Effects of cytokines and prolactin on the replication of Toxoplasma gondii in murine microglia

In the central nervous system, cytokine-activated microglia play a crucial role in host defence against Toxoplasma gondii infections. In this study, the effect of recombinant tumor necrosis factor (rTNF)-alpha and prolactin (PRL) on T. gondii infection in microglia was examined. Pretreatment of microglia with rTNF-alpha and PRL induced toxoplasmastatic activity, the intracellular killing of T. gondii and the release of interleukin (IL)-1 beta IL-3 and IL-6: 50% of the intracellular killing was abrogated by anti-ICAM-1 monoclonal antibodies, whereas more than 54 or 87% of toxoplasmastatic activity was reversed by anti-IL-3 or IL-6 monoclonal antibodies. In addition, the treatment of microglia with either rIL-3 or rIL-6, in the absence or presence of rTNF-alpha significantly limited T. gondii replication. Inasmuch as either NMA or S-M-ITU affected cytokine-activated toxoplasmastatic activity during the infection phase, the NO-dependent pathway itself appears not to be directly involved in the parasitostatic activity. These findings suggest that TNF-alpha and PRL up-regulate the expression of ICAM-1 and the production of endogenous IL-6 and IL-3 by microglia, which could induce anti-parasitic functions against T. gondii infection in the brain.     

Interactions between cytokines and growth hormone for resistance to experimental

Cytokine-activated human vein endothelial cells (HUVEC) may play an important role in resistance to Toxoplasma gondii infection. In this study, it was investigated the role of rTNF-alpha and GH in the induction of antitoxoplasmal activities in HUVEC. Co-treatment of HUVEC with rTNF-alpha plus GH induced both toxoplasmastatic activity and the intracellular killing of T. gondii (p <0.01 each vs untreated cells). Thus, these functions were inhibited by both neutralizing antibodies to IL-6 and GM-CSF (but not to IL-3) suggesting that these cytokines participate in the inhibitory process. Consistent with this hypothesis, the treatment of HUVEC with rIL-6 or rGM-CSF in the presence of rTNF-alpha, limited T. gondii multiplication in a dose-dependent manner (p <0.01 each vs untreated cells). In order to elucidate the inhibitory mechanism of HUVEC, it was assessed by L-arginine analogs (e.g., NG-monomethyl-arginine) whether NO2 molecules originating from HUVEC were directly or indirectly involved in the rTNF-alpha/GH-dependent induction of toxoplasmastatic activity. A good correlation was found between toxoplasmastatic activity and NO2 release during the activation phase, before infection of the HUVEC with T. gondii, but no correlation was found between the parasitostatic activity and NO2 release during the infection phase. These data indicate that NO2- itself does not directly affect toxoplasmastatic activity. Besides, the reduction of intracellular killing by monoclonal antibodies to ICAM-1 suggest that this adhesin plays a role in controlling T. gondii entry into cells.     

Anti-proliferative effect of IFN-gamma in immune regulation. II. IFN-gamma inhibits the proliferation of murine bone marrow cells stimulated with IL-3, IL-4, or granulocyte-macrophage colony-stimulating factor

A biphasic dose response curve was observed when the bone marrow-derived cell line FDCP1, used as an indicator line for IL-3 bioassays, was exposed to supernatants from some activated T cell clones but not others. The active component which inhibited proliferation at the higher supernatant concentrations appeared to be IFN-gamma, based on the following observations. 1) Only those culture supernatants which contained IFN-gamma gave a biphasic dose response curve; 2) with these supernatants, an anti-IFN-gamma mAb augmented the proliferation of FDCP1 cells at the higher supernatant concentrations; and 3) rIFN-gamma profoundly inhibited the proliferation of FDCP1 cells stimulated with rIL-3 or rIL-4. rTNF-alpha inhibited FDCP1 proliferation only to a modest extent, yet the combination of rTNF-alpha + rIFN-gamma provided greater inhibition than each agent alone. The proliferation of a second bone marrow-derived cell line, DA1, was not inhibited by rIFN-gamma or rIFN-gamma + rTNF-alpha when stimulated with rIL-3 or recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF). Fresh bone marrow cells also showed a suboptimal proliferative response when stimulated with T cell supernatants containing IFN-gamma, and this response was augmented considerably upon the addition of anti-IFN-gamma mAb. Bone marrow cell proliferation was observed upon exposure to rIL-3, rIL-4, or rGM-CSF, and these responses were inhibited by rIFN-gamma; rTNF-alpha also produced a synergistic effect with these cells. Bone marrow cell colony formation stimulated by rIL-3 or rGM-CSF also was inhibited by rIFN-gamma. Colony formation in bone marrow cell cultures was not observed in response to rIL-4. Collectively, these results suggest that Th1 cells, which in addition to IL-3 and GM-CSF also produce IFN-gamma, may regulate hemopoietic cell proliferation and colony formation differently from the way Th2 cells do, which do not produce IFN-gamma.     

Complex network of cytokines activating murine microglial cell activity against Acanthamoeba castellani

In the central nervous system, cytokine-primed microglia play a pivotal role in host defence against Acanthamoeba castellani infections. In this study, the effect of rIL-1beta, rIL-6 or rTNF-alpha, combined or not with rIFN-gamma, on A. castellani infection of murine microglia was examined. Priming of microglial cells with either rIL-1beta or rIL-6, in the presence or absence of rIFN-gamma, triggered amebastatic activity, while the treatment of microglia with rTNF-alpha plus rIFN-gamma additively triggered, in a dose-dependent fashion, amebicidal activity. Inasmuch as NGMA affected cytokine-triggered anti-parasitic activity during the priming process, the NO-dependent pathway itself appears not to be directly involved in the anti-amebic capacities. These data suggest that the proinflammatory cytokines IL-1beta, IL-6 or TNF-alpha could trigger anti-microbial activity against A. castellani infection in the brain.     

Cytokine regulation of interleukin 6 production by human endothelial cells

The influence of recombinant (r) human tumor necrosis factor alpha (rTNF-alpha), r human interleukin 1 beta (rIL-1 beta), and r human interferon gamma (rIFN-gamma) on the production of interleukin 6 (IL-6) by human endothelial cells (HEC) was investigated. The addition of 1-100 U/ml of either rTNF-alpha or rIL-1 beta to cultures of HEC monolayers caused a dose-related increase in IL-6 production as detected after 24 hr of incubation. In contrast to rIL-1 beta and rTNF-alpha, the use of up to 1000 U/ml of rIFN-gamma caused only a moderate increase in IL-6 production. However, significantly greater quantities of IL-6 were produced by HEC monolayers subjected to 1000 U/ml of rIFN-gamma in combination with 1-100 U/ml of rTNF-alpha. Furthermore, the addition of graded concentrations of human transforming growth factor beta (TGF-beta) to cultures resulted in a dose-related inhibition of rIL-1 beta- and rTNF-alpha-induced IL-6 production by HEC. The results demonstrate that rIL-1 beta and rTNF-alpha share the ability to stimulate HEC for production of IL-6 and indicate that TGF-beta may act as an immunosuppressive agent, at least partially, through its ability to inhibit the action of TNF-alpha and IL-1 on endothelial cells.     

Human endothelial cells are activated by IFN-gamma to inhibit Toxoplasma gondii replication. Inhibition is due to a different mechanism from that existing in mouse macrophages and human fibroblasts

Toxoplasma gondii invaded and proliferated in cultured human umbilical vein endothelial cells. Preincubation of the human umbilical vein endothelial cells with human rIFN-gamma induced a high degree of inhibition of T. gondii replication, with the effect being dose dependent. In order to try to elucidate the inhibitory mechanism, we tested the presence of several factors that are known to operate against intracellular parasites in other cell types. We found, by means of a competitive inhibitor, that L-arginine-dependent production of reactive nitrogen intermediates was not the cause of inhibition of T. gondii proliferation, thus contrasting with the inhibitory mechanism found in activated mouse macrophages. Furthermore, the inhibition of replication was not overcome by oxygen scavengers or by saturation of the system with tryptophan, suggesting that neither reactive oxygen intermediates nor the induction of tryptophan starvation was responsible.     

Enhancement of human B cell proliferation and differentiation by tumor necrosis factor-alpha and interleukin 1

The role of tumor necrosis factor-alpha (TNF-alpha) in human B cell responses was examined and compared with that of interleukin (IL) 1 by assessing the ability of each cytokine to support proliferation and differentiation. Recombinant TNF-alpha (rTNF-alpha) and recombinant IL-1 (rIL-1) each enhanced the generation of immunoglobulin-secreting cells (ISC) in cultures of pokeweed mitogen-stimulated B cells incubated with T cells. To examine the direct effect of rTNF-alpha and rIL-1 on the responding B cell, highly purified peripheral blood B cells were stimulated with Cowan I Staphylococcus aureus (SA). In the absence of T cell factors, proliferation was minimal and there was no generation of ISC. Recombinant IL-2 (rIL-2) supported both responses. Although rTNF-alpha alone did not support SA-stimulated generation of ISC, it did increase SA-stimulated B cell DNA synthesis by two- to eightfold. In addition, rTNF-alpha augmented B cell proliferation in rIL-2 supported SA-stimulated cultures. Moreover, rTNF-alpha enhanced the generation of ISC stimulated by rIL-2 alone or rIL-2 and SA. rIL-1 also augmented DNA synthesis and generation of ISC by B cells stimulated with SA and rIL-2. However, rTNF-alpha enhanced proliferation and ISC generation in SA + rIL-2-stimulated cultures even when they were supplemented with saturating concentrations of rIL-1. Utilizing a two-stage culture system, it was found that the major effect of rTNF-alpha was to enhance responsiveness of SA-activated B cells to rIL-2, whereas it exerted only a minimal effect during initial stimulation. These results indicate that TNF-alpha as well as IL-1 augment B cell responsiveness. Moreover, the ability of rTNF-alpha to enhance B cell responsiveness was not an indirect effect resulting from the induction of Il-1 production by contaminating monocytes, but rather resulted from the delivery of a signal by rTNF-alpha directly to the responding B cell that promoted both proliferation and differentiation after initial activation. The data therefore indicate that human B cell responsiveness can be independently regulated by the action of two separate monocyte-derived cytokines.     

Tumor necrosis factor-alpha-dependent production of reactive nitrogen intermediates mediates IFN-gamma plus IL-2-induced murine macrophage tumoricidal activity.

We have previously established that IFN-gamma plus IL-2 induces murine macrophage tumoricidal activity. The purpose of this study was to identify the effector molecules that account for the IFN-gamma plus IL-2-induced macrophage cytotoxicity against P815 mastocytoma cells. ANA-1 macrophages and normal thioglycollate-elicited mouse peritoneal macrophages produced little or no detectable nitrite (NO2-) after incubation with IFN-gamma alone or IL-2 alone; however, IL-2 synergized with IFN-gamma for the production of NO2-. IFN-gamma plus IL-2 did not induce NO2- production or tumoricidal activity in ANA-1 macrophages that were cultured in medium devoid of L-arginine or in ANA-1 macrophages that were incubated with NG-monomethyl-L-arginine. As observed previously with ANA-1 macrophage tumoricidal activity, IL-4 inhibited IFN-gamma plus IL-2-induced, but not IFN-gamma plus LPS-induced, NO2- production. IL-4 also selectively decreased the ability of IFN-gamma and/or IL-2 to augment TNF-alpha mRNA expression in ANA-1 macrophages. Lastly, incubation of ANA-1 macrophages with anti-TNF mAb selectively inhibited the ability of IFN-gamma plus IL-2 to induce NO2- production and tumoricidal activity. These results indicate that IFN-gamma plus IL-2-induced tumoricidal activity is dependent upon the metabolism of L-arginine to reactive nitrogen intermediates, and they establish a role for TNF-alpha as a required intermediate for IL-2-dependent NO2- production and tumoricidal activity.     

Human mononuclear phagocyte antiprotozoal mechanisms: oxygen-dependent vs oxygen-independent activity against intracellular Toxoplasma gondii

To determine if the oxygen-dependent and -independent antiprotozoal mechanisms with which the human mononuclear phagocyte is equipped to act against Leishmania donovani operate against other intracellular parasites, oxidatively intact and deficient cells were challenged with Toxoplasma gondii. Fresh monocytes and lymphokine- or gamma-interferon (IFN-gamma)-activated macrophages from normal individuals killed 35% and 50% of T. gondii within 6 hr, respectively, and each of these cell populations inhibited the replication of surviving parasites 20 hr after infection. This activity was associated with the capacity to release large amounts of H2O2 (572 to 971 nmol/mg) and to respond to toxoplasma ingestion with respiratory burst activity. Impairing the ability to generate oxygen intermediates by glucose deprivation or treatment with superoxide dismutase, catalase, or mannitol inhibited toxoplasmacidal activity by greater than 80% and permitted a 2.6- to 4.3-fold increase in the number of intracellular toxoplasmas. In contrast to normal cells, fresh monocytes from patients with chronic granulomatous disease (CGD) killed less than 8% of toxoplasmas and exerted 50% less toxoplasmastatic activity. However, although associated with the induction of only modest toxoplasmacidal effects (18 to 20% killing), lymphokine stimulation did induce CGD monocytes and macrophages as well as oxidatively inactive human endothelial cells to display near normal levels of toxoplasmastatic activity. Similar to oxygen-dependent mechanisms, the enhancement of oxygen-independent activity by crude lymphokines could be abolished by a monoclonal anti-IFN-gamma antibody and could be achieved by treatment with recombinant IFN-gamma alone. Unstimulated CGD monocytes, however, were found to lose all antitoxoplasma activity after two days in culture, whereas normal cells continued to effectively inhibit T. gondii replication, suggesting that oxygen-independent responses may not actually be required for the normal monocyte to act against T. gondii. Taken together with previous findings with L. donovani, these results indicate that the human mononuclear phagocyte possesses an oxygen-independent antiprotozoal mechanism and that its effects can be enhanced by lymphokines (IFN-gamma), but that nevertheless this cell's primary response to intracellular protozoa is largely oxygen dependent.     

Tumor necrosis factor-alpha synergizes with IFN-gamma in mediating killing of Leishmania major through the induction of nitric oxide

CBA mice develop cutaneous lesions when infected with Leishmania major. The disease development was significantly reduced by injecting into the lesion a combination of rIFN-gamma and rTNF-alpha. The doses of IFN-gamma and TNF-alpha used were suboptimal in that either cytokine alone did not have any effect. The therapeutic effect of IFN-gamma and TNF-alpha in vivo is reflected in their ability to activate macrophages to kill the intracellular parasites in vitro. The macrophage leishmanicidal activity induced by TNF-alpha and IFN-gamma can be completely inhibited by a specific inhibitor (L-NG monomethyl arginine) of nitric oxide synthesis. There was a direct correlation between the intracellular killing of the parasites and the production of nitric oxide by the macrophages. In contrast, there was no correlation between leishmanicidal activity and superoxide production by macrophages.     

Tumor necrosis factor-alpha triggers antitoxoplasmal activity of IFN-gamma primed macrophages

IFN-gamma is an important mediator of cellular resistance against microbial pathogens and tumor cells due in part to its potent capacity to activate macrophages for enhanced cytotoxicity. The present study demonstrates that TNF-alpha regulates the expression of enhanced antimicrobial activity by triggering IFN-gamma primed macrophages to kill or inhibit intracellular Toxoplasma gondii. Resident mouse macrophages stimulated with rIFN-gamma at levels up to 2500 U/ml failed to display enhanced antitoxoplasmal activity when cultured in vitro under low endotoxin conditions (less than 10 pg/ml), but were triggered by addition of small amounts of LPS (0.1 ng/ml). A similar requirement for LPS as a second signal necessary to trigger antitoxoplasmal activity was observed when IFN-gamma was administered to mice in vivo. The essential nature of this triggering step allowed us to explore the role of cytokines that act as endogenous regulators of macrophage activation. rTNF-alpha, although unable to confer antitoxoplasmal activity when used alone to treat macrophages, was capable of triggering IFN-gamma-primed macrophages cultured under low endotoxin conditions. The ability of TNF-alpha to trigger IFN-gamma-primed macrophages was blocked by rabbit anti-TNF-alpha polyclonal antisera but was not affected by polymyxin B indicating that TNF-alpha triggering was not due to contamination with LPS. Collectively, these findings demonstrate that TNF-alpha performs an important regulatory role in the expression of enhanced anti-microbial activity by IFN-gamma-primed macrophages.     

Cytotoxicity of lymphokine activated peritoneal macrophages against Trichomonas vaginalis]

Trichomonas vaginalis is a parasitic flagellate in the urogenital tract of human. Innate cytotoxicity of macrophages against T. vaginalis has been recognized, but any report on the cytotoxicity of lymphokine-activated macrophages to T. vaginalis is not yet available. The present study aimed to elucidate the lymphokine-activated cell mediated cytotoxic effect against T. vaginalis by mouse peritoneal macrophages. Cytotoxicity was measured by counting the release of 3H-thymidine from prelabeled protozoa, and tested in U-bottom microtiter plates. Nitrite concentration in culture supernatants was measured by standard Griess reaction. The results obtained are as follows: 1. The cytotoxicity of macrophages was increased by addition of rIL-2 or rIFN-gamma. 2. Cytotoxicity of macrophages was reduced by addition of rIL-4 to rGM-CSF, rIL-2 or rIFN-gamma. 3. Crude lymphokine mixed with anti-IL-2 decreased the cytotoxicity of macrophages. 4. In case of macrophages cultured with rIFN-gamma or rIL-4, the concentration of nitrite was related with cytotoxicity of macrophages against T. vaginalis, but the cytotoxicity of macrophages cultured with rIL-2 and rIFN-gamma was decreased in spite of its high production of nitrite. From the results obtained, it is assumed that rIL-2 and rIFN-gamma enhance the cytotoxicity of macrophages while rIL-4 inhibits the cytotoxicity against T. vaginalis, and that the production of nitrite does not relate with the cytotoxicity of macrophages, but nitric oxide may play a role as an inhibitory factor on the proliferation of T. vaginalis.     

A regulatory effect of the balance between TNF-alpha and IL-6 in the granulomatous and inflammatory response to Rhodococcus aurantiacus infection in mice

After i.v. inoculation with Rhodococcus aurantiacus, wild-type (WT) mice develop nonnecrotic, epithelioid granulomas. Because a high level of TNF-alpha is observed during the initial phase postinfection, we examined the extent to which TNF-alpha contributes to granulomatous inflammation using TNF-alpha gene-deficient (TNF-alpha(-/-)) mice. Despite a lack of R. aurantiacus proliferation, TNF-alpha(-/-) mice displayed high mortality rates within 5 days postinfection, as well as a high level of IL-6 in their spleens. Histological examination showed an absence of granuloma formation in TNF-alpha(-/-) mice. Pretreatment of TNF-alpha(-/-) mice with rTNF-alpha failed to restore this granuloma formation but accelerated bacterial removal and cellular recruitment. This rTNF-alpha administration also attenuated IL-6 production, resulting in increased survival rates of TNF-alpha(-/-) mice. Heat-killed R. aurantiacus induced in vitro enhanced mRNA expression and production of IL-6 in macrophages and DCs from TNF-alpha(-/-) mice when compared with WT controls, and treatment of TNF-alpha(-/-) mouse cells with rTNF-alpha decreased the IL-6 secretion. Moreover, anti-TNF-alpha or anti-IL-6 treatment increased IL-6 or TNF-alpha production by WT mouse cells, respectively. These data suggest that the production of TNF-alpha and IL-6 can be negatively regulated by each other. Administration of rIFN-gamma to TNF-alpha(-/-) mice caused immature granulomas in livers, and treatment with both rTNF-alpha and rIFN-gamma led to the formation of mature granulomas. Overall, TNF-alpha appears crucial for bacterial clearance, cellular recruitment, and granuloma formation. The balance between TNF-alpha and IL-6 during the early phase of infection controls the development of the inflammatory response to R. aurantiacus infection.     

Lung-specific delivery of cytokines induces sustained pulmonary and systemic immunomodulation in rats

The recombinant cytokines IFN-gamma and TNF-alpha stimulate several macrophage-mediated functions important in host defense. However, systemic administration of cytokines may be limited by significant host toxicity. We investigated whether aerosolized cytokines can stimulate alveolar macrophage and blood monocyte function, and whether they induce an inflammatory response in the lungs of normal rats. We found that aerosolized murine rIFN-gamma or recombinant human TNF-alpha increased IL-1 production by both alveolar macrophages and blood monocytes for at least 5 days after administration. Furthermore, murine rIFN-gamma increased the expression of Ia Ag on alveolar macrophages and human rTNF-alpha increased alveolar macrophage- and blood monocyte-mediated tumor lysis. Sequential aerosolization of IFN-gamma and TNF-alpha significantly increased both IL-1 release and Ia expression compared to either cytokine administered alone. Aerosolized human rTNF-alpha achieved lung levels comparable to those produced by an i.v. TNF-alpha dose reported to cause diffuse organ injury and death in rats. However, plasma TNF-alpha levels were several thousand-fold lower after aerosol administration. Aerosolized cytokines did not induce lung edema or an inflammatory cell infiltrate within the airways or alveoli. Aerosolized human rTNF-alpha alone, or murine rIFN-gamma and human rTNF-alpha, induced margination of leukocytes in pulmonary blood vessels 1 day after aerosolization, and a few small foci of pulmonary hemorrhage 5 days later. We conclude that aerosol administration of IFN-gamma or TNF-alpha enhances both pulmonary and systemic monocyte function, and that the combination of IFN-gamma and TNF-alpha produce additive or synergistic effects. Aerosolized cytokines induce only a minimal pulmonary inflammatory response. Aerosolized TNF-alpha produces high cytokine levels in the lung but very low uptake into the circulation.     

CD154 activates macrophage antimicrobial activity in the absence of IFN-gamma through a TNF-alpha-dependent mechanism

Protection against certain intracellular pathogens can take place in the absence of IFN-gamma through mechanisms dependent on TNF-alpha. In this regard, patients with partial defect in IFN-gamma receptor 1 are not susceptible to toxoplasmosis. Thus, we used a model of Toxoplasma gondii infection to investigate whether CD154 modulates IFN-gamma-independent mechanisms of host protection. Human monocyte-derived macrophages treated with recombinant CD154 exhibited increased anti-T. gondii activity. The number of tachyzoites per 100 macrophages at 20 h postinfection was lower in CD154-treated macrophages compared with controls. This was accompanied by a decrease in the percentage of infected cells in CD154-treated macrophages at 20 h compared with 1 h postinfection. CD154-bearing cells also induced antimicrobial activity in T. gondii-infected macrophages. CD154 enhanced macrophage anti-T. gondii activity independently of IFN-gamma. TNF-alpha mediated the effects of CD154 on macrophage anti-T. gondii activity. CD154 increased TNF-alpha production by T. gondii-infected macrophages, and neutralization of TNF-alpha inhibited the effect of CD154 on macrophage anti-T. gondii activity. These results demonstrate that CD154 triggers TNF-alpha-dependent antimicrobial activity in macrophages and suggest that CD154 regulates the mechanisms of host protection that take place when IFN-gamma signaling is deficient.     

Tumor necrosis factor-alpha as a proliferative signal for an IL-2-dependent T cell line: strict species specificity of action

The ability of TNF-alpha to stimulate T cell proliferation was examined. We demonstrate that murine rTNF-alpha induces the proliferation of CT6, a murine T cell line previously thought to be responsive only to IL-2. This activity appears to be the result of the direct action of murine rTNF-alpha on the CT6 cells because neither 1) murine IL-2 or murine IL-4, lymphokines also capable of inducing CT6 proliferation, were detected in culture supernatants from murine rTNF-alpha-treated CT6 cells nor 2) did antibodies specific for IL-2 or IL-4 inhibit murine rTNF-alpha-induced CT6 proliferation. Unlike many of the activities displayed by TNF-alpha, its ability to induce CT6 cell proliferation shows strict species specificity as indicated by the failure of human rTNF-alpha to stimulate these cells. Flow cytometric analysis and binding of radiolabeled TNF-alpha have indicated that receptors for TNF-alpha on these cells are specific for murine TNF-alpha. The ability of murine rTNF-alpha to induce the proliferation of certain T cell lines further indicates that this molecule plays an important role in regulation of T cell-mediated immune responses.     

Tumor necrosis factor-independent protective effect of recombinant IFN-gamma against acute toxoplasmosis in T cell-deficient mice

rIFN-gamma conferred remarkable resistance against acute infection with Toxoplasma gondii in T cell-deficient (athymic nude) mice. Mice that received an i.p. injection of rIFN-gamma every other day beginning 24 h before infection for a total of eight doses survived significantly longer than untreated control mice although all of the treated mice died after the lymphokine was discontinued. Mice that received 14 doses of rIFN-gamma survived significantly longer than those that received eight doses of the lymphokine although mice started dying soon after the final (14th) injection of rIFN-gamma and eventually all of the treated mice died. Histologic study revealed that the IFN-gamma treatment prevented proliferation of the organisms in all organs examined, including brain, lung, heart, liver, and spleen. The treatment was effective even when started 1 day after infection. Peritoneal macrophages obtained from mice injected with rIFN-gamma were activated and effectively killed tachyzoites of T. gondii in vitro. TNF activity could not be detected in sera of the infected mice during treatment with rIFN-gamma. Administration of anti-TNF antibody did not affect the protective effect of rIFN-gamma against T. gondii infection. These facts indicate that rIFN-gamma can confer resistance to acute infection with T. gondii without collaboration of lymphokines derived from T cells and TNF. This suggests that rIFN-gamma may be effective for therapy of toxoplasmosis in immunosuppressed patients who have impaired activity of T cell function, especially those with AIDS.