RESUMO
Systemic candidiasis is a common, high-mortality, nosocomial fungal infection. Unexpectedly, it has emerged as a complication of anti-complement C5-targeted monoclonal antibody treatment, indicating a critical niche for C5 in antifungal immunity. We identified transcription of complement system genes as the top biological pathway induced in candidemic patients and as predictive of candidemia. Mechanistically, C5a-C5aR1 promoted fungal clearance and host survival in a mouse model of systemic candidiasis by stimulating phagocyte effector function and ERK- and AKT-dependent survival in infected tissues. C5ar1 ablation rewired macrophage metabolism downstream of mTOR, promoting their apoptosis and enhancing mortality through kidney injury. Besides hepatocyte-derived C5, local C5 produced intrinsically by phagocytes provided a key substrate for antifungal protection. Lower serum C5a concentrations or a C5 polymorphism that decreases leukocyte C5 expression correlated independently with poor patient outcomes. Thus, local, phagocyte-derived C5 production licenses phagocyte antimicrobial function and confers innate protection during systemic fungal infection.
Assuntos
Antifúngicos , Candidíase , Animais , Camundongos , Complemento C5/metabolismo , Fagócitos/metabolismoRESUMO
BACKGROUND: Autoimmune polyendocrine syndrome type 1 (APS-1) is a life-threatening, autosomal recessive syndrome caused by autoimmune regulator (AIRE) deficiency. In APS-1, self-reactive T cells escape thymic negative selection, infiltrate organs, and drive autoimmune injury. The effector mechanisms governing T-cell-mediated damage in APS-1 remain poorly understood. METHODS: We examined whether APS-1 could be classified as a disease mediated by interferon-γ. We first assessed patients with APS-1 who were participating in a prospective natural history study and evaluated mRNA and protein expression in blood and tissues. We then examined the pathogenic role of interferon-γ using Aire-/-Ifng-/- mice and Aire-/- mice treated with the Janus kinase (JAK) inhibitor ruxolitinib. On the basis of our findings, we used ruxolitinib to treat five patients with APS-1 and assessed clinical, immunologic, histologic, transcriptional, and autoantibody responses. RESULTS: Patients with APS-1 had enhanced interferon-γ responses in blood and in all examined autoimmunity-affected tissues. Aire-/- mice had selectively increased interferon-γ production by T cells and enhanced interferon-γ, phosphorylated signal transducer and activator of transcription 1 (pSTAT1), and CXCL9 signals in multiple organs. Ifng ablation or ruxolitinib-induced JAK-STAT blockade in Aire-/- mice normalized interferon-γ responses and averted T-cell infiltration and damage in organs. Ruxolitinib treatment of five patients with APS-1 led to decreased levels of T-cell-derived interferon-γ, normalized interferon-γ and CXCL9 levels, and remission of alopecia, oral candidiasis, nail dystrophy, gastritis, enteritis, arthritis, Sjögren's-like syndrome, urticaria, and thyroiditis. No serious adverse effects from ruxolitinib were identified in these patients. CONCLUSIONS: Our findings indicate that APS-1, which is caused by AIRE deficiency, is characterized by excessive, multiorgan interferon-γ-mediated responses. JAK inhibition with ruxolitinib in five patients showed promising results. (Funded by the National Institute of Allergy and Infectious Diseases and others.).
Assuntos
Proteína AIRE , Interferon gama , Inibidores de Janus Quinases , Poliendocrinopatias Autoimunes , Adulto , Animais , Feminino , Humanos , Masculino , Camundongos , Proteína AIRE/deficiência , Proteína AIRE/genética , Proteína AIRE/imunologia , Autoanticorpos/sangue , Autoanticorpos/imunologia , Quimiocina CXCL9/genética , Interferon gama/genética , Interferon gama/imunologia , Inibidores de Janus Quinases/uso terapêutico , Camundongos Knockout , Nitrilas/uso terapêutico , Poliendocrinopatias Autoimunes/genética , Poliendocrinopatias Autoimunes/tratamento farmacológico , Poliendocrinopatias Autoimunes/imunologia , Pirazóis/uso terapêutico , Pirazóis/farmacologia , Pirimidinas/uso terapêutico , Linfócitos T/imunologia , Fatores de Transcrição/genética , Fatores de Transcrição/imunologia , Projetos Piloto , Modelos Animais de Doenças , Criança , Adolescente , Pessoa de Meia-IdadeRESUMO
Aspergillus fumigatus is an opportunistic mold that infects patients who are immunocompromised or have chronic lung disease, causing significant morbidity and mortality in these populations. While the factors governing the host response to A. fumigatus remain poorly defined, neutrophil recruitment to the site of infection is critical to clear the fungus. Galectin-3 is a mammalian ß-galactose-binding lectin with both antimicrobial and immunomodulatory activities, however the role of galectin-3 in the defense against molds has not been studied. Here we show that galectin-3 expression is markedly up-regulated in mice and humans with pulmonary aspergillosis. Galectin-3 deficient mice displayed increased fungal burden and higher mortality during pulmonary infection. In contrast to previous reports with pathogenic yeast, galectin-3 exhibited no antifungal activity against A. fumigatus in vitro. Galectin-3 deficient mice exhibited fewer neutrophils in their airways during infection, despite normal numbers of total lung neutrophils. Intravital imaging studies confirmed that galectin-3 was required for normal neutrophil migration to the airspaces during fungal infection. Adoptive transfer experiments demonstrated that stromal rather than neutrophil-intrinsic galectin-3 was necessary for normal neutrophil entry into the airspaces. Live cell imaging studies revealed that extracellular galectin-3 directly increases neutrophil motility. Taken together, these data demonstrate that extracellular galectin-3 facilitates recruitment of neutrophils to the site of A. fumigatus infection, and reveals a novel role for galectin-3 in host defense against fungal infections.
Assuntos
Aspergilose/imunologia , Aspergillus fumigatus/fisiologia , Galectina 3/imunologia , Pulmão/microbiologia , Neutrófilos/citologia , Animais , Aspergilose/genética , Aspergilose/microbiologia , Aspergilose/fisiopatologia , Aspergillus fumigatus/genética , Movimento Celular , Feminino , Galectina 3/genética , Humanos , Pulmão/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologiaRESUMO
Galactosaminogalactan and Pel are cationic heteropolysaccharides produced by the opportunistic pathogens Aspergillus fumigatus and Pseudomonas aeruginosa, respectively. These exopolysaccharides both contain 1,4-linked N-acetyl-d-galactosamine and play an important role in biofilm formation by these organisms. Proteins containing glycoside hydrolase domains have recently been identified within the biosynthetic pathway of each exopolysaccharide. Recombinant hydrolase domains from these proteins (Sph3h from A. fumigatus and PelAh from P. aeruginosa) were found to degrade their respective polysaccharides in vitro. We therefore hypothesized that these glycoside hydrolases could exhibit antibiofilm activity and, further, given the chemical similarity between galactosaminogalactan and Pel, that they might display cross-species activity. Treatment of A. fumigatus with Sph3h disrupted A. fumigatus biofilms with an EC50 of 0.4 nM. PelAh treatment also disrupted preformed A. fumigatus biofilms with EC50 values similar to those obtained for Sph3h In contrast, Sph3h was unable to disrupt P. aeruginosa Pel-based biofilms, despite being able to bind to the exopolysaccharide. Treatment of A. fumigatus hyphae with either Sph3h or PelAh significantly enhanced the activity of the antifungals posaconazole, amphotericin B, and caspofungin, likely through increasing antifungal penetration of hyphae. Both enzymes were noncytotoxic and protected A549 pulmonary epithelial cells from A. fumigatus-induced cell damage for up to 24 h. Intratracheal administration of Sph3h was well tolerated and reduced pulmonary fungal burden in a neutropenic mouse model of invasive aspergillosis. These findings suggest that glycoside hydrolases can exhibit activity against diverse microorganisms and may be useful as therapeutic agents by degrading biofilms and attenuating virulence.
Assuntos
Aspergilose/terapia , Aspergillus fumigatus/enzimologia , Proteínas de Bactérias/química , Biofilmes , Glicosídeo Hidrolases/química , Pseudomonas aeruginosa/enzimologia , Células A549 , Animais , Anti-Infecciosos/química , Antifúngicos/química , Aspergilose/microbiologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Polissacarídeos/química , Especificidade da Espécie , VirulênciaRESUMO
Background: Impaired delivery of antifungals to hyphae within necrotic lesions is thought to contribute to therapeutic failure in invasive pulmonary aspergillosis (IPA). We hypothesized that transfusion of leukocytes loaded ex vivo with the lipophilic antifungal posaconazole could improve delivery of antifungals to the sites of established infection and improve outcome in experimental IPA. Methods: The HL-60 leukemia cell line was differentiated to a neutrophil-like phenotype (differentiated HL-60 [dHL-60] cells) and then exposed to a range of posaconazole concentrations. The functional capacity and antifungal activity of these cells were assessed in vitro and in a mouse model of IPA. Results: Posaconazole levels in dHL-60 cells were 265-fold greater than the exposure concentration. Posaconazole-loaded cells were viable and maintained their capacity to undergo active chemotaxis. Contact-dependent transfer of posaconazole from dHL-60 cells to hyphae was observed in vitro, resulting in decreased fungal viability. In a neutropenic mouse model of IPA, treatment with posaconazole-loaded dHL-60 cells resulted in significantly reduced fungal burden in comparison to treatment with dHL-60 cells alone. Conclusions: Posaconazole accumulates at high concentrations in dHL-60 cells and increases their antifungal activity in vitro and in vivo. These findings suggest that posaconazole-loading of leukocytes may hold promise for the therapy of IPA.
Assuntos
Antifúngicos/uso terapêutico , Aspergilose Pulmonar Invasiva/tratamento farmacológico , Triazóis/uso terapêutico , Animais , Antifúngicos/farmacologia , Quimiotaxia/efeitos dos fármacos , Feminino , Células HL-60 , Humanos , Pulmão/microbiologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , Triazóis/farmacologiaRESUMO
Of the over 250 Aspergillus species, Aspergillus fumigatus accounts for up to 80% of invasive human infections. A. fumigatus produces galactosaminogalactan (GAG), an exopolysaccharide composed of galactose and N-acetyl-galactosamine (GalNAc) that mediates adherence and is required for full virulence. Less pathogenic Aspergillus species were found to produce GAG with a lower GalNAc content than A. fumigatus and expressed minimal amounts of cell wall-bound GAG. Increasing the GalNAc content of GAG of the minimally pathogenic A. nidulans, either through overexpression of the A. nidulans epimerase UgeB or by heterologous expression of the A. fumigatus epimerase Uge3 increased the amount of cell wall bound GAG, augmented adherence in vitro and enhanced virulence in corticosteroid-treated mice to levels similar to A. fumigatus. The enhanced virulence of the overexpression strain of A. nidulans was associated with increased resistance to NADPH oxidase-dependent neutrophil extracellular traps (NETs) in vitro, and was not observed in neutropenic mice or mice deficient in NADPH-oxidase that are unable to form NETs. Collectively, these data suggest that cell wall-bound GAG enhances virulence through mediating resistance to NETs.
Assuntos
Aspergillus/patogenicidade , Armadilhas Extracelulares , Neutrófilos/imunologia , Polissacarídeos/fisiologia , Animais , Biofilmes , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , VirulênciaRESUMO
Aspergillus fumigatus is the most virulent species within the Aspergillus genus and causes invasive infections with high mortality rates. The exopolysaccharide galactosaminogalactan (GAG) contributes to the virulence of A. fumigatus. A co-regulated five-gene cluster has been identified and proposed to encode the proteins required for GAG biosynthesis. One of these genes, sph3, is predicted to encode a protein belonging to the spherulin 4 family, a protein family with no known function. Construction of an sph3-deficient mutant demonstrated that the gene is necessary for GAG production. To determine the role of Sph3 in GAG biosynthesis, we determined the structure of Aspergillus clavatus Sph3 to 1.25 Å. The structure revealed a (ß/α)8 fold, with similarities to glycoside hydrolase families 18, 27, and 84. Recombinant Sph3 displayed hydrolytic activity against both purified and cell wall-associated GAG. Structural and sequence alignments identified three conserved acidic residues, Asp-166, Glu-167, and Glu-222, that are located within the putative active site groove. In vitro and in vivo mutagenesis analysis demonstrated that all three residues are important for activity. Variants of Asp-166 yielded the greatest decrease in activity suggesting a role in catalysis. This work shows that Sph3 is a glycoside hydrolase essential for GAG production and defines a new glycoside hydrolase family, GH135.
Assuntos
Aspergillus fumigatus/metabolismo , Coccidioidina/química , Proteínas Fúngicas/química , Glicosídeo Hidrolases/química , Polissacarídeos/biossíntese , Sequência de Aminoácidos , Aspergillus fumigatus/enzimologia , Aspergillus fumigatus/patogenicidade , Catálise , Domínio Catalítico , Parede Celular/enzimologia , Coccidioidina/genética , Coccidioidina/fisiologia , Sequência Conservada , Cristalografia por Raios X , Proteínas Fúngicas/genética , Proteínas Fúngicas/fisiologia , Glicosídeo Hidrolases/genética , Glicosídeo Hidrolases/fisiologia , Hidrólise , Dados de Sequência Molecular , Mutação , Polissacarídeos/genética , Conformação Proteica , Alinhamento de SequênciaRESUMO
In Aspergillus nidulans, the AcuK and AcuM transcription factors form a complex that regulates gluconeogenesis. In Aspergillus fumigatus, AcuM governs gluconeogenesis and iron acquisition in vitro and virulence in immunosuppressed mice. However, the function of AcuK was previously unknown. Through in vitro studies, we found that A. fumigatus ΔacuK single and ΔacuK ΔacuM double mutants had impaired gluconeogenesis and iron acquisition, similar to the ΔacuM mutant. Also, the ΔacuK, ΔacuM, and ΔacuK ΔacuM mutants had similar virulence defects in mice. However, the ΔacuK mutant had a milder defect in extracellular siderophore activity and induction of epithelial cell damage in vitro than did the ΔacuM mutant. Moreover, overexpression of acuM in the ΔacuK mutant altered expression of 3 genes and partially restored growth under iron-limited conditions, suggesting that AcuM can govern some genes independently of AcuK. Although the ΔacuK and ΔacuM mutants had very similar transcriptional profiles in vitro, their transcriptional profiles during murine pulmonary infection differed both from their in vitro profiles and from each other. While AcuK and AcuM governed the expression of only a few iron-responsive genes in vivo, they influenced the expression of other virulence-related genes, such as hexA and dvrA. Therefore, in A. fumigatus, while AcuK and AcuM likely function as part of the same complex, they can also function independently of each other. Furthermore, AcuK and AcuM have different target genes in vivo than in vitro, suggesting that in vivo infection stimulates unique transcriptional regulatory pathways in A. fumigatus.
Assuntos
Aspergilose/imunologia , Aspergillus fumigatus/genética , Aspergillus fumigatus/imunologia , Proteínas Fúngicas/genética , Regulação Fúngica da Expressão Gênica , Hospedeiro Imunocomprometido , Fatores de Transcrição/genética , Animais , Aspergilose/microbiologia , Aspergilose/patologia , Aspergillus fumigatus/crescimento & desenvolvimento , Aspergillus fumigatus/patogenicidade , Cortisona/administração & dosagem , Cortisona/análogos & derivados , Proteínas Fúngicas/metabolismo , Deleção de Genes , Perfilação da Expressão Gênica , Gluconeogênese/genética , Ferro/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Sideróforos/metabolismo , Fatores de Transcrição/metabolismo , Transcrição Gênica , VirulênciaRESUMO
Airway colonization by the mold Aspergillus fumigatus is common in patients with underlying lung disease and is associated with chronic airway inflammation. Studies probing the inflammatory response to colonization with A. fumigatus hyphae have been hampered by the lack of a model of chronic colonization in immunocompetent mice. By infecting mice intratracheally with conidia embedded in agar beads (Af beads), we have established an in vivo model to study the natural history of airway colonization with live A. fumigatus hyphae. Histopathological examination and galactomannan assay of lung homogenates demonstrated that hyphae exited beads and persisted in the lungs of mice up to 28 days postinfection without invasive disease. Fungal lesions within the airways were surrounded by a robust neutrophilic inflammatory reaction and peribronchial infiltration of lymphocytes. Whole-lung cytokine analysis from Af bead-infected mice revealed an increase in proinflammatory cytokines and chemokines early in infection. Evidence of a Th2 type response was observed only early in the course of colonization, including increased levels of interleukin-4 (IL-4), elevated IgE levels in serum, and a mild increase in airway responsiveness. Pulmonary T cell subset analysis during infection mirrored these results with an initial transient increase in IL-4-producing CD4(+) T cells, followed by a rise in IL-17 and Foxp3(+) cells by day 14. These results provide the first report of the evolution of the immune response to A. fumigatus hyphal colonization.
Assuntos
Hifas/imunologia , Aspergilose Pulmonar/imunologia , Aspergilose Pulmonar/patologia , Animais , Aspergillus fumigatus/imunologia , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Imunofenotipagem , Camundongos , Camundongos Endogâmicos C57BL , Subpopulações de Linfócitos T/imunologiaRESUMO
Aspergillus fumigatus is the most common cause of invasive mold disease in humans. The mechanisms underlying the adherence of this mold to host cells and macromolecules have remained elusive. Using mutants with different adhesive properties and comparative transcriptomics, we discovered that the gene uge3, encoding a fungal epimerase, is required for adherence through mediating the synthesis of galactosaminogalactan. Galactosaminogalactan functions as the dominant adhesin of A. fumigatus and mediates adherence to plastic, fibronectin, and epithelial cells. In addition, galactosaminogalactan suppresses host inflammatory responses in vitro and in vivo, in part through masking cell wall ß-glucans from recognition by dectin-1. Finally, galactosaminogalactan is essential for full virulence in two murine models of invasive aspergillosis. Collectively these data establish a role for galactosaminogalactan as a pivotal bifunctional virulence factor in the pathogenesis of invasive aspergillosis.
Assuntos
Aspergilose/imunologia , Aspergillus fumigatus/imunologia , Aspergillus fumigatus/patogenicidade , Polissacarídeos Fúngicos/imunologia , Polissacarídeos/imunologia , Fatores de Virulência/imunologia , beta-Glucanas/imunologia , Animais , Aspergilose/genética , Aspergilose/patologia , Aspergillus fumigatus/genética , Carboidratos Epimerases/genética , Carboidratos Epimerases/imunologia , Linhagem Celular , Modelos Animais de Doenças , Polissacarídeos Fúngicos/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/imunologia , Humanos , Hifas/genética , Hifas/imunologia , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Camundongos , Polissacarídeos/genética , Fatores de Virulência/genéticaRESUMO
The mold Aspergillus fumigatus and bacterium Pseudomonas aeruginosa form biofilms in the airways of individuals with cystic fibrosis. Biofilm formation by A. fumigatus depends on the self-produced cationic exopolysaccharide galactosaminogalactan (GAG), while P. aeruginosa biofilms can contain the cationic exopolysaccharide Pel. GAG and Pel are rendered cationic by deacetylation mediated by either the secreted deacetylase Agd3 (A. fumigatus) or the periplasmic deacetylase PelA (P. aeruginosa). Given the similarities between these polymers, the potential for biofilm interactions between these organisms were investigated. P. aeruginosa were observed to adhere to A. fumigatus hyphae in a GAG-dependent manner and to GAG-coated coverslips of A. fumigatus biofilms. In biofilm adherence assays, incubation of P. aeruginosa with A. fumigatus culture supernatants containing de-N-acetylated GAG augmented the formation of adherent P. aeruginosa biofilms, increasing protection against killing by the antibiotic colistin. Fluorescence microscopy demonstrated incorporation of GAG within P. aeruginosa biofilms, suggesting that GAG can serve as an alternate biofilm exopolysaccharide for this bacterium. In contrast, Pel-containing bacterial culture supernatants only augmented the formation of adherent A. fumigatus biofilms when antifungal inhibitory molecules were removed. This study demonstrates biofilm interaction via exopolysaccharides as a potential mechanism of co-operation between these organisms in chronic lung disease.
RESUMO
The use of mature neutrophil (granulocyte) transfusions for the treatment of neutropenic patients with invasive fungal infections (IFIs) has been the focus of multiple clinical trials. Despite these efforts, the transfusion of mature neutrophils has resulted in limited clinical benefit, likely owing to problems of insufficient numbers and the very short lifespan of these donor cells. In this report, we employed a system of conditionally immortalized murine neutrophil progenitors that are capable of continuous expansion, allowing for the generation of unlimited numbers of homogenous granulocyte-macrophage progenitors (GMPs). These GMPs were assayed in vivo to demonstrate their effect on survival in 2 models of IFI: candidemia and pulmonary aspergillosis. Mature neutrophils derived from GMPs executed all cardinal functions of neutrophils. Transfused GMPs homed to the bone marrow and spleen, where they completed normal differentiation to mature neutrophils. These neutrophils were capable of homing and extravasation in response to inflammatory stimuli using a sterile peritoneal challenge model. Furthermore, conditionally immortalized GMP transfusions significantly improved survival in models of candidemia and pulmonary aspergillosis. These data confirm the therapeutic benefit of prophylactic GMP transfusions in the setting of neutropenia and encourage development of progenitor cellular therapies for the management of fungal disease in high-risk patients.
Assuntos
Infecções Fúngicas Invasivas , Neutropenia , Neutrófilos , Animais , Candidemia , Terapia Baseada em Transplante de Células e Tecidos , Infecções Fúngicas Invasivas/prevenção & controle , Transfusão de Leucócitos , Camundongos , Neutropenia/terapia , Neutrófilos/transplante , Aspergilose PulmonarRESUMO
The lung naturally resists Aspergillus fumigatus (Af) in healthy individuals, but multiple conditions can disrupt this resistance, leading to lethal invasive infections. Core processes of natural resistance and its breakdown are undefined. We investigated three distinct conditions predisposing to lethal aspergillosis-severe SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) infection, influenza A viral pneumonia, and systemic corticosteroid use-in human patients and murine models. We found a conserved and essential coupling of innate B1a lymphocytes, Af-binding natural immunoglobulin G antibodies, and lung neutrophils. Failure of this axis concealed Af from neutrophils, allowing rapid fungal invasion and disease. Reconstituting the axis with immunoglobulin therapy reestablished resistance, thus representing a realistic pathway to repurpose currently available therapies. Together, we report a vital host resistance pathway that is responsible for protecting against life-threatening aspergillosis in the context of distinct susceptibilities.
Assuntos
COVID-19 , Neutrófilos , Humanos , Animais , Camundongos , SARS-CoV-2 , Esteroides/uso terapêuticoRESUMO
Pulmonary innate immunity is required for host defense; however, excessive neutrophil inflammation can cause life-threatening acute lung injury. B lymphocytes can be regulatory, yet little is known about peripheral transitional IgM+ B cells in terms of regulatory properties. Using single-cell RNA sequencing, we discovered eight IgM+ B cell subsets with unique gene regulatory networks in the lung circulation dominated by transitional type 1 B and type 2 B (T2B) cells. Lung intravital confocal microscopy revealed that T2B cells marginate in the pulmonary capillaries via CD49e and require CXCL13 and CXCR5. During lung inflammation, marginated T2B cells dampened excessive neutrophil vascular inflammation via the specialized proresolving molecule lipoxin A4 (LXA4). Exogenous CXCL13 dampened excessive neutrophilic inflammation by increasing marginated B cells, and LXA4 recapitulated neutrophil regulation in B cell-deficient mice during inflammation and fungal pneumonia. Thus, the lung microvasculature is enriched in multiple IgM+ B cell subsets with marginating capillary T2B cells that dampen neutrophil responses.
Assuntos
Linfócitos B/patologia , Pulmão/patologia , Neutrófilos/patologia , Pneumonia/patologia , Animais , Aspergilose/microbiologia , Aspergilose/patologia , Linfócitos B/fisiologia , Capilares/patologia , Adesão Celular , Quimiocina CXCL13/metabolismo , Integrina alfa5/metabolismo , Microscopia Intravital , Lipoxinas/metabolismo , Pulmão/irrigação sanguínea , Pulmão/diagnóstico por imagem , Camundongos Mutantes , Pneumonia/diagnóstico por imagem , Receptores CXCR5/metabolismo , Análise de Célula ÚnicaRESUMO
Aspergillus fumigatus airway infections are associated with increased rates of hospitalizations and declining lung function in patients with chronic lung disease. While the pathogenesis of invasive A. fumigatus infections is well studied, little is known about the development and progression of airway infections. Previous studies have demonstrated a critical role for the IL-1 cytokines, IL-1α and IL-1ß in enhancing pulmonary neutrophil recruitment during invasive aspergillosis. Here we use a mouse model of A. fumigatus airway infection to study the role of these IL-1 cytokines in immunocompetent mice. In the absence of IL-1 receptor signaling, mice exhibited reduced numbers of viable pulmonary neutrophils and increased levels of neutrophil apoptosis during fungal airway infection. Impaired neutrophil viability in these mice was associated with reduced pulmonary and systemic levels of G-CSF, and treatment with G-CSF restored both neutrophil viability and resistance to A. fumigatus airway infection. Taken together, these data demonstrate that IL-1 dependent G-CSF production plays a key role for host resistance to A. fumigatus airway infection through suppressing neutrophil apoptosis at the site of infection.
Assuntos
Aspergilose/imunologia , Aspergillus fumigatus/patogenicidade , Pulmão/imunologia , Neutrófilos/fisiologia , Aspergilose Pulmonar/imunologia , Receptores de Interleucina-1/fisiologia , Animais , Apoptose/imunologia , Quimiocinas/metabolismo , Fator Estimulador de Colônias de Granulócitos/metabolismo , Humanos , Interleucina-1alfa , Interleucina-1beta , Pulmão/patologia , Macrófagos , Camundongos , Camundongos Endogâmicos C57BL , Infiltração de Neutrófilos , Neutrófilos/imunologiaRESUMO
Background Chronic vascular disease atherosclerosis starts with an uptake of atherogenic modified low-density lipoproteins (LDLs) by resident macrophages, resulting in formation of arterial fatty streaks and eventually atheromatous plaques. Increased plasma sialic acid levels, increased neuraminidase activity, and reduced sialic acid LDL content have been previously associated with atherosclerosis and coronary artery disease in human patients, but the mechanism underlying this association has not been explored. Methods and Results We tested the hypothesis that neuraminidases contribute to development of atherosclerosis by removing sialic acid residues from glycan chains of the LDL glycoprotein and glycolipids. Atherosclerosis progression was investigated in apolipoprotein E and LDL receptor knockout mice with genetic deficiency of neuraminidases 1, 3, and 4 or those treated with specific neuraminidase inhibitors. We show that desialylation of the LDL glycoprotein, apolipoprotein B 100, by human neuraminidases 1 and 3 increases the uptake of human LDL by human cultured macrophages and by macrophages in aortic root lesions in Apoe-/- mice via asialoglycoprotein receptor 1. Genetic inactivation or pharmacological inhibition of neuraminidases 1 and 3 significantly delays formation of fatty streaks in the aortic root without affecting the plasma cholesterol and LDL levels in Apoe-/- and Ldlr-/- mouse models of atherosclerosis. Conclusions Together, our results suggest that neuraminidases 1 and 3 trigger the initial phase of atherosclerosis and formation of aortic fatty streaks by desialylating LDL and increasing their uptake by resident macrophages.
Assuntos
Aorta Abdominal/patologia , Aterosclerose/metabolismo , Doença da Artéria Coronariana/metabolismo , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , Neuraminidase/metabolismo , Animais , Aorta Abdominal/metabolismo , Aterosclerose/patologia , Biomarcadores/metabolismo , Células Cultivadas , Doença da Artéria Coronariana/patologia , Modelos Animais de Doenças , Humanos , Macrófagos/patologia , Camundongos , Camundongos Knockout , FagocitoseRESUMO
As the incidence rate of invasive fungal infections has increased with the use of modern medical interventions, so too has the occurrence of fungi invading the brain. Fungi such as Candida albicans, Cryptococcus neoformans, and Aspergillus fumigatus often infect immunocompromised individuals, and can use several strategies to invade the central nervous system (CNS) by penetrating the blood-brain barrier. Once in the brain parenchyma the specialized resident immune cells need to effectively recognize the fungus and mount an appropriate immune response to clear the infection, without causing debilitating immune-mediated toxicity and neuronal damage. Here we review the current knowledge pertaining to the antifungal response of the CNS and highlight areas where future research is required.
Assuntos
Encéfalo/imunologia , Fungos/fisiologia , Infecções Fúngicas Invasivas/imunologia , Animais , Encefalopatias/imunologia , Encefalopatias/microbiologia , Fungos/genética , Humanos , Imunidade , Infecções Fúngicas Invasivas/microbiologiaRESUMO
The exopolysaccharide galactosaminogalactan (GAG) is an important virulence factor of the fungal pathogen Aspergillus fumigatus. Deletion of a gene encoding a putative deacetylase, Agd3, leads to defects in GAG deacetylation, biofilm formation, and virulence. Here, we show that Agd3 deacetylates GAG in a metal-dependent manner, and is the founding member of carbohydrate esterase family CE18. The active site is formed by four catalytic motifs that are essential for activity. The structure of Agd3 includes an elongated substrate-binding cleft formed by a carbohydrate binding module (CBM) that is the founding member of CBM family 87. Agd3 homologues are encoded in previously unidentified putative bacterial exopolysaccharide biosynthetic operons and in other fungal genomes.
Assuntos
Amidoidrolases/química , Amidoidrolases/metabolismo , Aspergillus fumigatus/enzimologia , Aspergillus fumigatus/fisiologia , Biofilmes/crescimento & desenvolvimento , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Polissacarídeos/metabolismo , Acetilação , Sequência de Aminoácidos , Aspergillus fumigatus/genética , Domínio Catalítico , Sequência Conservada , Regulação Fúngica da Expressão Gênica , Glicosaminoglicanos/biossíntese , Metais/metabolismo , Domínios Proteicos , Homologia Estrutural de Proteína , Relação Estrutura-Atividade , Especificidade por Substrato , Fatores de TempoRESUMO
The incidence of fungal infections has dramatically increased in recent years, in large part due to increased use of immunosuppressive medications, as well as aggressive medical and surgical interventions that compromise natural skin and mucosal barriers. There are relatively few currently licensed antifungal drugs, and rising resistance to these agents has led to interest in the development of novel preventative and therapeutic strategies targeting these devastating infections. One approach to combat fungal infections is to augment the host immune response towards these organisms. The polysaccharide-rich cell wall is the initial point of contact between fungi and the host immune system, and therefore, represents an important target for immunotherapeutic approaches. This review highlights the advances made in our understanding of the mechanisms by which the immune system recognizes and interacts with exopolysaccharides produced by four of the most common fungal pathogens: Aspergillus fumigatus, Candida albicans, Cryptococcus neoformans, and Histoplasma capsulatum. Work to date suggests that inner cell wall polysaccharides that play an important structural role are the most conserved across diverse members of the fungal kingdom, and elicit the strongest innate immune responses. The immune system senses these carbohydrates through receptors, such as lectins and complement proteins. In contrast, a greater diversity of polysaccharides is found within the outer cell walls of pathogenic fungi. These glycans play an important role in immune evasion, and can even induce anti-inflammatory host responses. Further study of the complex interactions between the host immune system and the fungal polysaccharides will be necessary to develop more effective therapeutic strategies, as well as to explore the use of immunosuppressive polysaccharides as therapeutic agents to modulate inflammation.
RESUMO
Fungal biofilms are communities of adherent cells surrounded by an extracellular matrix. These biofilms are commonly found during infection caused by a variety of fungal pathogens. Clinically, biofilm infections can be extremely difficult to eradicate due to their resistance to antifungals and host defenses. Biofilm formation can protect fungal pathogens from many aspects of the innate immune system, including killing by neutrophils and monocytes. Altered immune recognition during this phase of growth is also evident by changes in the cytokine profiles of monocytes and macrophages exposed to biofilm. In this manuscript, we review the host response to fungal biofilms, focusing on how these structures are recognized by the innate immune system. Biofilms formed by Candida, Aspergillus, and Cryptococcus have received the most attention and are highlighted. We describe common themes involved in the resilience of fungal biofilms to host immunity and give examples of biofilm defenses that are pathogen-specific.