Recurrent superantigen exposure in vivo leads to highly suppressive CD4+CD25+ and CD4+CD25‐ T cells with anergic and suppressive genetic signatures

Staphylococcal enterotoxin B (SEB) activates T cells via non‐canonical signalling through the T cell receptor and is an established model for T cell unresponsiveness in vivo. In this study, we sought to characterize the suppressive qualities of SEB‐exposed CD4⁺ T cells and correlate this with genetic signatures of anergy and suppression. SEB‐exposed CD25⁺ and CD25^‐Vβ8⁺CD4⁺ T cells expressed forkhead box P3 (FoxP3) at levels comparable to naive CD25⁺ T regulatory cells and were enriched after exposure in vivo. Gene related to anergy in lymphocytes (GRAIL), an anergy‐related E3 ubiquitin ligase, was up‐regulated in the SEB‐exposed CD25⁺ and CD25^‐FoxP3⁺Vβ8⁺CD4⁺ T cells and FoxP3^‐CD25^‐Vβ8⁺CD4⁺ T cells, suggesting that GRAIL may be important for dominant and recessive tolerance. The SEB‐exposed FoxP3⁺GRAIL⁺ T cells were highly suppressive and non‐proliferative independent of CD25 expression level and via a glucocorticoid‐induced tumour necrosis factor R‐related protein‐independent mechanism, whereas naive T regulatory cells were non‐suppressive and partially proliferative with SEB activation in vitro. Lastly, adoptive transfer of conventional T cells revealed that induction of FoxP3⁺ regulatory cells is not operational in this model system. These data provide a novel paradigm for chronic non‐canonical T cell receptor engagement leading to highly suppressive FoxP3⁺GRAIL⁺CD4⁺ T cells.     

CD4+ CD25+ [corrected] regulatory T cells render naive CD4+ CD25- T cells anergic and suppressive

CD4(+) CD25(+) Foxp3(+) naturally occurring regulatory T cells (nTreg) are potent inhibitors of almost all immune responses. However, it is unclear how this minor population of cells is capable of exerting its powerful suppressor effects. To determine whether nTreg mediate part of their suppressor function by rendering naive T cells anergic or by converting them to the suppressor phenotype, we cocultured mouse nTreg with naive CD4(+) CD25(-) T cells from T-cell receptor (TCR) transgenic mice on a RAG deficient (RAG(-/-)) background in the presence of anti-CD3 and interleukin-4 (IL-4) to promote cell viability. Two distinct responder cell populations could be recovered from the cocultures. One population remained undivided in the coculture and was non-responsive to restimulation with anti-CD3 or exogenous IL-2, and could not up-regulate IL-2 mRNA or CD25 expression upon TCR restimulation. Those responder cells that had divided in the coculture were anergic to restimulation with anti-CD3 but responded to restimulation with IL-2. The undivided population was capable of suppressing the response of fresh CD4(+) CD25(-) T cells and CD8(+) T cells, while the divided population was only marginally suppressive. Although cell contact between the induced regulatory T cell (iTreg) and the responders was required for suppression to be observed, anti-transforming growth factor-beta partially abrogated their suppressive function. The iTreg did not express Foxp3. Therefore nTreg are not only able to suppress immune responses by inhibiting cytokine production by CD4(+) CD25(-) responder cells, but also appear to modulate the responder cells to render them both anergic and suppressive.     

IL-2 induces in vivo suppression by CD4(+)CD25(+)Foxp3(+) regulatory T cells

Interleukin-2 (IL-2) treatment is currently used to enhance T cell-mediated immune responses against tumors or in viral infections. At the same time, IL-2 is essential for the peripheral homeostasis of CD4(+)CD25(+)Foxp3(+ )regulatory T cells (Treg). In our study, we show that IL-2 is also an important activator of Treg suppressive activity in vivo. IL-2 treatment induces Treg expansion as well as IL-10 production and increases their suppressive potential in vitro. Importantly, in vivo application of IL-2 via gene-gun vaccination using IL-2 encoding DNA plasmids (pIL-2) inhibited naive antigen-specific T cell proliferation as well as a Th1-induced delayed type hypersensitivity response. The suppressive effect can be transferred onto naive animals by Treg from IL-2-treated mice and the suppression depends on the synergistic action of IL-10 and TGF-beta. These data highlight that during therapeutic treatment with IL-2 the concomitant activation of Treg may indeed counteract the intended activation of cellular immunity.     

CD25+ CD4+ T cells compete with naive CD4+ T cells for IL-2 and exploit it for the induction of IL-10 production

Maintenance of homeostasis in the immune system involves competition for resources between T lymphocytes, which avoids the development of immune pathology seen in lymphopenic mice. CD25+ CD4+ T cells are important for homeostasis, but there is as yet no consensus on their mechanisms of action. Although CD25+ CD4+ T cells cause substantial down-regulation of IL-2 mRNA in responder T cells in an in vitro co-culture system, the presence of IL-protein can be demonstrated by intracellular staining. As a consequence of competition for IL-2, CD25+ CD4+ T cells further up-regulate the IL-2R alpha chain (CD25), a process that is strictly dependent on IL-2, whereas responder T cells fail to up-regulate CD25. Similarly, adoptive transfer into lymphopenic mice showed that CD25+ CD4+ T cells interfere with CD25 up-regulation on co-transferred naive T cells, while increasing their own CD25 levels. IL-2 sequestration by CD25+ CD4+ T cells is not a passive phenomenon but instead initiates--in conjunction with signals through the TCR--their differentiation to IL-10 production. Although IL-10 is not required for in vitro suppression, it is vital for the in vivo function of regulatory T cells. Our data provide a link explaining the apparent difference in regulatory mechanisms in vitro and in vivo.     

Natural CD4 CD25(+) regulatory T cells control the burst of superantigen-induced cytokine production: the role of IL-10.

In normal mice a subpopulation of CD4 T cells constitutively expresses the IL-2 receptor alpha chain (CD25). This natural CD4 CD25(+) subset is thymus-born, constitutively expresses IL-10 mRNA,does not produce IL-2 and is resistant to apoptosis. These cells behave as regulatory T cells in the control of self-tolerance, inflammatory reactions and T cell homeostasis. The mechanisms by which natural CD4 CD25(+) cells control the immune response is unclear. We examined CD25-deficient mice, which over-express various cytokines, including proinflammatory molecules, after bacterial superantigen stimulation in vivo. We observed that this abnormal cytokine production could be controlled by the injection of natural CD4 CD25(+) T cells and that IL-10 production is needed, as CD4 CD25(+) T cells from IL-10 knockout mice do not correct cytokine over-production in vivo. As the circulating IL-10 produced by CD25-deficient mice was ineffective, we deduced that the key source of IL-10 was the regulatory T cell population. IL-10 is also involved in the control of cytokine production by normal T cells. However, the target of IL-10 in this control is undefined. Whether it acts directly on the effector T cells or on the regulatory CD4 CD25(+) T cells themselves to induce their functional maturation has to be clarified.     

Induction of IL-10 producing CD4+ T cells with regulatory activities by stimulation with IL-10 gene-modified bone marrow derived dendritic cells

Dendritic cells (DCs) can induce both tolergenic as well as effective immune responses in the lung. Pulmonary DCs producing interleukin (IL)-10 mediated tolerance induced by respiratory exposure to antigen. IL-10 is an important immunosuppressive cytokine, which inhibits maturation and function of DC. To assess whether IL-10 producing DCs can exert the tolergenic effect through the differentiation of regulatory T cells, bone marrow derived DCs were genetically modified by IL-10 expressing adenovirus. IL-10 gene modified DCs (Ad-IL-10-DC) displayed a characteristic phenotype of immature DCs. Here we showed that in vitro repetitive stimulation of naïve DO11·10 CD4⁺ T cells with Ad-IL-10-DCs resulted in a development of IL-10 producing T-cell regulatory cells. These T cells could not proliferate well but also lost their ability to produce interferon-γ upon restimulation with irradiated splenocytes and ovalbumin peptide. Furthermore, in co-culture experiments these T cells inhibited the antigen-driven proliferation of naïve CD4+ T cells in a dose-dependent manner. Our findings demonstrated that IL-10 producing DCs had the potential to induce the differentiation of Tr1-like cells and suggested their therapeutic use.     

The role of different subsets of T regulatory cells in controlling autoimmunity

T regulatory cells-in addition to clonal deletion and anergy-are essential for the downregulation of T cell responses to both foreign and self antigens, and for the prevention of autoimmunity. Recent progress has been made in characterising the different subsets of T regulatory cells, the factors that drive their differentiation, and their mode of action. The resolution of these mechanisms will make it possible to use T regulatory cells therapeutically in human autoimmune diseases.     

Impaired CD28-mediated co-stimulation in anergic T cells

We have investigated a CD28 co-stimulation in anergic T cellsin staphylococcal enterotoxin Binoculated mice by stimulatingthe cells with a plate-coated anti-TCR antibody in the presenceor absence of an anti-CD28 antibody. CD28 co-stimulation increasedthe levels of IL-2 and IL-4 mRNAs in nalve CD4⁺V_ß8⁺T cells. However, it did not increase the levels of IL-4 mRNAat all and only partially increased those of IL-2 mRNA in anergicT cells. It was demonstrated that CD28 co-stimulation was impairedso that it no longer stabilized cytoklne mRNAs in anergic cells.The levels of IL-4 mRNA in response to TCR stimulation werehigher in anergic T cells than those in nalve T cells in spiteof the defective CD28 co-stimulation in the former cells. Anergyinduction and generation of a T_h2-type immune response in vivoare discussed     

Naturally anergic and suppressive CD25(+)CD4(+) T cells as a functionally and phenotypically distinct immunoregulatory T cell subpopulation

A CD4(+) T cell subpopulation defined by the expression levels of a particular cell surface molecule (e.g. CD5, CD45RB, CD25, CD62L or CD38) bears an autoimmune-preventive activity in various animal models. Here we show that the expression of CD25 is highly specific, when compared with other molecules, in delineating the autoimmune-preventive immunoregulatory CD4(+) T cell population. Furthermore, although CD25 is an activation marker for T cells, the following findings indicate that immunoregulatory CD25(+)CD4(+) T cells are functionally distinct from activated or anergy-induced T cells derived from CD25(-)CD4(+) T cells. First, the former are autoimmune-preventive in vivo, naturally unresponsive (anergic) to TCR stimulation in vitro and, upon TCR stimulation, able to suppress the activation/proliferation of other T cells, whereas the latter scarcely exhibit the in vivo autoimmune-preventive activity or the in vitro suppressive activity. Second, such activated or anergy-induced CD25(-) spleen cells produce various autoimmune diseases when transferred to syngeneic athymic nude mice, whereas similarly treated normal spleen cells, which include CD25(+)CD4(+) T cells, do not. Third, upon polyclonal T cell stimulation, CD25(+)CD4(+) T cells express CD25 at higher levels and more persistently than CD25(-)CD4(+) T cell-derived activated T cells; moreover, when the stimulation is ceased, the former revert to the original levels of CD25 expression, whereas the latter lose the expression. These results collectively indicate that naturally anergic and suppressive CD25(+)CD4(+) T cells present in normal naive mice are functionally and phenotypically stable, distinct from other T cells, and play a key role in maintaining immunologic self-tolerance.     

CD25(+)CD4(+) regulatory T cells exert in vitro suppressive activity independent of CTLA-4

Cytotoxic T lymphocyte antigen-4 (CTLA-4) is constitutively expressed on CD25(+)CD4(+) regulatory T cells (Treg) and is suggested to play a role in Treg-mediated suppression. However, the results of analysis with anti-CTLA-4 have been controversial. We addressed this issue by analyzing mice over-expressing or deficient in CTLA-4. For over-expression, CTLA-4 transgenic mice expressing a full-length (FL) or a truncated (TL) mutant of CTLA-4 were analyzed. FL T cells expressed similar levels of CTLA-4 to Treg, whereas TL T cells expressed much higher levels on the cell surface. The number of Treg in both mice was decreased, although Foxp3 expression was not altered. Treg from both mice exerted suppressive activity, whereas CD25(-) T cells from FL mice showed no suppression. Furthermore, CD25(+)CD4 thymocytes from young CTLA-4-deficient mice were analyzed and found to exhibit suppressive activity. These results indicate that Treg exert in vitro suppressive activity independent of CTLA-4 expression.     

Differential production of IL-10 by T cells and monocytes of HIV-infected individuals: association of IL-10 production with CD28-mediated immune responsiveness

Immune unresponsiveness in HIV-1 infection can result from impaired signals delivered by the costimulatory CD28-B7 pathway and the altered production of immunoregulatory cytokines, in particular IL-10, whose production is altered in HIV-1 infection. In this study we investigate IL-10 regulation in T cells and monocytes from HIV⁺ individuals, and its association with CD28-mediated T cell proliferation. IL-10 production as analysed in T cell- and monocyte-depleted peripheral blood mononuclear cells (PBMC), and by intracellular staining at the single-cell level, reveals a defect in IL-10 production by CD4⁺ and CD8⁺ T cells, whereas monocytes constitute the major IL-10-producing cell type. To investigate the impact of IL-10 on immune responsiveness, CD28-mediated proliferative responses in HIV⁺ individuals were correlated with PHA-induced IL-10 production. CD4⁺ T cells expressed CD28, yet exhibited markedly reduced CD28-mediated cell proliferation. This CD28-mediated CD4⁺ T cell proliferation was found to be inversely associated with the levels of PHA-induced IL-10 production and could be restored, at least in part, by anti-IL-10 antibodies. These results suggest that IL-10 production is differentially regulated in T cells and monocytes of HIV⁺ individuals, and that IL-10 may have a role in inducing immune unresponsiveness by modulating the CD28-B7 pathway.     

Role of CD4(+)CD25(+) T regulatory cells in IL-2-induced vascular leak

T regulatory cells (CD4(+)CD25(+)) play an important role in the regulation of the immune response. However, little is known about the ability of T regulatory cells to regulate endothelial cell (EC) damage following activation of lymphocytes with IL-2. Therefore, in the current study, we examined the role of T regulatory cells and the subsequent T(h)1/T(h)2 bias in IL-2-mediated EC injury using the well-characterized C57BL/6 (T(h)1-biased) and BALB/c (T(h)2-biased) models. Following IL-2 treatment, BALB/c mice were less susceptible to IL-2-induced vascular leak syndrome (VLS) compared with C57BL/6 mice. Splenocytes from BALB/c mice displayed less cytotoxicity against ECs compared with those from C57BL/6 mice. Interestingly, BALB/c mice had significantly higher numbers of CD4(+)CD25(+) T regulatory cells, which proliferated more profoundly following IL-2 treatment, compared with CD4(+)CD25(+) T regulatory cells from C57BL/6 mice. In addition, T regulatory cells from naive BALB/c mice were more potent suppressors of anti-CD3 mAb-stimulated proliferation of T cells than similar cells from C57BL/6 mice. Depletion of T regulatory cells in both BALB/c and C57BL/6 mice led to a significant increase in IL-2-induced VLS. Together, the results from this study suggest that CD4(+)CD25(+) T regulatory cells play an important role in the regulation of IL-2-induced EC injury.     

HIV-1 induces IL-10 production in human monocytes via a CD4-independent pathway

In HIV-infected patients, increased levels of IL-10, mainly produced by virally infected monocytes, were reported to be associated with impaired cell-mediated immune responses. In this study, we investigated how HIV-1 induces IL-10 production in human monocytes. We found that CD14(+) monocytes infected by either HIV-1(213) (X4) or HIV-1(BaL) (R5) produced IL-10, IL-6, tumor necrosis factor-alpha (TNF-alpha), and to a lesser extent, IFN-gamma. However, the capacity of HIV-1 to induce these cytokines was not dependent on virus replication since UV-inactivated HIV-1 induced similar levels of these cytokines. In addition, soluble HIV-1 gp160 could induce CD14(+) monocytes to produce IL-10 but at lower levels. Cross-linking CD4 molecules (XLCD4) with anti-CD4 mAbs and goat anti-mouse IgG (GAM) resulted in high levels of IL-6, TNF-alpha and IFN-gamma but no IL-10 production by CD14(+) monocytes. Interestingly, neither anti-CD4 mAbs nor recombinant soluble CD4 (sCD4) receptor could block IL-10 secretion induced by HIV-1(213), HIV-1(BaL) or HIV-1 gp160 in CD14(+) monocytes, whereas anti-CD4 mAb or sCD4 almost completely blocked the secretion of the other cytokines. Furthermore, HIV-1(213) could induce IL-10 mRNA expression in CD14(+) monocytes while XLCD4 by anti-CD4 mAb and GAM failed to do so. As with IL-10 protein levels, HIV-1(213)-induced IL-10 mRNA expression in CD14(+) monocytes could not be inhibited by anti-CD4 mAb or sCD4. Taken together, HIV-1 binding to CD14(+) monocytes can induce CD4-independent IL-10 production at both mRNA and protein levels. This finding suggests that HIV induces the immunosuppressive IL-10 production in monocytes and is not dependent on CD4 molecules and that interference with HIV entry through CD4 molecules may have no impact on counteracting the effects of IL-10 during HIV infection.     

Two modes of immune suppression by Foxp3(+) regulatory T cells under inflammatory or non-inflammatory conditions

Foxp3-expressing regulatory T cells (Tregs) play a crucial role in maintaining immune tolerance and homeostasis. One of the key issues for understanding Treg immunobiology is to determine how they suppress excessive or aberrant immune responses. Although a number of molecules have been reported to contribute to Treg suppressive function, the importance and precise role of each molecule is not clear. In this review, we propose and discuss that two modes of suppression can be distinguished. In the physiological and steady state, activation of naïve T cells can be suppressed by natural Tregs via deprivation of activation signals including CD28 signal and IL-2 from antigen-reactive T cells, keeping the latter in a naïve state in lymphoid tissues. These deprivation mechanisms are transiently abrogated in inflammatory conditions, allowing T cells to respond to antigen. In contrast, in highly inflammatory environments, for example, in microbial infection, activated Tregs acquire the capacity to kill or inactivate effector T cells and antigen-presenting cells, for example, via granzyme/perforin formation and IL-10 secretion, thereby actively damping excessive immune responses. Understanding these processes will help effectively controlling physiological and pathological immune responses via Tregs.     

Total glucosides of paeony induces regulatory CD4(+)CD25(+) T cells by increasing Foxp3 demethylation in lupus CD4(+) T cells

Total glucosides of paeony (TGP), an active compound extracted from Paeony root, has been used in therapy for autoimmune diseases. However the molecular mechanism of TGP in the prevention of autoimmune response remains unclear. In this study, we found that TGP treatment significantly increased the percentage and number of Treg cells in lupus CD4(+) T cells. Further investigation revealed that treatment with TGP increased the expression of Foxp3 in lupus CD4(+) T cells by down-regulating Foxp3 promoter methylation levels. However, we couldn't observe similar results in healthy control CD4(+) T cells treated by TGP. Moreover, our results also showed that IFN-γ and IL-2 expression was enhanced in TGP-treated lupus CD4(+) T cells. These findings indicate that TGP inhibits autoimmunity in SLE patients possibly by inducing Treg cell differentiation, which may in turn be due to its ability to regulate the methylation status of the Foxp3 promoter and activate IFN-γ and IL-2 signaling.     

筛选反义寡核苷酸抑制哮喘大鼠CD4+T细胞IL-4表达的研究

目的探讨不同序列的反义寡核苷酸对哮喘大鼠CD4+T细胞IL-4表达的抑制作用.方法采用脂质体转染技术,将不同的反义寡核苷酸(AS-1反外显子-1;AS-2反外显子-2;AS-3反翻译终止部位)和空白对照分别转入经免疫磁珠阴性分离的哮喘大鼠CD4+T淋巴细胞,细胞培养28小时后,用ELISA和半定量RT-PCR分别检测细胞培养上清IL-4和细胞内IL-4 mRNA的水平.结果不同组RT-PCR结果(IL-4/β-actin相对吸光度)AS-1、AS-2、 AS-3及空白组分别为0.261 5±0.147 6、0.288 5±0.141 1、1.101 2±0.364 1及1.206 8±0.383 6(F=22.597,P＜0.01).ELISA检测培养上清液IL-4结果AS-1、AS-2、 AS-3及空白组分别为13.800±7.233、15.329±7.358、52.643±12.075及58.286±14.100(F=34.976,P＜0.01).经AS1、2干预后细胞内IL-4mRNA和细胞培养上清IL-4的水平均较AS-3和空白组干预后 IL-4的水平低(P＜0.01).结论IL-4反义寡核苷酸能够抑制哮喘大鼠CD4+T淋巴细胞IL-4和IL-4mRNA表达;不同序列IL-4反义寡核苷酸抑制作用存在差异.     

Long-term cultured CD40-activated B lymphocytes differentiate into plasma cells in response to IL-10 but not IL-4

We compared the effects of IL-10 and IL-4 on the functions ofB lymphocytes triggered through their CD40. During the initialphase, IL-10 was as potent as IL-4 in inducing the expansionof viable B cells. Then, cellular expansion slowed down andafter {small tilde}3 weeks the number of B cells started todecline. While the combination of IL-10 and IL-4 was synerglsticduring the first 2 weeks of culture, B cell recovery declinedafter 3 weeks, indicating that IL-10 prevails over IL-4. Thoseeffects were not restricted to a specific B cell subset as bothslgD⁺ B cells and slgD^– B cells behaved in a similar way,though the latter population responded with a slightly acceleratedkinetic. Inverted microscope examination and scanning electronmicroscopy showed that in response to IL-10, CD40-activatedB cell cultures were heterogeneous with loose aggregates ofcells as well as free floating large ovoid cells. In contrast,in the presence of IL-4, CD40-activated B cell cultures wereessentially composed of tight cell clumps. IL-10 progressivelyinduced all B cells to differentiate into non-replicating cellswith intracytoplasmic Ig that secreted Ig at a high rate. Cytologlcanalysis indicated that IL-10 cultured cells display a basophiliccytoplasm with an arcoplasm and a low nucleus/cytoplasm ratio.Transmission electron microscopy demonstrated that when IL-10was added to the culture, B cells displayed structures for excretionwith extended endoplasmic reticulum and dilated cisternae containingparacrystalline structures, typical of plasmablasts cells. Takentogether, these results indicate that IL-10 acts as a plasmacell differentiation factor for CD40-activated B cells.     

A model of suppression of the antigen-specific CD4 T cell response by regulatory CD25+CD4 T cells in vivo

Despite intense recent interest, the suppressive mechanisms of regulatory CD25+CD4 T cells remain poorly understood. One deficiency in the field is the lack of in vivo models where the effects of regulatory CD25+CD4 T cells on antigen-specific responder T cells can be measured quantitatively. We describe one such model here. We compared responses of adoptively transferred naive wild-type antigen-specific CD4 T cells in syngeneic CD28-/- and wild-type recipient mice toward a nominal antigen. The cells exhibited a greater degree of proliferation and differentiation in CD28-/- mice and could not be rendered functionally hyporesponsive by systemic exposure to adjuvant-free antigen. The only reason we were able to find to explain this difference was the deficiency of regulatory CD25+CD4 T cells in the CD28-/- mice. Use of CD28-/- mice as adoptive transfer recipients provides a simple model that reveals the contribution of regulatory CD25+CD4 T cells in controlling antigen-driven responses in vivo.     

IL-10 is involved in the suppression of experimental autoimmune encephalomyelitis by CD25+CD4+ regulatory T cells

CD25(+)CD4(+) regulatory T cells inhibit the activation of autoreactive T cells in vitro and in vivo, and suppress organ-specific autoimmune diseases. The mechanism of CD25(+)CD4(+) T cells in the regulation of experimental autoimmune encephalomyelitis (EAE) is poorly understood. To assess the role of CD25(+)CD4(+) T cells in EAE, SJL mice were immunized with myelin proteolipid protein (PLP)(139-151) to develop EAE and were treated with anti-CD25 mAb. Treatment with anti-CD25 antibody following immunization resulted in a significant enhancement of EAE disease severity and mortality. There was increased inflammation in the central nervous system (CNS) of anti-CD25 mAb-treated mice. Anti-CD25 antibody treatment caused a decrease in the percentage of CD25(+)CD4(+) T cells in blood, peripheral lymph node (LN) and spleen associated with increased production of IFN-gamma and a decrease in IL-10 production by LN cells stimulated with PLP(130-151) in vitro. In addition, transfer of CD25(+)CD4(+) regulatory T cells from naive SJL mice decreased the severity of active EAE. In vitro, anti-CD3-stimulated CD25(+)CD4(+) T cells from naive SJL mice secreted IL-10 and IL-10 soluble receptor (sR) partially reversed the in vitro suppressive activity of CD25(+)CD4(+) T cells. CD25(+)CD4(+) T cells from IL-10-deficient mice were unable to suppress active EAE. These findings demonstrate that CD25(+)CD4(+) T cells suppress pathogenic autoreactive T cells in actively induced EAE and suggest they may play an important natural regulatory function in controlling CNS autoimmune disease through a mechanism that involves IL-10.