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. 2004 Mar;72(3):1257-64.
doi: 10.1128/IAI.72.3.1257-1264.2004.

STAT1 is essential for antimicrobial effector function but dispensable for gamma interferon production during Toxoplasma gondii infection

L Cristina Gavrilescu  1 , Barbara A Butcher, Laura Del Rio, Gregory A Taylor, Eric Y Denkers
Affiliations

STAT1 is essential for antimicrobial effector function but dispensable for gamma interferon production during Toxoplasma gondii infection

L Cristina Gavrilescu et al. Infect Immun. 2004 Mar.

Abstract

The opportunistic protozoan Toxoplasma gondii is a prototypic Th1-inducing pathogen inducing strong gamma interferon (IFN-gamma) cytokine responses that are required to survive infection. Intracellular signaling intermediate STAT1 mediates many effects of IFN-gamma and is implicated in activation of T-bet, a master regulator of Th1 differentiation. Here, we show that T. gondii-infected STAT1-null mice fail to upregulate the IFN-gamma-dependent effector molecules inducible nitric oxide synthase (iNOS), IGTP, and LRG-47, which are required for mice to survive infection. Both T-bet and interleukin-12 receptor beta2 (IL-12Rbeta2) failed to undergo normal upregulation in response to T. gondii. Development of IFN-gamma-producing CD4(+) and CD8(+) T lymphocytes was severely curtailed in the absence of STAT1, but a substantial level of STAT1-independent non-T-cell-derived IFN-gamma was induced. Absence of STAT1 also resulted in increased IL-4, Arg1, Ym1, and Fizz1, markers of Th2 differentiation and alternative macrophage activation. Together, the results show that T. gondii induces STAT1-dependent T-lymphocyte and STAT1-independent non-T-cell IFN-gamma production, but that effector functions of this type 1 cytokine cannot operate in the absence of STAT1, resulting in extreme susceptibility to acute infection.

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Figures

FIG. 1.
FIG. 1.
STAT1−/− mice are highly susceptible to T. gondii infection. (A) STAT1−/− mice die by 10 days postinfection with 20 ME49 cysts, while WT 129Sv/Ev mice survive infection until termination of the experiment at 30 days. Data are representative of two independent experiments with 10 mice per group. (B) Increased parasitemia in macrophages of infected STAT1−/− mice. Peritoneal exudate cells were recovered at 0, 4, and 7 days postinfection, and the percentage of infected macrophages was determined at 4 and 7 days postinfection. Student's t test revealed statistically significant differences between WT (solid bars) and KO (open bars) samples (*, P = 0.008; and **, P = 0.014). Data were pooled from two independent experiments with three mice per group; bars represent standard error.
FIG. 2.
FIG. 2.
Defective macrophage responses to T. gondii infection. PMØ (A and C) and BMMØ (B and D) were infected with RH tachyzoites (TZ; 1:1 parasite/cell ratio) after 18 h of stimulation with 500-U/ml rIFN-γ or medium alone. IL-12 p40 (A and B) and NO (C and D) were measured in the culture supernatants after 24 h of infection. Bars represent the standard deviation of duplicate samples. Solid bars, WT; open bars, KO. Data are representative of four independent experiments.
FIG. 3.
FIG. 3.
Failure to induce IGTP and LRG-47 in STAT1−/− animals. At the indicated number of days postinfection (dpi), splenocytes were recovered and lysed. Samples were resolved by SDS-PAGE and transferred to a nitrocellulose membrane. Blots were then submitted to IGTP (upper panel) and LRG-47 (lower panel) Western blot analysis. Data are representative of two independent experiments, with independently processed samples from three mice per group at each time point.
FIG. 4.
FIG. 4.
Production of proinflammatory cytokines in infected STAT1−/− versus STAT1+/+ animals. Sera were collected from mice at 0 and 4 and 7 days postinfection and subjected to ELISA for IL-12 p40 (A) and IFN-γ (B) cytokine detection. Splenocytes were cultured for 72 h in the presence of STAg, supernatants were assayed for IL-12 p40 (C) and IFN-γ (D) production by ELISA, and NO was measured by the Greiss assay (E). Cytokines were not detected in medium-only cultures. Samples were run in duplicate. Bars represent the standard error of three mice per group. Solid bars, WT; open bars, KO. ND, not detected. Data are representative of two independent experiments.
FIG. 5.
FIG. 5.
Defective T-lymphocyte production of IFN-γ in STAT1−/− mice. Splenocytes were obtained from mice 7 days after infection, immediately stimulated with PMA and ionomycin, and stained for T-cell markers and IFN-γ as described in Materials and Methods. The numbers in each quadrant indicate the percentage of cells in that quadrant. This experiment was repeated three times with essentially identical results.
FIG. 6.
FIG. 6.
Lack of induction of Th1 differentiation markers, but activation of anti-inflammatory markers, in infected STAT1 KO mice. RNA was isolated from splenocytes at the indicated times after infection and subjected to real-time PCR analysis. RNA samples were individually processed (three mice per group). Real-time PCR was performed, and the results were analyzed as described in Materials and Methods and compared to the level of gene expression found in noninfected WT animals, arbitrarily assigned the value 1. Bars represent the fold increase in gene expression over the expression level in the noninfected WT samples. Solid bars, WT; open bars, KO. Data are representative of two independent experiments.
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References

    1. Afkarian, M., J. R. Sedy, J. Yang, N. G. Jacobson, N. Cereb, S. Y. Yang, T. L. Murphy, and K. M. Murphy. 2002. T-bet is a STAT1-induced regulator of IL-12R expression in naive CD4+ T cells. Nat. Immunol. 6:549-557. - PubMed
    1. Bliss, S. K., A. J. Marshall, Y. Zhang, and E. Y. Denkers. 1999. Human polymorphonuclear leukocytes produce IL-12, TNF-α, and the chemokines macrophage-inflammatory protein-1α and -1β in response to Toxoplasma gondii antigens. J. Immunol. 162:7369-7375. - PubMed
    1. Bliss, S. K., Y. Zhang, and E. Y. Denkers. 1999. Murine neutrophil stimulation by Toxoplasma gondii antigen drives high level production of IFN-γ-independent IL-12. J. Immunol. 163:2081-2088. - PubMed
    1. Chang, N. C., S. I. Hung, K. Y. Hwa, I. Kato, J. E. Chen, C. H. Liu, and A. C. Chang. 2001. A macrophage protein, Ym1, transiently expressed during inflammation is a novel mammalian lectin. J. Biol. Chem. 276:17497-17506. - PubMed
    1. Collazo, C. M., G. S. Yap, S. Hieny, P. Caspar, C. G. Feng, G. A. Taylor, and A. Sher. 2002. The function of gamma interferon-inducible GTP-binding protein IGTP in host resistance to Toxoplasma gondii is Stat1 dependent and requires expression in both hematopoietic and nonhematopoietic cellular compartments. Infect. Immun. 70:6933-6939. - PMC - PubMed

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