Both the Fas ligand and inducible nitric oxide synthase are needed for control of parasite replication within lesions in mice infected with Leishmania major whereas the contribution of tumor necrosis factor is minimal

Following infection with the protozoan parasite Leishmania major, C57BL/6 mice develop a small lesion that heals spontaneously. Resistance to infection is associated with the development of CD4(+) Th1 cells producing gamma interferon (IFN-gamma) and tumor necrosis factor (TNF), which synergize in activating macrophages to their microbicidal state. We show here that C57BL/6 mice lacking both TNF and Fas ligand (FasL) (gld TNF(-/-) mice) infected with L. major neither resolved their lesions nor controlled Leishmania replication despite the development of a strong Th1 response. Comparable inducible nitric oxide synthase (iNOS) activities were detected in lesions of TNF(-/-), gld TNF(-/-), and gld mice, but only gld and gld TNF(-/-) mice failed to control parasite replication. Parasite numbers were high in gld mice and even more elevated in gld TNF(-/-) mice, suggesting that, in addition to iNOS, the Fas/FasL pathway is required for successful control of parasite replication and that TNF contributes only a small part to this process. Furthermore, FasL was shown to synergize with IFN-gamma for the induction of leishmanicidal activity within macrophages infected with L. major in vitro. Interestingly, TNF(-/-) mice maintained large lesion size throughout infection, despite being able to largely control parasite numbers. Thus, IFN-gamma, FasL, and iNOS appear to be essential for the complete control of parasite replication, while the contribution of TNF is more important in controlling inflammation at the site of parasite inoculation.     

Activated T cells induce macrophages to produce NO and control Leishmania major in the absence of tumor necrosis factor receptor p55

The ability to activate macrophages in vitro for nitric oxide production and killing of Leishmania major parasites is dependent on tumor necrosis factor, although L. major-infected mice lacking the TNF receptor p55 (TNFRp55(-/-) mice) or both the TNFRp55 and TNFRp75 (TNFRp55p75(-/-) mice) are able to produce NO in vivo and eliminate the parasites. Here we report that activated T cells cocultured with macrophages results in TNFR-independent activation sufficient to control parasites and that both CD40/CD40L and LFA-1 contribute to T-cell-mediated macrophage activation. Thus, anti-CD3-stimulated T cells activated TNFR-deficient macrophages, while T cells from CD40L(-/-) mice were partially defective in triggering NO production by TNFRp55p75(-/-) macrophages. Moreover, in the presence of gamma interferon, anti-CD40 monoclonal antibody (MAb) activated TNFR-deficient macrophages. Finally, MAb blockade of LFA-1 completely inhibited macrophage NO production. Our data indicate that T cells can activate macrophages in the absence of TNF, thus providing a mechanism for how TNFR-deficient mice can control intracellular pathogens.     

Patterns of cytokine secretion in murine leishmaniasis: correlation with disease progression or resolution.

Susceptibility or resistance to infection with Leishmania major correlates with the ability of mice to produce characteristic panels of lymphokines in response to the parasite. To investigate the role of antigen-presenting cells in this phenomenon, we developed a model system which used congenic (H-2d) susceptible and resistant mice. L. major-specific T cells were isolated from infected BALB/c and B10.D2 mice, and the cells were restimulated in vitro on syngenic or congenic antigen-presenting cells. BALB/c L. major-reactive T cells restimulated with either antigen-presenting cell produced high levels of interleukin-4 and low levels of gamma interferon. In contrast, T cells from B10.D2 mice produced gamma interferon. Radiation-induced chimeras reconstituted with BALB/c bone marrow also produced more interleukin-4 in response to L. major than did chimeras reconstituted with B10.D2 bone marrow. To test whether this pattern of cytokine secretion was unique to infection with L. major, we infected the mice with a second intracellular pathogen, Mycobacterium bovis BCG. Mycobacterium-specific T cells from both BALB/c and B10.D2 mice produced interleukin-2 and no interleukin-4. Finally, when BALB/c mice were vaccinated with avirulent L. major, the induced resistance correlated with reduced production of interleukin-4 but no increase in gamma interferon production. Instead, T cells from the vaccinated mice produced high levels of tumor necrosis factor. This suggests that tumor necrosis factor, in addition to gamma interferon, may be involved in resistance to L. major and that interleukin-4 may inhibit the leishmanicidal activity of tumor necrosis factor and/or gamma interferon.     

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.     

Regulation of gamma interferon production by natural killer cells in scid mice: roles of tumor necrosis factor and bacterial stimuli.

CB-17 scid mice exhibit a T-cell-independent but gamma interferon (IFN-gamma)-dependent immunity to Listeria monocytogenes. In this study, we analyzed the specific cellular interactions involved in this process. scid mouse-derived natural killer (NK) cells cultured with heat-killed (HK) L. monocytogenes and macrophages secreted IFN-gamma. No IFN-gamma was produced in cultures containing HK L. monocytogenes but lacking macrophages. However, medium derived from macrophages incubated with HK L. monocytogenes or other microorganisms stimulated IFN-gamma production by isolated NK cells. Treatment of macrophage-conditioned supernatants with neutralizing monoclonal anti-tumor necrosis factor (TNF) significantly reduced their capacity to stimulate NK cells to produce IFN-gamma. Yet, purified recombinant TNF-alpha by itself was unable to stimulate NK cells. Thus, TNF was necessary but not sufficient to induce maximal IFN-gamma production by NK cells. Sonicated L. monocytogenes stimulated production of IFN-gamma by NK cells that was resistant to anti-TNF. Stimulation was markedly enhanced by the addition of recombinant TNF-alpha. These studies demonstrated that activation of scid NK cells for secretion of IFN-gamma requires two signals: TNF-alpha and a second product which may be of bacterial origin and may require processing by mononuclear phagocytes. We suggest that the T-cell-independent production of IFN-gamma by NK cells provides the host with a rapid mechanism to temporarily heighten nonspecific resistance to infection until such time as T-cell-dependent sterilizing immune responses can be generated.     

Induction of tumor necrosis factor by Legionella pneumophila.

Mice were inoculated with Legionella pneumophila via an intratracheal route to establish an experimental model of infection. Lung lavage fluid obtained from infected mice contained a cytolytic factor identified as tumor necrosis factor (TNF). Peak levels of TNF were produced at about 24 h postinfection and rapidly declined thereafter. Treatment of the mice with dextran sulfate before inoculation with the bacteria resulted in lowered amounts of TNF in the lung lavage fluid, suggesting that macrophages were responsible for production of the cytokine. Furthermore, cultures of adherent lung leukocytes and a macrophage cell line, PU 5-1.8, were stimulated to produce TNF by exposure to Legionella antigens. In addition, adherent lung leukocytes from Legionella-infected mice spontaneously released TNF into the culture supernatant. Inoculation of mice with saline or latex particles failed to induce TNF in vivo, indicating that bacterial antigens or products were the stimulating signals. Since there was no detectable TNF activity in sera at any time after intratracheal inoculation, TNF production appeared to be confined to the site of infection. Pretreatment of PU 5-1.8 cultures with gamma interferon, which was detected in the lung lavage fluid before TNF, resulted in augmented TNF production, suggesting cooperativity may exist between the two cytokines, either in the pathogenicity of the bacterium or in a possible immunomodulatory function of TNF and interferon during infection.     

Cytokine secretion induced by superantigens in peripheral blood mononuclear cells, lamina propria lymphocytes, and intraepithelial lymphocytes.

Superantigens are potent inducers of T-cell proliferation and induce a broad range of cytokines, including tumor necrosis factor (TNF), gamma interferon, and interleukin 2 (IL-2). In the present study, we compared the abilities of different staphylococcal superantigens (staphylococcal enterotoxin B [SEB], staphylococcal enterotoxin E [SEE], and toxic shock syndrome toxin 1 [TSST-1]) to stimulate distinct cytokine profiles in peripheral blood mononuclear cells (PBMC), lamina propria lymphocytes (LPL), and intraepithelial lymphocytes (IEL). One million PBMC, LPL, and IEL were stimulated with various concentrations of superantigen (10 to 0.001 ng/ml) for 24, 48, and 72 h. Maximum cytokine production by PBMC, LPL, and IEL was observed for all three superantigens at 48 h at a concentration of 1 ng/ml. In PBMC, SEE and TSST-1 stimulated more IL-2 and gamma interferon than SEB. SEE and TSST-1 also stimulated more TNF and IL-4 production than SEB. In contrast, SEB stimulated more IL-6 than either SEE or TSST-1. In LPL, there was no SEE-induced IL-2 or IL-4 production, but IL-6, TNF, and gamma interferon were induced. SEB similarly induced no IL-2 or gamma interferon from the LPL, but IL-4, IL-6, and TNF were detected. TSST-1 stimulation of LPL resulted in IL-2 and TNF production but no IL-4, IL-6, or gamma interferon. In IEL, SEE induced no IL-2, IL-4, or gamma interferon but produced IL-6 and TNF, while SEB stimulation resulted in no IL-2 or gamma interferon but did result in detectable IL-4, IL-6, and TNF. Taken together, these data indicate that there are significant differences in the cytokine profiles induced by superantigens in LPL and IEL compared with those in PBMC, and these differences may relate to differences in activation requirements.     

Effective clearance of intracellular Leishmania major in vivo requires Pten in macrophages

Leishmaniases are a major international public health problem, and macrophages are crucial for host resistance to this parasite. To determine if phosphatase and tensin homologue deleted on chromosome ten (Pten), a negative regulator of the PI3K pathway, plays a role in macrophage-mediated resistance to Leishmania, we generated C57BL/6 mice lacking Pten specifically in macrophages (LysMCrePten(flox/flox) mice). Examination of lesions resulting from Leishmania major infection showed that LysMCrePten(flox/flox) mice were more susceptible to the parasite than wild-type (WT) mice in the early phase of the infection, but were eventually able to eliminate the pathogen. In vitro Pten-deficient macrophages showed a reduced ability to kill parasites in response to IFN-gamma treatment, possibly because the mutant cells exhibited decreased TNF secretion that correlated with reductions in inducible nitric oxide synthase expression and nitric oxide production. In response to various TLR ligands, Pten-deficient macrophages produced less TNF and IL-12 but more IL-10 than WT cells. However, analysis of cells in the lymph nodes draining L. major inoculation sites indicated that both LysMCrePten(flox/flox) and WT mice developed normal Th1 responses following L. major infection, in line with the ability of LysMCrePten(flox/flox) mice to eventually eliminate the parasite. Our results indicate that the efficient clearance of intracellular parasites requires Pten in macrophages.     

A novel tumor necrosis factor (TNF) mimetic peptide prevents recrudescence of Mycobacterium bovis bacillus Calmette-Guerin (BCG) infection in CD4+ T cell-depleted mice

Tumor necrosis factor (TNF) is required to control mycobacterial infections, but its therapeutic value is limited by its in vivo instability and toxicity. The efficacy of a nontoxic TNF-mimetic peptide (TNF70-80) was tested in mice infected with Mycobacterium bovis bacillus Callette-Guerin (BCG). In vitro TNF70-80 and recombinant human TNF (hTNF) acted with interferon gamma (IFN-gamma) to reduce bacterial replication and to induce synthesis of bactericidal nitric oxide (NO) in BCG-infected, bone marrow-derived murine macrophages. The dose-dependent inhibitory effect on bacterial replication was blocked by neutralizing anti-IFN-gamma and anti-hTNF mAbs. Further, n-monomethyl-L-arginine (n-MMA) and a soluble TNF-receptor I (TNFRI-IgG) blocked bacterial growth and NO synthesis. Therefore, the peptide acted with IFN-gamma via induction of NO synthase and signaled through TNFRI receptors. Concomitant in vivo treatment with TNF70-80 or hTNF prevented reactivation of chronic BCG infection in mice depleted of CD4+ T cells by injecting anti-CD4 antibodies. Granuloma number and bacterial load were comparable in treated, T cell-depleted mice and in chronically infected, intact animals. Thus, TNF70-80 and hTNF can modulate recrudescent BCG infection in CD4+ T cell-deficient mice.     

An Essential Role for Gamma Interferon in Innate Resistance to Shigella flexneri Infection

Shigella spp. are the major cause of bacillary dysentery worldwide. To identify immune effectors associated with protection of the naive host during infection, the susceptibility to pulmonary Shigella infection of each of various mouse strains that have a targeted deletion in a specific aspect of the immune system was evaluated. Our results demonstrate that mice deficient in gamma interferon are 5 orders of magnitude more susceptible to Shigella than are wild-type mice, whereas mice deficient in B and T lymphocytes or in T lymphocytes alone exhibit no difference in susceptibility. Significantly lower numbers of shigellae were recovered from immunocompetent compared with gamma-interferon-deficient mice after infection. While immunocompetent mice were able to clear a sublethal Shigella inoculum by day 5 postinfection, progressively increasing numbers of shigellae were cultured from the lungs of gamma interferon-deficient mice over the same period. Histopathology of the lungs from immunocompetent mice infected with a sublethal Shigella inoculum showed mild inflammatory changes, whereas the lungs from gamma interferon-deficient mice demonstrated progressively worsening acute bronchiolitis with ulceration. Further, the time to death in gamma interferon-deficient mice correlates inversely with the size of the Shigella inoculum. To identify the cellular source of gamma interferon, we infected SCID mice, T-cell-receptor-deficient mice, beige mice (a mouse strain deficient in natural killer [NK] cell activity), and mice depleted of NK cells using anti-asialo-GM₁. Each NK cell-deficient mouse strain exhibited a 10-fold-greater susceptibility to Shigella infection than immunocompetent mice. To test the protective effects of gamma interferon in vitro, survival of intracellular Shigella was examined in primary macrophages from wild-type mice, primary macrophages from gamma interferon-deficient mice, a macrophage cell line, and a fibroblast cell line. Following activation with gamma interferon, each cell type eradicated intracellular Shigella, while nonactivated macrophages fostered Shigella replication and nonactivated fibroblast cells fostered both Shigella replication and intercellular spread. Taken together, these data establish that NK cell-mediated gamma interferon is essential to resistance following primary Shigella infection.     

Migration-inhibitory factor gene-deficient mice are susceptible to cutaneous Leishmania major infection

To determine the role of endogenous migration-inhibitory factor (MIF) in the development of protective immunity against cutaneous leishmaniasis, we analyzed the course of cutaneous Leishmania major infection in MIF gene-deficient mice (MIF(-/-)) and wild-type (MIF(+/+)) mice. Following cutaneous L. major infection, MIF(-/-) mice were susceptible to disease and developed significantly larger lesions and greater parasite burdens than MIF(+/+) mice. Interestingly, antigen-stimulated lymph node cells from MIF(-/-) mice produced more interleukin-4 (IL-4) and gamma interferon (IFN-gamma) than those from MIF(+/+) mice, although the differences were statistically not significant. IFN-gamma-activated resting peritoneal macrophages from MIF(-/-) mice showed impaired macrophage leishmanicidal activity and produced significantly lower levels of nitric oxide and superoxide in vitro. The macrophages from MIF(-/-) mice, however, produced much more IL-6 than macrophages from wild-type mice. These findings demonstrate that endogenous MIF plays an important role in the development of protective immunity against L. major in vivo. Furthermore, they indicate that the susceptibility of MIF(-/-) mice to L. major infection is due to impaired macrophage leishmanicidal activity rather than dysregulation of Th1 and Th2 responses.     

Infection of murine macrophages with Toxoplasma gondii is associated with release of transforming growth factor beta and downregulation of expression of tumor necrosis factor receptors.

Toxoplasma gondii is capable of invading and multiplying within murine peritoneal macrophages. Previous studies have shown that treatment of macrophage monolayers with recombinant gamma interferon but not tumor necrosis factor (TNF) is associated with intracellular killing of T. gondii by macrophages. Furthermore, infection of macrophages with T. gondii prevents their stimulation for mycobactericidal activity by TNF. Since transforming growth factor beta (TGF-beta) is known to suppress a number of functions in macrophages, we investigated the influence of infection with T. gondii on macrophage TNF receptors and on production of TGF-beta. Infection with T. gondii was associated with increased production of TGF-beta and downregulation of TNF receptors. This effect was observed early after infection and was partially inhibited by anti-TGF-beta 1 antibody.     

Production of tumor necrosis factor alpha by resident and activated murine macrophages.

Alveolar macrophages (Am phi s), resident peritoneal macrophages (RPm phi s), and thioglycolate-elicited peritoneal macrophages (TGPm phi s) were isolated from C57BL/6 mice and incubated with lipopolysaccharide (LPS), stimulated cell supernatant, or recombinant interferon gamma (IFN-gamma) for 24 h. Tumor necrosis factor (TNF) in cell-free supernatants was measured by enzyme-linked immunosorbent assay. Amo phi s incubated with 10(3) ng/ml LPS produced 50 times more TNF than RPm phi s and 5 times more than TGPm phi s, and LPS alone induced maximum TNF production by Am phi s. Stimulated cell supernatant or recombinant IFN-gamma alone did not induce TNF production. A combination of LPS with stimulated cell supernatant or IFN-gamma had only a limited synergistic effect on TNF production by Am phi s. However, both LPS and stimulated cell supernatant or recombinant IFN-gamma induced maximum TNF production by RPm phi s and TGPm phi s. TGPm phi s showed greater sensitivity to LPS and stimulated cell supernatant or IFN-gamma with regard to TNF production than the other macrophage populations investigated.     

Gamma interferon-mediated increase in the number of Ia-bearing macrophages during infection with Listeria monocytogenes.

The role of gamma interferon (IFN-gamma) in an increase in Ia-bearing macrophages during Listeria monocytogenes infection was studied. The peritoneal macrophages from L. monocytogenes-infected mice contained a high proportion of Ia. Intraperitoneal injection of the supernatant from a culture of spleen cells from L. monocytogenes-infected mice induced Ia-rich exudates in normal mice. The Ia-inducing activity in the culture supernatant was abrogated by the pretreatment of spleen cells with anti-Thy-1.2 antibody plus complement. Immunoadsorption of the culture supernatant with anti-recombinant IFN-gamma antibody and protein A-Sepharose CL-4B completely abrogated its Ia-inducing activity. These results suggested that an increase in Ia-bearing macrophages during L. monocytogenes infection was attributable to T-cell-derived IFN-gamma.     

Cyclosporin A-mediated killing of <Emphasis Type="Italic">Leishmania major</Emphasis> by macrophages is independent of reactive nitrogen and endogenous TNF-α and is not inhibited by IL-10 and 13

Murine macrophages were treated with various doses of cyclosporin A (CsA) to enhance the killing of Leishmania major parasites. CsA reduced the rate of infected cells from 75% in non-treated controls to less than 15% with 1 micro g CsA/ml in a dose-dependent manner. The leishmanicidal effect was also observed when CsA was added 48 h after the infection of macrophages. In contrast, FK506, another structural non-related immunosuppressive drug with antiparasitic activities, showed no effect on the ability of macrophages to kill intracellular Leishmania parasites. Since nitric oxide has been identified as a key molecule for the leishmanicidal function of macrophages, we analyzed the role of this molecule. There was no influence on the leishmanicidal effect of CsA when L- N-(1-iminoethyl)lysine, a potent and selective inhibitor of mouse inducible nitric oxide synthase, was added. Furthermore, the presence of the macrophage-inhibiting cytokines interleukin (IL)-10 and IL-13 simultaneously or prior to CsA did not inhibit leishmania killing, while both cytokines completely prevented parasite killing by macrophages activated with gamma interferon and tumor necrosis factor (TNF). CsA was fully active on macrophages from TNF-receptor p55 knockout mice arguing against autocrine activation by TNF. We therefore conclude that the antileishmanial effect of CsA is independent of effector mechanisms employed by macrophage-activating cytokines.     

Interleukin-12 secretion by Mycobacterium tuberculosis-infected macrophages.

Infection with Mycobacterium tuberculosis or phagocytosis of large latex beads induced interleukin-12 (IL-12) production in macrophages. In contrast, tumor necrosis factor (TNF) was produced only in response to M. tuberculosis infection, not after phagocytosis of latex beads. Comparable results were obtained with cells from immunocompetent C57BL/6 and gamma interferon receptor-deficient mutant mice. Thus, phagocytosis by mechanisms not specific for M. tuberculosis was a sufficient trigger for IL-12 secretion, emphasizing the central role of this cytokine in the initiation of anti-infective immunity.     

Expansion of gamma interferon-producing CD8+ T cells following secondary infection of mice immune to Leishmania major.

Reinfection of immune mice with Leishmania major elicits a secondary gamma interferon (IFN-gamma) response to which specific CD8+ T cells are essential. We have shown previously that specific CD8+ T cells from reinfected immune mice release substantially higher levels of IFN-gamma, a cytokine essential for the efficient activation of parasitized macrophages to kill intracellular L. major. By using an ELISPOT assay, which allows the detection of IFN-gamma production by individual cells, it is shown here that this elevated IFN-gamma response is the result of an increase of up to 50-fold in the frequency of parasite-specific CD8+ T lymphocytes in the spleens and draining lymph nodes of both immune reinfected CBA and BALB/c mice. This observation is additional evidence of the role that CD8+ T cells play in immunity to reinfection with L. major.     

Enhanced replication of Leishmania amazonensis amastigotes in gamma interferon-stimulated murine macrophages: implications for the pathogenesis of cutaneous leishmaniasis

During Leishmania major infection in mice, gamma interferon (IFN-gamma) plays an essential role in controlling parasite growth and disease progression. In studies designed to ascertain the role of IFN-gamma in Leishmania amazonensis infection, we were surprised to find that IFN-gamma could promote L. amazonensis amastigote replication in macrophages (Mphis), although it activated Mphis to kill promastigotes. The replication-promoting effect of IFN-gamma on amastigotes was independent of the source and genetic background of Mphis, was apparently not affected by surface opsonization of amastigotes, was not mediated by interleukin-10 or transforming growth factor beta, and was observed at different temperatures. Consistent with the different fates of promastigotes and amastigotes in IFN-gamma-stimulated Mphis, L. amazonensis-specific Th1 transfer helped recipient mice control L. amazonensis infection established by promastigotes but not L. amazonensis infection established by amastigotes. On the other hand, IFN-gamma could stimulate Mphis to limit amastigote replication when it was coupled with lipopolysaccharides but not when it was coupled with tumor necrosis factor alpha. Thus, IFN-gamma may play a bidirectional role at the level of parasite-Mphi interactions; when it is optimally coupled with other factors, it has a protective effect against infection, and in the absence of such synergy it promotes amastigote growth. These results reveal a quite unexpected aspect of the L. amazonensis parasite and have important implications for understanding the pathogenesis of the disease and for developing vaccines and immunotherapies.     

Modulation of tumor necrosis factor production by macrophages in Entamoeba histolytica infection.

The macrophage-derived mediator tumor necrosis factor alpha (TNF) is a cytokine with pleiotropic effects. TNF exhibits potent immunologic and inflammatory properties in parasitic diseases. The present study examined the production of TNF by macrophages isolated from gerbils infected with Entamoeba histolytica and by naive macrophages in response to amoebae in vitro. Amoebic liver abscess-derived macrophages produced low constitutive basal levels of TNF; in response to lipopolysaccharide (LPS) stimulation, TNF production was enhanced by 14-, 11-, and 6-fold at 10, 20, and 30 days postinfection, respectively. Amoebic liver abscess-derived macrophages pretreated with either recombinant gamma interferon (IFN-gamma) or the cyclooxygenase inhibitor indomethacin augmented TNF production in response to soluble amoebic proteins and LPS. Kupffer cells and peritoneal and spleen macrophages from infected animals did not release TNF constitutively in vitro. However, TNF production in response to LPS stimulation was significantly higher at 10 and 20 days postinfection. Macrophages from infected and naive animals pretreated with recombinant IFN-gamma or indomethacin produced increased amounts of TNF in response to LPS but not in response to soluble amoebic protein stimulation. Pretreatment of naive macrophages with amoebic proteins inhibited LPS-induced TNF production by 69 to 79%; the effect of the amoebic proteins was partially reversed by indomethacin pretreatment. In contrast, IFN-gamma- and LPS-activated naive macrophages produced enhanced levels of TNF in response to live amoebae and soluble amoebic proteins. Our results demonstrate that TNF production by macrophages is altered during E. histolytica infection and in response to amoebae and suggest a role for IFN-gamma and prostaglandin E2 in regulating TNF production during the infection.     

Endogenous tumor necrosis factor, interleukin-6, and gamma interferon levels during Listeria monocytogenes infection in mice.

Mice were infected intravenously with a sublethal dose of Listeria monocytogenes cells and then levels of tumor necrosis factor (TNF), interleukin-6 (IL-6), and gamma interferon (IFN-gamma) in the bloodstreams, spleens, and livers were monitored. The maximum level of TNF was detected at 72 h in the spleens and livers, but TNF was never detected in the bloodstreams. IL-6 appeared in the bloodstreams and spleens and peaked at 48 h. The maximum level of IFN-gamma could be detected in all three specimens, and the highest titer was shown in the spleens. Endogenous TNF production was suppressed by in vivo administration of anti-CD4 monoclonal antibody (MAb) or anti-asialo GM1 antibody but not by anti-CD8 MAb, whereas none of these antibodies suppressed endogenous IL-6 production. Endogenous production of neither IL-6 nor IFN-gamma was inhibited in rabbit anti-recombinant mouse TNF-alpha antibody-treated mice. Similarly, production of TNF and IL-6 did not decrease in anti-mouse IFN-gamma MAb-treated animals, but TNF production was augmented in these animals. These results suggest that the these endogenous cytokines are produced by different mechanisms in L. monocytogenes infection.