokines during TEM. In conclusion, T(reg) cell-mediated reduction of T cell TEM may reduce T cell recruitment in patients with epithelial malignancies, thereby hampering anti-tumour responses.     

CD4-mediated functional activation of human CD4+CD25+ regulatory T cells

Naturally occurring CD4(+)CD25(+)FoxP3(+) regulatory T cells (CD25(+) Tregs) constitute a specialized population of T cells that is essential for the maintenance of peripheral self-tolerance. The immune regulatory function of CD25(+) Tregs depends upon their activation. We found that anti-CD4 antibodies activate the suppressive function of human CD25(+) Tregs in a dose-dependent manner. We demonstrate that CD4-activated CD25(+) Tregs suppress the proliferation of CD4(+) and CD8(+) T cells, their IL-2 and IFN-gamma production as well as the capacity of CD8(+) T cells to re-express CD25. By contrast, anti-CD4 stimulation did not induce suppressive activity in conventional CD4(+) T cells. These results identify CD4 as a trigger for the suppressive function of CD25(+) Tregs and suggest a possible CD4-mediated exploitation of these cells.     

CD28 signals the differential control of regulatory T cells and effector T cells

One possible means of driving antigen‐specific immune suppression is to expand or induce antigen‐specific FoxP3‐expressing Treg cells. One way of activating and expanding these specialized cells, both in vitro and in vivo, is by strong costimulation via CD28 with an agonistic anti‐CD28 monoclonal antibody, called anti‐CD28 superagonist (CD28SA). However, CD28SA also strongly activates conventional T (Tconv) cells to secrete proinflammatory cytokines and, under certain conditions, causes serious cytokine release syndrome. In this issue of European Journal of Immunology, Tabares et al. [Eur. J. Immunol. 2014. 44: 1225–1236] address how CD28SA can be used for the differential control of human Treg and Tconv cells to suppress immune responses without serious adverse effects. They show that, depending on the dose of the antibody or by comedication of cortico‐steroid, the selective expansion of Treg cells can be achieved without significantly activating Tconv cells to produce inflammatory cytokines. This difference in CD28 signal sensitivity between the two populations can be exploited for better control of immune responses.     

Selective expansion of memory CD4(+) T cells by mitogenic human CD28 generates inflammatory cytokines and regulatory T cells

Costimulatory signals are important for development of effector and regulatory T cells. In this case, CD28 signaling is usually considered inert in the absence of signaling through the TCR. By contrast, mitogenic rat CD28 mAb reportedly expand regulatory T cells without TCR stimulation. We found that a commercially available human CD28 mAb (ANC28) stimulated PBMC without TCR co-ligation or cross-linking; ANC28 selectively expanded CD4(+)CD25(+)FOXP3(-) (Teff) and CD4(+)CD25(+)FOXP3(+) (Treg) cells. ANC28 stimulated the CD45RO(+) CD4(+) (memory) population, whereas CD45RA(+)CD4(+) (naive) cells did not respond. ANC28 also induced inflammatory cytokines. Treg induced by ANC28 retain the Treg phenotype longer than costimulated Treg. Treg induced by ANC28 suppressed CD25(-) T cells through a contact-dependent mechanism. Purity influenced the response of CD4(+)CD25(+ )cells because bead-purified CD4(+)CD25(+ )cells (85-90% pure) responded strongly to ANC28, whereas 98% pure FACS-sorted CD4(+)CD25(bright) (Treg) did not respond. Purified CD4(+)CD25(int) cells responded similarly to the bead-purified CD4(+)CD25(+) cells. Thus, pre-activated CD4(+) T cells (CD25(int)) respond to ANC28 rather than Treg (CD25(bright)). The ability of ANC28 to expand both effectors producing inflammatory cytokines as well as suppressive regulatory T cells might be useful for ex vivo expansion of therapeutic T cells.     

子痫前期孕妇外周血及脐血中CD4＋CD25＋T细胞比例的变化

目的 研究子痫前期孕妇外周血及脐血中CD4＋CD25＋T细胞的比例变化，探讨CD4＋CD25＋T细胞在妊娠免疫中的作用。方法 应用流式细胞技术检测孕妇外周血及脐血中CD4＋CD25＋T细胞的比例变化。结果 子痫前期孕妇外周血及脐血中CD4＋CD25＋T细胞比例较低，低于同孕期的正常妊娠组P〈0.05）。结论 CD4＋CD25＋T细胞在妊娠期高血压疾病的发生发展中发挥一定作用。     

多发性硬化免疫调节细胞的变化及与疾病复发的关系

目的观察复发与缓解期多发性硬化患者免疫调节细胞(CD4+CD25+细胞及Th2细胞)的变化及与疾病复发的关系.方法用流式细胞仪分别对复发期(n=26)及缓解期(n=22)MS患者的外周血标本进行CD4+CD25+细胞及Th2细胞的百分数进行检测,并与疾病复发进行相关性比较.结果复发期与缓解期CD4+CD25+细胞及Th2细胞均少于正常对照标本;而CD4+CD25-细胞及Th1细胞与对照标本比较无统计学意义的差异.CD4+CD25+细胞及Th2细胞的数量与疾病复发具有良好的相关性(P值分别等于0.000及0.003).结论MS患者在复发期与缓解期均存在有免疫调节细胞数量减少,免疫调节细胞的减少程度与疾病复发具有良好的相关性.     

Peripheral CD4loCD40+ auto-aggressive T cell expansion during insulin-dependent diabetes mellitus

The generation of auto-aggressive T cells involves failure of central or peripheral tolerance. We previously demonstrated that peripheral CD4(lo)CD40(+) T cells give rise to pathogenic T cells in the non-obese diabetic (NOD) model. Here we show that peripheral CD4(+)CD40(+) T cells from diabetic or pre-diabetic NOD mice induce insulin-dependent diabetes mellitus. Consistent with breach of peripheral tolerance, CD4(lo)CD40(+) T cells expand with age in NOD mice but not in MHC-matched non-obese resistant (NOR) or BALB/c controls. Suggestive of a causal role for CD40 in autoimmunity, blocking CD40-CD154 interactions early during NOD development prevents autoaggressive T cell expansion while promoting increases in CD4(+)CD25(+) regulatory T cells. Importantly, CD40 signals promote expansion of V alpha 3.2(+) and V alpha 8.3(+) T cells. Furthermore, peripheral V alpha 3.2(+)CD40(+) T cells induce diabetes in NOD.scid recipients while V alpha 8.3(+) T cells or V alpha 3.2(+)-depleted T cell populations do not. This is the first demonstration that primary T cells transfer disease with the kinetics of auto-aggressive T cell clones and that specific TCR V alpha expansion promotes diabetes.