RESUMO
Staphylococcus aureus bacteremia (SaB) causes significant disease in humans, carrying mortality rates of â¼25%. The ability to rapidly predict SaB patient responses and guide personalized treatment regimens could reduce mortality. Here, we present a resource of SaB prognostic biomarkers. Integrating proteomic and metabolomic techniques enabled the identification of >10,000 features from >200 serum samples collected upon clinical presentation. We interrogated the complexity of serum using multiple computational strategies, which provided a comprehensive view of the early host response to infection. Our biomarkers exceed the predictive capabilities of those previously reported, particularly when used in combination. Last, we validated the biological contribution of mortality-associated pathways using a murine model of SaB. Our findings represent a starting point for the development of a prognostic test for identifying high-risk patients at a time early enough to trigger intensive monitoring and interventions.
Assuntos
Bacteriemia/sangue , Bacteriemia/mortalidade , Infecções Estafilocócicas/sangue , Infecções Estafilocócicas/mortalidade , Staphylococcus aureus/patogenicidade , Animais , Bacteriemia/metabolismo , Biomarcadores/sangue , Biomarcadores/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Metabolômica/métodos , Camundongos , Pessoa de Meia-Idade , Prognóstico , Proteômica/métodos , Fatores de Risco , Infecções Estafilocócicas/metabolismoRESUMO
Pseudomonas aeruginosa (PA) CbpD belongs to the lytic polysaccharide monooxygenases (LPMOs), a family of enzymes that cleave chitin or related polysaccharides. Here, we demonstrate a virulence role of CbpD in PA pneumonia linked to impairment of host complement function and opsonophagocytic clearance. Following intratracheal challenge, a PA ΔCbpD mutant was more easily cleared and produced less mortality than the wild-type parent strain. The x-ray crystal structure of the CbpD LPMO domain was solved to subatomic resolution (0.75Å) and its two additional domains modeled by small-angle X-ray scattering and Alphafold2 machine-learning algorithms, allowing structure-based immune epitope mapping. Immunization of naive mice with recombinant CbpD generated high IgG antibody titers that promoted human neutrophil opsonophagocytic killing, neutralized enzymatic activity, and protected against lethal PA pneumonia and sepsis. IgG antibodies generated against full-length CbpD or its noncatalytic M2+CBM73 domains were opsonic and protective, even in previously PA-exposed mice, while antibodies targeting the AA10 domain were not. Preexisting antibodies in PA-colonized cystic fibrosis patients primarily target the CbpD AA10 catalytic domain. Further exploration of LPMO family proteins, present across many clinically important and antibiotic-resistant human pathogens, may yield novel and effective vaccine antigens.
Assuntos
Oxigenases de Função Mista , Pneumonia , Humanos , Camundongos , Animais , Oxigenases de Função Mista/metabolismo , Pseudomonas aeruginosa/metabolismo , Polissacarídeos/metabolismo , ImunizaçãoRESUMO
Acne vulgaris, rosacea, and hidradenitis suppurativa are enduring inflammatory skin conditions that frequently manifest with akin clinical attributes, posing a considerable challenge for their distinctive diagnosis. While these conditions do exhibit certain resemblances, they also demonstrate distinct underlying pathophysiological mechanisms and treatment modalities. Delving into both the molecular parallels and disparities among these three disorders can yield invaluable insights for refined diagnostics, effective management, and targeted therapeutic interventions. In this report, we present a comparative analysis of transcriptomic data across these three diseases, elucidating differentially expressed genes and enriched pathways specific to each ailment, as well as those shared among them. Specifically, we identified multiple zinc-binding proteins (SERPINA1, S100A7, S100A8, S100A9 and KRT16) as consistently highly upregulated genes across all three diseases. Our hypothesis suggests that these proteins could bind and sequester zinc, potentially leading to localized zinc deficiency and heightened inflammation. We identified high-dose dietary zinc as a promising therapeutic approach and confirmed its effectiveness through validation in an acne mouse model.
Assuntos
Acne Vulgar , Perfilação da Expressão Gênica , Hidradenite Supurativa , Rosácea , Zinco , Acne Vulgar/tratamento farmacológico , Acne Vulgar/genética , Zinco/uso terapêutico , Zinco/metabolismo , Rosácea/tratamento farmacológico , Rosácea/genética , Hidradenite Supurativa/tratamento farmacológico , Hidradenite Supurativa/genética , Animais , Camundongos , Humanos , Proteína A7 Ligante de Cálcio S100/metabolismo , Proteína A7 Ligante de Cálcio S100/genética , Calgranulina A/genética , Calgranulina A/metabolismo , Calgranulina B/genética , Calgranulina B/metabolismo , Transcriptoma , Proteínas S100/genética , Proteínas S100/metabolismo , Proteínas de Transporte/genética , Proteínas de Transporte/metabolismo , Modelos Animais de Doenças , Regulação para CimaRESUMO
Staphylococcus aureus (S. aureus) is one of the most common bacterial infections worldwide, and antibiotic resistant strains such as Methicillin-Resistant S. aureus (MRSA) are a major threat and burden to public health. MRSA not only infects immunocompromised patients but also healthy individuals and has rapidly spread from the healthcare setting to the outside community. However, all vaccines tested in clinical trials to date have failed. Immunocompromised individuals such as patients with HIV or decreased levels of CD4+ T cells are highly susceptible to S. aureus infections, and they are also at increased risk of developing fungal infections. We therefore wondered whether stimulation of antifungal immunity might promote the type of immune responses needed for effective host defense against S. aureus. Here we show that vaccination of mice with a fungal ß-glucan particle (GP) loaded with S. aureus antigens provides protective immunity to S. aureus. We generated glucan particles loaded with the four S. aureus proteins ClfA, IsdA, MntC, and SdrE, creating the 4X-SA-GP vaccine. Vaccination of mice with three doses of 4X-SA-GP promoted protection in a systemic model of S. aureus infection with a significant reduction in the bacterial burden in the spleen and kidneys. 4X-SA-GP vaccination induced antigen-specific Th1 and Th17 CD4+ T cell and antibody responses and provided long-term protection. This work suggests that the GP vaccine system has potential as a novel approach to developing vaccines for S. aureus.
Assuntos
Saccharomyces cerevisiae/imunologia , Infecções Estafilocócicas/imunologia , Vacinas Antiestafilocócicas/imunologia , Staphylococcus aureus/imunologia , Animais , Anticorpos Antibacterianos/imunologia , Coagulase/administração & dosagem , Coagulase/genética , Coagulase/imunologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Saccharomyces cerevisiae/química , Infecções Estafilocócicas/microbiologia , Vacinas Antiestafilocócicas/administração & dosagem , Vacinas Antiestafilocócicas/genética , Staphylococcus aureus/genética , Células Th1/imunologia , Células Th17/imunologia , Vacinação , beta-Glucanas/administração & dosagem , beta-Glucanas/imunologiaRESUMO
A successful Staphylococcus aureus vaccine remains elusive, and one controversy in the field is whether humans generate a protective adaptive immune response to infection. We developed a bacterial challenge murine assay that directly assesses the protective capacity of adoptively transferred human serum samples. We first validated the model by showing that postpneumococcal vaccine serum samples from humans induced effective clearance of Streptococcus pneumoniae in mice. We then found that human serum samples adoptively transferred from children with invasive S. aureus infections exhibited protection from disease in a murine model, with some samples conferring near complete protection. These findings demonstrate that human serum samples are capable of conferring a protective adaptive response generated by humans during invasive staphylococcal disease, allowing for the study of protective factors in a murine model. Identification of the protective factors present in the most efficacious serum samples would be of high interest as potential staphylococcal vaccine candidates or passive therapeutics.
Assuntos
Transferência Adotiva , Anticorpos Antibacterianos/imunologia , Sepse , Infecções Estafilocócicas , Animais , Criança , Modelos Animais de Doenças , Humanos , Camundongos , Sepse/prevenção & controle , Infecções Estafilocócicas/prevenção & controle , Staphylococcus aureusRESUMO
Angiotensin-converting enzyme (ACE) can hydrolyze many peptides and plays a central role in controlling blood pressure. Moreover, ACE overexpression in monocytes and macrophages increases resistance of mice to tumor growth. ACE is composed of two independent catalytic domains. Here, to investigate the specific role of each domain in tumor resistance, we overexpressed either WT ACE (Tg-ACE mice) or ACE lacking N- or C-domain catalytic activity (Tg-NKO and Tg-CKO mice) in the myeloid cells of mice. Tg-ACE and Tg-NKO mice exhibited strongly suppressed growth of B16-F10 melanoma because of increased ACE expression in macrophages, whereas Tg-CKO mice resisted melanoma no better than WT animals. The effect of ACE overexpression reverted to that of the WT enzyme with an ACE inhibitor but not with an angiotensin II type 1 (AT1) receptor antagonist. ACE C-domain overexpression in macrophages drove them toward a pronounced M1 phenotype upon tumor stimulation, with increased activation of NF-κB and signal transducer and activator of transcription 1 (STAT1) and decreased STAT3 and STAT6 activation. Tumor necrosis factor α (TNFα) is important for M1 activation, and TNFα blockade reverted Tg-NKO macrophages to a WT phenotype. Increased ACE C-domain expression increased the levels of reactive oxygen species (ROS) and of the transcription factor C/EBPß in macrophages, important stimuli for TNFα expression, and decreased expression of several M2 markers, including interleukin-4Rα. Natural ACE C-domain-specific substrates are not well-described, and we propose that the peptide(s) responsible for the striking ACE-mediated enhancement of myeloid function are substrates/products of the ACE C-domain.
Assuntos
Polaridade Celular , Macrófagos/citologia , Melanoma Experimental/patologia , Peptidil Dipeptidase A/metabolismo , Animais , Catálise , Linhagem Celular Tumoral , Sobrevivência Celular , Regulação Neoplásica da Expressão Gênica , Macrófagos/imunologia , Melanoma Experimental/enzimologia , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Camundongos , Camundongos Transgênicos , NF-kappa B/metabolismo , Peptidil Dipeptidase A/química , Fator de Transcrição STAT1/metabolismo , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Staphyloxanthin, a carotenoid in S. aureus, is a powerful antioxidant against oxidative stresses. The crtOPQMN operon driving pigment synthesis is under the control of σB . CspA, a cold shock protein, is known to control σB activity. To ascertain genes that regulate cspA, we screened a transposon library that exhibited reduced cspA expression and pigmentation. We found that the adaptor protein YjbH activates cspA expression. Spx, the redox-sensitive transcriptional regulator and a proteolytic target for YjbH and ClpXP, complexes with αCTD of RNAP prior to binding the cspA promoter to repress cspA activity. Increased cspA expression in trans in the inactive spx C10A mutant of JE2 did not enhance pigment production while it did in JE2, suggesting that cspA is downstream to Spx in pigmentation control. As the staphyloxanthin pigment is critical to S. aureus survival in human hosts, we demonstrated that the cspA and yjbH mutants survived less well than the parent in whole blood killing assay. Collectively, our studies suggest a pathway wherein YjbH and ClpXP proteolytically cleave Spx, a repressor of cspA transcription, to affect σB -dependent carotenoid expression, thus providing a critical link between intracellular redox sensing by Spx and carotenoid production to improve S. aureus survival during infections.
Assuntos
Proteínas de Bactérias/metabolismo , Carotenoides/metabolismo , Proteínas e Peptídeos de Choque Frio/metabolismo , Regulação Bacteriana da Expressão Gênica , Fator sigma/metabolismo , Staphylococcus aureus/metabolismo , Proteínas de Bactérias/genética , Proteínas e Peptídeos de Choque Frio/genética , Endopeptidase Clp/genética , Endopeptidase Clp/metabolismo , Óperon , Oxirredução , Proteólise , Fator sigma/genética , Staphylococcus aureus/genéticaRESUMO
The intracellular redox environment of Staphylococcus aureus is mainly buffered by bacillithiol (BSH), a low molecular weight thiol. The identity of enzymes responsible for the recycling of oxidized bacillithiol disulfide (BSSB) to the reduced form (BSH) remains elusive. We examined YpdA, a putative bacillithiol reductase, for its role in maintaining intracellular redox homeostasis. The ypdA mutant showed increased levels of BSSB and a lower bacillithiol redox ratio vs. the isogenic parent, indicating a higher level of oxidative stress within the bacterial cytosol. We showed that YpdA consumed NAD(P)H; and YpdA protein levels were augmented in response to stress. Wild type strains overexpressing YpdA showed increased tolerance to oxidants and electrophilic agents. Importantly, YpdA overexpression in the parental strain caused an increase in BSH levels accompanied by a decrease in BSSB concentration in the presence of stress, resulting in an increase in bacillithiol redox ratio vs. the vector control. Additionally, the ypdA mutant exhibited decreased survival in human neutrophils (PMNs) as compared with the parent, while YpdA overexpression protected the resulting strain from oxidative stress in vitro and from killing by human neutrophils ex vivo. Taken together, these data present a new role for YpdA in S. aureus physiology and virulence through the bacillithiol system.
Assuntos
Proteínas de Bactérias/metabolismo , Proteínas Quinases/metabolismo , Staphylococcus aureus/enzimologia , Staphylococcus aureus/patogenicidade , Células Cultivadas , Homeostase , Humanos , Mutação , Neutrófilos/microbiologia , Oxirredução , Proteínas Quinases/genética , Staphylococcus aureus/genética , VirulênciaRESUMO
Hyper-immunoglobulin E syndrome (HIES) is a primary immunodeficiency disease characterized by recurrent Staphylococcus aureus (S. aureus) infections, eczema, skeletal abnormalities and high titers of serum immunoglobulin E. Although the genetic basis of HIES was not known for almost a half century, HIES most frequently exhibits autosomal dominant trait that is transmitted with variable expressivity. Careful genetic studies in recent years identified dominant-negative mutations in human signal transducer and activator of transcription 3 (STAT3) gene as the cause of sporadic and dominant forms of HIES. The STAT3 mutations were localized to DNA-binding, SRC homology 2 (SH2) and transactivating domains and disrupted T helper 17 (TH17) cell differentiation and downstream expression of TH17 cytokines IL-17 and IL-22. Deficiency of IL-17 and IL-22 in turn is responsible for suboptimal expression of anti-staphylococcal host factors, such as neutrophil-recruiting chemokines and antimicrobial peptides, by human keratinocytes and bronchial epithelial cells. TH17 cytokines deficiency thereby explains the recurrent staphylococcal lung and skin infections of HIES patients.
Assuntos
Suscetibilidade a Doenças , Síndrome de Job/complicações , Infecções Estafilocócicas/etiologia , Staphylococcus aureus/imunologia , Animais , Biomarcadores , Citocinas/metabolismo , Dermatite/diagnóstico , Dermatite/etiologia , Suscetibilidade a Doenças/imunologia , Interações Hospedeiro-Patógeno , Humanos , Síndrome de Job/diagnóstico , Síndrome de Job/etiologia , Síndrome de Job/metabolismo , Proteínas Citotóxicas Formadoras de Poros/biossíntese , Fator de Transcrição STAT3 , Infecções Estafilocócicas/diagnóstico , Células Th17/imunologia , Células Th17/metabolismoRESUMO
Angiotensin-converting enzyme (ACE) inhibitors are widely used to reduce blood pressure. Here, we examined if an ACE is important for the antibacterial effectiveness of neutrophils. ACE knockout mice or mice treated with an ACE inhibitor were more susceptible to bacterial infection by methicillin-resistant Staphylococcus aureus (MRSA). In contrast, mice overexpressing ACE in neutrophils (NeuACE mice) have increased resistance to MRSA and better in vitro killing of MRSA, Pseudomonas aeruginosa, and Klebsiella pneumoniae ACE overexpression increased neutrophil production of reactive oxygen species (ROS) following MRSA challenge, an effect independent of the angiotensin II AT1 receptor. Specifically, as compared with wild-type (WT) mice, there was a marked increase of superoxide generation (>twofold, P < .0005) in NeuACE neutrophils following infection, whereas ACE knockout neutrophils decreased superoxide production. Analysis of membrane p47-phox and p67-phox indicates that ACE increases reduced NAD phosphate oxidase activity but does not increase expression of these subunits. Increased ROS generation mediates the enhanced bacterial resistance of NeuACE mice because the enhanced resistance is lost with DPI (an inhibitor of ROS production by flavoenzymes) inhibition. NeuACE granulocytes also have increased neutrophil extracellular trap formation and interleukin-1ß release in response to MRSA. In a mouse model of chemotherapy-induced neutrophil depletion, transfusion of ACE-overexpressing neutrophils was superior to WT neutrophils in treating MRSA infection. These data indicate a previously unknown function of ACE in neutrophil antibacterial defenses and suggest caution in the treatment of certain individuals with ACE inhibitors. ACE overexpression in neutrophils may be useful in boosting the immune response to antibiotic-resistant bacterial infection.
Assuntos
Resistência à Doença/genética , Imunidade Inata , Neutrófilos/imunologia , Peptidil Dipeptidase A/imunologia , Infecções Estafilocócicas/imunologia , Superóxidos/imunologia , Animais , Membrana Celular , Armadilhas Extracelulares/imunologia , Feminino , Regulação da Expressão Gênica , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Klebsiella pneumoniae , Masculino , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , Staphylococcus aureus Resistente à Meticilina/imunologia , Camundongos , Camundongos Knockout , NADPH Oxidases/genética , NADPH Oxidases/imunologia , Neutrófilos/citologia , Neutrófilos/transplante , Peptidil Dipeptidase A/deficiência , Peptidil Dipeptidase A/genética , Fosfoproteínas/genética , Fosfoproteínas/imunologia , Pseudomonas aeruginosa , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/imunologia , Transdução de Sinais , Infecções Estafilocócicas/enzimologia , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/prevenção & controle , Superóxidos/metabolismoRESUMO
Type I IFNs are a cytokine family essential for antiviral defense. More recently, type I IFNs were shown to be important during bacterial infections. In this article, we show that, in addition to known cytokine functions, IFN-ß is antimicrobial. Parts of the IFN-ß molecular surface (especially helix 4) are cationic and amphipathic, both classic characteristics of antimicrobial peptides, and we observed that IFN-ß can directly kill Staphylococcus aureus Further, a mutant S. aureus that is more sensitive to antimicrobial peptides was killed more efficiently by IFN-ß than was the wild-type S. aureus, and immunoblotting showed that IFN-ß interacts with the bacterial cell surface. To determine whether specific parts of IFN-ß are antimicrobial, we synthesized IFN-ß helix 4 and found that it is sufficient to permeate model prokaryotic membranes using synchrotron x-ray diffraction and that it is sufficient to kill S. aureus These results suggest that, in addition to its well-known signaling activity, IFN-ß may be directly antimicrobial and be part of a growing family of cytokines and chemokines, called kinocidins, that also have antimicrobial properties.
Assuntos
Antibacterianos/farmacologia , Membrana Celular/efeitos dos fármacos , Interferon beta/fisiologia , Staphylococcus aureus/efeitos dos fármacos , Animais , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Catiônicos Antimicrobianos/farmacologia , Humanos , Interferon beta/química , Interferon beta/metabolismo , Interferon beta/farmacologia , Camundongos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/genética , Difração de Raios XRESUMO
Staphylococcus aureus is a leading cause of skin and soft-tissue infections worldwide. Mice are the most commonly used animals for modeling human staphylococcal infections. However a supra-physiologic S. aureus inoculum is required to establish gross murine skin pathology. Moreover, many staphylococcal factors, including Panton-Valentine leukocidin (PVL) elaborated by community-associated methicillin-resistant S. aureus (CA-MRSA), exhibit selective human tropism and cannot be adequately studied in mice. To overcome these deficiencies, we investigated S. aureus infection in non-obese diabetic (NOD)/severe combined immune deficiency (SCID)/IL2rγnull (NSG) mice engrafted with human CD34+ umbilical cord blood cells. These "humanized" NSG mice require one to two log lower inoculum to induce consistent skin lesions compared with control mice, and exhibit larger cutaneous lesions upon infection with PVL+ versus isogenic PVL- S. aureus. Neutrophils appear important for PVL pathology as adoptive transfer of human neutrophils alone to NSG mice was sufficient to induce dermonecrosis following challenge with PVL+ S. aureus but not PVL- S. aureus. PMX53, a human C5aR inhibitor, blocked PVL-induced cellular cytotoxicity in vitro and reduced the size difference of lesions induced by the PVL+ and PVL- S. aureus, but PMX53 also reduced recruitment of neutrophils and exacerbated the infection. Overall, our findings establish humanized mice as an important translational tool for the study of S. aureus infection and provide strong evidence that PVL is a human virulence factor.
Assuntos
Toxinas Bacterianas/farmacologia , Suscetibilidade a Doenças/imunologia , Exotoxinas/farmacologia , Leucocidinas/farmacologia , Infecções Cutâneas Estafilocócicas/microbiologia , Staphylococcus aureus , Animais , Modelos Animais de Doenças , Humanos , Camundongos , Infecções Cutâneas Estafilocócicas/tratamento farmacológicoRESUMO
In Staphylococcus aureus, the low-molecular-weight thiol called bacillithiol (BSH), together with cognate S-transferases, is believed to be the counterpart to the glutathione system of other organisms. To explore the physiological role of BSH in S. aureus, we constructed mutants with the deletion of bshA (sa1291), which encodes the glycosyltransferase that catalyzes the first step of BSH biosynthesis, and fosB (sa2124), which encodes a BSH-S-transferase that confers fosfomycin resistance, in several S. aureus strains, including clinical isolates. Mutation of fosB or bshA caused a 16- to 60-fold reduction in fosfomycin resistance in these S. aureus strains. High-pressure liquid chromatography analysis, which quantified thiol extracts, revealed some variability in the amounts of BSH present across S. aureus strains. Deletion of fosB led to a decrease in BSH levels. The fosB and bshA mutants of strain COL and a USA300 isolate, upon further characterization, were found to be sensitive to H2O2 and exhibited decreased NADPH levels compared with those in the isogenic parents. Microarray analyses of COL and the isogenic bshA mutant revealed increased expression of genes involved in staphyloxanthin synthesis in the bshA mutant relative to that in COL under thiol stress conditions. However, the bshA mutant of COL demonstrated decreased survival compared to that of the parent in human whole-blood survival assays; likewise, the naturally BSH-deficient strain SH1000 survived less well than its BSH-producing isogenic counterpart. Thus, the survival of S. aureus under oxidative stress is facilitated by BSH, possibly via a FosB-mediated mechanism, independently of its capability to produce staphyloxanthin.
Assuntos
Proteínas de Bactérias/fisiologia , Cisteína/análogos & derivados , Glucosamina/análogos & derivados , Estresse Oxidativo/fisiologia , Staphylococcus aureus/metabolismo , Amidoidrolases/deficiência , Análise de Variância , Antibacterianos/farmacologia , Proteínas de Bactérias/genética , Cisteína/genética , Cisteína/fisiologia , Glucosamina/genética , Glucosamina/fisiologia , Glicosiltransferases/genética , Peróxido de Hidrogênio/farmacologia , Análise em Microsséries , Testes de Sensibilidade Microbiana , Mutação , NADP/metabolismo , Peroxidase/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/genética , Staphylococcus aureus/crescimento & desenvolvimento , Xantofilas/biossínteseRESUMO
We have synthesized 39 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] analogs having two side chains attached to carbon-20 (Gemini) with various modifications and compared their anticancer activities. Five structure-function rules emerged to identify analogs with enhanced anticancer activity. One of these active analogs, BXL-01-0126, was more potent than 1,25(OH)2D3 in mediating 50% clonal inhibition of cancer cell growth. Murine studies found that BXL-01-0126 and 1,25(OH)2D3 had nearly the same potency to raise serum calcium levels. Taken together, BXL-01-0126 when compared to 1,25(OH)2D3 has greater anticancer potency, but similar toxicity causing hypercalcemia. We focused on the effect of these compounds on the stimulation of expression of human cathelicidin antimicrobial peptide (CAMP) whose gene has a vitamin D response element in its promoter. Expression of CAMP mRNA and protein increased in a dose-response fashion after exposure of acute myeloid leukemia (AML) cells to the Gemini analog, BXL-01-126, in vitro. A xenograft model of AML was developed using U937 AML cells injected into NSG-immunodeficient mice. Administration of vitamin D3 compounds to these mice resulted in substantial levels of CAMP in the systemic circulation. This suggests a unique prophylactic treatment at diagnosis or during induction chemotherapy for AML patients to provide them with protection against various microbial infections through CAMP induction.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Calcitriol/análogos & derivados , Catelicidinas/biossíntese , Colecalciferol/farmacologia , Animais , Peptídeos Catiônicos Antimicrobianos , Antineoplásicos/química , Calcitriol/síntese química , Calcitriol/química , Calcitriol/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Colecalciferol/análogos & derivados , Colecalciferol/síntese química , Citometria de Fluxo , Xenoenxertos , Humanos , Camundongos , Reação em Cadeia da Polimerase em Tempo Real , Relação Estrutura-AtividadeRESUMO
Several groups have shown that detection of microbial components by TLRs on hematopoietic stem and progenitor cells (HSPCs) instructs myeloid cell generation, raising interest in the possibility of targeting TLRs on HSPCs to boost myelopoiesis. However, although "TLR-derived" cells exhibit myeloid cell characteristics (phagocytosis, cytokine production, antigen presentation), it is not clear whether they are functionally equivalent to macrophages derived in the absence of TLR activation. Our in vitro and in vivo studies show that macrophages derived from mouse and human HSPC subsets (including stem cells) exposed to a TLR2 agonist prior to or during macrophage differentiation produce lower levels of inflammatory cytokines (TNF-α, IL-6, and IL-1ß) and reactive oxygen species. This is in contrast to prior exposure of differentiated macrophages to the TLR2 agonist ("tolerance"), which suppresses inflammatory cytokine production, but elevates reactive oxygen species. Soluble factors produced following exposure of HSPCs to a TLR2 agonist can also act in a paracrine manner to influence the function of macrophages derived from unexposed HSPCs. Our data demonstrate that macrophage function can be influenced by TLR signaling in the HSPCs from which they are derived, and that this may impact the clinical utility of targeting TLRs on HSPCs to boost myelopoiesis.
Assuntos
Células-Tronco Hematopoéticas/metabolismo , Macrófagos/metabolismo , Receptor 2 Toll-Like/agonistas , Animais , Diferenciação Celular , Células Cultivadas , Células-Tronco Hematopoéticas/efeitos dos fármacos , Interleucina-1beta/biossíntese , Interleucina-6/biossíntese , Lipopeptídeos/farmacologia , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Mieloides , Mielopoese , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/imunologia , Fator de Necrose Tumoral alfa/biossínteseRESUMO
Group B Streptococcus (GBS) is a leading cause of invasive bacterial infections in human newborns and immune-compromised adults. The pore-forming toxin (PFT) ß hemolysin/cytolysin (ßh/c) is a major virulence factor for GBS, which is generally attributed to its cytolytic functions. Here we show ßh/c has immunomodulatory properties on macrophages at sub-lytic concentrations. ßh/c-mediated activation of p38 MAPK drives expression of the anti-inflammatory and immunosuppressive cytokine IL-10, and inhibits both IL-12 and NOS2 expression in GBS-infected macrophages, which are critical factors in host defense. Isogenic mutant bacteria lacking ßh/c fail to activate p38-mediated IL-10 production in macrophages and promote increased IL-12 and NOS2 expression. Furthermore, targeted deletion of p38 in macrophages increases resistance to invasive GBS infection in mice, associated with impaired IL-10 induction and increased IL-12 production in vivo. These data suggest p38 MAPK activation by ßh/c contributes to evasion of host defense through induction of IL-10 expression and inhibition of macrophage activation, a new mechanism of action for a PFT and a novel anti-inflammatory role for p38 in the pathogenesis of invasive bacterial infection. Our studies suggest p38 MAPK may represent a new therapeutic target to blunt virulence and improve clinical outcome of invasive GBS infection.
Assuntos
Proteínas de Bactérias/metabolismo , Proteínas Hemolisinas/metabolismo , Interleucina-10/biossíntese , Macrófagos/imunologia , Macrófagos/microbiologia , Infecções Estreptocócicas/imunologia , Streptococcus agalactiae/patogenicidade , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Imunidade Inata , Interleucina-12/biossíntese , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Ativação de Macrófagos , Macrófagos/metabolismo , Camundongos , Óxido Nítrico Sintase/biossíntese , Infecções Estreptocócicas/microbiologia , Streptococcus agalactiae/imunologia , Streptococcus agalactiae/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/genéticaRESUMO
The importance of type I IFNs in the host response to viral infection is well established; however, their role in bacterial infection is not fully understood. Several bacteria (both Gram-positive and -negative) have been shown to induce IFN-ß production in myeloid cells, but this IFN-ß is not always beneficial to the host. We examined whether Staphylococcus aureus induces IFN-ß from myeloid phagocytes, and if so, whether it is helpful or harmful to the host to do so. We found that S. aureus poorly induces IFN-ß production compared with other bacteria. S. aureus is highly resistant to degradation in the phagosome because it is resistant to lysozyme. Using a mutant that is more sensitive to lysozyme, we show that phagosomal degradation and release of intracellular ligands is essential for induction of IFN-ß and inflammatory chemokines downstream of IFN-ß. Further, we found that adding exogenous IFN-ß during S. aureus infection (in vitro and in vivo) was protective. Together, the data demonstrate that failure to induce IFN-ß production during S. aureus infection contributes to pathogenicity.
Assuntos
Interferon beta , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/microbiologia , Staphylococcus aureus/imunologia , Staphylococcus aureus/patogenicidade , Animais , Células Cultivadas , Modelos Animais de Doenças , Humanos , Interferon beta/antagonistas & inibidores , Interferon beta/biossíntese , Interferon beta/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mutação/imunologia , Infecções Estafilocócicas/sangue , Staphylococcus aureus/genéticaRESUMO
Vaccination arguably remains the only long-term strategy to limit the spread of S. aureus infections and its related antibiotic resistance. To date, however, all staphylococcal vaccines tested in clinical trials have failed. In this review, we propose that the failure of S. aureus vaccines is intricately linked to prior host exposure to S. aureus and the pathogen's capacity to evade adaptive immune defenses. We suggest that non-protective immune imprints created by previous exposure to S. aureus are preferentially recalled by SA vaccines, and IL-10 induced by S. aureus plays a unique role in shaping these non-protective anti-staphylococcal immune responses. We discuss how S. aureus modifies the host immune landscape, which thereby necessitates alternative approaches to develop successful staphylococcal vaccines.
RESUMO
The failure of the Staphylococcus aureus (SA) IsdB vaccine trial can be explained by the recall of non-protective immune imprints from prior SA exposure. Here, we investigate natural human SA humoral imprints to understand their broader impact on SA immunizations. We show that antibody responses against SA cell-wall-associated antigens (CWAs) are non-opsonic, while antibodies against SA toxins are neutralizing. Importantly, the protective characteristics of the antibody imprints accurately predict the failure of corresponding vaccines against CWAs and support vaccination against toxins. In passive immunization platforms, natural anti-SA human antibodies reduce the efficacy of the human monoclonal antibodies suvratoxumab and tefibazumab, consistent with the results of their respective clinical trials. Strikingly, in the absence of specific humoral memory responses, active immunizations are efficacious in both naive and SA-experienced mice. Overall, our study points to a practical and predictive approach to evaluate and develop SA vaccines based on pre-existing humoral imprint characteristics.
Assuntos
Staphylococcus aureus Resistente à Meticilina , Vacinas Antiestafilocócicas , Vacinas , Animais , Humanos , Camundongos , Imunização , Staphylococcus aureus , Ensaios Clínicos como AssuntoRESUMO
IMPORTANCE: The prevalence of multidrug-resistant Staphylococcus aureus is of global concern, and vaccines are urgently needed. The iron-regulated surface determinant protein B (IsdB) of S. aureus was investigated as a vaccine candidate because of its essential role in bacterial iron acquisition but failed in clinical trials despite strong immunogenicity. Here, we reveal an unexpected second function for IsdB in pathogen-host interaction: the bacterial fitness factor IsdB triggers a strong inflammatory response in innate immune cells via Toll-like receptor 4 and the inflammasome, thus acting as a novel pathogen-associated molecular pattern of S. aureus. Our discovery contributes to a better understanding of how S. aureus modulates the immune response, which is necessary for vaccine development against the sophisticated pathogen.