RESUMEN
The ability of pathogens to influence host cell survival is a crucial virulence factor. Listeria monocytogenes (Lm) infection is known to be associated with severe apoptosis of hepatocytes and spleen cells. This impairs host defense mechanisms and thereby facilitates the spread of intracellular pathogens. The general mechanisms of apoptosis elicited by Lm infection are understood, however, the roles of BH3-only proteins during primary Lm infection have not been examined. To explore the roles of BH3-only proteins in Lm-induced apoptosis, we studied Listeria infections in mice deficient in Bim, Bid, Noxa or double deficient in BimBid or BimNoxa. We found that BimNoxa double knockout mice were highly resistant to high-dose challenge with Listeria. Decreased bacterial burden and decreased host cell apoptosis were found in the spleens of these mice. The ability of the BH3-deficient mice to clear bacterial infection more efficiently than WT was correlated with increased concentrations of ROS, neutrophil extracellular DNA trap release and downregulation of TNF-α. Our data show a novel pathway of infection-induced apoptosis that enhances our understanding of the mechanism by which BH3-only proteins control apoptotic host cell death during Listeria infection.
Asunto(s)
Apoptosis , Listeria monocytogenes , Listeriosis/etiología , Listeriosis/metabolismo , Proteínas Mitocondriales/metabolismo , Animales , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/deficiencia , Proteína 11 Similar a Bcl2/deficiencia , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Resistencia a la Enfermedad/genética , Resistencia a la Enfermedad/inmunología , Susceptibilidad a Enfermedades , Trampas Extracelulares/inmunología , Femenino , Expresión Génica , Listeriosis/mortalidad , Listeriosis/patología , Masculino , Ratones , Ratones Noqueados , Infiltración Neutrófila/genética , Infiltración Neutrófila/inmunología , Neutrófilos/inmunología , Neutrófilos/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/deficiencia , Especies Reactivas de Oxígeno/metabolismo , Bazo/inmunología , Bazo/metabolismo , Bazo/patología , Tasa de SupervivenciaRESUMEN
Experimental autoimmune myocarditis (EAM) is a CD4(+) T-cell-mediated model of human inflammatory dilated cardiomyopathies. Heart-specific CD4(+) T-cell activation is dependent on autoantigens presented by MHC class II (MHCII) molecules expressed on professional APCs. In this study, we addressed the role of inflammation-induced MHCII expression by cardiac nonhematopoietic cells on EAM development. EAM was induced in susceptible mice lacking inducible expression of MHCII molecules on all nonhematopoietic cells (pIV-/- K14 class II transactivator (CIITA) transgenic (Tg) mice) by immunization with α-myosin heavy chain peptide in CFA. Lack of inducible nonhematopoietic MHCII expression in pIV-/- K14 CIITA Tg mice conferred EAM resistance. In contrast, cardiac pathology was induced in WT and heterozygous mice, and correlated with elevated cardiac endothelial MHCII expression. Control mice with myocarditis displayed an increase in infiltrating CD4(+) T cells and in expression of IFN-γ, which is the major driver of nonhematopoietic MHCII expression. Mechanistically, IFN-γ neutralization in WT mice shortly before disease onset resulted in reduced cardiac MHCII expression and pathology. These findings reveal a previously overlooked contribution of IFN-γ to induce endothelial MHCII expression in the heart and to progress cardiac pathology during myocarditis.
Asunto(s)
Enfermedades Autoinmunes/inmunología , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/inmunología , Miocarditis/inmunología , Animales , Autoantígenos , Linfocitos T CD4-Positivos , Modelos Animales de Enfermedad , Endotelio/inmunología , Inflamación , Interferón gamma/inmunología , Activación de Linfocitos , Ratones , Ratones Transgénicos , Miocardio/patología , Miocardio/ultraestructura , Proteínas Nucleares/genética , Regiones Promotoras Genéticas , Transactivadores/genéticaRESUMEN
Leishmania major infection induces self-healing cutaneous lesions in C57BL/6 mice. Both IL-12 and IFN-γ are essential for the control of infection. We infected Jun dimerization protein p21SNFT (Batf3(-/-) ) mice (C57BL/6 background) that lack the major IL-12 producing and cross-presenting CD8α(+) and CD103(+) DC subsets. Batf3(-/-) mice displayed enhanced susceptibility with larger lesions and higher parasite burden. Additionally, cells from draining lymph nodes of infected Batf3(-/-) mice secreted less IFN-γ, but more Th2- and Th17-type cytokines, mirrored by increased serum IgE and Leishmania-specific immunoglobulin 1 (Th2 indicating). Importantly, CD8α(+) DCs isolated from lymph nodes of L. major-infected mice induced significantly more IFN-γ secretion by L. major-stimulated immune T cells than CD103(+) DCs. We next developed CD11c-diptheria toxin receptor: Batf3(-/-) mixed bone marrow chimeras to determine when the DCs are important for the control of infection. Mice depleted of Batf-3-dependent DCs from day 17 or wild-type mice depleted of cross-presenting DCs from 17-19 days after infection maintained significantly larger lesions similar to mice whose Batf-3-dependent DCs were depleted from the onset of infection. Thus, we have identified a crucial role for Batf-3-dependent DCs in protection against L. major.
Asunto(s)
Presentación de Antígeno , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/inmunología , Reactividad Cruzada , Células Dendríticas/inmunología , Leishmania major/inmunología , Leishmaniasis Cutánea/inmunología , Proteínas Represoras/inmunología , Animales , Anticuerpos Antiprotozoarios/sangre , Anticuerpos Antiprotozoarios/genética , Anticuerpos Antiprotozoarios/inmunología , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/biosíntesis , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Reactividad Cruzada/inmunología , Células Dendríticas/metabolismo , Células Dendríticas/patología , Inmunoglobulina E/sangre , Inmunoglobulina E/genética , Inmunoglobulina E/inmunología , Interferón gamma , Leishmania major/metabolismo , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/genética , Leishmaniasis Cutánea/patología , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas Represoras/biosíntesis , Proteínas Represoras/genética , Células Th2/inmunología , Células Th2/metabolismo , Células Th2/patologíaRESUMEN
Immune responses against intestinal microbiota contribute to the pathogenesis of inflammatory bowel diseases (IBD) and involve CD4(+) T cells, which are activated by major histocompatibility complex class II (MHCII) molecules on antigen-presenting cells (APCs). However, it is largely unexplored how inflammation-induced MHCII expression by intestinal epithelial cells (IEC) affects CD4(+) T cell-mediated immunity or tolerance induction in vivo. Here, we investigated how epithelial MHCII expression is induced and how a deficiency in inducible epithelial MHCII expression alters susceptibility to colitis and the outcome of colon-specific immune responses. Colitis was induced in mice that lacked inducible expression of MHCII molecules on all nonhematopoietic cells, or specifically on IECs, by continuous infection with Helicobacter hepaticus and administration of interleukin (IL)-10 receptor-blocking antibodies (anti-IL10R mAb). To assess the role of interferon (IFN)-γ in inducing epithelial MHCII expression, the T cell adoptive transfer model of colitis was used. Abrogation of MHCII expression by nonhematopoietic cells or IECs induces colitis associated with increased colonic frequencies of innate immune cells and expression of proinflammatory cytokines. CD4(+) T-helper type (Th)1 cells - but not group 3 innate lymphoid cells (ILCs) or Th17 cells - are elevated, resulting in an unfavourably altered ratio between CD4(+) T cells and forkhead box P3 (FoxP3)(+) regulatory T (Treg) cells. IFN-γ produced mainly by CD4(+) T cells is required to upregulate MHCII expression by IECs. These results suggest that, in addition to its proinflammatory roles, IFN-γ exerts a critical anti-inflammatory function in the intestine which protects against colitis by inducing MHCII expression on IECs. This may explain the failure of anti-IFN-γ treatment to induce remission in IBD patients, despite the association of elevated IFN-γ and IBD.
Asunto(s)
Colitis/inmunología , Colitis/prevención & control , Enterocitos/metabolismo , Interferón gamma/metabolismo , Proteínas Nucleares/metabolismo , Transactivadores/metabolismo , Animales , Anticuerpos Monoclonales/farmacología , Quimiocinas/metabolismo , Colitis/patología , Colon/efectos de los fármacos , Colon/inmunología , Colon/patología , Enterocitos/efectos de los fármacos , Enterocitos/patología , Factores de Transcripción Forkhead/metabolismo , Helicobacter/efectos de los fármacos , Helicobacter/fisiología , Infecciones por Helicobacter/inmunología , Inmunidad Innata/efectos de los fármacos , Mediadores de Inflamación/metabolismo , Queratina-14/genética , Recuento de Linfocitos , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Nucleares/genética , Regiones Promotoras Genéticas/genética , Receptores de Interleucina-10/inmunología , Células TH1/efectos de los fármacos , Células TH1/inmunología , Transactivadores/genética , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Research in vitro facilitates discovery, screening, and pilot experiments, often preceding research in vivo. Several technical difficulties render Dendritic Cell (DC) research particularly challenging, including the low frequency of DC in vivo, thorough isolation requirements, and the vulnerability of DC ex vivo. Critically, there is not as yet a widely accepted human or murine DC line and in vitro systems of DC research are limited. In this study, we report the generation of new murine DC lines, named MutuDC, originating from cultures of splenic CD8α conventional DC (cDC) tumors. By direct comparison to normal WT splenic cDC subsets, we describe the phenotypic and functional features of the MutuDC lines and show that they have retained all the major features of their natural counterpart in vivo, the splenic CD8α cDC. These features include expression of surface markers Clec9A, DEC205, and CD24, positive response to TLR3 and TLR9 but not TLR7 stimuli, secretion of cytokines, and chemokines upon activation, as well as cross-presentation capacity. In addition to the close resemblance to normal splenic CD8α cDC, a major advantage is the ease of derivation and maintenance of the MutuDC lines, using standard culture medium and conditions, importantly without adding supplementary growth factors or maturation-inducing stimuli to the medium. Furthermore, genetically modified MutuDC lines have been successfully obtained either by lentiviral transduction or by culture of DC tumors originating from genetically modified mice. In view of the current lack of stable and functional DC lines, these novel murine DC lines have the potential to serve as an important auxiliary tool for DC research.