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1.
Proc Natl Acad Sci U S A ; 121(33): e2406492121, 2024 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-39361877

RESUMO

Toll-like receptors (TLRs) on macrophages sense microbial components and trigger the production of numerous cytokines and chemokines that mediate the inflammatory response to infection. Although many of the components required for the activation of the TLR pathway have been identified, the mechanisms that appropriately regulate the magnitude and duration of the response and ultimately restore homeostasis are less well understood. Furthermore, a growing body of work indicates that TLR signaling reciprocally interacts with other fundamental cellular processes, including lipid metabolism but only a few specific molecular links between immune signaling and the macrophage lipidome have been studied in detail. Oxysterol-binding protein (Osbp) is the founding member of a family of lipid-binding proteins with diverse functions in lipid sensing, lipid transport, and cell signaling but its role in TLR responses is not well defined. Here, we demonstrate that altering the state of Osbp with its natural ligand, 25-hydroxycholesterol (25HC), or pharmacologically, sustains and thereby amplifies Tlr4-induced cytokine production in vitro and in vivo. CRISPR-induced knockdown of Osbp abrogates the ability of these ligands to sustain TLR responses. Lipidomic analysis suggested that the effect of Osbp on TLR signaling may be mediated by alterations in triglyceride production and treating cells with a Dgat1 inhibitor, which blocks triglyceride production and completely abrogates the effect of Osbp on TLR signaling. Thus, Osbp is a sterol sensor that transduces perturbations of the lipidome to modulate the resolution of macrophage inflammatory responses.


Assuntos
Citocinas , Hidroxicolesteróis , Macrófagos , Receptores de Esteroides , Transdução de Sinais , Animais , Macrófagos/metabolismo , Macrófagos/imunologia , Camundongos , Citocinas/metabolismo , Receptores de Esteroides/metabolismo , Receptores de Esteroides/genética , Hidroxicolesteróis/metabolismo , Receptores Toll-Like/metabolismo , Receptor 4 Toll-Like/metabolismo , Camundongos Endogâmicos C57BL , Metabolismo dos Lipídeos , Células RAW 264.7
2.
Proc Natl Acad Sci U S A ; 117(27): 15789-15798, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32581129

RESUMO

Patients infected with influenza are at high risk of secondary bacterial infection, which is a major proximate cause of morbidity and mortality. We have shown that in mice, prior infection with influenza results in increased inflammation and mortality upon Staphylococcus aureus infection, recapitulating the human disease. Lipidomic profiling of the lungs of superinfected mice revealed an increase in CYP450 metabolites during lethal superinfection. These lipids are endogenous ligands for the nuclear receptor PPARα, and we demonstrate that Ppara-/- mice are less susceptible to superinfection than wild-type mice. PPARα is an inhibitor of NFκB activation, and transcriptional profiling of cells isolated by bronchoalveolar lavage confirmed that influenza infection inhibits NFκB, thereby dampening proinflammatory and prosurvival signals. Furthermore, network analysis indicated an increase in necrotic cell death in the lungs of superinfected mice compared to mice infected with S. aureus alone. Consistent with this, we observed reduced NFκB-mediated inflammation and cell survival signaling in cells isolated from the lungs of superinfected mice. The kinase RIPK3 is required to induce necrotic cell death and is strongly induced in cells isolated from the lungs of superinfected mice compared to mice infected with S. aureus alone. Genetic and pharmacological perturbations demonstrated that PPARα mediates RIPK3-dependent necroptosis and that this pathway plays a central role in mortality following superinfection. Thus, we have identified a molecular circuit in which infection with influenza induces CYP450 metabolites that activate PPARα, leading to increased necrotic cell death in the lung which correlates with the excess mortality observed in superinfection.


Assuntos
Inflamação/genética , Influenza Humana/genética , PPAR alfa/genética , Infecções Estafilocócicas/genética , Superinfecção/genética , Animais , Lavagem Broncoalveolar/métodos , Coinfecção/genética , Coinfecção/microbiologia , Coinfecção/mortalidade , Sistema Enzimático do Citocromo P-450/genética , Modelos Animais de Doenças , Suscetibilidade a Doenças , Humanos , Inflamação/microbiologia , Inflamação/mortalidade , Influenza Humana/microbiologia , Influenza Humana/mortalidade , Pulmão/microbiologia , Pulmão/patologia , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Camundongos , Camundongos Knockout , Necroptose/genética , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/mortalidade , Superinfecção/mortalidade
3.
J Infect Dis ; 225(10): 1832-1840, 2022 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-33693706

RESUMO

Previous studies have identified whole-blood transcriptional risk and disease signatures for tuberculosis; however, several lines of evidence suggest that these signatures primarily reflect bacterial burden, which increases before symptomatic disease. We found that the peripheral blood transcriptome of mice with contained Mycobacterium tuberculosis infection (CMTI) has striking similarities to that of humans with active tuberculosis and that a signature derived from these mice predicts human disease with accuracy comparable to that of signatures derived directly from humans. A set of genes associated with immune defense are up-regulated in mice with CMTI but not in humans with active tuberculosis, suggesting that their up-regulation is associated with bacterial containment. A signature comprising these genes predicts both protection from tuberculosis disease and successful treatment at early time points where current signatures are not predictive. These results suggest that detailed study of the CMTI model may enable identification of biomarkers for human tuberculosis.


Assuntos
Mycobacterium tuberculosis , Tuberculose , Animais , Biomarcadores , Humanos , Camundongos , Transcriptoma
4.
PLoS Pathog ; 16(7): e1008655, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32673357

RESUMO

Progress in tuberculosis vaccine development is hampered by an incomplete understanding of the immune mechanisms that protect against infection with Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis. Although the M72/ASOE1 trial yielded encouraging results (54% efficacy in subjects with prior exposure to Mtb), a highly effective vaccine against adult tuberculosis remains elusive. We show that in a mouse model, establishment of a contained and persistent yet non-pathogenic infection with Mtb ("contained Mtb infection", CMTB) rapidly and durably reduces tuberculosis disease burden after re-exposure through aerosol challenge. Protection is associated with elevated activation of alveolar macrophages, the first cells that respond to inhaled Mtb, and accelerated recruitment of Mtb-specific T cells to the lung parenchyma. Systems approaches, as well as ex vivo functional assays and in vivo infection experiments, demonstrate that CMTB reconfigures tissue resident alveolar macrophages via low grade interferon-γ exposure. These studies demonstrate that under certain circumstances, the continuous interaction of the immune system with Mtb is beneficial to the host by maintaining elevated innate immune responses.


Assuntos
Modelos Animais de Doenças , Mycobacterium tuberculosis/imunologia , Vacinas contra a Tuberculose/imunologia , Tuberculose/imunologia , Tuberculose/virologia , Animais , Macrófagos Alveolares/imunologia , Camundongos
5.
EMBO J ; 34(9): 1244-58, 2015 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-25755249

RESUMO

LXR-cofactor complexes activate the gene expression program responsible for cholesterol efflux in macrophages. Inflammation antagonizes this program, resulting in foam cell formation and atherosclerosis; however, the molecular mechanisms underlying this antagonism remain to be fully elucidated. We use promoter enrichment-quantitative mass spectrometry (PE-QMS) to characterize the composition of gene regulatory complexes assembled at the promoter of the lipid transporter Abca1 following downregulation of its expression. We identify a subset of proteins that show LXR ligand- and binding-dependent association with the Abca1 promoter and demonstrate they differentially control Abca1 expression. We determine that NCOA5 is linked to inflammatory Toll-like receptor (TLR) signaling and establish that NCOA5 functions as an LXR corepressor to attenuate Abca1 expression. Importantly, TLR3-LXR signal crosstalk promotes recruitment of NCOA5 to the Abca1 promoter together with loss of RNA polymerase II and reduced cholesterol efflux. Together, these data significantly expand our knowledge of regulatory inputs impinging on the Abca1 promoter and indicate a central role for NCOA5 in mediating crosstalk between pro-inflammatory and anti-inflammatory pathways that results in repression of macrophage cholesterol efflux.


Assuntos
Transportador 1 de Cassete de Ligação de ATP/genética , Colesterol/metabolismo , Macrófagos/metabolismo , Coativadores de Receptor Nuclear/genética , Receptores Nucleares Órfãos/genética , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Animais , Feminino , Regulação da Expressão Gênica , Inflamação/genética , Inflamação/metabolismo , Receptores X do Fígado , Espectrometria de Massas/métodos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Coativadores de Receptor Nuclear/metabolismo , Receptores Nucleares Órfãos/metabolismo , Regiões Promotoras Genéticas , RNA Polimerase II/metabolismo , Transdução de Sinais , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo
6.
Proc Natl Acad Sci U S A ; 111(29): 10666-71, 2014 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-24994901

RESUMO

Cross-talk between sterol regulatory pathways and inflammatory pathways has been demonstrated to significantly impact the development of both atherosclerosis and infectious disease. The oxysterol 25-hydroxycholesterol (25HC) plays multiple roles in lipid biosynthesis and immunity. We recently used a systems biology approach to identify 25HC as an innate immune mediator that had a predicted role in atherosclerosis and we demonstrated a role for 25HC in foam cell formation. Here, we show that this mediator also has several complex roles in the antiviral response. The host response to viruses involves gene regulatory circuits with multiple feedback loops and we show here that 25HC acts as an amplifier of inflammatory signaling in macrophages. We determined that 25HC amplifies inflammatory signaling, at least in part, by mediating the recruitment of the AP-1 components FBJ osteosarcoma oncogene (FOS) and jun proto-oncogene (JUN) to the promoters of a subset of Toll-like receptor-responsive genes. Consistent with previous reports, we found that 25HC inhibits in vitro infection of airway epithelial cells by influenza. Surprisingly, we found that deletion of Ch25h, the gene encoding the enzyme responsible for 25HC production, is protective in a mouse model of influenza infection as a result of decreased inflammatory-induced pathology. Thus, our study demonstrates, for the first time to our knowledge, that in addition to its direct antiviral role, 25HC also regulates transcriptional responses and acts as an amplifier of inflammation via AP-1 and that the resulting alteration in inflammatory response leads to increased tissue damage in mice following infection with influenza.


Assuntos
Hidroxicolesteróis/farmacologia , Inflamação/metabolismo , Inflamação/patologia , Transdução de Sinais/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Retroalimentação Fisiológica/efeitos dos fármacos , Humanos , Influenza Humana/metabolismo , Influenza Humana/patologia , Receptores X do Fígado , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores Nucleares Órfãos/metabolismo , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/patologia , Poli I-C/farmacologia , Proto-Oncogene Mas , Esteroide Hidroxilases/metabolismo , Fator de Transcrição AP-1/metabolismo , Transcrição Gênica/efeitos dos fármacos
7.
PLoS Genet ; 10(12): e1004828, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25474352

RESUMO

We report the first systems biology investigation of regulators controlling arterial plaque macrophage transcriptional changes in response to lipid lowering in vivo in two distinct mouse models of atherosclerosis regression. Transcriptome measurements from plaque macrophages from the Reversa mouse were integrated with measurements from an aortic transplant-based mouse model of plaque regression. Functional relevance of the genes detected as differentially expressed in plaque macrophages in response to lipid lowering in vivo was assessed through analysis of gene functional annotations, overlap with in vitro foam cell studies, and overlap of associated eQTLs with human atherosclerosis/CAD risk SNPs. To identify transcription factors that control plaque macrophage responses to lipid lowering in vivo, we used an integrative strategy--leveraging macrophage epigenomic measurements--to detect enrichment of transcription factor binding sites upstream of genes that are differentially expressed in plaque macrophages during regression. The integrated analysis uncovered eight transcription factor binding site elements that were statistically overrepresented within the 5' regulatory regions of genes that were upregulated in plaque macrophages in the Reversa model under maximal regression conditions and within the 5' regulatory regions of genes that were upregulated in the aortic transplant model during regression. Of these, the TCF/LEF binding site was present in promoters of upregulated genes related to cell motility, suggesting that the canonical Wnt signaling pathway may be activated in plaque macrophages during regression. We validated this network-based prediction by demonstrating that ß-catenin expression is higher in regressing (vs. control group) plaques in both regression models, and we further demonstrated that stimulation of canonical Wnt signaling increases macrophage migration in vitro. These results suggest involvement of canonical Wnt signaling in macrophage emigration from the plaque during lipid lowering-induced regression, and they illustrate the discovery potential of an epigenome-guided, systems approach to understanding atherosclerosis regression.


Assuntos
Hipolipemiantes/uso terapêutico , Macrófagos/metabolismo , Macrófagos/patologia , Placa Aterosclerótica/tratamento farmacológico , Placa Aterosclerótica/genética , Transcriptoma , Via de Sinalização Wnt , Animais , Células Cultivadas , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/fisiologia , Feminino , Perfilação da Expressão Gênica , Genoma/efeitos dos fármacos , Hipolipemiantes/farmacologia , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Análise em Microsséries , Placa Aterosclerótica/metabolismo , Placa Aterosclerótica/patologia , Receptores de LDL/genética , Indução de Remissão , Transcriptoma/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos , Via de Sinalização Wnt/genética
8.
Proc Natl Acad Sci U S A ; 108(28): 11536-41, 2011 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-21709223

RESUMO

Precise control of the innate immune response is essential to ensure host defense against infection while avoiding inflammatory disease. Systems-level analyses of Toll-like receptor (TLR)-stimulated macrophages suggested that SHANK-associated RH domain-interacting protein (SHARPIN) might play a role in the TLR pathway. This hypothesis was supported by the observation that macrophages derived from chronic proliferative dermatitis mutation (cpdm) mice, which harbor a spontaneous null mutation in the Sharpin gene, exhibited impaired IL-12 production in response to TLR activation. Systems biology approaches were used to define the SHARPIN-regulated networks. Promoter analysis identified NF-κB and AP-1 as candidate transcription factors downstream of SHARPIN, and network analysis suggested selective attenuation of these pathways. We found that the effects of SHARPIN deficiency on the TLR2-induced transcriptome were strikingly correlated with the effects of the recently described hypomorphic L153P/panr2 point mutation in Ikbkg [NF-κB Essential Modulator (NEMO)], suggesting that SHARPIN and NEMO interact. We confirmed this interaction by co-immunoprecipitation analysis and furthermore found it to be abrogated by panr2. NEMO-dependent signaling was affected by SHARPIN deficiency in a manner similar to the panr2 mutation, including impaired p105 and ERK phosphorylation and p65 nuclear localization. Interestingly, SHARPIN deficiency had no effect on IκBα degradation and on p38 and JNK phosphorylation. Taken together, these results demonstrate that SHARPIN is an essential adaptor downstream of the branch point defined by the panr2 mutation in NEMO.


Assuntos
Proteínas de Transporte/imunologia , Proteínas de Transporte/metabolismo , Receptor 2 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Animais , Sequência de Bases , Proteínas de Transporte/genética , Primers do DNA/genética , Imunidade Inata/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação , NF-kappa B/metabolismo , Mapeamento de Interação de Proteínas , Transdução de Sinais , Análise de Sistemas , Biologia de Sistemas , Receptor 2 Toll-Like/genética , Fator de Transcrição AP-1/metabolismo
9.
Front Immunol ; 15: 1427846, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39007152

RESUMO

To investigate how host and pathogen diversity govern immunity against Mycobacterium tuberculosis (Mtb), we performed a large-scale screen of vaccine-mediated protection against aerosol Mtb infection using three inbred mouse strains [C57BL/6 (B6), C3HeB/FeJ (C3H), Balb/c x 129/SvJ (C129F1)] and three Mtb strains (H37Rv, CDC1551, SA161) representing two lineages and distinct virulence properties. We compared three protective modalities, all of which involve inoculation with live mycobacteria: Bacillus Calmette-Guérin (BCG), the only approved TB vaccine, delivered either subcutaneously or intravenously, and concomitant Mtb infection (CoMtb), a model of pre-existing immunity in which a low-level Mtb infection is established in the cervical lymph node following intradermal inoculation. We examined lung bacterial burdens at early (Day 28) and late (Day 98) time points after aerosol Mtb challenge and histopathology at Day 98. We observed substantial heterogeneity in the reduction of bacterial load afforded by these modalities at Day 28 across the combinations and noted a strong positive correlation between bacterial burden in unvaccinated mice and the degree of protection afforded by vaccination. Although we observed variation in the degree of reduction in bacterial burdens across the nine mouse/bacterium strain combinations, virtually all protective modalities performed similarly for a given strain-strain combination. We also noted dramatic variation in histopathology changes driven by both host and bacterial genetic backgrounds. Vaccination improved pathology scores for all infections except CDC1551. However, the most dramatic impact of vaccination on lesion development occurred for the C3H-SA161 combination, where vaccination entirely abrogated the development of the large necrotic lesions that arise in unvaccinated mice. In conclusion, we find that substantial TB heterogeneity can be recapitulated by introducing variability in both host and bacterial genetics, resulting in changes in vaccine-mediated protection as measured both by bacterial burden as well as histopathology. These differences can be harnessed in future studies to identify immune correlates of vaccine efficacy.


Assuntos
Mycobacterium tuberculosis , Animais , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Camundongos , Variação Genética , Feminino , Tuberculose/prevenção & controle , Tuberculose/imunologia , Tuberculose/microbiologia , Vacinas contra a Tuberculose/imunologia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos BALB C , Interações Hospedeiro-Patógeno/imunologia , Vacina BCG/imunologia , Pulmão/microbiologia , Pulmão/patologia , Pulmão/imunologia , Modelos Animais de Doenças , Carga Bacteriana , Vacinação
10.
Bioinformatics ; 26(17): 2071-5, 2010 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-20663846

RESUMO

MOTIVATION: Histone acetylation (HAc) is associated with open chromatin, and HAc has been shown to facilitate transcription factor (TF) binding in mammalian cells. In the innate immune system context, epigenetic studies strongly implicate HAc in the transcriptional response of activated macrophages. We hypothesized that using data from large-scale sequencing of a HAc chromatin immunoprecipitation assay (ChIP-Seq) would improve the performance of computational prediction of binding locations of TFs mediating the response to a signaling event, namely, macrophage activation. RESULTS: We tested this hypothesis using a multi-evidence approach for predicting binding sites. As a training/test dataset, we used ChIP-Seq-derived TF binding site locations for five TFs in activated murine macrophages. Our model combined TF binding site motif scanning with evidence from sequence-based sources and from HAc ChIP-Seq data, using a weighted sum of thresholded scores. We find that using HAc data significantly improves the performance of motif-based TF binding site prediction. Furthermore, we find that within regions of high HAc, local minima of the HAc ChIP-Seq signal are particularly strongly correlated with TF binding locations. Our model, using motif scanning and HAc local minima, improves the sensitivity for TF binding site prediction by approximately 50% over a model based on motif scanning alone, at a false positive rate cutoff of 0.01. AVAILABILITY: The data and software source code for model training and validation are freely available online at http://magnet.systemsbiology.net/hac.


Assuntos
Imunoprecipitação da Cromatina/métodos , Ativação de Macrófagos , Fatores de Transcrição/metabolismo , Acetilação , Animais , Sítios de Ligação , Genoma , Histonas/metabolismo , Camundongos , Modelos Biológicos , Software
11.
J Biomed Inform ; 44(1): 75-9, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-20123131

RESUMO

The Cell Ontology (CL) aims for the representation of in vivo and in vitro cell types from all of biology. The CL is a candidate reference ontology of the OBO Foundry and requires extensive revision to bring it up to current standards for biomedical ontologies, both in its structure and its coverage of various subfields of biology. We have now addressed the specific content of one area of the CL, the section of the ontology dealing with hematopoietic cells. This section has been extensively revised to improve its content and eliminate multiple inheritance in the asserted hierarchy, and the groundwork has been laid for structuring the hematopoietic cell type terms as cross-products incorporating logical definitions built from relationships to external ontologies, such as the Protein Ontology and the Gene Ontology. The methods and improvements to the CL in this area represent a paradigm for improvement of the entire ontology over time.


Assuntos
Células Sanguíneas/citologia , Hematopoese , Informática Médica , Vocabulário Controlado , Animais , Humanos
12.
Cell Rep ; 35(9): 109195, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-34077724

RESUMO

Metabolic reprogramming powers and polarizes macrophage functions, but the nature and regulation of this response during infection with pathogens remain controversial. In this study, we characterize the metabolic and transcriptional responses of murine macrophages to Mycobacterium tuberculosis (Mtb) in order to disentangle the underlying mechanisms. We find that type I interferon (IFN) signaling correlates with the decreased glycolysis and mitochondrial damage that is induced by live, but not killed, Mtb. Macrophages lacking the type I IFN receptor (IFNAR) maintain glycolytic flux and mitochondrial function during Mtb infection in vitro and in vivo. IFNß itself restrains the glycolytic shift of inflammatory macrophages and initiates mitochondrial stress. We confirm that type I IFN acts upstream of mitochondrial damage using macrophages lacking the protein STING. We suggest that a type I IFN-mitochondrial feedback loop controls macrophage responses to mycobacteria and that this could contribute to pathogenesis across a range of diseases.


Assuntos
Metabolismo Energético , Interferon Tipo I/metabolismo , Macrófagos/metabolismo , Mycobacterium tuberculosis/fisiologia , Tuberculose/metabolismo , Animais , Glicólise , Temperatura Alta , Proteínas de Membrana , Camundongos , Mitocôndrias/metabolismo , Transdução de Sinais , Estresse Fisiológico , Transcrição Gênica
13.
J Exp Med ; 200(5): 581-6, 2004 Sep 06.
Artigo em Inglês | MEDLINE | ID: mdl-15337791

RESUMO

Macrophages play a critical role in both innate and acquired immunity because of their unique ability to internalize, kill, and degrade bacterial pathogens through the process of phagocytosis. The adaptor protein, amphiphysin IIm, participates in phagocytosis and is transiently associated with early phagosomes. Certain pathogens, including Chlamydia pneumoniae, have evolved mechanisms to subvert macrophage phagosome maturation and, thus, are able to survive within these cells. We report here that, although amphiphysin IIm is usually only transiently associated with the phagosome, it is indefinitely retained on vacuoles containing C. pneumoniae. Under these wild-type conditions, C. pneumoniae do not elicit significant nitric oxide (NO) production and are not killed. Abrogation of amphiphysin IIm function results in C. pneumoniae-induced NO production and in the sterilization of the vacuole. The data suggest that C. pneumoniae retains amphiphysin IIm on the vacuole to survive within the macrophage.


Assuntos
Chlamydophila pneumoniae/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Proteínas do Tecido Nervoso/fisiologia , Animais , Células da Medula Óssea/citologia , Separação Celular , Sobrevivência Celular , Infecções por Chlamydia/patologia , Chlamydophila pneumoniae/patogenicidade , DNA/química , Relação Dose-Resposta a Droga , Citometria de Fluxo , Fluoresceína-5-Isotiocianato , Vetores Genéticos , Camundongos , Microscopia Eletrônica , Microscopia de Fluorescência , Proteínas do Tecido Nervoso/metabolismo , Óxido Nítrico/química , Óxido Nítrico/metabolismo , Nitritos , Fagocitose , Fagossomos/metabolismo , Fatores de Tempo , Transfecção
14.
PLoS Comput Biol ; 4(3): e1000021, 2008 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-18369420

RESUMO

Macrophages are versatile immune cells that can detect a variety of pathogen-associated molecular patterns through their Toll-like receptors (TLRs). In response to microbial challenge, the TLR-stimulated macrophage undergoes an activation program controlled by a dynamically inducible transcriptional regulatory network. Mapping a complex mammalian transcriptional network poses significant challenges and requires the integration of multiple experimental data types. In this work, we inferred a transcriptional network underlying TLR-stimulated murine macrophage activation. Microarray-based expression profiling and transcription factor binding site motif scanning were used to infer a network of associations between transcription factor genes and clusters of co-expressed target genes. The time-lagged correlation was used to analyze temporal expression data in order to identify potential causal influences in the network. A novel statistical test was developed to assess the significance of the time-lagged correlation. Several associations in the resulting inferred network were validated using targeted ChIP-on-chip experiments. The network incorporates known regulators and gives insight into the transcriptional control of macrophage activation. Our analysis identified a novel regulator (TGIF1) that may have a role in macrophage activation.


Assuntos
Ativação de Macrófagos/fisiologia , Macrófagos/fisiologia , Modelos Biológicos , Transdução de Sinais/fisiologia , Receptores Toll-Like/metabolismo , Fatores de Transcrição/fisiologia , Ativação Transcricional/fisiologia , Motivos de Aminoácidos , Animais , Simulação por Computador , Regulação da Expressão Gênica/fisiologia , Humanos , Cinética , Relação Estrutura-Atividade , Integração de Sistemas
15.
Sci Immunol ; 4(37)2019 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-31350281

RESUMO

Alveolar macrophages (AMs) are the first cells to be infected during Mycobacterium tuberculosis (M.tb.) infection. Thus, the AM response to infection is the first of many steps leading to initiation of the adaptive immune response required for efficient control of infection. A hallmark of M.tb. infection is the slow initiation of the adaptive response, yet the mechanisms responsible for this are largely unknown. To study the initial AM response to infection, we developed a system to identify, sort, and analyze M.tb.-infected AMs from the lung within the first 10 days of infection. In contrast to what has been previously described using in vitro systems, M.tb.-infected AMs up-regulate a cell-protective antioxidant transcriptional signature that is dependent on the lung environment but not bacterial virulence. Computational approaches including pathway analysis and transcription factor motif enrichment analysis identify NRF2 as a master regulator of the response. Using knockout mouse models, we demonstrate that NRF2 drives expression of the cell-protective signature in AMs and impairs the control of early bacterial growth. AMs up-regulate a substantial pro-inflammatory response to M.tb. infection only 10 days after infection, yet comparisons with bystander AMs from the same infected animals demonstrate that M.tb.-infected AMs generate a less robust inflammatory response than the uninfected cells around them. Our findings demonstrate that the initial macrophage response to M.tb. in the lung is far less inflammatory than has previously been described by in vitro systems and may impede the overall host response to infection.


Assuntos
Macrófagos Alveolares/imunologia , Macrófagos Alveolares/metabolismo , Mycobacterium tuberculosis/imunologia , Fator 2 Relacionado a NF-E2/metabolismo , Transcrição Gênica , Tuberculose Pulmonar/genética , Tuberculose Pulmonar/imunologia , Animais , Feminino , Macrófagos Alveolares/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia
16.
J Exp Med ; 209(4): 807-17, 2012 Apr 09.
Artigo em Inglês | MEDLINE | ID: mdl-22473958

RESUMO

Atherosclerosis is a chronic inflammatory disease characterized by the accumulation of lipid-loaded macrophages in the arterial wall. We demonstrate that macrophage lipid body formation can be induced by modified lipoproteins or by inflammatory Toll-like receptor agonists. We used an unbiased approach to study the overlap in these pathways to identify regulators that control foam cell formation and atherogenesis. An analysis method integrating epigenomic and transcriptomic datasets with a transcription factor (TF) binding site prediction algorithm suggested that the TF ATF3 may regulate macrophage foam cell formation. Indeed, we found that deletion of this TF results in increased lipid body accumulation, and that ATF3 directly regulates transcription of the gene encoding cholesterol 25-hydroxylase. We further showed that production of 25-hydroxycholesterol (25-HC) promotes macrophage foam cell formation. Finally, deletion of ATF3 in Apoe(-/-) mice led to in vivo increases in foam cell formation, aortic 25-HC levels, and disease progression. These results define a previously unknown role for ATF3 in controlling macrophage lipid metabolism and demonstrate that ATF3 is a key intersection point for lipid metabolic and inflammatory pathways in these cells.


Assuntos
Fator 3 Ativador da Transcrição/fisiologia , Aterosclerose/prevenção & controle , Hidroxicolesteróis/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Animais , Apolipoproteínas E/fisiologia , Células Cultivadas , Feminino , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regiões Promotoras Genéticas , Esteroide Hidroxilases/genética
17.
EMBO Mol Med ; 2(3): 79-89, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20201031

RESUMO

Atherosclerosis, a chronic inflammatory disease of the vascular system, presents significant challenges to developing effective molecular diagnostics and novel therapies. A systems biology approach integrating data from large-scale measurements (e.g. transcriptomics, proteomics and genomics) is successfully contributing to deciphering regulatory networks underlying the response of many different cellular systems to perturbations. Such a network analysis strategy using pathway information and data from multiple measurement platforms, tissues and species is a promising approach to elucidate the mechanistic underpinnings of complex diseases. Here, we present our views on the contributions that a systems approach can bring to the study of atherosclerosis, propose ways to tackle the complexity of the disease in a systems manner and review recent systems-level studies of the disease.


Assuntos
Aterosclerose/patologia , Aterosclerose/fisiopatologia , Regulação da Expressão Gênica , Biologia de Sistemas/métodos , Aterosclerose/diagnóstico , Aterosclerose/terapia , Perfilação da Expressão Gênica , Genômica , Humanos , Modelos Biológicos , Proteômica
18.
J Immunol ; 172(12): 7377-84, 2004 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-15187114

RESUMO

The 3-hydroxyl-3-methylglutaryl-coenzyme A reductase inhibitors, or statins, are a widely used class of drugs for cholesterol reduction. The reduction in mortality and morbidity in statin-treated patients is incompletely explained by their effects on cholesterol, and an anti-inflammatory role for the drug has been proposed. We report in this work that, unexpectedly, simvastatin enhances LPS-induced IL-12p40 production by murine macrophages, and that it does so by activating the IL-12p40 promoter. Mutational analysis and dominant-negative expression studies indicate that both C/EBP and AP-1 transcription factors have a crucial role in promoter activation. This occurs via a c-Fos- and c-Jun-based mechanism; we demonstrate that ectopic expression of c-Jun activates the IL-12p40 promoter, whereas expression of c-Fos inhibits IL-12p40 promoter activity. Simvastatin prevents LPS-induced c-Fos expression, thereby relieving the inhibitory effect of c-Fos on the IL-12p40 promoter. Concomitantly, simvastatin induces the phosphorylation of c-Jun by the c-Jun N-terminal kinase, resulting in c-Jun-dependent activation of the IL-12p40 promoter. This appears to be a general mechanism because simvastatin also augments LPS-dependent activation of the TNF-alpha promoter, perhaps because the TNF-alpha promoter has C/EBP and AP-1 binding sites in a similar configuration to the IL-12p40 promoter. The fact that simvastatin potently augments LPS-induced IL-12p40 and TNF-alpha production has implications for the treatment of bacterial infections in statin-treated patients.


Assuntos
Regulação da Expressão Gênica/imunologia , Inflamação/induzido quimicamente , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Sinvastatina/farmacologia , Fatores de Transcrição/genética , Adjuvantes Imunológicos/farmacologia , Animais , Proteínas Estimuladoras de Ligação a CCAAT/fisiologia , Sinergismo Farmacológico , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-12/biossíntese , Subunidade p40 da Interleucina-12 , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Regiões Promotoras Genéticas/efeitos dos fármacos , Subunidades Proteicas/biossíntese , Proteínas Proto-Oncogênicas c-fos/genética , Proteínas Proto-Oncogênicas c-fos/fisiologia , Proteínas Proto-Oncogênicas c-jun/genética , Proteínas Proto-Oncogênicas c-jun/fisiologia , Fator de Transcrição AP-1/fisiologia , Fatores de Transcrição/fisiologia
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