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1.
Nat Commun ; 12(1): 1399, 2021 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-33658521

RESUMO

Staphylococcus aureus is a prominent human pathogen that readily adapts to host immune defenses. Here, we show that, in contrast to Gram-negative pathogens, S. aureus induces a distinct airway immunometabolic response dominated by the release of the electrophilic metabolite, itaconate. The itaconate synthetic enzyme, IRG1, is activated by host mitochondrial stress, which is induced by staphylococcal glycolysis. Itaconate inhibits S. aureus glycolysis and selects for strains that re-direct carbon flux to fuel extracellular polysaccharide (EPS) synthesis and biofilm formation. Itaconate-adapted strains, as illustrated by S. aureus isolates from chronic airway infection, exhibit decreased glycolytic activity, high EPS production, and proficient biofilm formation even before itaconate stimulation. S. aureus thus adapts to the itaconate-dominated immunometabolic response by producing biofilms, which are associated with chronic infection of the human airway.


Assuntos
Interações Hospedeiro-Patógeno/fisiologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Staphylococcus aureus/patogenicidade , Succinatos/metabolismo , Adulto , Animais , Biofilmes/crescimento & desenvolvimento , Líquido da Lavagem Broncoalveolar , Metabolismo dos Carboidratos , Fibrose Cística/microbiologia , Regulação Bacteriana da Expressão Gênica , Glicólise/efeitos dos fármacos , Glicólise/fisiologia , Interações Hospedeiro-Patógeno/imunologia , Humanos , Hidroliases/metabolismo , Camundongos Endogâmicos C57BL , Infecções por Pseudomonas/imunologia , Infecções por Pseudomonas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Escarro/microbiologia , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/isolamento & purificação , Estresse Fisiológico , Succinatos/farmacologia , Ácido Succínico/metabolismo , Adulto Jovem
2.
Int J Mol Sci ; 22(4)2021 Feb 04.
Artigo em Inglês | MEDLINE | ID: mdl-33557346

RESUMO

Antimicrobial peptides (AMPs) are of interest as alternatives to antibiotics or immunomodulators. We generated and characterized the phenotypes of transgenic mice overexpressing protegrin 1 (PG1), a potent porcine cathelicidin. No obvious differences were observed between PG1 transgenic and wild-type mice in terms of growth, development, general behaviour, and the major immune cell population. However, PG1 transgenic mice intranasally infected with Staphylococcus aureus resulted in a reduction in microscopic pulmonary injury, improved clearance of bacteria, and lower proinflammatory cytokine secretion, compared to those of wild-type mice. On the other hand, approximately 25% of PG1 transgenic mice (n = 54/215) showed corneal opacity and developed inflammation in the eye, resulting ultimately in phthisis bulbi. Immunohistochemical analyses revealed that PG1 and its activator, neutrophil elastase, localized to the basal cells of the cornea and glands in eyelids, respectively. In addition, apoptosis indicated by a Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL)-positive signal was detected from flat cells of the cornea. Our study suggests that the expression regulation or localization of AMPs such as PG1 is important to prevent their adverse effects. However, our results also showed that the cytotoxic effects of PG1 on cells could be tolerated in animals, except for the eyes.


Assuntos
Anti-Infecciosos/metabolismo , Peptídeos Catiônicos Antimicrobianos/fisiologia , Opacidade da Córnea/patologia , Oftalmopatias/patologia , Inflamação/patologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/fisiologia , Animais , Opacidade da Córnea/etiologia , Opacidade da Córnea/metabolismo , Oftalmopatias/etiologia , Oftalmopatias/metabolismo , Feminino , Inflamação/etiologia , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mucina-1/genética , Regiões Promotoras Genéticas , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/patologia , Suínos
3.
Int J Mol Sci ; 21(24)2020 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-33327533

RESUMO

The function of natural killer (NK) cell-derived interferon-γ (IFN-γ) expands to remove pathogens by increasing the ability of innate immune cells. Here, we identified the critical role of thioredoxin-interacting protein (TXNIP) in the production of IFN-γ in NK cells during bacterial infection. TXNIP inhibited the production of IFN-γ and the activation of transforming growth factor ß-activated kinase 1 (TAK1) activity in primary mouse and human NK cells. TXNIP directly interacted with TAK1 and inhibited TAK1 activity by interfering with the complex formation between TAK1 and TAK1 binding protein 1 (TAB1). Txnip-/- (KO) NK cells enhanced the activation of macrophages by inducing IFN-γ production during Pam3CSK4 stimulation or Staphylococcus aureus (S. aureus) infection and contributed to expedite the bacterial clearance. Our findings suggest that NK cell-derived IFN-γ is critical for host defense and that TXNIP plays an important role as an inhibitor of NK cell-mediated macrophage activation by inhibiting the production of IFN-γ during bacterial infection.


Assuntos
Proteínas de Transporte/metabolismo , Interferon gama/metabolismo , Células Matadoras Naturais/metabolismo , Tiorredoxinas/metabolismo , Animais , Proteínas de Transporte/genética , Ensaio de Imunoadsorção Enzimática , Imunidade Inata/efeitos dos fármacos , Imunidade Inata/genética , Células Matadoras Naturais/imunologia , Lipopeptídeos/farmacologia , Camundongos , Camundongos Knockout , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/patogenicidade , Tiorredoxinas/genética , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Fator de Transcrição AP-1/genética , Fator de Transcrição AP-1/metabolismo
4.
Nat Commun ; 11(1): 5431, 2020 10 27.
Artigo em Inglês | MEDLINE | ID: mdl-33110079

RESUMO

Physical forces have profound effects on cellular behavior, physiology, and disease. Perhaps the most intruiguing and fascinating example is the formation of catch-bonds that strengthen cellular adhesion under shear stresses. Today mannose-binding by the Escherichia coli FimH adhesin remains one of the rare microbial catch-bond thoroughly characterized at the molecular level. Here we provide a quantitative demonstration of a catch-bond in living Gram-positive pathogens using force-clamp spectroscopy. We show that the dock, lock, and latch interaction between staphylococcal surface protein SpsD and fibrinogen is strong, and exhibits an unusual catch-slip transition. The bond lifetime first grows with force, but ultimately decreases to behave as a slip bond beyond a critical force (~1 nN) that is orders of magnitude higher than for previously investigated complexes. This catch-bond, never reported for a staphylococcal adhesin, provides the pathogen with a mechanism to tightly control its adhesive function during colonization and infection.


Assuntos
Adesinas Bacterianas/química , Infecções Estafilocócicas/microbiologia , Staphylococcus/metabolismo , Adesinas Bacterianas/genética , Adesinas Bacterianas/metabolismo , Aderência Bacteriana , Fibrinogênio/química , Fibrinogênio/metabolismo , Humanos , Ligação Proteica , Análise Espectral , Infecções Estafilocócicas/metabolismo , Staphylococcus/química , Staphylococcus/genética , Staphylococcus/crescimento & desenvolvimento
5.
PLoS Pathog ; 16(6): e1007806, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32497104

RESUMO

Coagulase-positive staphylococci, which frequently colonize the mucosal surfaces of animals, also cause a spectrum of opportunistic infections including skin and soft tissue infections, urinary tract infections, pneumonia, and bacteremia. However, recent advances in bacterial identification have revealed that these common veterinary pathogens are in fact zoonoses that cause serious infections in human patients. The global spread of multidrug-resistant zoonotic staphylococci, in particular the emergence of methicillin-resistant organisms, is now a serious threat to both animal and human welfare. Accordingly, new therapeutic targets that can be exploited to combat staphylococcal infections are urgently needed. Enzymes of the methylerythritol phosphate pathway (MEP) of isoprenoid biosynthesis represent potential targets for treating zoonotic staphylococci. Here we demonstrate that fosmidomycin (FSM) inhibits the first step of the isoprenoid biosynthetic pathway catalyzed by deoxyxylulose phosphate reductoisomerase (DXR) in staphylococci. In addition, we have both enzymatically and structurally determined the mechanism by which FSM elicits its effect. Using a forward genetic screen, the glycerol-3-phosphate transporter GlpT that facilitates FSM uptake was identified in two zoonotic staphylococci, Staphylococcus schleiferi and Staphylococcus pseudintermedius. A series of lipophilic ester prodrugs (termed MEPicides) structurally related to FSM were synthesized, and data indicate that the presence of the prodrug moiety not only substantially increased potency of the inhibitors against staphylococci but also bypassed the need for GlpT-mediated cellular transport. Collectively, our data indicate that the prodrug MEPicides selectively and robustly inhibit DXR in zoonotic staphylococci, and further, that DXR represents a promising, druggable target for future development.


Assuntos
Antibacterianos , Farmacorresistência Bacteriana Múltipla , Pró-Fármacos , Infecções Estafilocócicas , Staphylococcus , Zoonoses , Animais , Antibacterianos/química , Antibacterianos/farmacologia , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Farmacorresistência Bacteriana Múltipla/genética , Humanos , Pró-Fármacos/química , Pró-Fármacos/farmacologia , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/metabolismo , Staphylococcus/genética , Staphylococcus/crescimento & desenvolvimento , Zoonoses/tratamento farmacológico , Zoonoses/genética , Zoonoses/metabolismo , Zoonoses/microbiologia
6.
PLoS One ; 15(6): e0235006, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32559258

RESUMO

Distal limb wounds are common injuries sustained by horses and their healing is fraught with complications due to equine anatomy, prevalence of infection, and challenges associated with wound management. Gallium is a semi-metallic element that has been shown to possess antimicrobial properties and aid in wound healing in various preclinical models. The effects of Gallium have not been studied in equine wound healing. Therefore, the objective of this study was to compare healing rates between gallium-treated and untreated wounds of equine distal limbs and to demonstrate the antimicrobial effects of gallium on wounds inoculated with S. aureus. Using an established model of equine wound healing we demonstrated beneficial effects of 0.5% topical gallium maltolate on equine wound healing. Specifically we documented reduced healing times, reduced bioburden, and reduced formation of exuberant granulation tissue in wounds treated with gallium maltolate as compared with untreated wounds. Gallium appeared to exert its beneficial effects via its well-described antimicrobial actions as well as by altering the expression of specific genes known to be involved in wound healing of horses and other animals. Specifically, gallium maltolate appeared to increase expression of transforming growth factor-ß in both infected and un-infected wounds. Further work is needed to document the effects of gallium on naturally occurring equine wounds and to compare the effects of gallium with other wound treatment options. These data, however, suggest that gallium may be an attractive and novel means of improving equine distal limb wound healing.


Assuntos
Antibacterianos/uso terapêutico , Doenças dos Cavalos/tratamento farmacológico , Traumatismos da Perna/tratamento farmacológico , Compostos Organometálicos/uso terapêutico , Pironas/uso terapêutico , Infecções Estafilocócicas/tratamento farmacológico , Administração Tópica , Animais , Antibacterianos/administração & dosagem , Carga Bacteriana , Citocinas/genética , Citocinas/metabolismo , Doenças dos Cavalos/metabolismo , Cavalos , Traumatismos da Perna/metabolismo , Traumatismos da Perna/veterinária , Compostos Organometálicos/administração & dosagem , Pironas/administração & dosagem , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/veterinária , Cicatrização
7.
PLoS Pathog ; 16(6): e1008511, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32555671

RESUMO

The clinical importance of microbiomes to the chronicity of wounds is widely appreciated, yet little is understood about patient-specific processes shaping wound microbiome composition. Here, a two-cohort microbiome-genome wide association study is presented through which patient genomic loci associated with chronic wound microbiome diversity were identified. Further investigation revealed that alternative TLN2 and ZNF521 genotypes explained significant inter-patient variation in relative abundance of two key pathogens, Pseudomonas aeruginosa and Staphylococcus epidermidis. Wound diversity was lowest in Pseudomonas aeruginosa infected wounds, and decreasing wound diversity had a significant negative linear relationship with healing rate. In addition to microbiome characteristics, age, diabetic status, and genetic ancestry all significantly influenced healing. Using structural equation modeling to identify common variance among SNPs, six loci were sufficient to explain 53% of variation in wound microbiome diversity, which was a 10% increase over traditional multiple regression. Focusing on TLN2, genotype at rs8031916 explained expression differences of alternative transcripts that differ in inclusion of important focal adhesion binding domains. Such differences are hypothesized to relate to wound microbiomes and healing through effects on bacterial exploitation of focal adhesions and/or cellular migration. Related, other associated loci were functionally enriched, often with roles in cytoskeletal dynamics. This study, being the first to identify patient genetic determinants for wound microbiomes and healing, implicates genetic variation determining cellular adhesion phenotypes as important drivers of infection type. The identification of predictive biomarkers for chronic wound microbiomes may serve as risk factors and guide treatment by informing patient-specific tendencies of infection.


Assuntos
Microbiota , Polimorfismo de Nucleotídeo Único , Infecções por Pseudomonas , Pseudomonas aeruginosa , Infecções Estafilocócicas , Staphylococcus epidermidis , Cicatrização/genética , Infecção dos Ferimentos , Animais , Doença Crônica , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Estudo de Associação Genômica Ampla , Humanos , Masculino , Camundongos , Infecções por Pseudomonas/genética , Infecções por Pseudomonas/metabolismo , Infecções por Pseudomonas/microbiologia , Infecções por Pseudomonas/patologia , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/patologia , Talina/genética , Talina/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Infecção dos Ferimentos/genética , Infecção dos Ferimentos/metabolismo , Infecção dos Ferimentos/microbiologia , Infecção dos Ferimentos/patologia
8.
Science ; 369(6504): 706-712, 2020 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-32527925

RESUMO

Viral infections of the lower respiratory tract are a leading cause of mortality. Mounting evidence indicates that most severe cases are characterized by aberrant immune responses and do not depend on viral burden. In this study, we assessed how type III interferons (IFN-λ) contribute to the pathogenesis induced by RNA viruses. We report that IFN-λ is present in the lower, but not upper, airways of patients with coronavirus disease 2019 (COVID-19). In mice, we demonstrate that IFN-λ produced by lung dendritic cells in response to a synthetic viral RNA induces barrier damage, causing susceptibility to lethal bacterial superinfections. These findings provide a strong rationale for rethinking the pathophysiological role of IFN-λ and its possible use in clinical practice against endemic viruses, such as influenza virus as well as the emerging severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.


Assuntos
Betacoronavirus , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/metabolismo , Células Dendríticas/metabolismo , Interferons/fisiologia , Pulmão/metabolismo , Pulmão/patologia , Pneumonia Viral/imunologia , Pneumonia Viral/metabolismo , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Proliferação de Células , Citocinas/metabolismo , Humanos , Interferon Tipo I/metabolismo , Interferons/metabolismo , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Nasofaringe/imunologia , Pandemias , Poli I-C/administração & dosagem , Mucosa Respiratória/patologia , Transdução de Sinais , Infecções Estafilocócicas/metabolismo , Superinfecção , Receptor 3 Toll-Like/metabolismo
9.
Proc Natl Acad Sci U S A ; 117(20): 10989-10999, 2020 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-32354997

RESUMO

Staphylococcus aureus infections can lead to diseases that range from localized skin abscess to life-threatening toxic shock syndrome. The SrrAB two-component system (TCS) is a global regulator of S. aureus virulence and critical for survival under environmental conditions such as hypoxic, oxidative, and nitrosative stress found at sites of infection. Despite the critical role of SrrAB in S. aureus pathogenicity, the mechanism by which the SrrAB TCS senses and responds to these environmental signals remains unknown. Bioinformatics analysis showed that the SrrB histidine kinase contains several domains, including an extracellular Cache domain and a cytoplasmic HAMP-PAS-DHp-CA region. Here, we show that the PAS domain regulates both kinase and phosphatase enzyme activity of SrrB and present the structure of the DHp-CA catalytic core. Importantly, this structure shows a unique intramolecular cysteine disulfide bond in the ATP-binding domain that significantly affects autophosphorylation kinetics. In vitro data show that the redox state of the disulfide bond affects S. aureus biofilm formation and toxic shock syndrome toxin-1 production. Moreover, with the use of the rabbit infective endocarditis model, we demonstrate that the disulfide bond is a critical regulatory element of SrrB function during S. aureus infection. Our data support a model whereby the disulfide bond and PAS domain of SrrB sense and respond to the cellular redox environment to regulate S. aureus survival and pathogenesis.


Assuntos
Proteínas de Bactérias/metabolismo , Cisteína/metabolismo , Proteínas Repressoras/metabolismo , Staphylococcus aureus/metabolismo , Animais , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Toxinas Bacterianas , Sequência de Bases , Biofilmes , Domínio Catalítico , Modelos Animais de Doenças , Endocardite , Enterotoxinas , Feminino , Regulação Bacteriana da Expressão Gênica , Histidina Quinase/metabolismo , Masculino , Modelos Moleculares , Mutação , Oxirredução , Domínios Proteicos , Coelhos , Proteínas Repressoras/química , Proteínas Repressoras/genética , Sepse , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/genética , Staphylococcus aureus/patogenicidade , Superantígenos , Thermotoga maritima , Virulência/genética , Virulência/fisiologia
10.
PLoS One ; 15(5): e0231168, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32365127

RESUMO

Staphylococcus aureus is one of the most important pathogens causing mastitis in dairy cows. The objective of this study was to establish a rat model of mastitis induced by S. aureus infection and to explore changes in the proteomes of mammary tissue in different udder states, providing a better understanding of the host immune response to S. aureus mastitis. On day 3 post-partum, 6 rats were randomly divided into two groups (n = 3), with either 100 µL of PBS (blank group) or a S. aureus suspension containing 2×107 CFU·mL-1 (challenge group) infused into the mammary gland duct. After 24 h of infection, the rats were sacrificed, and mammary gland tissue was collected. Tandem mass tag (TMT)-based technology was applied to compare the proteomes of healthy and mastitic mammary tissues. Compared with the control group, the challenge group had 555 proteins with significant differences in expression, of which 428 were significantly upregulated (FC>1.2 and p<0.05) and 127 were downregulated (FC>0.83 and p<0.05 or p<0.01). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses revealed that upregulated differentially significant expressed proteins (DSEPs) were associated with mainly immune responses, including integrin alpha M, inter-α-trypsin inhibitor heavy chain 4, and alpha-2-macroglobulin. This study is the first in which a rat model of S. aureus-induced mastitis was used to explore the proteins related to mastitis in dairy cows by TMT technology, providing a model for replication of dairy cow S. aureus-induced mastitis experiments.


Assuntos
Glândulas Mamárias Animais/metabolismo , Mastite/metabolismo , Proteoma/análise , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/fisiologia , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Feminino , Glândulas Mamárias Animais/microbiologia , Glândulas Mamárias Animais/patologia , Mastite/microbiologia , Mastite/patologia , Gravidez , Proteoma/metabolismo , Proteômica/métodos , Ratos , Ratos Sprague-Dawley , Infecções Estafilocócicas/patologia , Espectrometria de Massas em Tandem
11.
J Neuroimmunol ; 344: 577262, 2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32450430

RESUMO

Microglial inflammation plays a pivotal role in the pathogenesis of S. aureus induced brain abscesses. The objective of this study was to regulate microglial activation by the combinatorial treatment of ciprofloxacin either with dexamethasone or celecoxib via targeting M1 and M2 polarization. The antibiotic-immunomodulator combinations were applied either by opening both TLR-2 and GR or neutralizing each of them. Our results confirmed that dexamethasone along with ciprofloxacin attenuated bacterial burden along with ROS production more efficiently than celecoxib combination during TLR-2 neutralization. FACS data indicated microglial M1 to M2 switching that was responsible for the better resolution of microglial inflammation.


Assuntos
Ciprofloxacino/administração & dosagem , Dexametasona/administração & dosagem , Microglia/metabolismo , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus , Receptor 2 Toll-Like/metabolismo , Animais , Antibacterianos/administração & dosagem , Anti-Inflamatórios/administração & dosagem , Células Cultivadas , Quimioterapia Combinada , Masculino , Camundongos , Microglia/efeitos dos fármacos , Infecções Estafilocócicas/tratamento farmacológico , Receptor 2 Toll-Like/antagonistas & inibidores
12.
Proc Natl Acad Sci U S A ; 117(22): 12394-12401, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32414924

RESUMO

The bacterial pathogen Staphylococcus aureus is capable of infecting a broad spectrum of host tissues, in part due to flexibility of metabolic programs. S. aureus, like all organisms, requires essential biosynthetic intermediates to synthesize macromolecules. We therefore sought to determine the metabolic pathways contributing to synthesis of essential precursors during invasive S. aureus infection. We focused specifically on staphylococcal infection of bone, one of the most common sites of invasive S. aureus infection and a unique environment characterized by dynamic substrate accessibility, infection-induced hypoxia, and a metabolic profile skewed toward aerobic glycolysis. Using a murine model of osteomyelitis, we examined survival of S. aureus mutants deficient in central metabolic pathways, including glycolysis, gluconeogenesis, the tricarboxylic acid (TCA) cycle, and amino acid synthesis/catabolism. Despite the high glycolytic demand of skeletal cells, we discovered that S. aureus requires glycolysis for survival in bone. Furthermore, the TCA cycle is dispensable for survival during osteomyelitis, and S. aureus instead has a critical need for anaplerosis. Bacterial synthesis of aspartate in particular is absolutely essential for staphylococcal survival in bone, despite the presence of an aspartate transporter, which we identified as GltT and confirmed biochemically. This dependence on endogenous aspartate synthesis derives from the presence of excess glutamate in infected tissue, which inhibits aspartate acquisition by S. aureus Together, these data elucidate the metabolic pathways required for staphylococcal infection within bone and demonstrate that the host nutrient milieu can determine essentiality of bacterial nutrient biosynthesis pathways despite the presence of dedicated transporters.


Assuntos
Ácido Aspártico/biossíntese , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Nutrientes/metabolismo , Osteomielite/metabolismo , Osteomielite/microbiologia , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/genética
13.
PLoS Pathog ; 16(3): e1008354, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32142554

RESUMO

Biofilm-associated prosthetic joint infections (PJIs) cause significant morbidity due to their recalcitrance to immune-mediated clearance and antibiotics, with Staphylococcus aureus (S. aureus) among the most prevalent pathogens. We previously demonstrated that S. aureus biofilm-associated monocytes are polarized to an anti-inflammatory phenotype and the adoptive transfer of pro-inflammatory macrophages attenuated biofilm burden, highlighting the critical role of monocyte/macrophage inflammatory status in dictating biofilm persistence. The inflammatory properties of leukocytes are linked to their metabolic state, and here we demonstrate that biofilm-associated monocytes exhibit a metabolic bias favoring oxidative phosphorylation (OxPhos) and less aerobic glycolysis to facilitate their anti-inflammatory activity and biofilm persistence. To shift monocyte metabolism in vivo and reprogram cells to a pro-inflammatory state, a nanoparticle approach was utilized to deliver the OxPhos inhibitor oligomycin to monocytes. Using a mouse model of S. aureus PJI, oligomycin nanoparticles were preferentially internalized by monocytes, which significantly reduced S. aureus biofilm burden by altering metabolism and promoting the pro-inflammatory properties of infiltrating monocytes as revealed by metabolomics and RT-qPCR, respectively. Injection of oligomycin alone had no effect on monocyte metabolism or biofilm burden, establishing that intracellular delivery of oligomycin is required to reprogram monocyte metabolic activity and that oligomycin lacks antibacterial activity against S. aureus biofilms. Remarkably, monocyte metabolic reprogramming with oligomycin nanoparticles was effective at clearing established biofilms in combination with systemic antibiotics. These findings suggest that metabolic reprogramming of biofilm-associated monocytes may represent a novel therapeutic approach for PJI.


Assuntos
Biofilmes/efeitos dos fármacos , Reprogramação Celular/efeitos dos fármacos , Implantes Experimentais/microbiologia , Monócitos/metabolismo , Oligomicinas/farmacologia , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/fisiologia , Animais , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Camundongos , Monócitos/patologia , Fosforilação Oxidativa/efeitos dos fármacos , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/patologia
14.
Med Sci Sports Exerc ; 52(7): 1485-1494, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32168105

RESUMO

PURPOSE: To screen for candidate hub genes associated with the effects of exercise on melanoma tumor tissues and to review the potential signaling pathways involved in this process using bioinformatics analysis. METHODS: The GSE62628 expression profile was downloaded from Gene Expression Omnibus database. This data set contains 10 melanoma tumor tissues from two groups of exercise and nonexercise mice. The R software was utilized to identify differentially expressed genes between samples, and functional annotation and pathway analysis were performed. Results were visualized using Cytoscape software. RESULTS: In total, 315 differentially expressed genes were obtained, including 294 upregulated and 21 downregulated genes. The functional analysis showed that these genes were mainly enriched in immune response, inflammatory response, and positive regulation of the ERK1/2 cascade in biological process functional groups. The top 10 candidate hub genes were C3, Kng1, C3ar1, Ptafr, Fgg, Alb, Pf4, Orm1, Aldh3b1, and Apob. The pathway analysis of the most significant module identified from the protein-protein interaction network revealed that the complement and coagulation cascades, Staphylococcus aureus infection, cytokine-cytokine receptor interaction, chemokine signaling pathway and phagosome were mainly involved. C3, C3ar1, Kng1, Ptafr, and Fgg may be the critical genes in the complement and coagulation cascades pathway, and S. aureus in the infection pathway. CONCLUSIONS: Exercise may ameliorate the immune response and inflammatory response in melanoma tissue, and further studies exploring their relationships are warranted.


Assuntos
Regulação Neoplásica da Expressão Gênica , Melanoma Experimental/genética , Condicionamento Físico Animal/fisiologia , Transdução de Sinais , Animais , Coagulação Sanguínea/fisiologia , Quimiocinas/metabolismo , Proteínas do Sistema Complemento/metabolismo , Biologia Computacional , Regulação para Baixo , Feminino , Perfilação da Expressão Gênica , Camundongos , Análise em Microsséries , Domínios e Motivos de Interação entre Proteínas/genética , Mapas de Interação de Proteínas , Receptores de Citocinas/metabolismo , Infecções Estafilocócicas/metabolismo , Regulação para Cima
15.
Exp Eye Res ; 193: 107994, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32147399

RESUMO

Staphylococcus aureus is a common bacterial isolate from cases of microbial keratitis. The virulence factors that contribute to its pathogenicity during this disease have not been fully resolved. The aim of the current study was to examine the effects of the extracellular protease Staphopain A on corneal virulence. Two strains were used, one Staph 38 that gives a high pathology score during keratitis and a less virulent strain ATCC 8325-4. The effect of inhibition of Staphopain by general or specific protease inhibitors on adhesion of strains to fibronectin-coated glass or PMMA was determined. This was followed by an analysis of the effect of Staphopain A on the ability of the bacteria to adhere to and invade corneal epithelial cells. Finally, the effect of inhibiting Staphopain A on pathogenesis in a mouse model of keratitis was studied. Staphopain A increased the adhesion of strains to fibronectin-coated substrata and inhibition of Staphopain A reduced adhesion. The inhibition of Staphopain A by staphostatin A significantly decreased both association with and invasion into human corneal epithelial cells by 15-fold for strain Saur38. Inhibition of Staphopain A significantly reduced the pathology associated with S. aureus keratitis, reducing the infecting numbers of bacteria from 1.8x105 to <1x104 cells/cornea (p ≤ 0.001), significantly reducing the corneal pathology score (p ≤ 0.038) and reducing the numbers of infiltrating PMNs. This study shows that Staphopain increases adhesion and invasion of corneal cells due to increasing fibronectin binding and its inhibition has a significant impact on pathogenicity of S. aureus during keratitis.


Assuntos
Cisteína Endopeptidases/metabolismo , Infecções Oculares Bacterianas/microbiologia , Ceratite/microbiologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/metabolismo , Animais , Modelos Animais de Doenças , Infecções Oculares Bacterianas/metabolismo , Infecções Oculares Bacterianas/patologia , Humanos , Ceratite/metabolismo , Ceratite/patologia , Masculino , Camundongos , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/patologia
16.
Sci Adv ; 6(11): eaax7515, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32195339

RESUMO

Many pathogens produce virulence factors that are specific toward their natural host. Clinically relevant methicillin-resistant Staphylococcus aureus (MRSA) isolates are highly adapted to humans and produce an array of human-specific virulence factors. One such factor is LukAB, a recently identified pore-forming toxin that targets human phagocytes by binding to the integrin component CD11b. LukAB exhibits strong tropism toward human, but not murine, CD11b. Here, phylogenetics and biochemical studies lead to the identification of an 11-residue domain required for the specificity of LukAB toward human CD11b, which is sufficient to render murine CD11b compatible with toxin binding. CRISPR-mediated gene editing was used to replace this domain, resulting in a "humanized" mouse. In vivo studies revealed that the humanized mice exhibit enhanced susceptibility to MRSA bloodstream infection, a phenotype mediated by LukAB. Thus, these studies establish LukAB as an important toxin for MRSA bacteremia and describe a new mouse model to study MRSA pathobiology.


Assuntos
Proteínas de Bactérias/metabolismo , Toxinas Bacterianas/metabolismo , Antígeno CD11b/metabolismo , Leucocidinas/metabolismo , Staphylococcus aureus Resistente à Meticilina/metabolismo , Infecções Estafilocócicas/metabolismo , Fatores de Virulência/metabolismo , Animais , Proteínas de Bactérias/genética , Toxinas Bacterianas/genética , Antígeno CD11b/genética , Células HL-60 , Humanos , Leucocidinas/genética , Staphylococcus aureus Resistente à Meticilina/genética , Camundongos , Camundongos Transgênicos , Infecções Estafilocócicas/genética , Fatores de Virulência/genética
17.
Infect Immun ; 88(5)2020 04 20.
Artigo em Inglês | MEDLINE | ID: mdl-32041788

RESUMO

Implanted medical device-associated infections pose significant health risks, as they are often the result of bacterial biofilm formation. Staphylococcus aureus is a leading cause of biofilm-associated infections which persist due to mechanisms of device surface adhesion, biofilm accumulation, and reprogramming of host innate immune responses. We found that the S. aureus fibronectin binding protein A (FnBPA) is required for normal biofilm development in mammalian serum and that the SaeRS two-component system is required for functional FnBPA activity in serum. Furthermore, serum-developed biofilms deficient in FnBPA were more susceptible to macrophage invasion, and in a model of biofilm-associated implant infection, we found that FnBPA is crucial for the establishment of infection. Together, these findings show that S. aureus FnBPA plays an important role in physical biofilm development and represents a potential therapeutic target for the prevention and treatment of device-associated infections.


Assuntos
Adesinas Bacterianas/metabolismo , Proteínas de Bactérias/metabolismo , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/metabolismo , Staphylococcus aureus/patogenicidade , Animais , Aderência Bacteriana/fisiologia , Biofilmes/crescimento & desenvolvimento , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica/fisiologia
18.
Sci Adv ; 6(4): eaax8820, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-32010784

RESUMO

Extreme pathophysiological stressors induce expansion of otherwise infrequent leukocyte populations. Here, we found a previously unidentified CD11b+Gr-1+ myeloid cell population that expresses stem cell antigen-1 (Sca-1) induced upon experimental infection with Staphylococcus aureus. Although CD11b+Gr-1+Sca-1+ cells have impaired migratory capacity and superoxide anion-producing activity, they secrete increased levels of several cytokines and chemokines compared to Sca-1- counterparts. The generation of CD11b+Gr-1+Sca-1+ cells is dependent on IFN-γ in vivo, and in vitro stimulation of bone marrow cells or granulocyte-macrophage progenitors with IFN-γ generated CD11b+Gr-1+Sca-1+ cells. Depletion of CD11b+Gr-1+Sca-1+ cells by administrating anti-Sca-1 antibody strongly increased survival rates in an S. aureus infection model by reducing organ damage and inflammatory cytokines. However, adoptive transfer of CD11b+Gr-1+Sca-1+ cells decreased survival rates by worsening the pathogenesis of S. aureus infection. Together, we found a previously unidentified pathogenic CD11b+Gr-1+Sca-1+ population that plays an essential role in mortality during bacterial infection.


Assuntos
Infecções Bacterianas/etiologia , Infecções Bacterianas/mortalidade , Biomarcadores , Células Mieloides/imunologia , Transferência Adotiva , Animais , Antígenos Ly/metabolismo , Infecções Bacterianas/metabolismo , Antígeno CD11b/metabolismo , Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Proteínas de Membrana/metabolismo , Camundongos , Células Mieloides/metabolismo , Fenótipo , Prognóstico , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/mortalidade , Staphylococcus aureus/imunologia , Superóxidos/metabolismo
19.
Microb Pathog ; 140: 103963, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31911194

RESUMO

Mitochondria are essential organelles involved in abundant cellular functions ranging from energy metabolism to cell survival. The inhibition of these mitochondrial functions by bacterial toxin proteins promotes disease and inhibits cell growth. Prominent evidence proposes that mitochondria provide a platform for innate immune response signalling pathways. To investigate how a bacterial toxin manipulates the mitochondrial environment of the host Caenorhabditis elegans at the molecular level, a quantitative proteomic study of mitochondria following exposure to the PemKSa toxin was performed. In this study, we purified C. elegans mitochondria and performed a comprehensive proteomic analysis using a shotgun proteomic approach (LC-MS/MS). LC-MS/MS data were analysed using various bioinformatics tools, which revealed the role and involvement of several regulatory proteins and pathways associated with mitochondrial functions. We detected variation in protein expression in key metabolic pathways, including oxidative phosphorylation (OXPHOS), the tricarboxylic acid (TCA) cycle, carbon metabolism, glycolysis and apoptosis, which suggests global reprogramming of host mitochondria metabolism by the toxin. Our results provide new horizons for mitochondria-associated protein functions and the classification of mitochondrial diseases during host-toxin interactions.


Assuntos
Toxinas Bacterianas/toxicidade , Caenorhabditis elegans/metabolismo , Mitocôndrias/metabolismo , Proteoma/metabolismo , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/metabolismo , Animais , Toxinas Bacterianas/metabolismo , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/genética , Caenorhabditis elegans/microbiologia , Cromatografia Líquida , Ciclo do Ácido Cítrico , Modelos Animais de Doenças , Humanos , Mitocôndrias/química , Mitocôndrias/genética , Proteoma/química , Proteoma/genética , Proteômica , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/genética , Espectrometria de Massas em Tandem
20.
Appl Radiat Isot ; 158: 109047, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31989931

RESUMO

The specific uptake of 99mTc radiolabeled Staphylococcus aureus aptamers in the infectious foci was evaluated by scintigraphic imaging of infection-bearing mice. The radiotracer uptake was inhibited by non-radiolabeled aptamers in a competition assay. In addition, when a different number of bacterial cells was used to infect mice an increase in the target/non-target ratios of images correlated with the increase of CFU per gram of tissue was verified. These results confirmed that 99mTc-aptamers were specific to bacterial focus and the level of uptake was dependent on the number of bacterial cells.


Assuntos
Aptâmeros de Nucleotídeos/farmacocinética , Compostos de Organotecnécio/farmacocinética , Infecções Estafilocócicas/diagnóstico por imagem , Staphylococcus aureus/metabolismo , Animais , Contagem de Colônia Microbiana , Eletroforese em Gel de Ágar , Estudos de Avaliação como Assunto , Camundongos , Cintilografia , Infecções Estafilocócicas/metabolismo , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/isolamento & purificação
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