RESUMO
Toxoplasma gondii is an orally acquired pathogen that induces strong IFN-γ based immunity conferring protection but that can also be the cause of immunopathology. The response in mice is driven in part by well-characterized MyD88-dependent signaling pathways. Here we focus on induction of less well understood immune responses that do not involve this Toll-like receptor (TLR)/IL-1 family receptor adaptor molecule, in particular as they occur in the intestinal mucosa. Using eYFP-IL-12p40 reporter mice on an MyD88-/- background, we identified dendritic cells, macrophages, and neutrophils as cellular sources of MyD88-independent IL-12 after peroral T. gondii infection. Infection-induced IL-12 was lower in the absence of MyD88, but was still clearly above noninfected levels. Overall, this carried through to the IFN-γ response, which while generally decreased was still remarkably robust in the absence of MyD88. In the latter mice, IL-12 was strictly required to induce type I immunity. Type 1 and type 3 innate lymphoid cells (ILC), CD4+ T cells, and CD8+ T cells each contributed to the IFN-γ pool. We report that ILC3 were expanded in infected MyD88-/- mice relative to their MyD88+/+ counterparts, suggesting a compensatory response triggered by loss of MyD88. Furthermore, bacterial flagellin and Toxoplasma specific CD4+ T cell populations in the lamina propria expanded in response to infection in both WT and KO mice. Finally, we show that My88-independent IL-12 and T cell mediated IFN-γ production require the presence of the intestinal microbiota. Our results identify MyD88-independent intestinal immune pathways induced by T. gondii including myeloid cell derived IL-12 production, downstream type I immunity and IFN-γ production by ILC1, ILC3, and T lymphocytes. Collectively, our data reveal an underlying network of immune responses that do not involve signaling through MyD88.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Microbioma Gastrointestinal/imunologia , Imunidade nas Mucosas/imunologia , Subunidade p40 da Interleucina-12/imunologia , Toxoplasmose Animal/imunologia , Animais , Mucosa Intestinal/imunologia , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/deficiência , Fator 88 de Diferenciação Mieloide/imunologia , Transdução de Sinais/imunologia , Receptores Toll-Like/deficiência , Receptores Toll-Like/imunologia , Toxoplasma/imunologiaRESUMO
The apicomplexan Toxoplasma gondii induces strong protective immunity dependent upon recognition by Toll-like receptors (TLR)11 and 12 operating in conjunction with MyD88 in the murine host. However, TLR11 and 12 proteins are not present in humans, inspiring us to investigate MyD88-independent pathways of resistance. Using bicistronic IL-12-YFP reporter mice on MyD88+/+ and MyD88-/- genetic backgrounds, we show that CD11c+MHCII+F4/80- dendritic cells, F4/80+ macrophages, and Ly6G+ neutrophils were the dominant cellular sources of IL-12 in both wild type and MyD88 deficient mice after parasite challenge. Parasite dense granule protein GRA24 induces p38 MAPK activation and subsequent IL-12 production in host macrophages. We show that Toxoplasma triggers an early and late p38 MAPK phosphorylation response in MyD88+/+ and MyD88-/- bone marrow-derived macrophages. Using the uracil auxotrophic Type I T. gondii strain cps1-1, we demonstrate that the late response does not require active parasite proliferation, but strictly depends upon GRA24. By i. p. inoculation with cps1-1 and cps1-1:Δgra24, we identified unique subsets of chemokines and cytokines that were up and downregulated by GRA24. Finally, we demonstrate that cps1-1 triggers a strong host-protective GRA24-dependent Th1 response in the absence of MyD88. Our data identify GRA24 as a major mediator of p38 MAPK activation, IL-12 induction and protective immunity that operates independently of the TLR/MyD88 cascade.
Assuntos
Interleucina-12/imunologia , Sistema de Sinalização das MAP Quinases/imunologia , Macrófagos/imunologia , Fator 88 de Diferenciação Mieloide/imunologia , Proteínas de Protozoários/imunologia , Toxoplasma/imunologia , Toxoplasmose/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologia , Animais , Ativação Enzimática/genética , Ativação Enzimática/imunologia , Interleucina-12/genética , Sistema de Sinalização das MAP Quinases/genética , Macrófagos/parasitologia , Macrófagos/patologia , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Proteínas de Protozoários/genética , Toxoplasma/genética , Toxoplasmose/genética , Toxoplasmose/patologia , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
Diet is an important regulator of the gastrointestinal microbiota. Vitamin A and vitamin D deficiencies result in less diverse, dysbiotic microbial communities and increased susceptibility to infection or injury of the gastrointestinal tract. The vitamin A and vitamin D receptors are nuclear receptors expressed by the host, but not the microbiota. Vitamin A- and vitamin D-mediated regulation of the intestinal epithelium and mucosal immune cells underlies the effects of these nutrients on the microbiota. Vitamin A and vitamin D regulate the expression of tight junction proteins on intestinal epithelial cells that are critical for barrier function in the gut. Other shared functions of vitamin A and vitamin D include the support of innate lymphoid cells that produce IL-22, suppression of IFN-γ and IL-17 by T cells, and induction of regulatory T cells in the mucosal tissues. There are some unique functions of vitamin A and D; for example, vitamin A induces gut homing receptors on T cells, while vitamin D suppresses gut homing receptors on T cells. Together, vitamin A- and vitamin D-mediated regulation of the intestinal epithelium and mucosal immune system shape the microbial communities in the gut to maintain homeostasis.
Assuntos
Microbioma Gastrointestinal , Imunidade nas Mucosas , Vitamina A/imunologia , Vitamina D/imunologia , Animais , Homeostase , Humanos , Mucosa Intestinal/imunologia , Mucosa Intestinal/microbiologiaRESUMO
Toxoplasma gondii induces strong IFN-γ-based immunity. Innate lymphoid cells (ILC), in particular ILC1, are an important innate source of this protective cytokine during infection. Our objective was to determine how MyD88-dependent signaling influences ILC function during peroral compared with i.p. infection with T. gondii. MyD88 +/+ and MyD88 -/- mice were orally inoculated with ME49 cysts, and small intestinal lamina propria ILC were assessed using flow cytometry. We observed T-bet+ ILC1, retinoic acid-related orphan receptor γt+ ILC3, and a population of T-bet+retinoic acid-related orphan receptor γt+ double-positive ILC. In MyD88 -/- mice, IFN-γ-producing T-bet+ ILC1 frequencies were reduced compared with wild-type. Treatment of MyD88 -/- mice with an antibiotic mixture to deplete microflora reduced IFN-γ+ ILC1 frequencies. To examine ILC responses outside of the mucosal immune system, peritoneal exudate cells were collected from wild-type and knockout mice after i.p. inoculation with ME49 cysts. In this compartment, ILC were highly polarized to the ILC1 subset that increased significantly and became highly positive for IFN-γ over the course of infection. Increased ILC1 was associated with expression of the Ki67 cell proliferation marker, and the response was driven by IL-12p40. In the absence of MyD88, IFN-γ expression by ILC1 was not maintained, but proliferation remained normal. Collectively, these data reveal new aspects of ILC function that are influenced by location of infection and shaped further by MyD88-dependent signaling.
Assuntos
Microbiota , Toxoplasmose , Animais , Imunidade Inata , Linfócitos , Camundongos , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , TretinoínaRESUMO
Vitamin A (VA) deficiency and diarrheal diseases are both serious public health issues worldwide. VA deficiency is associated with impaired intestinal barrier function and increased risk of mucosal infection-related mortality. The bioactive form of VA, retinoic acid, is a well-known regulator of mucosal integrity. Using Citrobacter rodentium-infected mice as a model for diarrheal diseases in humans, previous studies showed that VA-deficient (VAD) mice failed to clear C. rodentium as compared to their VA-sufficient (VAS) counterparts. However, the distinct intestinal gene responses that are dependent on the host's VA status still need to be discovered. The mRNAs extracted from the small intestine (SI) and the colon were sequenced and analyzed on three levels: differential gene expression, enrichment, and co-expression. C. rodentium infection interacted differentially with VA status to alter colon gene expression. Novel functional categories downregulated by this pathogen were identified, highlighted by genes related to the metabolism of VA, vitamin D, and ion transport, including improper upregulation of Cl- secretion and disrupted HCO3- metabolism. Our results suggest that derangement of micronutrient metabolism and ion transport, together with the compromised immune responses in VAD hosts, may be responsible for the higher mortality to C. rodentium under conditions of inadequate VA.
Assuntos
Infecções por Enterobacteriaceae , Deficiência de Vitamina A , Animais , Citrobacter rodentium , Colo/metabolismo , Diarreia/complicações , Mucosa Intestinal/metabolismo , Intestino Delgado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Vitamina A/metabolismo , Deficiência de Vitamina A/complicaçõesRESUMO
Vitamin A (VA) deficiency remains prevalent in resource limited areas. Using Citrobacter rodentium infection in mice as a model for diarrheal diseases, previous reports showed reduced pathogen clearance and survival due to vitamin A deficient (VAD) status. To characterize the impact of preexisting VA deficiency on gene expression patterns in the intestines, and to discover novel target genes in VA-related biological pathways, VA deficiency in mice were induced by diet. Total mRNAs were extracted from small intestine (SI) and colon, and sequenced. Differentially Expressed Gene (DEG), Gene Ontology (GO) enrichment, and co-expression network analyses were performed. DEGs compared between VAS and VAD groups detected 49 SI and 94 colon genes. By GO information, SI DEGs were significantly enriched in categories relevant to retinoid metabolic process, molecule binding, and immune function. Three co-expression modules showed significant correlation with VA status in SI; these modules contained four known retinoic acid targets. In addition, other SI genes of interest (e.g., Mbl2, Cxcl14, and Nr0b2) in these modules were suggested as new candidate genes regulated by VA. Furthermore, our analysis showed that markers of two cell types in SI, mast cells and Tuft cells, were significantly altered by VA status. In colon, "cell division" was the only enriched category and was negatively associated with VA. Thus, these data suggested that SI and colon have distinct networks under the regulation of dietary VA, and that preexisting VA deficiency could have a significant impact on the host response to a variety of disease conditions.
Assuntos
Colo/metabolismo , Intestino Delgado/metabolismo , RNA-Seq/métodos , Deficiência de Vitamina A/genética , Animais , Citrobacter rodentium , Infecções por Enterobacteriaceae/genética , Infecções por Enterobacteriaceae/microbiologia , Perfilação da Expressão Gênica/métodos , Ontologia Genética , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/genética , Transcriptoma , Tretinoína/metabolismo , Vitamina A/genética , Vitamina A/metabolismoRESUMO
The gastrointestinal tract is a major portal of entry for many pathogens, including the protozoan parasite Toxoplasma gondii. Billions of people worldwide have acquired T. gondii at some point in their life, and for the vast majority this has led to latent infection in the central nervous system. The first line of host defense against Toxoplasma is located within the intestinal mucosa. Appropriate coordination of responses by the intestinal epithelium, intraepithelial lymphocytes, and lamina propria cells results in an inflammatory response that controls acute infection. Under some conditions, infection elicits bacterial dysbiosis and immune-mediated tissue damage in the intestine. Here, we discuss the complex interactions between the microbiota, the epithelium, as well as innate and adaptive immune cells in the intestinal mucosa that induce protective immunity, and that sometimes switch to inflammatory pathology as T. gondii encounters tissues of the gut.
Assuntos
Linfócitos Intraepiteliais , Microbiota , Toxoplasma , Disbiose , Humanos , Mucosa IntestinalRESUMO
Vitamin A deficiency (A-) increases morbidity and mortality to gastrointestinal (GI) infection. Blocking retinoid signaling (dominant negative retinoic acid receptor, dnRAR) in intestinal epithelial cells (IEC, IECdnRAR) had no effect on vitamin A absorption, the expression of tight junction proteins or the integrity of the barrier. Immune cells in the gut were present in normal frequencies in the IECdnRAR mice, with the exception of the T cell receptor (TCR)αß+/CD8αα cells, which were significantly lower than in wildtype littermates. Challenging the IECdnRAR mice with dextran sodium sulfate to induce colitis or Citrobacter rodentium infection resulted in similar disease to wildtype littermates. Feeding mice vitamin A deficient diets reduced vitamin A status and the A- IECdnRAR mice developed more severe colitis and C. rodentium infection. In particular, retinoid signaling in the IEC was crucial for the A- host to survive early infection following C. rodentium. Treating A- mice with retinoic acid (RA) beginning on the day of infection protects most mice from early lethality. However, RA treatment of the A- IECdnRAR mice was ineffective for preventing lethality following C. rodentium infection. Retionid signaling in IEC is critical, especially when there are reduced levels of dietary vitamin A. IEC are direct targets of vitamin A for mounting early defense against infection.
Assuntos
Colite/metabolismo , Colite/mortalidade , Mucosa Intestinal/metabolismo , Receptores do Ácido Retinoico/metabolismo , Retinoides/metabolismo , Transdução de Sinais , Animais , Biomarcadores , Citrobacter rodentium , Colite/etiologia , Colite/patologia , Sulfato de Dextrana/efeitos adversos , Modelos Animais de Doenças , Suscetibilidade a Doenças , Infecções por Enterobacteriaceae/etiologia , Infecções por Enterobacteriaceae/metabolismo , Infecções por Enterobacteriaceae/mortalidade , Infecções por Enterobacteriaceae/patologia , Mucosa Intestinal/microbiologia , Camundongos , Camundongos Knockout , Prognóstico , Receptores do Ácido Retinoico/genética , Índice de Gravidade de Doença , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Vitamin A deficiency affects over 250 million preschool-age children worldwide and is associated with increased childhood mortality and risk of developing enteric infections. Vitamin A deficient (A-) mice developed chronic Citrobacter rodentium infection. A single oral dose of retinoic acid (RA) at d7 post-infection was sufficient to induce clearance of the pathogen in A- mice. RA treatment of A- mice induced il17 expression in the colon. In A- mice, colonic IL-17 was primarily produced by CD11b+ cells; however, in A+ mice, the major source of colonic IL-17 was CD4+ T cells. To determine the cellular targets of vitamin A required for host resistance to C. rodentium, mice that express a dominant negative (dn) retinoic acid receptor (RAR) in T cells (T-dnRAR) or macrophage/neutrophils (LysM-dnRAR) were used. T-dnRAR mice had T cells that produced a robust intestinal IL-17 response and for 40% of the mice was enough to clear the infection. The remainder of the T-dnRAR mice developed a chronic infection. A- LysM-dnRAR mice developed early lethal infections with surviving mice becoming chronically infected. RA treatment of A- LysM-dnRAR mice was ineffective for inducing colonic IL-17 or clearing C. rodentium. Retinoid signaling is required in T cells and CD11b+ cells for complete elimination of enteric pathogens.
Assuntos
Antígeno CD11b/metabolismo , Citrobacter rodentium/efeitos dos fármacos , Infecções por Enterobacteriaceae/tratamento farmacológico , Linfócitos T/metabolismo , Tretinoína/uso terapêutico , Deficiência de Vitamina A/tratamento farmacológico , Análise de Variância , Animais , Citrobacter rodentium/metabolismo , Colo/imunologia , Infecções por Enterobacteriaceae/etiologia , Interleucina-17/metabolismo , Fígado/imunologia , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Tretinoína/metabolismo , Deficiência de Vitamina A/induzido quimicamente , Deficiência de Vitamina A/complicaçõesRESUMO
Neurodegeneration is a critical problem in aging populations and is characterized by severe central nervous system (CNS) inflammation. Macrophages closely regulate inflammation in the CNS and periphery by taking on different activation states. The source of inflammation in many neurodegenerative diseases has been preliminarily linked to a decrease in the CNS M2 macrophage population and a subsequent increase in M1-mediated neuroinflammation. The Recepteur D'Origine Nantais (Ron) is a receptor tyrosine kinase expressed on tissue-resident macrophages including microglia. Activation of Ron by its ligand, macrophage-stimulating protein, attenuates obesity-mediated inflammation in the periphery. An in vivo deletion of the ligand binding domain of Ron (Ron-/-) promotes inflammatory (M1) and limits a reparative (M2) macrophage activation. However, whether or not this response influences CNS inflammation has not been determined. In this study, we demonstrate that in homeostasis Ron-/- mice developed an inflammatory CNS niche with increased tissue expression of M1-associated markers when compared to age-matched wild-type (WT) mice. Baseline metabolic analysis of CNS tissue indicates exacerbated levels of metabolic stress in Ron-/- CNS. In a disease model of multiple sclerosis, experimental autoimmune encephalomyelitis, Ron-/- mice exhibit higher disease severity when compared to WT mice associated with increased CNS tissue inflammation. In a model of diet-induced obesity (DIO), Ron-/- mice exhibit exacerbated CNS inflammation with decreased expression of the M2 marker Arginase-1 (Arg-1) and a robust increase in M1 markers compared to WT mice following 27 weeks of DIO. Collectively, these results illustrate that activation of Ron in the CNS could be a potential therapeutic approach to treating various grades of CNS inflammation underlying neurodegeneration.
Assuntos
Sistema Nervoso Central/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Macrófagos/imunologia , Esclerose Múltipla/metabolismo , Inflamação Neurogênica/metabolismo , Obesidade/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Animais , Diferenciação Celular , Linhagem Celular , Citocinas/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/patologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Esclerose Múltipla/patologia , Neuroproteção , Obesidade/patologia , Receptores Proteína Tirosina Quinases/genética , Células Th1/imunologiaRESUMO
Vitamin D is a direct and indirect regulator of T cells. The mechanisms by which vitamin D directly regulates T cells are reviewed and new primary data on the effects of 1,25 dihydroxyvitamin D (1,25(OH)2D) on human invariant natural killer (iNK)T cells is presented. The in vivo effects of vitamin D on murine T cells include inhibition of T cell proliferation, inhibition of IFN-γ, IL-17 and induction of IL-4. Experiments in mice demonstrate that the effectiveness of 1,25(OH)2D requires NKT cells, IL-10, the IL-10R and IL-4. Comparisons of mouse and human T cells show that 1,25(OH)2D inhibits IL-17 and IFN-γ, and induces T regulatory cells and IL-4. IL-4 was induced by 1,25(OH)2D in mouse and human iNKT cells. Activation for 72 h was required for optimal expression of the vitamin D receptor (VDR) in human and mouse T and iNKT cells. In addition, T cells are potential autocrine sources of 1,25(OH)2D but again only 48-72 h after activation. Together the data support the late effects of vitamin D on diseases like inflammatory bowel disease and multiple sclerosis where reducing IL-17 and IFN-γ, while inducing IL-4 and IL-10, would be beneficial.