Cells and cytokines in resistance to Toxoplasma gondii.
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Animals↗
Biomedical subjects
C A Hunter
Publications and source records attributed to C A Hunter.
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.
Animals↗
IL-1 beta is required for IL-12 to induce production of IFN-gamma by NK cells. A role for IL-1 beta in the T cell-independent mechanism of resistance against intracellular pathogens.
Mice with the severe combined immunodeficiency (SCID) possess an IFN-gamma-dependent mechanism of resistance to the intracellular pathogens Toxoplasma gondii and Listeria monocytogenes that is dependent on IL-12-induced production of IFN-gamma by NK cells. In this report we demonstrate that IL-1 beta is required for IL-12 to stimulate production of IFN-gamma by NK cells, and that IL-1 is important in IL-12-mediated resistance to T. gondii in vivo. Stimulation of SCID mouse splenocytes with tachyzoites of T. gondii resulted in production of IFN-gamma. Addition of neutralizing Ab specific for IL-1 beta to these cultures inhibited completely the production of IFN-gamma. Similar results were obtained when LPS or L. monocytogenes were used to stimulate production of IFN-gamma by SCID mouse splenocytes. Addition of a neutralizing Ab to IL-1 alpha did not affect production of IFN-gamma by SCID mouse splenocytes stimulated with T. gondii, L. monocytogenes, or LPS. Stimulation of SCID mouse splenocytes with IL-1 beta or IL-1 alpha did not result in production of IFN-gamma but enhanced remarkably the ability of T. gondii or IL-12 to stimulate production of IFN-gamma. Furthermore, production of IFN-gamma by SCID mouse splenocytes stimulated with IL-12 plus TNF-alpha was completely ablated by anti-IL-1 beta, but not by anti-IL-1 alpha. Analysis of the culture supernatants of spleen cells from SCID mice stimulated with T. gondii or IL-12 plus TNF-alpha detected low levels of IL-1 beta; addition of a neutralizing Ab to IFN-gamma resulted in a 5- to 10-fold increase in levels of IL-1 beta. Furthermore, stimulation of SCID mouse splenocytes with IL-12, in the presence of anti-IFN-gamma, resulted in an increase in detectable levels of IL-1 beta. To determine the in vivo relevance of our in vitro data, SCID mice were infected with T. gondii and treated with IL-12 alone or IL-12 in combination with an Ab specific for the type I IL-1 receptor. This Ab reduced production of IFN-gamma by SCID mouse splenocytes stimulated with either T. gondii, LPS, L. monocytogenes, or IL-12 plus IL-1 beta. In vivo administration of this Ab antagonized significantly the ability of exogenous IL-12 to delay the time to death of SCID mice infected with T. gondii.(ABSTRACT TRUNCATED AT 400 WORDS)
Adjuvants, Immunologic↗
Transforming growth factor-beta inhibits interleukin-12-induced production of interferon-gamma by natural killer cells: a role for transforming growth factor-beta in the regulation of T cell-independent resistance to Toxoplasma gondii.
Severe-combined immune deficient (SCID) mice have been found to resist infection with the intracellular protozoan parasite Toxoplasma gondii via interleukin (IL)-12 stimulation of interferon (IFN)-gamma production by natural killer (NK) cells. Previously, we demonstrated the presence of increased levels of transcripts for transforming growth factor-beta (TGF-beta) in the brains and lungs of SCID mice infected with T. gondii, leading us to investigate the role of TGF-beta in the mechanism of resistance to T. gondii in these mice. Stimulation of splenocytes from SCID mice with heat-killed T. gondii resulted in production of low levels of IFN-gamma and a two to threefold increase in levels of TGF-beta in the culture supernatants. Production of IFN-gamma in these cultures was increased three to fourfold by addition of anti-TGF-beta antibody. Stimulation of splenocytes from SCID mice with IL-12 in combination with either TNF-alpha or IL-1 beta resulted in production of high levels of IFN-gamma. Addition of TGF-beta to these cultures inhibited production of IFN-gamma in a dose-dependent manner. Immunohistochemical studies revealed increased levels of TGF-beta protein in the spleens of SCID mice 5 days after oral infection with the ME49 strain of T gondii, and brains of SCID mice at 18 days post-infection. However, no difference was detected in the levels of TGF-beta transcripts in the spleens of uninfected mice or mice infected for 5 days. To test whether TGF-beta could antagonize IL-12 mediated resistance to T. gondii in vivo, we administered TGF-beta to SCID mice infected with T. gondii. This treatment resulted in earlier mortality of infected mice and significantly reduced the ability of exogenous IL-12 to delay time-to-death. Administration of anti-TGF-beta to SCID mice, beginning 24 h prior to infection and every 2 days thereafter, delayed significantly time-to-death. Together, our data demonstrate that TGF-beta antagonizes the ability of IL-12 to stimulate production of IFN-gamma by splenocytes from SCID mice, and suggest a role for TGF-beta in regulation of T cell-independent resistance to T. gondii.
Animals↗
Roles of gamma interferon and other cytokines in suppression of the spleen cell proliferative response to concanavalin A and toxoplasma antigen during acute toxoplasmosis.
Suppressed splenocyte proliferation in response to mitogen and toxoplasma lysate antigen (TLA) is observed in mice acutely infected with Toxoplasma gondii. Recently, we reported that NG-monomethyl-L-arginine (NMMA), an inhibitor of reactive nitrogen intermediate (RNI) production, partially restored proliferative responses of splenocytes from infected mice. In the present study we have examined the effect of NMMA on production of cytokines by splenocytes from mice acutely infected with T. gondii and assessed the role of gamma interferon (IFN-gamma) and interleukin-10 (IL-10) in the RNI-mediated suppression. Stimulation with concanavalin A (ConA) or TLA of splenocytes from CBA/Ca mice infected for 7 days resulted in increased production of IFN-gamma, IL-4, and IL-10 but reduced levels of IL-2 when compared with cultures of splenocytes from uninfected mice. Whereas addition of NMMA did not alter levels of cytokines produced by splenocytes from uninfected mice, splenocytes from infected mice stimulated with ConA produced significantly higher levels of IL-10 and reduced levels of IL-2 and IL-4. Addition of anti-IFN-gamma monoclonal antibodies to cultures of spleen cells from mice infected for 7 or 14 days remarkably decreased the levels of nitrite and resulted in a 47- and 4-fold increase in proliferation induced by stimulation with ConA or TLA, respectively. Anti-IL-10 did not reduce levels of nitrite produced in culture but did result in a fourfold increase in the proliferative response of splenocytes from mice infected for 14 days. In vivo administration of anti-IFN-gamma or anti-IL-10 monoclonal antibodies to infected mice partially restored ex vivo spleen cell proliferative responses by approximately 40 and 15%, respectively. Our data indicate that IFN-gamma is important in inducing the RNI-mediated immunosuppression, which, in turn, affects production of cytokines by splenocytes. Our data also demonstrate that IL-10 is involved in the suppression observed but that this activity is independent of RNI.
Acute Disease↗
Preferential activation and expansion of human peripheral blood gamma delta T cells in response to Toxoplasma gondii in vitro and their cytokine production and cytotoxic activity against T. gondii-infected cells.
Studies were conducted to determine if gamma delta T cells participate in the immune response to Toxoplasma gondii. Preferential expansion of human gamma delta T cells occurred when peripheral blood T cells from either T. gondii-seronegative or seropositive individuals were incubated with autologous PBMC infected with the parasite. That gamma delta T cells proliferated after incubation with infected cells was confirmed using purified of gamma delta T cells. These T. gondii-induced gamma delta T cell responses did not require prior exposure to the parasite since T cells obtained from umbilical cord blood from seronegative newborns also exhibited preferential expansion of gamma delta T cells. Cytofluorometric analysis of T cells obtained from either umbilical cord blood or peripheral blood from adults revealed that V gamma 9+ and V delta 2+ gamma delta T cells responded to stimulation with infected cells. Preferential expansion of gamma delta T cells was not restricted by polymorphic determinants of MHC molecules. PBMC that had internalized killed parasites but not PBMC incubated with T. gondii lysate antigens also stimulated preferential expansion and activation of gamma delta T cells as assessed by expression of CD25 and HLA-DR molecules. V gamma 9+V delta 2+ gamma delta T cells were cytotoxic for T. gondii-infected cells in an MHC-unrestricted manner, and produced IFN-gamma, IL-2, TNF-alpha, but not IL-4 when incubated with cells infected with the parasite. These results suggest that rapid induction of a remarkable primary gamma delta T cell response may be important in the early protective immune response to T. gondii.
Animals↗
Definitive identification of a gene that confers resistance against Toxoplasma cyst burden and encephalitis.
Control of resistance to cyst burden following per-oral infection with Toxoplasma gondii has been mapped previously to a region of mouse chromosome 17 of approximately 140 kb. This region is contiguous with and contains the class I gene, Ld. Resistance to development of toxoplasmic encephalitis has also been reported to be controlled by genes in this region of H-2. TNF-alpha, D and L genes, as well as unidentified genes, are also in this region. The work described here was performed to identify definitively the gene(s) in this 140 kb region that confers resistance to cysts and encephalitis. The study demonstrates that relative resistance to T. gondii organisms and cyst burden in brain, and toxoplasmic encephalitis, 30 days following per-oral T. gondii infection is correlated absolutely with the presence of the Ld gene in inbred, recombinant, mutant and C3H.Ld transgenic mice. Mice with the Ld gene had lower cyst burdens and less encephalitis than those without the Ld gene. Specifically, 30 days after infection mice with the Ld gene had minimal perivascular inflammation and meningeal inflammation and very few Toxoplasma cysts or organisms in immunoperoxidase-stained preparations of their brains. Mice without the Ld gene had a similar pattern of inflammation, but in addition they had collections of inflammatory cells in the brain parenchyma. Free tachyzoites were found within these foci of inflammation and cysts were present in these areas as well as in contiguous areas without inflammatory cells. There were CD4+ and CD8+ T lymphocytes in the areas of inflammation and throughout the brain parenchyma. Mice that were resistant to cysts and encephalitis had little detectable brain cytokine mRNA expression, while mice that were susceptible had elevated levels of mRNA for a wide range of cytokines, consistent with their greater amounts of inflammation. The present work definitively demonstrates that a Ld-restricted response decreases the number of organisms and cysts within the brain and thereby limits toxoplasmic encephalitis and levels of interferon-gamma (IFN-gamma), tumour necrosis factor-alpha (TNF-alpha), interleukin-2 (IL-2), IL-6, IL-10, transforming growth factor-beta (TGF-beta), IL-1 alpha, IL1 beta and macrophage inhibiting protein (MIP) mRNA in the brain 30 days after per-oral infection.
Animals↗
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.
Acute Disease↗
Enhancement of intracellular replication of Toxoplasma gondii by IL-6. Interactions with IFN-gamma and TNF-alpha.
Toxoplasma gondii is a major pathogen of immunocompromised hosts, and one defense mechanism against the parasite is activation of macrophages (M phi) for toxoplasmacidal activity by IFN-gamma after triggering by TNF-alpha. IL-6, IFN-gamma, and TNF-alpha are cytokines involved in inflammatory responses and are induced by T. gondii infection. We studied the interaction of these three cytokines using an in vitro model of T. gondii infection. Pretreatment (but not post-treatment) of unelicited murine peritoneal M phi with IL-6 enhanced T. gondii replication in a dose-dependent manner. Pretreatment with IFN-gamma resulted in active killing of parasites whereas the addition of IL-6 to IFN-gamma pretreatment resulted in a reversal of IFN-gamma-mediated toxoplasmacidal activity. Combining TNF-alpha with IL-6 and IFN-gamma pretreatment resulted in restoration of toxoplasmacidal activity. Addition of a polyclonal anti-TNF-alpha Ab to IL-6 and IFN-gamma pretreatment resulted in enhancement in the IL-6-mediated impairment of IFN-gamma function. These data taken together suggest that IL-6 enhances intracellular replication of T. gondii and reverses IFN-gamma mediated activation of murine peritoneal M phi, and that certain of the interactions between these two cytokines may be at the level of TNF-alpha triggering.
Animals↗
The role of cytokines in toxoplasmosis.
Infection with Toxoplasma gondii is normally asymptomatic, but as a consequence of the AIDS epidemic the incidence of symptomatic disease and especially toxoplasmic encephalitis (TE) has grown in frequency. The high frequency of adverse reactions to conventional therapeutic regimens for toxoplasmosis highlight the need to develop new strategies for the management of this disease. The importance of cytokines in resistance against T. gondii has been shown primarily in murine models of toxoplasmosis and a number of cytokines (e.g., IFN-gamma, TNF-alpha, IL-2 and IL-12) have been proposed for trials in patients with TE. One mechanism by which these cytokines produce their effects is through stimulation of macrophages and/or NK cells. However, there are problems with immunological intervention in immunocompromised patients with TE since the infection is present primarily in the central nervous system (CNS), an immunoprivileged site, and because certain cytokines may down regulate the immune response. While much valuable information has been obtained from studies conducted in immunocompetent strains of mice their relevance to an immunocompromised host is unknown. The development of genetically altered mice with immune deficiencies offers promising new models that may allow for more rational development of new treatment regimens.
AIDS-Related Opportunistic Infections↗
Immunocytochemical detection of cytokines in the lymph nodes and brains of mice resistant or susceptible to toxoplasmic encephalitis.
BALB/c mice develop toxoplasmic encephalitis (TE) but the pathology resolves; in contrast, CBA/Ca mice develop progressive TE. Immunohistochemical analysis revealed peak staining for interferon-gamma (IFN-gamma) and tumor necrosis factor (TNF) in lymph nodes of both strains on day 7 of infection, with elevated levels of IFN-gamma and TNF persisting in CBA/Ca mice. TNF was present in the central nervous systems (CNS) of both strains of mice and correlated with severity of pathology. Interleukin (IL)-2 was detected in lymph nodes and CNS of CBA/Ca mice after day 52 of infection and only in the lymph nodes of BALB/c mice between days 14 and 21 after infection. Highest levels of IL-10 were observed in lymph nodes and CNS of infected CBA/Ca mice. Variations in cytokine levels in lymph nodes and CNS of these mouse strains may account for differences in their susceptibility to TE.
Animals↗
Immunopathogenesis of toxoplasmic encephalitis.
The incidence of toxoplasmic encephalitis (TE) has increased with the increasing numbers of patients with immunodeficiencies, in whom reactivation of latent Toxoplasma infection may occur. This highlights the important role of the immune response in maintaining infection with Toxoplasma gondii in a latent form. Because the brain is the most commonly affected site of latent infection and because it is anatomically unique in regard to the immune system, understanding the systemic immune response to infection within the brain is important. Murine models have proven useful for the study of the immune response to T. gondii and identified the importance of cytokines and NK and T cells in the regulation of protective immunity to T. gondii. Further studies on the development of TE have indicated the possible importance of the interactions of glial cells, such as microglia and astrocytes, with infiltrating T cells to mediate immunity to T. gondii within the brain.
Animals↗
Temporal differences in the expression of mRNA for IL-10 and IFN-gamma in the brains and spleens of C57BL/10 mice infected with Toxoplasma gondii.
C57BL/10 Sc Sn (B10) mice infected orally with Toxoplasma gondii tissue cysts were killed at regular intervals up to day 116 post infection (p.i.) and their brains excised. These were used either to count the total number of cysts in the brain, for RNA purification or histopathological studies. Mortality levels in a parallel group of T. gondii infected B10 mice were also monitored and regular plasma samples taken to measure specific antibody production. Seventy per cent of mice died within the first 35 days of infection. Thereafter deaths were infrequent. Inflammation in the brain was apparent from day 10 onwards and by day 25 there was widespread astrocyte activation, perivascular cuffing, meningitis and extensive encephalitis. Total cyst numbers increased rapidly from day 15 to day 35 when they peaked. By day 60, however, cyst numbers had dropped dramatically and this decrease continued through to day 116. Using the polymerase chain reaction mRNA transcripts for IFN-gamma were detected from the first time point sampled, day 25 p.i., until the end of the study. Transcripts for IL-10, an inhibitor of IFN-gamma production, release and activity, were not detected until day 70. The predominant antibody detected against T. gondii was IgG2a but not IgG1. Significantly transcripts for IFN-gamma were found in the spleens of infected but not non-infected animals. Our results suggest that an inflammatory response associated with IFN-gamma production in B10 mice eventually controls T. gondii infection. After the cyst burden has dropped dramatically transcripts for IL-10 are detected in the brain, perhaps to suppress inflammation, and limit pathology.
Animals↗
Mitogen- and antigen-specific proliferation of T cells in murine toxoplasmosis is inhibited by reactive nitrogen intermediates.
Acute and chronic infections with Toxoplasma gondii result in a nonspecific suppression of immunologic function in mice and humans. Proliferation of spleen cells in response to concanavalin A (ConA) and toxoplasma lysate antigen (TLA) was studied during the course of infection in mice susceptible (CBA/Ca) and resistant (BALB/c) to development of toxoplasmic encephalitis to determine if reactive nitrogen intermediates (RNI) are involved in the suppression of the proliferative responses. Maximal suppression of proliferation of spleen cells in response to ConA and TLA was observed on days 7 and 14 after infection and correlated with elevated levels of nitrite in spleen cell culture supernatants. By day 68 postinfection in BALB/c mice, proliferative responses returned to normal levels, whereas in CBA/Ca mice, they remained suppressed. The addition of an inhibitor of production of RNI (NG-monomethyl-L-arginine) increased proliferation of spleen cells in response to both ConA and TLA at days 7, 14, and 21 after infection. Depletion of adherent cells from spleen cell preparations obtained from acutely infected mice followed by their repletion with adherent spleen cells from uninfected mice resulted in increased proliferation of spleen cells from infected mice and a significant decrease in nitrite in the cultures. These results indicate that production of RNI by macrophages contributes significantly to the suppression of the spleen cell proliferation observed in the acute stage of toxoplasmosis.
Animals↗
Production of gamma interferon by natural killer cells from Toxoplasma gondii-infected SCID mice: regulation by interleukin-10, interleukin-12, and tumor necrosis factor alpha.
Previous studies of mice have implicated natural killer (NK) cells as mediators of protective activity against Toxoplasma gondii through their production of gamma interferon (IFN-gamma). In the present study, we have compared NK-cell activity in infected and uninfected SCID mice. Our data reveal that infection results in increased levels of IFN-gamma in serum and elevated NK-cell activity but that these NK cells were not cytotoxic for T. gondii-infected P815 cells. Treatment with anti-IFN-gamma antibody abrogated the increase in NK-cell activity and resulted in earlier mortality of infected mice. In vivo treatment with anti-asialo GM1 antiserum reduced NK cell activity and levels of IFN-gamma in serum but did not alter time to death. Spleen cells from infected mice produced higher levels of IFN-gamma than those from uninfected mice when stimulated in vitro with live T. gondii or parasite antigen preparations. Further analysis revealed that interleukin 10 (IL-10) inhibited, whereas tumor necrosis factor alpha (TNF-alpha) and IL-12 enhanced, IFN-gamma production by spleen cells from infected or uninfected mice. The combination of IL-12 and TNF-alpha induced higher levels of IFN-gamma from whole spleen cells of infected mice than from those of uninfected mice. Depletion of the adherent cell population from the spleen cells of infected mice led to a significant reduction in the levels of IFN-gamma produced after stimulation with IL-12 plus TNF-alpha. Similar results did not occur with cells from uninfected mice. These data indicate that other cytokines produced by the adherent cell population from infected mice may be involved in maximal production of IFN-gamma by NK cells stimulated with IL-12 and TNF-alpha. To assess the importance of endogenous IL-12, a polyclonal anti-IL-12 was administered to infected SCID mice. This treatment led to earlier mortality, indicating that endogenous IL-12 mediates resistance to T. gondii.
Animals↗
Sequence-dependent DNA structure. The role of base stacking interactions.
The sequence-dependent structure of DNA is analysed on the basis of the energetics of the base stacking (pi-pi) interactions. The conformational preferences of the ten possible base-pair steps in double-helical DNA have been calculated and compared with experimental data from X-ray fibre diffraction, X-ray crystal structures and gel-running experiments. The calculations account for many features of sequence-dependent DNA structure, including polymorphism in DNA, the lack of polymorphism in RNA, the structure of Z-DNA, bistability in pyrimidine-purine (YR) steps, the origin of propeller twist and buckle and the role of TATA sequences at the sites of origin of replication. The computational model used specifically allows for the charge distribution associated with the out-of-plane pi-electron density of the bases. The results obtained are rationalized on the basis of the shapes and charge distributions of the bases. Calladine's cross-strand steric clashes at pyrimidine-purine (YR) and CX/XG steps are reproduced. In AX/XT steps, same-strand steric clashes occur between the thymine methyl group and the 5'-neighbouring sugar. They are the cause of the large negative propeller twist observed in A.T base-pairs. Steric clashes between the thymine methyl group and the 5'-neighbouring base block A-DNA conformations in AX/XT steps. Electrostatic interactions between partial atomic charges are most important for C.G base-pairs which are highly polarized. They lead to strong preferences for positive slide and negative slide conformations in CG and GC steps, respectively. This combination can be accommodated in poly(CG) by left-handed Z-DNA. Many of the more subtle sequence-dependent effects are caused by electrostatic interactions between the partial atomic charges on one base-pair and the out-of-plane pi-electron density on another. The effect is most important in CX-XG steps and leads to bistability. In general, electrostatic interactions cause non-zero slide. This is opposed by the hydrophobic effect which favours the zero slide B-type conformation. Thus, B-DNA is observed at high water content in fibres and electrostatic interactions force high or low slide A or C-DNA conformations at low water content. If two juxtaposed steps have very different conformational preferences, this incompatibility can lead to unusual structures such as Z-DNA or strain such as in TATA sequences.
Base Sequence↗
Cytokine mRNA in the central nervous system of SCID mice infected with Toxoplasma gondii: importance of T-cell-independent regulation of resistance to T. gondii.
Levels of cytokine mRNA were studied in the central nervous system (CNS) of SCID mice infected with Toxoplasma gondii. This infection led to 100% mortality by day 23 postinfection. Inflammation was observed in the lungs on day 7 and in the heart, liver, and kidneys on days 14 and 18 of infection. In the CNS, necrotic, acellular lesions that contained numerous parasites, accompanied by a localized astrocyte activation, were evident on day 14. Polymerase chain reaction-assisted amplification of RNA revealed that, although transcripts for interleukin-1 alpha (IL-1 alpha) and IL-1 beta were present in the brains of uninfected mice, increased levels of these transcripts were detected on day 7 of infection. Transcripts for macrophage inflammatory protein 1 and transforming growth factor beta were also detected in brains of infected mice at this time point. On days 14 and 18, levels of these transcripts had increased and transcripts for IL-6, IL-10, gamma interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and granulocyte-macrophage colony-stimulating factor (GM-CSF) were also detected. Transcripts for IL-2 or IL-4 were not detected at any of the time points. Detection of locally produced cytokine transcripts may reflect involvement of the cytokines in the immunopathogenesis of this infection or involvement in mediating antitoxoplasma activity. To assess the possible role of endogenous IFN-gamma, TNF-alpha, IL-10, IL-6, and GM-CSF, cytokine-neutralizing monoclonal antibodies were administered to infected SCID mice. Neutralization of IFN-gamma or TNF-alpha led to earlier mortality than that in controls. In contrast, treatment with antibody to IL-10 and IL-6 increased survival time. Treatment with anti-GM-CSF did not alter the time to death. These results indicate that TNF-alpha and IFN-gamma are both involved in T-cell-independent mechanisms of resistance to T. gondii in SCID mice and that IL-10 and IL-6 may downregulate the immune response to this pathogen.
Animals↗
Regulation of tumor necrosis factor production by adrenaline and beta-adrenergic agonists.
The effects of adrenaline and isoproterenol, a specific beta-adrenergic agonist, on TNF production were investigated. Both agents inhibited the production of TNF by human blood and THP-1 cells stimulated by LPS. The effect of adrenaline was prevented by a beta-receptor antagonist, but not by an alpha-receptor antagonist. Levels of TNF mRNA were not reduced by adrenaline. Inhibition of TNF production was observed only if cells were first exposed to adrenaline or isoproterenol at about the same time as to LPS; incubation of THP-1 cells with isoproterenol for 24 h before LPS stimulation dramatically increased response, and prevented suppression of TNF production by a second dose of isoproterenol. Intracellular cAMP levels were increased by adrenaline and isoproterenol, at concentrations that inhibited TNF production. However, prolonged incubation of THP-1 cells with isoproterenol resulted in depression of cAMP concentrations to below basal levels. These data suggest that TNF production can be regulated by beta-receptor stimulation, that such regulation is mediated by changes in intracellular cAMP concentrations and is exerted at a posttranscriptional level. Adrenaline may be an important endogenous regulator of TNF production in sepsis.
Adrenergic beta-Agonists↗