Immunosuppression during acute Trypanosoma cruzi infection: involvement of Ly6G (Gr1(+))CD11b(+ )immature myeloid suppressor cells

Trypanosoma cruzi infection is associated with a severe unresponsiveness of spleen cells (SC) to antigens and mitogens. A high production of NO by concanavalin A (Con A)-stimulated SC from infected but not from control mice was observed. Neutralization of endogenous IFN-gamma production or treatment with NO synthase (NOS) inhibitor, L-N-monomethyl-arginine, blocked Con A-induced NO production and greatly restored proliferation by SC from infected mice. This was confirmed by using IFN-gammaR(-/-) and inducible NOS (iNOS)(-/- )knockout mice, since unresponsiveness to mitogens of SC from those infected mice was much less pronounced than in control littermates. Interestingly, SC unresponsiveness was associated with a huge increase in CD11b(+) cells that express Ly-6G (Gr1)(+) and other immature myeloid markers These cells were absent in infected IFN-gammaR(-/-) spleens. Purified immature Gr1(+)CD11b(+) cells produced NO and expressed iNOS upon IFN-gamma treatment, and were able to inhibit T cell proliferation. In addition, depletion of myeloid CD11b(+ )cells abrogated NO production and restored mitogen-induced proliferation, but not IL-2 synthesis, in SC from infected mice. IL-2 production and CD25 cell surface expression by mitogen-activated T cells were greatly depressed in SC from IFN-gammaR(-/-) and iNOS(-/- )mice, confirming that Gr1(+)CD11b(+) cells were not involved in their down-regulation. In contrast, IL-5, tumor necrosis factor and IFN-gamma production, and CD69 expression by T cells were not depressed in infected SC. The results indicate the existence of an immunosuppressive mechanism during T. cruzi infection, mediated through IFN-gamma-dependent NO secretion by immature Ly-6G (Gr1)(+)CD11b(+ )myeloid cells.     

Early IFN-gamma production is related to the presence of interleukin (IL)-18 and the absence of IL-13 in experimental Trypanosoma cruzi infections

Gamma-interferon (IFN-gamma) production, the hallmark of the Th1 immune response, has been shown to play a central role in the resistance to Trypanosoma cruzi infections, in particular when produced in the very early acute infection. BALB/c mice infected with T. cruzi, Tulahuén strain, reach high parasitemias during the acute phase, and their spleen cells release IFN-gamma in the second week of the infection, while those of the resistant C3H strain produce the cytokine earlier, at 2 days post-infection (pi). We studied in the spleen cells supernatants of infected BALB/c and C3H mice, the spontaneous production of cytokines involved in the induction, interleukin (IL)-18 and IL-12 p70, as well as in the downregulation, IL-13 and IL-10, of the Th1 immune response. We found that, at 2 days pi, only C3H mice produced IL-18, while IL-12 p70 was detected in both mouse strains. Moreover, at this time pi splenocytes from BALB/c mice spontaneously produced high amounts of IL-13. At 14 days pi, despite the increased levels of IL-13 and IL-10 detected in C3H mice, they still showed high concentrations of IL-18 and IL-12 p70. In contrast, spleen cells from BALB/c mice did not secrete IL-18, IL-12 p70 and IL-13 at this time pi, but produced higher amounts of IL-10 than C3H mice. Non of these cytokines was found increased in the cell supernatants of chronically infected mice. The addition of lipopolysaccharide (LPS) or Concanavalin A (Con A) to the cell cultures did not enhance the production of IL-18 and IL-12 at the time points tested. On the other hand, at 21 days pi, when parasitemia peaked, an inhibition of both the LPS induced IL-10 release and the IL-13 production upon Con A stimulation was observed in C3H, but not in BALB/c mice. We did not find an increase of IL-18, IL-10, or IL-12 p70 in the serum of the infected mice, despite the high seric IL-12 p40 concentrations reached during the infection. The data show that the different kinetics of the production of these cytokines in the spleen of both mouse strains could have a key role in the in vivo regulation of IFN-gamma production. In these experimental models, early IFN-gamma release and thus resistance to T. cruzi infection, could be related to the combined effect of both IL-18 and IL-12p70 in the absence of IL-13.     

Effects of interleukin-4 deprivation and treatment on resistance to Trypanosoma cruzi

Trypanosoma cruzi (Y strain)-infected interleukin-4(-/-) (IL-4(-/-)) mice of strains 129/J, BALB/c, and C57BL/6 showed no significant difference in parasitemia levels or end point mortality rates compared to wild-type (WT) mice. Higher production of gamma interferon (IFN-gamma) by parasite antigen (Ag)-stimulated splenocytes was observed only for C57BL/6 IL-4(-/-) mice. Treatment of 129/J WT mice with recombinant IL-4 (rIL-4), rIL-10, anti-IL-4, and/or anti-IL-10 monoclonal antibodies (MAbs) did not modify parasitism. However, WT mice treated with rIL-4 and rIL-10 had markedly increased parasitism and suppressed IFN-gamma synthesis by spleen cells stimulated with parasite Ag, concanavalin A, or anti-CD3. Addition of anti-IL-4 MAbs to splenocyte cultures from infected WT 129/J, BALB/c, or C57BL/6 mice failed to modify IFN-gamma synthesis levels; in contrast, IL-10 neutralization increased IFN-gamma production and addition of rIL-4 and/or rIL-10 diminished IFN-gamma synthesis. We conclude that endogenous IL-4 is not a major determinant of susceptibility to Y strain T. cruzi infection but that IL-4 can, in association with IL-10, modulate IFN-gamma production and resistance.     

Studies on the role of interleukin-12 in acute murine toxoplasmosis.

Interleukin-12 (IL-12) is important in the regulation of resistance to Toxoplasma gondii in mice with severe combined immunodeficiency (SCID). The protective ability of IL-12 in SCID mice appears to be through its activity on natural killer (NK) cells to induce production of interferon-gamma (IFN-gamma). In this study we assessed the role of IL-12 in the acute stage of toxoplasmosis in immunocompetent mice. Administration of IL-12 to BALB/c mice infected with the virulent C56 strain of T. gondii remarkably delayed time to death. The protective activity of IL-12 was abrogated by administration of monoclonal antibodies to IFN-gamma or tumour necrosis factor-alpha (TNF-alpha), and by depletion of NK cells using an antisera against asialoGM1. Whereas BALB/c mice infected with the ME49 strain of T. gondii survived infection, administration of anti-IL-12 to infected mice resulted in 100% mortality accompanied by decreased serum levels of IFN-gamma. Furthermore, this treatment significantly reversed the suppression of spleen cell proliferation to concanavalin A (Con A), which is associated with the acute stage of infection, and resulted in decreased ex vivo production of IFN-gamma, IL-2, IL-4 and IL-10 in response to Con A. Our results indicate an important role for IL-12 in mediating resistance to T. gondii during acute infection in immunocompetent mice, that NK cells are required for this protective activity, and that IL-12 is involved in the immunosuppression which accompanies this infection.     

Endogenous IL-10 and IFN-γ Production Controls Thymic Cell Proliferation in Mice Acutely Infected by Trypanosoma cruzi

Thymocytes from mice with experimental Trypanosoma cruzi infection respond poorly to Con-A stimulation. However, the proliferative capacity of these cells is not impaired, as demonstrated by the fact that at high doses, exogenous rIL-2 restores thymidine uptake. This finding could be explained either by insufficient IL-2 production or by the appearance of inhibitory factors during T. cruzi infection. This paper shows that in response to Con A, IL-2 production is decreased in the model. Furthermore, the whole profile of cylokine production is modified, with a striking increase in IL-10, IFN-γ, IL-4, IL-5 and IL-6 production. The results indicate that IL-10 it plus IFN-γ are responsible for the decrease in the Con A induced proliferation since a normal proliferative response as well as normal IL-2 production can be restored if both cytokines are neutralized by adding their monoclonal antibodies (MoAbs). Evidence is provided also for an enhanced non-specific cytotoxicity of thymic cells from infected mice that might involve IL-4, IL-5 and IL-6. This is the first study demonstrating an alteration of thymic cell function by T. cruzi infection which results from overstimulation of IL-10 and IFN-γ production.     

RIL—2对急性克氏锥虫感染小鼠的抗SRBC直接溶血空斑反应及抗虫保护性免疫影响的初步研究

为研究ＩＬ—２在克氏锥虫感染过程中出现的免疫抑制机制中的作用，８只克氏锥虫感染的Ｃ３Ｈ／ＨｅＪｎ小鼠每日皮下注射两次，每日总量５０００μ重组人ｒＩＬ－２；对照组相同数量小鼠皮下注射等量的无钙、镁离子的ＰＢＳ。实验结果表明ｒＩＬ－２注射组小鼠脾细胞对羊红细胞的反应能力由ＰＢＳ注射鼠的１５。２％提高到９３。５％（Ｐ＜０．０５）。相当接近于正常的未感染小鼠的空斑形成细胞数。ｒＩＬ－２注射组小鼠仅表现轻度降低的寄生虫血症，其动态变化在两组间未显示出差异。除ＰＢＳ组的１只小鼠在感染后第３６ｄ死亡外，ｒＩＬ－２以及ＰＢＳ注射组的其它动物都在克氏锥虫急性感染期存活。尽管ｒＩＬ－２注射并未明显地提高克氏锥虫急性感染期Ｃ３Ｈ／ＨｅＪｎ小鼠的抗虫保护性免疫，本实验提示ＩＬ－２不足是克氏锥虫急性感染期Ｃ３Ｈ／ＨｅＪｎ小鼠非特异性免疫抑制的原因之一。     

Interference of natural mouse hepatitis virus infection with cytokine production and susceptibility to Trypanosoma cruzi

Mouse hepatitis virus (MHV) infection can have a pronounced impact on several investigation areas. Reports on natural MHV outbreaks are rare and most studies have been conducted by deliberately infecting mice with MHV laboratory strains that cause moderate to severe disturbances to the immune system. We have investigated the effects of a natural acute outbreak of MHV in our otherwise specific-pathogen-free (SPF) inbred mouse colonies, and of enzootic chronic MHV infection on cytokine production and resistance to the intracellular pathogen Trypanosoma cruzi. We found that BALB/c and/or C57BL/6 SPF mice that had been injected with T. cruzi blood trypomastigotes from recently MHV-contaminated (MHV+) mice developed significantly higher parasite blood counts, accelerated death, and showed higher IL-10 production by spleen cells than their counterparts whose T. cruzi inoculum was derived from MHV-negative (MHV-) donors. Interferon-gamma (IFN-gamma) production by MHV+ and MHV- mice was not significantly different. In contrast, T. cruzi infection of chronically MHV-infected mice did not result in major changes in the course of infection when compared with that observed in mice from MHV- colonies, although a trend to higher parasitaemia levels was observed in BALB/c MHV+ mice. Nevertheless, both BALB/c and C57BL/6 T. cruzi-infected MHV+ mice had diminished IFN-gamma production to parasite-antigen stimulation in comparison with similarly infected MHV- mice. Interleukin-10 (IL-10) production levels by spleen cells did not differ between chronic MHV+ and MHV- mice, but IFN-gamma neutralization by monoclonal antibody treatment of anti-CD3-stimulated spleen cell cultures showed higher levels of IL-10 synthesis in MHV+ BALB/c mice.     

IL-12 and IFN-gamma production, and NK cell activity, in acute and chronic experimental Trypanosoma cruzi infections

Resistance to acute Trypanosoma cruzi infection is mainly associated with a Th1 immune response, characterized by gamma-interferon (IFN-gamma) production and activation of macrophages. The outcome of the Th1 response in the spleen and serum of BALB/c and C3H mice infected with T. cruzi, Tulahuén strain was studied. The levels of interleukin-12 p40 (IL-12 p40) and IFN-gamma, as well as natural killer (NK) cell cytotoxicity were determined at different time-points during the acute phase, and the production of cytokines was also studied in the chronic infection. At 2 days post-infection (pi), spleen cells from C3H mice increased their NK cell activity and the ex vivo spontaneous release of both IL-12 p40 and IFN-gamma. On the other hand, BALB/c mice reached low levels of NK cell cytotoxicity and no IFN-gamma production was detected at this time pi, but the cytokine was released at high amounts in the second week of the infection. Seric IL-12 p40 concentrations showed a 3-fold increase in both mouse strains on the second day pi and remained high throughout the acute phase. However, seric IFN-gamma levels increased during the late acute infection and were higher in BALB/c than in C3H mice. In chronically infected mice IL-12 p40 was as high as in the acute phase in the serum of both strains, but only BALB/c mice still produced IFN-gamma. To the authors' knowledge this is the first report showing the protein levels of IL-12 p40 determined in vivo in acute and chronic T. cruzi infections. The results reveal differences between both mouse strains in the mechanisms controlling the onset and fate of the Th1 response triggered by the parasite and a long lasting pro-inflammatory stimuli.     

Gamma interferon suppresses acute and chronic Trypanosoma cruzi infection in cyclosporin-treated mice.

To determine if exogenous gamma interferon is effective in immunosuppressed mice infected with Trypanosoma cruzi, recombinant murine gamma interferon was administered to cyclosporin-treated mice with either acute or chronic T. cruzi infection. Gamma interferon significantly decreased parasitemia and prevented death in acutely infected mice. Parasitemias and mortality of mice treated with both gamma interferon and cyclosporin were similar to those of immunocompetent controls. In chronically infected mice, cyclosporin treatment produced significantly more organ explant cultures positive for T. cruzi. Fewer positive cultures, particularly for spleen and heart, were obtained from cyclosporin-treated mice when they also received gamma interferon. Ketoconazole treatment of mice resulted in no positive cultures. Cyclosporin treatment did not prevent activation of peritoneal macrophages by parenteral gamma interferon, nor did it have a consistent effect on serum titers of alpha/beta or gamma interferon in response to a second challenge inoculum of T. cruzi. These data indicate that exogenous gamma interferon suppresses acute and chronic T. cruzi infection in cyclosporin-treated mice but that gamma interferon is not as effective as the relatively specific antimicrobial ketoconazole. Gamma interferon activates macrophages despite cyclosporin treatment, and its effects appear to be tissue specific.     

Gene gun-mediated delivery of an interleukin-12 expression plasmid protects against infections with the intracellular protozoan parasites Leishmania major and Trypanosoma cruzi in mice

An interleukin-12 (IL-12) expression plasmid was transferred, using a gene gun, to mice infected with Leishmania major or Trypanosoma cruzi. Transfer of the IL-12 gene to susceptible BALB/c mice resulted in regression of lesion size and reduced the number of parasites in draining lymph nodes (LN) at the site of L. major infection. Coincident with these protective effects, the T-helper type (Th) response shifted towards Th1, as evaluated by cytokine production in vitro and L. major-specific antibody responses. Protective effects of the IL-12 gene were also observed in T. cruzi infection. Treatment of BALB/c mice infected with T. cruzi enhanced the production of interferon-gamma (IFN-gamma) by spleen cells, while suppressed production of interleukin-10 (IL-10) compared with control mice. Administration of anti-CD4 or anti-CD8 monoclonal antibody (mAb) abolished the protective immunity against T. cruzi infection, and treatment with the IL-12 gene could not restore the resistance in these mice. Mice depleted of natural killer (NK) cells with anti-asialo GM1 also became susceptible to infection, while the resistance was restored when these mice were treated with the IL-12 gene. Thus, target cells for the treatment appear to be CD4+ and CD8+ T cells, which are ordinarily activated by NK cells. These results suggest that the transfer of cytokine genes using a gene gun is an effective method for investigating the roles of cytokines and gene therapy in infectious diseases.     

Experimental autoimmune encephalomyelitis can be prevented and cured by infection with Trypanosoma cruzi

Trypanosoma cruzi is an intracellular parasite that induces a strong Th1-type response and immunosuppression during the acute phase of infection. To study how the infection with T. cruzi would modulate the development of an autoimmune disease, we immunized C57BL/6 mice and IL-10 or iNOS knock-out mice of the same background with the encephalitogenic MOG 35-55 peptide and infected them with T. cruzi. Our results demonstrate that infection with T. cruzi completely prevents EAE development and furthermore induces complete and lasting remission in mice that were infected with this parasite after they had developed clinical EAE. Nitric oxide and IL-10 participate in triggering the mechanisms associated with EAE suppression by the infection. Decreased lymphoproliferation and increased frequencies of Annexin-positive cells and of T cells bearing CD95, CD95L or CTLA-4 were observed in the spleen from immunized/infected mice, as well as lower IL-2 and increased TGF-beta production in comparison with only immunized mice. Our results indicate that several effector and regulatory mechanisms of the immune response that arise during the acute phase of T. cruzi infection lastingly affect the expansion and/or effector functions of encephalitogenic cells, preventing the onset or inducing complete remission of EAE.     

Interleukin-12 mediates resistance to Trypanosoma cruzi in mice and is produced by murine macrophages in response to live trypomastigotes.

Host resistance to infection by Trypanosoma cruzi is dependent on both natural and acquired immune responses. During the first week of infection in mice, NK cell-derived gamma interferon (IFN-gamma) is involved in controlling intracellular parasite replication, mainly through the induction of NO biosynthesis by activated macrophages. Interleukin-12 (IL-12) has been shown to be a powerful cytokine in inducing IFN-gamma synthesis by NK cells, as well as in mediating resistance to different intracellular protozoa. We have therefore studied the ability of T. cruzi to elicit IL-12 synthesis by macrophages and the role of this cytokine in controlling parasite replication during acute infection in mice. Our results show that macrophages cultured in the presence of live trypomastigote forms (but not epimastigotes) release IL-12 that can induce IFN-gamma production by normal spleen cells. IL-12 was detected in as little as 12 h after the addition of the trypomastigotes, and the level of IL-12 peaked at 48 h after the initial macrophage-parasite incubation. The addition of anti-IL-12 monoclonal antibody to macrophage-trypomastigote supernatants dose-dependently inhibited IFN-gamma production by naive splenocytes. Finally, the in vivo role of IL-12 in resistance to infection by T. cruzi was analyzed. Mice treated with anti-IL-12 monoclonal antibody had significantly increased parasitemia and mortality in comparison with those of control infected mice treated with control antibody. Together, these results suggest that macrophage-derived IL-12 plays a major role in controlling the parasitemia in T. cruzi-infected mice and that the animal's resistance during the acute phase of infection may, at least in part, be a consequence of postinfection levels of IL-12.     

Pivotal role of interleukin-12 and interferon-gamma axis in controlling tissue parasitism and inflammation in the heart and central nervous system during Trypanosoma cruzi infection.

The role of cytokines in the control of tissue parasitism and pathogenesis of experimental Chagas' disease was investigated. Wild-type and different cytokine as well as inducible nitric oxide synthase (iNOS) knockout mice were infected with the Colombian strain of Trypanosoma cruzi, and the kinetics of tissue parasitism, inflammatory reaction, parasitemia, and mortality were determined. We demonstrate the pivotal role of the interleukin (IL)-12/interferon (IFN)-gamma/iNOS axis and the antagonistic effect of IL-4 in controlling heart tissue parasitism, inflammation, and host resistance to acute infection with T. cruzi. Further, the heart and central nervous system were shown the main sites of reactivation of T. cruzi infection in mice lacking functional genes for IFN-gamma and IL-12, respectively. Our results also show that in contrast to IFN-gamma knockout (KO) mice, splenocytes from IL-12 KO mice infected with T. cruzi produced low levels of IFN-gamma upon stimulation with antigen. Consistently, high levels of anti-T. cruzi IgG2a antibodies were detected in the sera from IL-12 KO, but not from IFN-gamma KO mice, infected with the Colombian strain of T. cruzi. Thus, our results suggest that the level of IFN-gamma deficiency is a major determinant of the site of reactivation of T. cruzi infection in immunocompromised host.     

During Trypanosoma cruzi Infection CD1d-Restricted NK T Cells Limit Parasitemia and Augment the Antibody Response to a Glycophosphoinositol-Modified Surface Protein

Trypanosoma cruzi is a protozoan parasite that chronically infects many mammalian species and in humans causes Chagas' disease, a chronic inflammatory disease. The parasite expresses glycophosphoinositol (GPI), which potently stimulates interleukin 12 (IL-12) production. During T. cruzi infection IL-12, and possibly GPI, might stimulate NK T cells to affect the protective and chronic inflammatory responses. Here we report that during T. cruzi infection CD1d-restricted NK T cells are stimulated as NK T-cell-deficient mice have greater parasitemia. Furthermore, during T. cruzi infection the percentages of NK T cells in the liver and spleen become decreased for prolonged periods of time, and in vitro stimulation of NK T cells derived from livers of chronically infected mice, compared to uninfected mice, results in increased gamma interferon and IL-4 secretion. Moreover, in NK T-cell-deficient mice the chronic-phase antibody response to a GPI-modified surface protein is decreased. These results indicate that, during the acute infection, NK T cells limit parasitemia and that, during the chronic phase, NK T cells augment the antibody response. Thus, during T. cruzi infection the quality of an individual's NK T-cell response can affect the level of parasitemia and parasite tissue burden, the intensity of the chronic inflammatory responses, and possibly the outcome of Chagas' disease.     

Restoration of immunological responses by THF, a thymic hormone, in mice infected with murine cytomegalovirus (MCMV).

Cytomegalovirus causes T cell immune impairment in infected mice, reflected by a decreased response to the T cell mitogens PHA and Con A, and reduction of Con A-induced IL-2 secretion. Concomitantly, a marked increase in the spleen weight of infected mice was found. Histological examination of the liver revealed focal hepatitis. These signs of infection lasted from day 5 to day 14 after infection when mice slowly started to recover. Systemic treatment of MCMV-infected mice with thymic humoral factor (THF) resulted in a reconstitution of the mitogenic responses, IL-2 secretion, normalization of spleen weight and recovery of liver inflammation. Unlike other thymic hormones, THF did not affect interferon synthesis and NK cytotoxicity in infected mice. It is concluded that THF restores immune competence of mice immunosuppressed as a result of MCMV infection, through modulation of the T cell compartment.     

Interleukin-12-mediated resistance to Trypanosoma cruzi is dependent on tumor necrosis factor alpha and gamma interferon.

The aim of this study was to determine if interleukin-12 (IL-12) has a role in the immune response to Trypanosoma cruzi. Infection of BALB/c mice with the virulent Tulahuen strain of T. cruzi is characterized by a high-level parasitemia, pathology in the heart associated with the presence of amastigotes, and death during the acute phase of the disease. Administration of IL-12 to BALB/c mice infected with T. cruzi resulted in a reduced parasitemia and a significant delay in the time to death compared with those for infected controls. This protective effect was correlated with increased levels of gamma interferon (IFN-gamma) and tumor necrosis factor alpha (TNF-alpha) in serum. To determine if these cytokines were involved in the protective effects of IL-12, we treated infected mice with IL-12 alone or in combination with monoclonal antibodies specific for IFN-gamma or TNF-alpha. These antibodies antagonized the protective effect of exogenous IL-12. Treatment of infected mice with a polygonal antibody specific for IL-12 resulted in a significant increase in parasitemia but did not affect the time to death. These latter studies demonstrate a role for endogenous IL-12 in resistance to T. cruzi. Together, our data identify an IL-12-mediated mechanism of resistance to T. cruzi, which is dependent on IFN-gamma and TNF-alpha.     

Effect of hydroxyurea on the concanavalin-A proliferative response of Trypanosoma cruzi infected mice

Mice infected with Trypanosoma cruzi develop immunosuppression with a deficient production of interleukin-2 (IL-2). In this situation the deficient concanavalin A (Con A) T-cell response is not corrected by addition of exogenous IL-2. Here we show that elimination of cycling cells by treatment of infected mice with hydroxyurea (HU) fully restored the ability of spleen cells to respond to IL-2. Further, capacity for IL-2 production was restored to HU treated infected mice, but not as completely as the response to IL-2.     

Modulation of cellular immune responses in mice with disseminated histoplasmosis by recombinant interleukin-2.

Depression of the cellular immune responses in mice with disseminated histoplasmosis is associated with deficient production of interleukin-2 (IL-2) by splenocytes. Therefore, we examined whether a highly purified preparation of IL-2, recombinant human IL-2 (rIL-2), could modify the cellular immune responses in infected mice and whether this lymphokine could alter the severity of histoplasmosis in animals. Exogenous rIL-2, at concentrations of up to 1,000 U/ml, failed to augment the proliferative responses to concanavalin A by unfractionated splenocytes or splenic T cells from mice infected for 1 week. In addition, rIL-2 did not modulate the plaque-forming cell response to sheep erythrocytes by splenocytes from these same mice. However, at week 3, rIL-2 in concentrations ranging from 10 to 1,000 U/ml considerably augmented the proliferative response to concanavalin A and plaque-forming cell response to sheep erythrocytes by splenocytes from infected mice. Kinetics studies demonstrated that rIL-2 exerted maximal immunoregulatory activity when added on day 0 or 1 to cultures of splenocytes. In vivo administration of rIL-2, 200 to 20,000 U/day, for 10 days to normal and 3-week-infected mice did not alter the proliferative activity of splenocytes to concanavalin A; 200,000 U of rIL-2 per day actually depressed the proliferative responses of splenocytes from normal and infected mice. In vivo, rIL-2 did not modify delayed-type hypersensitivity responses to sheep erythrocytes or to histoplasmin by normal and infected mice. Moreover, treatment with rIL-2 in vivo did not reduce the number of Histoplasma CFU in spleens of mice. Thus, despite the immunoenhancing effect of rIL-2 in vitro, this lymphokine failed to exert similar effects in vivo.