Coordinated control of immunity to muscle stage Trichinella spiralis by IL-10, regulatory T cells, and TGF-beta

We previously demonstrated that IL-10 is critical in the control of acute inflammation during development of Trichinella spiralis in the muscle. In this study, we use gene-targeted knockout mice, adoptive transfer of specific T cell populations, and in vivo Ab treatments to determine the mechanisms by which inflammation is controlled and effector T cell responses are moderated during muscle infection. We report that CD4(+)CD25(-) effector T cells, rather than CD4(+)CD25(+) regulatory T cells, suppress inflammation by an IL-10-dependent mechanism that limits IFN-gamma production and local inducible NO synthase induction. Conversely, we show that depletion of regulatory T cells during infection results in exaggerated Th2 responses. Finally, we provide evidence that, in the absence of IL-10, TGF-beta participates in control of local inflammation in infected muscle and promotes parasite survival.     

Suppression of murine allergic airway disease by IL-2:anti-IL-2 monoclonal antibody-induced regulatory T cells

Regulatory T cells (Treg) play a decisive role in many diseases including asthma and allergen-induced lung inflammation. However, little progress has been made developing new therapeutic strategies for pulmonary disorders. In the current study we demonstrate that cytokine:antibody complexes of IL-2 and anti-IL-2 mAb reduce the severity of allergen-induced inflammation in the lung by expanding Tregs in vivo. Unlike rIL-2 or anti-IL-2 mAb treatment alone, IL-2:anti-IL-2 complexes dampened airway inflammation and eosinophilia while suppressing IL-5 and eotaxin-1 production. Mucus production, airway hyperresponsiveness to methacholine, and parenchymal tissue inflammation were also dramatically reduced following IL-2:anti-IL-2 treatment. The suppression in allergic airway disease was associated with a marked expansion of Tregs (IL-10(+)CD4(+)CD25(+) and Foxp3(+)CD4(+)CD25(+)) in the tissues, with a corresponding decrease in effector T cell responses. The ability of IL-2:anti-IL-2 complexes to suppress airway inflammation was dependent on Treg-derived IL-10, as IL-10(+/+), but not IL-10(-/-) Tregs, were capable of mediating the suppression. Furthermore, a therapeutic protocol using a model of established airway allergy highlighted the ability of IL-2:anti-IL-2 complexes to expand Tregs and prevent successive airway inflammation and airway hyperresponsiveness. This study suggests that endogenous Treg therapy may be a useful tool to combat the rising incidence of allergic airway disease.     

Severe CD4 T cell-mediated immunopathology in murine schistosomiasis is dependent on IL-12p40 and correlates with high levels of IL-17

C57BL/6 mice infected with the helminth Schistosoma mansoni develop small hepatic granulomas around parasite eggs, but concomitant immunization with soluble schistosome egg Ags (SEA) in CFA (SEA/CFA) causes marked exacerbation of the lesions in a Th1-dominated environment characterized by high levels of IFN-gamma. We explored the cause of the severe immunopathology by using IL-12p40(-/-) and IL-12p35(-/-) mice. SEA/CFA-immunized IL-12p40(-/-) mice, incapable of making IL-12 or IL-23, were completely resistant to high pathology, and their SEA-stimulated lymphoid cells failed to secrete significant IFN-gamma or IL-17. In contrast, SEA/CFA-immunized IL-12p35(-/-) mice, able to make IL-23 but not IL-12, developed severe lesions that correlated with high levels of IL-17, low IFN-gamma, and an expansion of activated CD4 T cells with a CD44(high)/CD62L(low) memory phenotype. In vivo administration of neutralizing anti-IL-17 mAb markedly inhibited hepatic granulomatous inflammation. Importantly, CBA mice, a naturally high pathology strain, also displayed elevated IL-17 levels comparable to those seen in the SEA/CFA-immunized BL/6 mice, and their lesions were similarly reduced by in vivo treatment with anti-IL-17. Our findings indicate that an IL-17-producing T cell population, likely driven by IL-23, significantly contributes to severe immunopathology in schistosomiasis.     

IL-10 synergizes with IL-4 and transforming growth factor-beta to inhibit macrophage cytotoxic activity.

After activation with IFN-gamma, thioglycollate-elicited murine peritoneal macrophages kill schistosomula of Schistosoma mansoni in vitro by an L-arginine-dependent mechanism which involves the production of reactive nitrogen oxides (NO). In the present study we demonstrate that the regulatory cytokines IL-10, IL-4, and transforming growth factor-beta (TGF-beta) are potent inhibitors of this extracellular killing function of activated macrophages. Each cytokine was found to suppress killing of schistosomula in a dose-dependent fashion. The activity of IL-10 was not permanent, because subsequent treatment with additional IFN-gamma 2 to 6 h later reversed the inhibition of macrophage larval killing. More importantly, the combination of suboptimal levels of any two of these three cytokines was found to give a potent synergistic suppression of schistosomulum killing by IFN-gamma-treated macrophages. Similarly, IL-10, IL-4, or TGF-beta alone blocked the production of NO, and when used in combination these cytokines exhibited an enhanced inhibitory effect on nitrite production. Macrophage-mediated killing of schistosomula through the generation of NO has been shown previously to be a major effector mechanism of schistosome immunity. The results presented here suggest that the suppression of this mechanism by induction of the regulatory cytokines IL-10, IL-4, and TGF-beta, which are known to be produced during schistosome infection, may be an important strategy used by the parasite to evade macrophage-mediated immune destruction.     

Production of IL-10 by CD4+ T lymphocytes correlates with down-regulation of Th1 cytokine synthesis in helminth infection.

After the onset of parasite egg deposition, mice infected with the helminth Schistosoma mansoni mount strong Th2 cytokine responses in the absence of significant Th1 cytokine synthesis. To examine the basis of this immunoregulatory state, spleen or lymph node cells from schistosome-infected mice were stimulated with parasite-specific Ag and the supernatants tested for their capacity to suppress IFN-gamma synthesis by a Th1 cell line. Strong inhibition was observed that was neutralized by a mAb against IL-10, a cytokine previously shown to down-regulate Th1 cytokine synthesis. By means of ELISA measurements the production of IL-10 in schistosome infection was confirmed and shown to depend on CD4+ T cells. IL-10 synthesis stimulated by either mitogen or Ag was observed only at those stages of infection when Th2 response induction and Th1 cytokine down-regulation also occurred and was not detected in mice vaccinated with attenuated parasites. Moreover, the addition of the neutralizing anti-IL-10 mAb to Ag-stimulated spleen cell cultures from infected mice caused a dramatic augmentation in IFN-gamma synthesis. These findings suggest that IL-10 is responsible for the down-regulation of Th1 responses observed in schistosome infections, a phenomenon that may enable the parasite to escape potentially harmful cell-mediated responses.     

Memory T(H)2 cells induce alternatively activated macrophages to mediate protection against nematode parasites

Although primary and memory responses against bacteria and viruses have been studied extensively, T helper type 2 (T(H)2) effector mechanisms leading to host protection against helminthic parasites remain elusive. Examination of the intestinal epithelial submucosa of mice after primary and secondary infections by a natural gastrointestinal parasite revealed a distinct immune-cell infiltrate after challenge, featuring interleukin-4-expressing memory CD4(+) T cells that induced IL-4 receptor(hi) (IL-4R(hi)) CD206(+) alternatively activated macrophages. In turn, these alternatively activated macrophages (AAMacs) functioned as important effector cells of the protective memory response contributing to parasite elimination, demonstrating a previously unknown mechanism for host protection against intestinal helminths.     

IL-27 promotes IL-10 production by effector Th1 CD4+ T cells: a critical mechanism for protection from severe immunopathology during malaria infection

Infection with the malaria parasite, Plasmodium, is characterized by excessive inflammation. The establishment of a precise balance between the pro- and anti-inflammatory responses is critical to guarantee control of the parasite and survival of the host. IL-10, a key regulatory cytokine produced by many cells of the immune system, has been shown to protect mice against pathology during acute Plasmodium0 chabaudi chabaudi AS model of malaria. However, the critical cellular source of IL-10 is still unknown. In this article, we demonstrate that T cell-derived IL-10 is necessary for the control of pathology during acute malaria, as mice bearing specific deletion of Il10 in T cells fully reproduce the phenotype observed in Il10(-)(/)(-) mice, with significant weight loss, decline in temperature, and increased mortality. Furthermore, we show that IFN-γ(+) Th1 cells are the main producers of IL-10 throughout acute infection, expressing high levels of CD44 and ICOS, and low levels of CD127. Although Foxp3(+) regulatory CD4(+) T cells produce IL-10 during infection, highly activated IFN-γ(+) Th1 cells were shown to be the essential and sufficient source of IL-10 to guarantee protection against severe immune-mediated pathology. Finally, in this model of malaria, we demonstrate that the generation of protective IL10(+)IFN-γ(+) Th1 cells is dependent on IL-27 signaling and independent of IL-21.     

Using eggs from Schistosoma mansoni as an in vivo model of helminth-induced lung inflammation

Schistosoma parasites are blood flukes that infect an estimated 200 million people worldwide. In chronic infection with Schistosoma, the severe pathology, including liver fibrosis and splenomegaly, is caused by the immune response to the parasite eggs rather than the parasite itself. Parasite eggs induce a Th2 response characterized by the production of IL-4, IL-5 and IL-13, the alternative activation of macrophages and the recruitment of eosinophils. Here, we describe injection of Schistosoma mansoni eggs as a model to examine parasite-specific Th2 cytokine responses in the lung and draining lymph nodes, the formation of pulmonary granulomas surrounding the egg, and airway inflammation. Following intraperitoneal sensitization and intravenous challenge, S. mansoni eggs are transported to the lung via the pulmonary arteries where they are trapped within the lung parenchyma by granulomas composed of lymphocytes, eosinophils and alternatively activated macrophages. Associated with granuloma formation, inflammation in the broncho-alveolar spaces, expansion of the draining lymph nodes and CD4 T cell activation can be observed. Here we detail the protocol for isolating Schistosoma mansoni eggs from infected livers (modified from), sensitizing and challenging mice, and recovering the organs (broncho-alveolar lavage (BAL), lung and draining lymph nodes) for analysis. We also include representative histologic and immunologic data and suggestions for additional immunologic analysis. Overall, this method provides an in vivo model to investigate helminth-induced immunologic responses in the lung, which is broadly applicable to the study of Th2 inflammatory diseases including helminth infection, fibrotic diseases, allergic inflammation and asthma. Advantages of this model for the study of type 2 inflammation in the lung include the reproducibility of a potent Th2 inflammatory response in the lung and draining lymph nodes, the ease of assessment of inflammation by histologic examination of the granulomas surrounding the egg, and the potential for long-term storage of the parasite eggs.     

Schistosoma mansoni worms induce anergy of T cells via selective up-regulation of programmed death ligand 1 on macrophages

Infectious pathogens can selectively stimulate activation or suppression of T cells to facilitate their survival within humans. In this study we demonstrate that the trematode parasite Schistosoma mansoni has evolved with two distinct mechanisms to suppress T cell activation. During the initial 4- to 12-wk acute stages of a worm infection both CD4(+) and CD8(+) T cells are anergized. In contrast, infection with male and female worms induced T cell anergy at 4 wk, which was replaced after egg laying by T cell suppression via a known NO-dependent mechanism, that was detected for up to 40 wk after infection. Worm-induced anergy was mediated by splenic F4/80(+) macrophages (Mphi) via an IL-4-, IL-13-, IL-10-, TGF-beta-, and NO-independent, but cell contact-dependent, mechanism. F4/80(+) Mphi isolated from worm-infected mice were shown to induce anergy of naive T cells in vitro. Furthermore, naive Mphi exposed to live worms in vitro also induced anergy in naive T cells. Flow cytometry on in vivo and in vitro worm-modulated Mphi revealed that of the family of B7 costimulatory molecules, only programmed death ligand 1 (PD-L1) was selectively up-regulated. The addition of inhibitory mAb against PD-L1, but not PD-L2, to worm-modulated Mphi completely blocked the ability of these cells to anergize T cells. These data highlight a novel mechanism through which S. mansoni worms have usurped the natural function of PD-L1 to reduce T cell activation during early acute stages of infection before the subsequent emergence of egg-induced T cell suppression in the chronic stages of infection.     

IL-22 mediates host defense against an intestinal intracellular parasite in the absence of IFN-γ at the cost of Th17-driven immunopathology

The roles of Th1 and Th17 responses as mediators of host protection and pathology in the intestine are the subjects of intense research. In this study, we investigated a model of intestinal inflammation driven by the intracellular apicomplexan parasite Eimeria falciformis. Although IFN-γ was the predominant cytokine during E. falciformis infection in wild-type mice, it was found to be dispensable for host defense and the development of intestinal inflammation. E. falciformis-infected IFN-γR(-/-) and IFN-γ(-/-) mice developed dramatically exacerbated body weight loss and intestinal pathology, but they surprisingly harbored fewer parasites. This was associated with a striking increase in parasite-specific IL-17A and IL-22 production in the mesenteric lymph nodes and intestine. CD4(+) T cells were found to be the source of IL-17A and IL-22, which drove the recruitment of neutrophils and increased tissue expression of anti-microbial peptides (RegIIIβ, RegIIIγ) and matrix metalloproteinase 9. Concurrent neutralization of IL-17A and IL-22 in E. falciformis-infected IFN-γR(-/-) mice resulted in a reduction in infection-induced body weight loss and inflammation and significantly increased parasite shedding. In contrast, neutralization of IL-22 alone was sufficient to increase parasite burden, but it had no effect on body weight loss. Treatment of an E. falciformis-infected intestinal epithelial cell line with IFN-γ, IL-17A, or IL-22 significantly reduced parasite development in vitro. Taken together, to our knowledge these data demonstrate for the first time an antiparasite effect of IL-22 during an intestinal infection, and they suggest that IL-17A and IL-22 have redundant roles in driving intestinal pathology in the absence of IFN-γ signaling.     

Naturally occurring CD4+Foxp3+ regulatory T cells are an essential, IL-10-independent part of the immunoregulatory network in Schistosoma mansoni egg-induced inflammation

In acute and chronic schistosomiasis, survival of the host requires a carefully balanced immune response against highly immunogenic parasite eggs. We characterized the phenotype, distribution, and functional role of CD4(+)Foxp3(+) naturally occurring regulatory T cells (naTregs) in schistosome egg-induced inflammation. In adoptive transfer experiments and by intracellular staining for Foxp3, we demonstrate significant frequencies of naTregs in hepatic granulomas and draining lymphoid tissues of mice infected with the trematode Schistosoma mansoni. Strikingly, egg-induced inflammation does not change the normal ratio between naTregs and effector CD4(+) T cells at the inflammatory site or in lymphoid organs in acute or chronic disease. However, increasing frequencies of CD103-expressing cells in the naTreg compartment indicate a change in phenotype for naTregs with disease progression. Because CD103 was described recently as an activation marker for naTregs, we speculate that naTregs in chronic schistosomiasis are potentially more suppressive. Furthermore, we found that most naTregs do not contribute to egg-induced IL-4 and IL-10 production. Importantly, depletion of CD25(+) naTregs strongly enhances the frequency of IL-4-producing effector T cells in acute egg-induced inflammation. It does not change clonal expansion of activated CD4(+) T cells. This regulation of egg-induced cytokine production does not require the presence of IL-10. These data demonstrate that naTregs limit egg-induced effector-cytokine production in our model. Our results identify naTregs as an important, IL-10-independent part of the regulatory network in schistosome egg-induced inflammation.     

IL-4R alpha expression by bone marrow-derived cells is necessary and sufficient for host protection against acute schistosomiasis

IL 4 receptor alpha (IL-4Ralpha) expression by non-bone marrow (BM)-derived cells is required to protect hosts against several parasitic helminth species. In contrast, we demonstrate that IL-4Ralpha expression by BM-derived cells is both necessary and sufficient to prevent Schistosoma mansoni-infected mice from developing severe inflammation directed against parasite ova, whereas IL-4Ralpha expression by non-BM-derived cells is neither necessary nor sufficient. Chimeras that express IL-4Ralpha only on non-BM-derived cells still produce Th2 cytokines, but overproduce IL-12p40, TNF, and IFN-gamma, fail to generate alternatively activated macrophages, and develop endotoxemia and severe hepatic and intestinal pathology. In contrast, chimeras that express IL-4Ralpha only on BM-derived cells have extended survival, even though the granulomas that they develop around parasite eggs are small and devoid of collagen. These observations identify distinct roles for IL-4/IL-13 responsive cell lineages during schistosomiasis: IL-4Ralpha-mediated signaling in non-BM-derived cells regulates granuloma size and fibrosis, whereas signaling in BM-derived cells suppresses parasite egg-driven inflammation within the liver and intestine.     

IL-4 influences IL-2 production and granulomatous inflammation in murine schistosomiasis mansoni.

In previous studies the dynamics of IL-2 production by splenic cells of Schistosoma mansoni infected mice was correlated with the intensity of hepatic granulomatous inflammation. To extend those observations, the present studies examined the role of IL-4 on the immune responsiveness of infected mice. The dynamics of IL-4 production by soluble egg Ag-stimulated splenic cells was similar to that of IL-2: minimal levels at the pre-oviposition or early worm egg deposition stages (4 to 6 wk) peak production coincident with maximal granulomatous response (8 wk) followed by a concurrent decline at the chronic stage (18 to 20 wk) in both parameters. Addition of murine rIL-4 to splenocyte cultures of acutely or chronically infected mice did not significantly enhance the soluble egg Ag-elicited proliferative response. Daily injections of rIL-4 (10 to 1000 U) given for 14 days to groups of mice with acute infection, at the high dose-enhanced IL-2, but not IL-4, production. Similar treatment given to chronically infected mice did not augment diminished lymphokine production. Chronically infected mice treated with 10 to 1000 U of rIL-4 showed significantly enhanced liver granulomatous responses compared with untreated animals and the augmented granulomas contained more enlarged macrophages and connective tissue matrix. Repeated injections of anti-IL-4 mAb (11B11) given to acutely infected mice significantly suppressed splenic cell proliferation, IL-2 and IL-4 production, and hepatic granulomatous inflammation. Similar treatment given to chronically infected mice also diminished the down-modulated granulomatous response. These data demonstrate that IL-4 plays an important role in the egg-directed granulomatous response and participates in the regulation of Ag-specific lymphoproliferation, and IL-2 and IL-4 production during the course of the infection.     

Infection with a helminth parasite prevents experimental colitis via a macrophage-mediated mechanism

The propensity of a range of parasitic helminths to stimulate a Th2 or regulatory cell-biased response has been proposed to reduce the severity of experimental inflammatory bowel disease. We examined whether infection with Schistosoma mansoni, a trematode parasite, altered the susceptibility of mice to colitis induced by dextran sodium sulfate (DSS). Mice infected with schistosome worms were refractory to DSS-induced colitis. Egg-laying schistosome infections or injection of eggs did not render mice resistant to colitis induced by DSS. Schistosome worm infections prevent colitis by a novel mechanism dependent on macrophages, and not by simple modulation of Th2 responses, or via induction of regulatory CD4+ or CD25+ cells, IL-10, or TGF-beta. Infected mice had marked infiltration of macrophages (F4/80+CD11b+CD11c(-)) into the colon lamina propria and protection from DSS-induced colitis was shown to be macrophage dependent. Resistance from colitis was not due to alternatively activated macrophages. Transfer of colon lamina propria F4/80+ macrophages isolated from worm-infected mice induced significant protection from colitis in recipient mice treated with DSS. Therefore, we propose a new mechanism whereby a parasitic worm suppresses DSS-induced colitis via a novel colon-infiltrating macrophage population.     

A role for CD44 in the production of IFN-gamma and immunopathology during infection with Toxoplasma gondii

The interaction of activated CD44 with its ligand, low m.w. hyaluronan, is involved in inflammation, but no role has been identified for this interaction in the regulation of an immune response to infection. In these studies, infection of C57BL/6 mice with Toxoplasma gondii resulted in increased expression of CD44 on T cells, B cells, NK cells, and macrophages, and a small percentage of CD4(+) T cells express an activated form of CD44. Administration of anti-CD44 to infected mice prevented the development of a CD4(+) T cell-dependent, infection-induced inflammatory response in the small intestine characterized by the overproduction of IFN-gamma. The protective effect of anti-CD44 treatment was associated with reduced production of IFN-gamma, but not IL-12, in vivo and in vitro. Furthermore, the addition of low m.w. hyaluronan to cultures of splenocytes or purified CD4(+) T cells from infected mice resulted in the production of high levels of IFN-gamma, which was dependent on IL-12 and TCR stimulation. Together, these results identify a novel role for CD44 in the regulation of IFN-gamma production by CD4(+) T cells during infection and demonstrate a role for CD44 in the regulation of infection-induced immune pathology.     

CD4+ natural regulatory T cells prevent experimental cerebral malaria via CTLA-4 when expanded in vivo

Studies in malaria patients indicate that higher frequencies of peripheral blood CD4(+) Foxp3(+) CD25(+) regulatory T (Treg) cells correlate with increased blood parasitemia. This observation implies that Treg cells impair pathogen clearance and thus may be detrimental to the host during infection. In C57BL/6 mice infected with Plasmodium berghei ANKA, depletion of Foxp3(+) cells did not improve parasite control or disease outcome. In contrast, elevating frequencies of natural Treg cells in vivo using IL-2/anti-IL-2 complexes resulted in complete protection against severe disease. This protection was entirely dependent upon Foxp3(+) cells and resulted in lower parasite biomass, impaired antigen-specific CD4(+) T and CD8(+) T cell responses that would normally promote parasite tissue sequestration in this model, and reduced recruitment of conventional T cells to the brain. Furthermore, Foxp3(+) cell-mediated protection was dependent upon CTLA-4 but not IL-10. These data show that T cell-mediated parasite tissue sequestration can be reduced by regulatory T cells in a mouse model of malaria, thereby limiting malaria-induced immune pathology.     

Goblet cell-derived resistin-like molecule beta augments CD4+ T cell production of IFN-gamma and infection-induced intestinal inflammation

The secreted goblet cell-derived protein resistin-like molecule beta (RELMbeta) has been implicated in divergent functions, including a direct effector function against parasitic helminths and a pathogenic function in promoting inflammation in models of colitis and ileitis. However, whether RELMbeta influences CD4(+) T cell responses in the intestine is unknown. Using a natural model of intestinal inflammation induced by chronic infection with gastrointestinal helminth Trichuris muris, we identify dual functions for RELMbeta in augmenting CD4(+) Th1 cell responses and promoting infection-induced intestinal inflammation. Following exposure to low-dose Trichuris, wild-type C57BL/6 mice exhibit persistent infection associated with robust IFN-gamma production and intestinal inflammation. In contrast, infected RELMbeta(-/-) mice exhibited a significantly reduced expression of parasite-specific CD4(+) T cell-derived IFN-gamma and TNF-alpha and failed to develop Trichuris-induced intestinal inflammation. In in vitro T cell differentiation assays, recombinant RELMbeta activated macrophages to express MHC class II and secrete IL-12/23p40 and enhanced their ability to mediate Ag-specific IFN-gamma expression in CD4(+) T cells. Taken together, these data suggest that goblet cell-macrophage cross-talk, mediated in part by RELMbeta, can promote adaptive CD4(+) T cell responses and chronic inflammation following intestinal helminth infection.     

Anti-IL-2 treatment impairs the expansion of T(reg) cell population during acute malaria and enhances the Th1 cell response at the chronic disease

Plasmodium chabaudi infection induces a rapid and intense splenic CD4(+) T cell response that contributes to both disease pathogenesis and the control of acute parasitemia. The subsequent development of clinical immunity to disease occurs concomitantly with the persistence of low levels of chronic parasitemia. The suppressive activity of regulatory T (T(reg)) cells has been implicated in both development of clinical immunity and parasite persistence. To evaluate whether IL-2 is required to induce and to sustain the suppressive activity of T(reg) cells in malaria, we examined in detail the effects of anti-IL-2 treatment with JES6-1 monoclonal antibody (mAb) on the splenic CD4(+) T cell response during acute and chronic P. chabaudi AS infection in C57BL/6 mice. JES6-1 treatment on days 0, 2 and 4 of infection partially inhibits the expansion of the CD4(+)CD25(+)Foxp3(+) cell population during acute malaria. Despite the concomitant secretion of IL-2 and expression of high affinity IL-2 receptor by large CD4(+) T cells, JES6-1 treatment does not impair effector CD4(+) T cell activation and IFN-γ production. However, at the chronic phase of the disease, an enhancement of cellular and humoral responses occurs in JES6-1-treated mice, with increased production of TNF-α and parasite-specific IgG2a antibodies. Furthermore, JES6-1 mAb completely blocked the in vitro proliferation of CD4(+) T cells from non-treated chronic mice, while it further increased the response of CD4(+) T cells from JES6-1-treated chronic mice. We conclude that JES6-1 treatment impairs the expansion of T(reg) cell population during early P. chabaudi malaria and enhances the Th1 cell response in the late phase of the disease.     

Despite increased CD4+Foxp3+ cells within the infection site, BALB/c IL-4 receptor-deficient mice reveal CD4+Foxp3-negative T cells as a source of IL-10 in Leishmania major susceptibility

BALB/c IL-4Ralpha(-/-) mice, despite the absence of IL-4/IL-13 signaling and potent Th2 responses, remain highly susceptible to Leishmania major substain LV39 due exclusively to residual levels of IL-10. To address the contribution of CD4(+)CD25(+) T regulatory (Treg) cells to IL-10-mediated susceptibility, we depleted CD4(+)CD25(+) cells in vivo and reconstituted IL-4Ralpha x RAG2 recipients with purified CD4(+)CD25(-) T cells. Although anti-CD25 mAb treatment significantly decreased parasite numbers in IL-4Ralpha(-/-) mice, treatment with anti-IL-10R mAb virtually eliminated L. major parasites in both footpad and dermal infection sites. In addition, IL-4Ralpha x RAG2 mice reconstituted with CD4(+) cells depleted of CD25(+) Treg cells remained highly susceptible to infection. Analysis of L. major-infected BALB/c and IL-4Ralpha(-/-) inflammatory sites revealed that the majority of IL-10 was secreted by the CD4(+)Foxp3(-) population, with a fraction of IL-10 coming from CD4(+)Foxp3(+) Treg cells. All T cell IFN-gamma production was also derived from the CD4(+)Foxp3(-) population. Nevertheless, the IL-4Ralpha(-/-)-infected ear dermis, but not draining lymph nodes, consistently displayed 1.5- to 2-fold greater percentages of CD4(+)CD25(+) and CD4(+)Foxp3(+) Treg cells compared with the BALB/c-infected dermis. Thus, CD4(+)Foxp3(-) T cells are a major source of IL-10 that disrupts IFN-gamma activity in L. major-susceptible BALB/c mice. However, the increase in CD4(+)Foxp3(+) T cells within the IL-4Ralpha(-/-) dermis implies a possible IL-10-independent role for Treg cells within the infection site, and may indicate a novel immune escape mechanism used by L. major parasites in the absence of IL-4/IL-13 signaling.