RESUMEN
Variation between inbred mice of susceptibility to experimental Trypanosoma cruzi infection has frequently been described, but the immunogenetic background is poorly understood. The outcross of the susceptible parental mouse strains C57BL/6 (B6) and DBA/2 (D2), B6D2F1 (F1) mice, is highly resistant to this parasite. In the present study we show by quantitative PCR that the increase of tissue parasitism during the early phase of infection is comparable up to day 11 between susceptible B6 and resistant F1 mice. A reduction of splenic parasite burdens occurs thereafter in both strains but is comparatively retarded in susceptible mice. Splenic microarchitecture is progressively disrupted with loss of follicles and B lymphocytes in B6 mice, but not in F1 mice. By genotyping of additional backcross offspring we corroborate our earlier findings that susceptibility maps to three loci on Chromosomes 5, 13 and 17. Analysis of gene expression of spleen cells from infected B6 and F1 mice with microarrays identifies about 0.3% of transcripts that are differentially expressed. Assuming that differential susceptibility is mediated by altered gene expression, we propose that the following differentially expressed transcripts from these loci are strong candidates for the observed phenotypic variation: H2-Ealpha, H2-D1, Ng23, Msh5 and Tubb5 from Chromosome 17; and Cxcl11, Bmp2k and Spp1 from Chromosome 5. Our results indicate that innate mechanisms are not of primary relevance to resistance of F1 mice to T. cruzi infection, and that differential susceptibility to experimental infection with this protozoan pathogen is not paralleled by extensive variation of the transcriptome.
Asunto(s)
Enfermedad de Chagas/genética , Enfermedad de Chagas/inmunología , Trypanosoma cruzi/inmunología , Trypanosoma cruzi/patogenicidad , Animales , Secuencia de Bases , Enfermedad de Chagas/parasitología , Enfermedad de Chagas/patología , Mapeo Cromosómico , Cartilla de ADN/genética , Modelos Animales de Enfermedad , Expresión Génica , Predisposición Genética a la Enfermedad , Interacciones Huésped-Parásitos/genética , Interacciones Huésped-Parásitos/inmunología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Análisis de Secuencia por Matrices de Oligonucleótidos , Especificidad de la Especie , Bazo/inmunología , Bazo/parasitología , Bazo/patología , Factores de TiempoRESUMEN
The protozoan parasite Trypanosoma cruzi circulates in the blood as trypomastigotes and invades a variety of cells to multiply intracellularly as amastigotes. The acute phase leads to an immune response that restricts the proliferation of the parasite. However, parasites are able to persist in different tissues, which causes the pathology of Chagas' disease. Natural killer (NK) cells play an important role in innate resistance to a variety of pathogens. In the present study we analyzed whether NK cells participated in the control of experimental T. cruzi infection. NK cells were depleted from C57BL/6 mice by antiasialo antibodies. This treatment caused an increased parasitemia during the acute phase, but tissue parasite burdens were not significantly altered according to quantitative real-time PCR. Our results demonstrated that NK cells were activated during the initial phase of a T. cruzi infection and exhibited a contact-dependent antiparasitic activity against extracellular parasites that was independent from perforin. Thus, NK cells limit the propagation of the parasite by acting on circulating T. cruzi trypomastigotes.
Asunto(s)
Enfermedad de Chagas/inmunología , Células Asesinas Naturales/inmunología , Glicoproteínas de Membrana/fisiología , Parasitemia/inmunología , Trypanosoma cruzi/inmunología , Animales , Interleucina-12/biosíntesis , Activación de Linfocitos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Perforina , Proteínas Citotóxicas Formadoras de PorosRESUMEN
Protective immunity to the parasite Trypanosoma cruzi in mice depends on a pro-inflammatory T cell response involving the production of interferon-gamma (IFN-gamma). In conjunction with interleukin-12 (IL-12), IL-18 promotes the synthesis of IFN-gamma and a T helper type 1 immune response. We investigated the requirements of IL-12 and IL-18 in murine T. cruzi infection by use of C57BL/6 mice genetically deficient in either cytokine. IL-12p40(-/-) mice succumbed to infection at doses of 100 parasites, whereas IL-18(-/-) and wild-type mice resisted infectious doses up to 1000 parasites to the same extent. Levels of parasitemia were comparable between the latter groups, as were tissue parasite burdens according to quantitative real-time PCR. In contrast, IL-12p40(-/-) mice displayed vastly increased levels of parasites both in blood and in tissue. IFN-gamma concentrations in the serum of infected mice and in supernatants of splenocytes stimulated in vitro were decreased in IL-18(-/-) mice, whereas in IL-12p40(-/-) mice, IFN-gamma was undetectable in the serum and drastically reduced in cell supernatants. Levels of IL-12 production were generally comparable between wild-type and IL-18(-/-) mice, as were levels of IL-4, IL-2 and nitric oxide. Thus, the requirement for endogenous pro-inflammatory cytokines for a protective murine immune response against T. cruzi is satisfied by the expression of IL-12, while IL-18 is dispensable.
Asunto(s)
Enfermedad de Chagas/inmunología , Interleucina-12/inmunología , Interleucina-18/inmunología , Trypanosoma cruzi/inmunología , Animales , Enfermedad de Chagas/parasitología , Enfermedad de Chagas/patología , Enfermedad de Chagas/fisiopatología , Interferón gamma/biosíntesis , Interferón gamma/sangre , Interleucina-12/biosíntesis , Interleucina-12/sangre , Interleucina-12/deficiencia , Interleucina-18/deficiencia , Interleucina-4/sangre , Hígado/parasitología , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Músculos/parasitología , Óxido Nítrico/sangre , Parasitemia , Bazo/parasitología , Linfocitos T/inmunologíaRESUMEN
The 90-kDa heat shock proteins (HSP90) are important in the regulation of numerous intracellular processes in eukaryotic cells. In particular, HSP90 has been shown to be involved in the control of the cellular differentiation of the protozoan parasite Leishmania donovani. We investigated the role of HSP90 in the related parasite Trypanosoma cruzi by inhibiting its function using geldanamycin (GA). GA induced a dose-dependent increase in heat shock protein levels and a dose-dependent arrest of proliferation. Epimastigotes were arrested in G(1) phase of the cell cycle, but no stage differentiation occurred. Blood form trypomastigotes showed conversion towards spheromastigote-like forms when they were cultivated with GA, but differentiation into epimastigotes was permanently blocked. We conclude that, similar to leishmanial HSP90, functional HSP90 is essential for cell division in T. cruzi and serves as a feedback inhibitor in the cellular stress response. In contrast to L. donovani cells, however, T. cruzi cells treated with GA do not begin to differentiate into relevant life cycle stages.
Asunto(s)
Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Trypanosoma cruzi/metabolismo , Animales , Benzoquinonas , Ciclo Celular , Diferenciación Celular , División Celular , Relación Dosis-Respuesta a Droga , Citometría de Flujo , Técnica del Anticuerpo Fluorescente Indirecta , Fase G1 , Immunoblotting , Lactamas Macrocíclicas , Microscopía Electrónica de Rastreo , Quinonas/farmacología , Factores de TiempoRESUMEN
Cytolytic T lymphocyte-associated Ag-4 (CD152) is a negatively regulating molecule, which is primarily expressed on T cells following their activation. In this study, we have examined the role of CTLA-4 expression in experimental blood-stage malaria. Similar to human malaria, CTLA-4 is expressed on CD4(+) T cells of C57BL/6 mice after infection with Plasmodium berghei. A kinetic analysis revealed that CTLA-4 expression was increased on day 5 postinfection and reached a peak on day 9 postinfection, when almost 10% of splenic CD4(+) T cells expressed CTLA-4. Blockade of CTLA-4 in vivo by a specific mAb and subsequent challenge with P. berghei caused neurological signs reminiscent of murine cerebral malaria and earlier death. Histologic examination of brain sections from anti-CTLA-4-treated mice revealed pathologic changes such as hemorrhages and edema, which were absent in control mice. Furthermore, treatment with anti-CTLA-4 also reversed the extensive loss of CD4(+) T cells and the suppressed T cell response occurring during blood-stage malaria. Our data suggest that CTLA-4 expression prevents immune pathology by restricting T cell activation during malaria. They also indicate that the development of cerebral malaria is mediated by a failure to down-regulate T cell activation.