RESUMEN
The cynomolgus macaque model of low-dose Mycobacterium tuberculosis infection recapitulates clinical aspects of human tuberculosis pathology, but it is unknown whether the 2 systems are sufficiently similar that host-based signatures of tuberculosis will be predictive across species. By blind prediction, we demonstrate that a subset of genes comprising a human signature for tuberculosis risk is simultaneously predictive in humans and macaques and prospectively discriminates progressor from controller animals 3-6 weeks after infection. Further analysis yielded a 3-gene signature involving PRDX2 that predicts tuberculosis progression in macaques 10 days after challenge, suggesting novel pathways that define protective responses to M. tuberculosis.
Asunto(s)
Macaca fascicularis , Mycobacterium tuberculosis/inmunología , ARN Bacteriano/sangre , Tuberculosis Pulmonar/microbiología , Animales , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Pulmón/patología , Mycobacterium tuberculosis/genética , Tuberculosis Pulmonar/patologíaRESUMEN
Whole blood transcriptional profiling offers great diagnostic and prognostic potential. Although studies identified signatures for pulmonary tuberculosis (TB) and transcripts that predict the risk for developing active TB in humans, the early transcriptional changes immediately following Mycobacterium tuberculosis infection have not been evaluated. We evaluated the gene expression changes in the cynomolgus macaque model of TB, which recapitulates all clinical aspects of human M. tuberculosis infection, using a human microarray and analytics platform. We performed genome-wide blood transcriptional analysis on 38 macaques at 11 postinfection time points during the first 6 mo of M. tuberculosis infection. Of 6371 differentially expressed transcripts between preinfection and postinfection, the greatest change in transcriptional activity occurred 20-56 d postinfection, during which fluctuation of innate and adaptive immune response-related transcripts was observed. Modest transcriptional differences between active TB and latent infection were observed over the time course with substantial overlap. The pattern of module activity previously published for human active TB was similar in macaques with active disease. Blood transcript activity was highly correlated with lung inflammation (lung [18F]fluorodeoxyglucose [FDG] avidity) measured by positron emission tomography and computed tomography at early time points postinfection. The differential signatures between animals with high and low lung FDG were stronger than between clinical outcomes. Analysis of preinfection signatures of macaques revealed that IFN signatures could influence eventual clinical outcomes and lung FDG avidity, even before infection. Our data support that transcriptional changes in the macaque model are translatable to human M. tuberculosis infection and offer important insights into early events of M. tuberculosis infection.
Asunto(s)
Células Sanguíneas/fisiología , Pulmón/diagnóstico por imagen , Macaca fascicularis/inmunología , Mycobacterium tuberculosis/inmunología , Neumonía/inmunología , Transcriptoma/inmunología , Tuberculosis Pulmonar/inmunología , Inmunidad Adaptativa/genética , Animales , Células Cultivadas , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Fluorodesoxiglucosa F18/metabolismo , Perfilación de la Expresión Génica , Humanos , Inmunidad Innata/genética , Pulmón/microbiología , Neumonía/genética , Tomografía de Emisión de Positrones , Tomografía Computarizada por Rayos X , Tuberculosis Pulmonar/genéticaRESUMEN
Despite the availability of annually formulated vaccines, influenza virus infection remains a worldwide public health burden. Therefore, it is important to develop preclinical challenge models that enable the evaluation of vaccine candidates while elucidating mechanisms of protection. Here, we report that naive rhesus macaques challenged with 2009 pandemic H1N1 (pH1N1) influenza virus do not develop observable clinical symptoms of disease but develop a subclinical biphasic fever on days 1 and 5 to 6 postchallenge. Whole blood microarray analysis further revealed that interferon activity was associated with fever. We then tested whether type I interferon activity in the blood is a correlate of vaccine efficacy. The animals immunized with candidate vaccines carrying hemagglutinin (HA) or nucleoprotein (NP) exhibited significantly reduced interferon activity on days 5 to 6 postchallenge. Supported by cellular and serological data, we conclude that blood interferon activity is a prominent marker that provides a convenient metric of influenza virus vaccine efficacy in the subclinical rhesus macaque model.
Asunto(s)
Anticuerpos Antivirales/sangre , Subtipo H1N1 del Virus de la Influenza A/inmunología , Vacunas contra la Influenza/inmunología , Gripe Humana/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Animales , Humanos , Inmunización , Macaca mulatta , VacunaciónRESUMEN
BACKGROUND: HIV-1-infected individuals with higher viral set points progress to AIDS more rapidly than those with lower set points. Predicting viral set point early following infection can contribute to our understanding of early control of HIV-1 replication, to predicting long-term clinical outcomes, and to the choice of optimal therapeutic regimens. METHODS: In a longitudinal study of 10 untreated HIV-1-infected patients, we used gene expression profiling of peripheral blood mononuclear cells to identify transcriptional networks for viral set point prediction. At each sampling time, a statistical analysis inferred the optimal transcriptional network that best predicted viral set point. We then assessed the accuracy of this transcriptional model by predicting viral set point in an independent cohort of 10 untreated HIV-1-infected patients from Malawi. RESULTS: The gene network inferred at time of enrollment predicted viral set point 24 weeks later in the independent Malawian cohort with an accuracy of 87.5%. As expected, the predictive accuracy of the networks inferred at later time points was even greater, exceeding 90% after week 4. The composition of the inferred networks was largely conserved between time points. The 12 genes comprising this dynamic signature of viral set point implicated the involvement of two major canonical pathways: interferon signaling (p<0.0003) and membrane fraction (p<0.02). A silico knockout study showed that HLA-DRB1 and C4BPA may contribute to restricting HIV-1 replication. CONCLUSIONS: Longitudinal gene expression profiling of peripheral blood mononuclear cells from patients with acute HIV-1 infection can be used to create transcriptional network models to early predict viral set point with a high degree of accuracy.
Asunto(s)
Perfilación de la Expresión Génica , Redes Reguladoras de Genes , Infecciones por VIH/diagnóstico , VIH-1 , Análisis de Secuencia por Matrices de Oligonucleótidos , Carga Viral/genética , Simulación por Computador , Progresión de la Enfermedad , Técnicas de Inactivación de Genes , Humanos , Leucocitos Mononucleares , Estudios Longitudinales , Malaui , Valor Predictivo de las Pruebas , Pronóstico , ARN Viral , Análisis de Regresión , Estados UnidosRESUMEN
Human T-lymphotropic virus type 1 (HTLV-1) is a retrovirus that persists lifelong in the host. In â¼4% of infected people, HTLV-1 causes a chronic disabling neuroinflammatory disease known as HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The pathogenesis of HAM/TSP is unknown and treatment remains ineffective. We used gene expression microarrays followed by flow cytometric and functional assays to investigate global changes in blood transcriptional profiles of HTLV-1-infected and seronegative individuals. We found that perturbations of the p53 signaling pathway were a hallmark of HTLV-1 infection. In contrast, a subset of interferon (IFN)-stimulated genes was over-expressed in patients with HAM/TSP but not in asymptomatic HTLV-1 carriers or patients with the clinically similar disease multiple sclerosis. The IFN-inducible signature was present in all circulating leukocytes and its intensity correlated with the clinical severity of HAM/TSP. Leukocytes from patients with HAM/TSP were primed to respond strongly to stimulation with exogenous IFN. However, while type I IFN suppressed expression of the HTLV-1 structural protein Gag it failed to suppress the highly immunogenic viral transcriptional transactivator Tax. We conclude that over-expression of a subset of IFN-stimulated genes in chronic HTLV-1 infection does not constitute an efficient host response but instead contributes to the development of HAM/TSP.
Asunto(s)
Regulación de la Expresión Génica , Virus Linfotrópico T Tipo 1 Humano/metabolismo , Interferón Tipo I/metabolismo , Leucocitos/metabolismo , Paraparesia Espástica Tropical/metabolismo , Transducción de Señal , Proteína p53 Supresora de Tumor/metabolismo , Femenino , Perfilación de la Expresión Génica/métodos , Productos del Gen gag/metabolismo , Productos del Gen tax/metabolismo , Humanos , Leucocitos/virología , Masculino , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Biología de Sistemas/métodosRESUMEN
Dysregulation of professional APC has been postulated as a major mechanism underlying Ag-specific T cell hyporesponsiveness in patients with patent filarial infection. To address the nature of this dysregulation, dendritic cells (DC) and macrophages generated from elutriated monocytes were exposed to live microfilariae (mf), the parasite stage that circulates in blood and is responsible for most immune dysregulation in filarial infections. DC exposed to mf for 24-96 h showed a marked increase in cell death and caspase-positive cells compared with unexposed DC, whereas mf exposure did not induce apoptosis in macrophages. Interestingly, 48-h exposure of DC to mf induced mRNA expression of the proapoptotic gene TRAIL and both mRNA and protein expression of TNF-alpha. mAb to TRAIL-R2, TNF-R1, or TNF-alpha partially reversed mf-induced cell death in DC, as did knocking down the receptor for TRAIL-R2 using small interfering RNA. The mf also induced gene expression of BH3-interacting domain death agonist and protein expression of cytochrome c in DC; mf-induced cleavage of BH3-interacting domain death agonist could be shown to induce release of cytochrome c, leading to activation of caspase 9. Our data suggest that mf induce DC apoptosis in a TRAIL- and TNF-alpha-dependent fashion.
Asunto(s)
Apoptosis/inmunología , Células Dendríticas/inmunología , Filariasis/inmunología , Microfilarias/inmunología , Ligando Inductor de Apoptosis Relacionado con TNF/inmunología , Animales , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/biosíntesis , Brugia Malayi/inmunología , Citocromos c/biosíntesis , Células Dendríticas/metabolismo , Citometría de Flujo , Expresión Génica , Regulación de la Expresión Génica , Humanos , Immunoblotting , Macrófagos/inmunología , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Mensajero/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Bordetella bronchiseptica is a Gram-negative bacterium equipped with several colonization factors that allow it to establish a persistent infection of the murine respiratory tract. Previous studies indicate that B. bronchiseptica adenylate cyclase toxin (ACT) and the type III secretion system (TTSS) synergize to drive dendritic cells into an altered phenotype to down-regulate the host immune response. In this study, we examined the effects of B. bronchiseptica ACT and TTSS on murine bone marrow-derived macrophages. We demonstrate that ACT and TTSS are required for the inhibition of Ag-driven CD4+ T cell proliferation by bacteria-infected macrophages. We identify PGE2 as the mediator of this inhibition, and we show that ACT and the TTSS synergize to increase macrophage production of PGE2. We further demonstrate that B. bronchiseptica can modulate normal macrophage function and drive the immune response toward a Th17 phenotype classified by the significant production of IL-17. In this study, we show that B. bronchiseptica-infected macrophages can induce IL-17 production from naive CD4+ splenocytes, and that lung tissues from B. bronchiseptica-infected mice exhibit a strong Th17 immune response. ACT inhibited surface expression of CD40 and CD86, suppressed TNF-alpha production, and up-regulated IL-6 production. TTSS also synergized with ACT to up-regulate IL-10 and PGE2 secretion. These findings indicate that persistent colonization by B. bronchiseptica may rely on the ability of the bacteria to differentially modulate both macrophage and dendritic cell function leading to an altered adaptive immune response and subsequent bacterial colonization.
Asunto(s)
Bordetella bronchiseptica/inmunología , Linfocitos T CD4-Positivos/citología , Linfocitos T CD4-Positivos/inmunología , Proliferación Celular , Inhibidores de Crecimiento/inmunología , Interleucina-17/biosíntesis , Macrófagos/inmunología , Macrófagos/microbiología , Toxina de Adenilato Ciclasa/farmacología , Animales , Bordetella bronchiseptica/patogenicidad , Linfocitos T CD4-Positivos/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Dinoprostona/biosíntesis , Femenino , Inmunofenotipificación , Interleucina-17/metabolismo , Interleucina-17/fisiología , Activación de Linfocitos/inmunología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Bazo/citología , Bazo/inmunología , Bazo/metabolismoRESUMEN
Chronic bacterial infection reflects a balance between the host immune response and bacterial factors that promote colonization and immune evasion. Bordetella bronchiseptica uses a type III secretion system (TTSS) to persist in the lower respiratory tract of mice. We hypothesize that colonization is facilitated by bacteria-driven modulation of dendritic cells (DCs), which leads to an immunosuppressive adaptive host response. Migration of DCs to the draining lymph nodes of the respiratory tract was significantly increased in mice infected with wild-type B. bronchiseptica compared with mice infected with TTSS mutant bacteria. Reduced colonization by TTSS-deficient bacteria was evident by 7 days after infection, whereas colonization by wild-type bacteria remained high. This decrease in colonization correlated with peak IFN-gamma production by restimulated splenocytes from infected animals. Wild-type bacteria also elicited peak IFN-gamma production on day 7, but the quantity was significantly lower than that elicited by TTSS mutant bacteria. Additionally, wild-type bacteria elicited higher levels of the immunosuppressive cytokine IL-10 compared with the TTSS mutant bacteria. B. bronchiseptica colonization in IL-10(-/-) mice was significantly reduced compared with infections in wild-type mice. These findings suggest that B. bronchiseptica use the TTSS to rapidly drive respiratory DCs to secondary lymphoid tissues where these APCs stimulate an immunosuppressive response characterized by increased IL-10 and decreased IFN-gamma production that favors bacterial persistence.
Asunto(s)
Infecciones por Bordetella/inmunología , Infecciones por Bordetella/microbiología , Bordetella bronchiseptica/inmunología , Movimiento Celular/inmunología , Células Dendríticas/inmunología , Terapia de Inmunosupresión , Animales , Proteínas Bacterianas/fisiología , Infecciones por Bordetella/metabolismo , Bordetella bronchiseptica/crecimiento & desarrollo , Bordetella bronchiseptica/patogenicidad , Células Dendríticas/citología , Regulación hacia Abajo/inmunología , Interferón Tipo I/metabolismo , Interleucina-10/deficiencia , Interleucina-10/genética , Interleucina-10/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Infecciones del Sistema Respiratorio/inmunología , Infecciones del Sistema Respiratorio/metabolismo , Infecciones del Sistema Respiratorio/microbiología , Regulación hacia Arriba/inmunologíaRESUMEN
Bordetella bronchiseptica utilizes a type III secretion system (TTSS) to establish a persistent infection of the murine respiratory tract. Previous studies have shown that the Bordetella TTSS mediated cytotoxicity in different cell types, inhibition of NF-kappaB in epithelial cells, and differentiation of dendritic cells into a semimature state. Here we demonstrate modulation of mitogen-activated protein kinase (MAPK) signaling pathways and altered cytokine production in macrophages and dendritic cells by the Bordetella TTSS. In macrophages, the MAPKs ERK and p38 were downregulated. This resulted in attenuated production of interleukin- (IL-)6 and IL-10. In contrast, the Th-1-polarizing cytokine IL-12 was produced at very low levels and remained unmodulated by the Bordetella TTSS. In dendritic cells, ERK was transiently activated, but this failed to alter cytokine profiles. These results suggest that the Bordetella TTSS modulates antigen-presenting cells in a cell type-specific manner and the secretion of high levels of IL-6 and IL-10 by macrophages might be important for pathogen clearance.
Asunto(s)
Bordetella bronchiseptica/metabolismo , Activación de Macrófagos , Proteínas Quinasas Activadas por Mitógenos/fisiología , Toxina de Adenilato Ciclasa/fisiología , Animales , Bordetella bronchiseptica/inmunología , Células Cultivadas , Regulación hacia Abajo , Interleucina-10/biosíntesis , Interleucina-6/biosíntesis , Sistema de Señalización de MAP Quinasas , Ratones , FosforilaciónRESUMEN
Bordetella bronchiseptica establishes persistent infection of the murine respiratory tract. We hypothesize that long-term colonization is mediated in part by bacteria-driven modulation of dendritic cells (DCs) leading to altered adaptive immune responses. Bone marrow-derived DCs (BMDCs) from C57BL/6 mice infected with live B. bronchiseptica exhibited high surface expression of MHCII, CD86, and CD80. However, B. bronchiseptica-infected BMDCs did not exhibit significant increases in CD40 surface expression and IL-12 secretion compared with BMDCs treated with heat-killed B. bronchiseptica. The B. bronchiseptica type III secretion system (TTSS) mediated the increase in MHCII, CD86, and CD80 surface expression, while the inhibition of CD40 and IL-12 expression was mediated by adenylate cyclase toxin (ACT). IL-6 secretion was independent of the TTSS and ACT. These phenotypic changes may result from differential regulation of MAPK signaling in DCs. Wild-type B. bronchiseptica activated the ERK 1/2 signaling pathway in a TTSS-dependent manner. Additionally, ACT was found to inhibit p38 signaling. These data suggest that B. bronchiseptica drive DC into a semimature phenotype by altering MAPK signaling. These semimature DCs may induce tolerogenic immune responses that allow the persistent colonization of B. bronchiseptica in the host respiratory tract.
Asunto(s)
Toxina de Adenilato Ciclasa/fisiología , Infecciones por Bordetella/inmunología , Bordetella bronchiseptica/inmunología , Células Dendríticas/inmunología , Animales , Antígenos CD/biosíntesis , Antígeno B7-1/biosíntesis , Antígeno B7-2 , Infecciones por Bordetella/microbiología , Bordetella bronchiseptica/fisiología , Antígenos CD40/biosíntesis , Diferenciación Celular/efectos de los fármacos , Células Dendríticas/metabolismo , Células Dendríticas/patología , Antígenos de Histocompatibilidad Clase II/biosíntesis , Tolerancia Inmunológica , Interleucina-12/biosíntesis , Interleucina-12/genética , Interleucina-12/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Glicoproteínas de Membrana/biosíntesis , Ratones , Ratones Endogámicos C57BL , FagocitosisRESUMEN
Virulence factors secreted by Listeria monocytogenes are known to interfere with host cellular signalling pathways. We investigated whether L. monocytogenes modulates T-cell receptor signalling by examining surface expression of proteins known to be upregulated on activated T cells. In vitro culture of murine splenocytes with L. monocytogenes resulted in a specific and dose-dependent upregulation of Fas ligand (FasL). Induction of FasL expression was also observed for pathogenic Listeria ivanovii but not for non-pathogenic Listeria innocua, indicating involvement of Listeria virulence protein(s). Examination of L. monocytogenes strains deficient in different virulence genes demonstrated that FasL upregulation was dependent on the expression of two secreted proteins: listeriolysin O (LLO) and phosphatidylcholine-preferring phospholipase C (PC-PLC). Treatment of cells with purified proteins demonstrated that LLO was sufficient for inducing FasL, while PC-PLC synergized with LLO for the induction of FasL expression. FasL-expressing cells induced by L. monocytogenes were capable of killing Fas-expressing target cells. Furthermore, L. monocytogenes infection results in upregulation of FasL on T cells in mice. These results describe a novel function for LLO and PC-PLC and suggest that L. monocytogenes may use these virulence factors to modulate the host immune response.