RESUMEN
Fungal-mediated disease progression and antifungal drug efficacy are significantly impacted by the dynamic infection microenvironment. At the site of infection, oxygen often becomes limiting and induces a hypoxia response in both the fungal pathogen and host cells. The fungal hypoxia response impacts several important aspects of fungal biology that contribute to pathogenesis, virulence, antifungal drug susceptibility, and ultimately infection outcomes. In this review, we summarize recent advances in understanding the molecular mechanisms of the hypoxia response in the most common human fungal pathogens, discuss potential therapeutic opportunities, and highlight important areas for future research.
Asunto(s)
Antifúngicos , Hipoxia , Humanos , Antifúngicos/farmacología , Antifúngicos/uso terapéutico , Virulencia , Progresión de la EnfermedadRESUMEN
Aspergillus fumigatus is a deadly agent of human fungal disease where virulence heterogeneity is thought to be at least partially structured by genetic variation between strains. While population genomic analyses based on reference genome alignments offer valuable insights into how gene variants are distributed across populations, these approaches fail to capture intraspecific variation in genes absent from the reference genome. Pan-genomic analyses based on de novo assemblies offer a promising alternative to reference-based genomics with the potential to address the full genetic repertoire of a species. Here, we evaluate 260 genome sequences of A. fumigatus including 62 newly sequenced strains, using a combination of population genomics, phylogenomics, and pan-genomics. Our results offer a high-resolution assessment of population structure and recombination frequency, phylogenetically structured gene presence-absence variation, evidence for metabolic specificity, and the distribution of putative antifungal resistance genes. Although A. fumigatus disperses primarily via asexual conidia, we identified extraordinarily high levels of recombination with the lowest linkage disequilibrium decay value reported for any fungal species to date. We provide evidence for 3 primary populations of A. fumigatus, with recombination occurring only rarely between populations and often within them. These 3 populations are structured by both gene variation and distinct patterns of gene presence-absence with unique suites of accessory genes present exclusively in each clade. Accessory genes displayed functional enrichment for nitrogen and carbohydrate metabolism suggesting that populations may be stratified by environmental niche specialization. Similarly, the distribution of antifungal resistance genes and resistance alleles were often structured by phylogeny. Altogether, the pan-genome of A. fumigatus represents one of the largest fungal pan-genomes reported to date including many genes unrepresented in the Af293 reference genome. These results highlight the inadequacy of relying on a single-reference genome-based approach for evaluating intraspecific variation and the power of combined genomic approaches to elucidate population structure, genetic diversity, and putative ecological drivers of clinically relevant fungi.
Asunto(s)
Antifúngicos , Aspergillus fumigatus , Aspergillus fumigatus/genética , Farmacorresistencia Fúngica , Genómica , Recombinación Genética/genéticaRESUMEN
Aspergillus fumigatus is a saprophytic, filamentous fungus found in soils and compost and the causative agent of several pulmonary diseases in humans, birds, and other mammals. A. fumigatus and other filamentous fungi grow as networks of filamentous hyphae that have characteristics of a classic microbial biofilm. These characteristics include production of an extracellular matrix (ECM), surface adhesion, multicellularity, and increased antimicrobial drug resistance. A. fumigatus biofilm growth occurs in vivo at sites of infection, highlighting the importance of defining mechanisms underlying biofilm development and associated emergent properties. We propose that there are 3 distinct phases in the development of A. fumigatus biofilms: biofilm initiation, immature biofilm, and mature biofilm. These stages are defined both temporally and by unique genetic and structural changes over the course of development. Here, we review known mechanisms within each of these stages that contribute to biofilm structure, ECM production, and increased resistance to contemporary antifungal drugs. We highlight gaps in our understanding of biofilm development and function that when addressed are expected to aid in the development of novel antifungal therapies capable of killing filamentous fungal biofilms.
Asunto(s)
Antifúngicos/farmacología , Aspergilosis/microbiología , Aspergillus fumigatus/crecimiento & desarrollo , Biopelículas/crecimiento & desarrollo , Farmacorresistencia Fúngica , Animales , Aspergilosis/tratamiento farmacológico , Aspergilosis/patología , Aspergillus fumigatus/efectos de los fármacos , Aspergillus fumigatus/fisiología , Biopelículas/efectos de los fármacos , Progresión de la Enfermedad , Humanos , Viabilidad MicrobianaRESUMEN
Human fungal infections may fail to respond to contemporary antifungal therapies in vivo despite in vitro fungal isolate drug susceptibility. Such a discrepancy between in vitro antimicrobial susceptibility and in vivo treatment outcomes is partially explained by microbes adopting a drug-resistant biofilm mode of growth during infection. The filamentous fungal pathogen Aspergillus fumigatus forms biofilms in vivo, and during biofilm growth it has reduced susceptibility to all three classes of contemporary antifungal drugs. Specific features of filamentous fungal biofilms that drive antifungal drug resistance remain largely unknown. In this study, we applied a fluorescence microscopy approach coupled with transcriptional bioreporters to define spatial and temporal oxygen gradients and single-cell metabolic activity within A. fumigatus biofilms. Oxygen gradients inevitably arise during A. fumigatus biofilm maturation and are both critical for, and the result of, A. fumigatus late-stage biofilm architecture. We observe that these self-induced hypoxic microenvironments not only contribute to filamentous fungal biofilm maturation but also drive resistance to antifungal treatment. Decreasing oxygen levels toward the base of A. fumigatus biofilms increases antifungal drug resistance. Our results define a previously unknown mechanistic link between filamentous fungal biofilm physiology and contemporary antifungal drug resistance. Moreover, we demonstrate that drug resistance mediated by dynamic oxygen gradients, found in many bacterial biofilms, also extends to the fungal kingdom. The conservation of hypoxic drug-resistant niches in bacterial and fungal biofilms is thus a promising target for improving antimicrobial therapy efficacy.
Asunto(s)
Antifúngicos/farmacología , Aspergillus fumigatus , Biopelículas/efectos de los fármacos , Microambiente Celular , Farmacorresistencia Fúngica , Aspergillus fumigatus/efectos de los fármacos , Aspergillus fumigatus/fisiología , Hipoxia de la Célula , Microambiente Celular/efectos de los fármacos , Microambiente Celular/fisiología , Oxígeno/farmacologíaRESUMEN
Beyond their canonical roles in hemostasis and thrombosis, platelets function in the innate immune response by interacting directly with pathogens and by regulating the recruitment and activation of immune effector cells. Thrombocytopenia often coincides with neutropenia in patients with hematologic malignancies and in allogeneic hematopoietic cell transplant recipients, patient groups at high risk for invasive fungal infections. While neutropenia is well established as a major clinical risk factor for invasive fungal infections, the role of platelets in host defense against human fungal pathogens remains understudied. Here, we examined the role of platelets in murine Aspergillus fumigatus infection using two complementary approaches to induce thrombocytopenia without concurrent neutropenia. Thrombocytopenic mice were highly susceptible to A. fumigatus challenge and rapidly succumbed to infection. Although platelets regulated early conidial phagocytosis by neutrophils in a spleen tyrosine kinase (Syk)-dependent manner, platelet-regulated conidial phagocytosis was dispensable for host survival. Instead, our data indicated that platelets primarily function to maintain hemostasis and lung integrity in response to exposed fungal antigens, since thrombocytopenic mice exhibited severe hemorrhage into the airways in response to fungal challenge in the absence of overt angioinvasion. Challenge with swollen, heat-killed, conidia was lethal in thrombocytopenic hosts and could be reversed by platelet transfusion, consistent with the model that fungus-induced inflammation in platelet-depleted mice was sufficient to induce lethal hemorrhage. These data provide new insights into the role of platelets in the anti-Aspergillus host response and expand their role to host defense against filamentous molds.
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Aspergillus fumigatus/inmunología , Plaquetas/inmunología , Trasplante de Células Madre Hematopoyéticas , Neutropenia/inmunología , Aspergilosis Pulmonar/inmunología , Quimera por Trasplante/inmunología , Aloinjertos , Animales , Ratones , Neutropenia/microbiología , Neutropenia/patología , Aspergilosis Pulmonar/patología , Quimera por Trasplante/microbiologíaRESUMEN
The initiation of Aspergillus fumigatus infection occurs via dormant conidia deposition into the airways. Therefore, conidial germination and subsequent hyphal extension and growth occur in a sustained heat shock (HS) environment promoted by the host. The cell wall integrity pathway (CWIP) and the essential eukaryotic chaperone Hsp90 are critical for fungi to survive HS. Although A. fumigatus is a thermophilic fungus, the mechanisms underpinning the HS response are not thoroughly described and important to define its role in pathogenesis, virulence and antifungal drug responses. Here, we investigate the contribution of the CWIP in A. fumigatus thermotolerance. We observed that the CWIP components PkcA, MpkA and RlmA are Hsp90 clients and that a PkcAG579R mutation abolishes this interaction. PkcAG579R also abolishes MpkA activation in the short-term response to HS. Biochemical and biophysical analyses indicated that Hsp90 is a dimeric functional ATPase, which has a higher affinity for ADP than ATP and prevents MpkA aggregation in vitro. Our data suggest that the CWIP is constitutively required for A. fumigatus to cope with the temperature increase found in the mammalian lung environment, emphasising the importance of this pathway in supporting thermotolerance and cell wall integrity.
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Adaptación Fisiológica , Aspergillus fumigatus/fisiología , Pared Celular/fisiología , Proteínas HSP90 de Choque Térmico/metabolismo , Respuesta al Choque Térmico , Aspergilosis/microbiología , Proteínas Fúngicas/metabolismo , Regulación Fúngica de la Expresión Génica , Interacciones Microbiota-Huesped , Mutación , Proteína Quinasa C/metabolismo , Transducción de Señal , Esporas Fúngicas/crecimiento & desarrollo , VirulenciaRESUMEN
RIG-I-like receptors (RLR) are cytosolic RNA sensors that signal through the MAVS adaptor to activate IFN responses against viruses. Whether the RLR family has broader effects on host immunity against other pathogen families remains to be fully explored. In this study, we demonstrate that MDA5/MAVS signaling was essential for host resistance against pulmonary Aspergillus fumigatus challenge through the regulation of antifungal leukocyte responses in mice. Activation of MDA5/MAVS signaling was driven by dsRNA from live A. fumigatus serving as a key vitality-sensing pattern recognition receptor. Interestingly, induction of type I IFNs after A. fumigatus challenge was only partially dependent on MDA5/MAVS signaling, whereas type III IFN expression was entirely dependent on MDA5/MAVS signaling. Ultimately, type I and III IFN signaling drove the expression of CXCL10. Furthermore, the MDA5/MAVS-dependent IFN response was critical for the induction of optimal antifungal neutrophil killing of A. fumigatus spores. In conclusion, our data broaden the role of the RLR family to include a role in regulating antifungal immunity against A. fumigatus.
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Aspergillus fumigatus/inmunología , Helicasa Inducida por Interferón IFIH1/inmunología , Helicasa Inducida por Interferón IFIH1/metabolismo , Moléculas de Patrón Molecular Asociado a Patógenos/inmunología , Moléculas de Patrón Molecular Asociado a Patógenos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/inmunología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Quimiocina CXCL10/inmunología , Quimiocina CXCL10/metabolismo , Femenino , Interferones/inmunología , Interferones/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Receptores de Reconocimiento de Patrones/inmunología , Receptores de Reconocimiento de Patrones/metabolismo , Transducción de Señal/inmunologíaRESUMEN
Growing evidence demonstrates that human mesenchymal stromal cells (MSCs) modify their in vivo anti-inflammatory actions depending on the specific inflammatory environment encountered. Understanding this better is crucial to refine MSC-based cell therapies for lung and other diseases. Using acute exacerbations of cystic fibrosis (CF) lung disease as a model, the effects of ex vivo MSC exposure to clinical bronchoalveolar lavage fluid (BALF) samples, as a surrogate for the in vivo clinical lung environment, on MSC viability, gene expression, secreted cytokines, and mitochondrial function were compared with effects of BALF collected from healthy volunteers. CF BALF samples that cultured positive for Aspergillus sp. (Asp) induced rapid MSC death, usually within several hours of exposure. Further analyses suggested the fungal toxin gliotoxin as a potential mediator contributing to CF BALF-induced MSC death. RNA sequencing analyses of MSCs exposed to either Asp+ or Asp- CF BALF samples identified a number of differentially expressed transcripts, including those involved in interferon signaling, antimicrobial gene expression, and cell death. Toxicity did not correlate with bacterial lung infections. These results suggest that the potential use of MSC-based cell therapies for CF or other lung diseases may not be warranted in the presence of Aspergillus.
Asunto(s)
Antiinflamatorios/uso terapéutico , Fibrosis Quística/terapia , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/citología , Líquido del Lavado Bronquioalveolar/microbiología , Fibrosis Quística/metabolismo , Humanos , Pulmón/metabolismo , Pulmón/microbiología , Trasplante de Células Madre Mesenquimatosas/métodosRESUMEN
It is estimated that fungal infections, caused most commonly by Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans, result in more deaths annually than malaria or tuberculosis. It has long been hypothesized the fungal metabolism plays a critical role in virulence though specific nutrient sources utilized by human pathogenic fungi in vivo has remained enigmatic. However, the metabolic utilisation of preferred carbon and nitrogen sources, encountered in a host niche-dependent manner, is known as carbon catabolite and nitrogen catabolite repression (CCR, NCR), and has been shown to be important for virulence. Several sensory and uptake systems exist, including carbon and nitrogen source-specific sensors and transporters, that allow scavenging of preferred nutrient sources. Subsequent metabolic utilisation is governed by transcription factors, whose functions and essentiality differ between fungal species. Furthermore, additional factors exist that contribute to the implementation of CCR and NCR. The role of the CCR and NCR-related factors in virulence varies greatly between fungal species and a substantial gap in knowledge exists regarding specific pathways. Further elucidation of carbon and nitrogen metabolism mechanisms is therefore required in a fungal species- and animal model-specific manner in order to screen for targets that are potential candidates for anti-fungal drug development.
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Represión Catabólica/genética , Represión Catabólica/fisiología , Virulencia/fisiología , Aspergillus fumigatus/metabolismo , Candida albicans/metabolismo , Carbono/metabolismo , Cryptococcus neoformans/metabolismo , Regulación Fúngica de la Expresión Génica/genética , Humanos , Micosis/metabolismo , Nitrógeno/metabolismo , Factores de Transcripción/metabolismo , Factores de Virulencia/metabolismoRESUMEN
The in vitro activities of two antifungal drugs in combination with four nonantifungal ophthalmic agents were evaluated using a broth microdilution method and a collection of eight Fusarium ocular isolates that exhibited resistance to both natamycin (MICs, 14 to 32 µg/ml) and voriconazole (MICs, 4 to >128 µg/ml). Synergistic and indifferent interactions were observed for natamycin and 5-fluorouracil and natamycin with timolol dependent on the Fusarium isolate tested. Isolate-dependent synergistic and indifferent interactions were also observed for natamycin with EDTA and natamycin with dorzolamide. Synergistic or indifferent interactions were observed for voriconazole with timolol and voriconazole with 5-fluorouracil depending on Fusarium isolate. Taken together, these data suggest that commonly used ophthalmic agents enhance the in vitro activity of antifungal drugs against drug-recalcitrant ocular fungal pathogens.
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Antifúngicos/farmacología , Infecciones Fúngicas del Ojo/tratamiento farmacológico , Fusarium/efectos de los fármacos , Natamicina/farmacología , Voriconazol/farmacología , Sinergismo Farmacológico , Infecciones Fúngicas del Ojo/microbiología , Hongos/efectos de los fármacos , Humanos , Itraconazol/farmacología , Pruebas de Sensibilidad Microbiana/métodosRESUMEN
Aspergillus fumigatus is responsible for a disproportionate number of invasive mycosis cases relative to other common filamentous fungi. While many fungal factors critical for infection establishment are known, genes essential for disease persistence and progression are ill defined. We propose that fungal factors that promote navigation of the rapidly changing nutrient and structural landscape characteristic of disease progression represent untapped clinically relevant therapeutic targets. To this end, we find that A. fumigatus requires a carbon catabolite repression (CCR) mediated genetic network to support in vivo fungal fitness and disease progression. While CCR as mediated by the transcriptional repressor CreA is not required for pulmonary infection establishment, loss of CCR inhibits fungal metabolic plasticity and the ability to thrive in the dynamic infection microenvironment. Our results suggest a model whereby CCR in an environmental filamentous fungus is dispensable for initiation of pulmonary infection but essential for infection maintenance and disease progression. Conceptually, we argue these data provide a foundation for additional studies on fungal factors required to support fungal fitness and disease progression and term such genes and factors, DPFs (disease progression factors).
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Aspergilosis/microbiología , Aspergillus fumigatus/genética , Carbono/metabolismo , Represión Catabólica , Proteínas Fúngicas/metabolismo , Redes Reguladoras de Genes , Aspergilosis/patología , Aspergillus fumigatus/fisiología , Progresión de la Enfermedad , Proteínas Fúngicas/genética , Regulación Fúngica de la Expresión Génica/efectos de los fármacos , Modelos Biológicos , Proteínas Represoras/genética , Proteínas Represoras/metabolismo , Estrés FisiológicoRESUMEN
AIMS/HYPOTHESIS: Recent studies have identified intracellular metabolism as a fundamental determinant of macrophage function. In obesity, proinflammatory macrophages accumulate in adipose tissue and trigger chronic low-grade inflammation, that promotes the development of systemic insulin resistance, yet changes in their intracellular energy metabolism are currently unknown. We therefore set out to study metabolic signatures of adipose tissue macrophages (ATMs) in lean and obese conditions. METHODS: F4/80-positive ATMs were isolated from obese vs lean mice. High-fat feeding of wild-type mice and myeloid-specific Hif1α-/- mice was used to examine the role of hypoxia-inducible factor-1α (HIF-1α) in ATMs part of obese adipose tissue. In vitro, bone marrow-derived macrophages were co-cultured with adipose tissue explants to examine adipose tissue-induced changes in macrophage phenotypes. Transcriptome analysis, real-time flux measurements, ELISA and several other approaches were used to determine the metabolic signatures and inflammatory status of macrophages. In addition, various metabolic routes were inhibited to determine their relevance for cytokine production. RESULTS: Transcriptome analysis and extracellular flux measurements of mouse ATMs revealed unique metabolic rewiring in obesity characterised by both increased glycolysis and oxidative phosphorylation. Similar metabolic activation of CD14+ cells in obese individuals was associated with diabetes outcome. These changes were not observed in peritoneal macrophages from obese vs lean mice and did not resemble metabolic rewiring in M1-primed macrophages. Instead, metabolic activation of macrophages was dose-dependently induced by a set of adipose tissue-derived factors that could not be reduced to leptin or lactate. Using metabolic inhibitors, we identified various metabolic routes, including fatty acid oxidation, glycolysis and glutaminolysis, that contributed to cytokine release by ATMs in lean adipose tissue. Glycolysis appeared to be the main contributor to the proinflammatory trait of macrophages in obese adipose tissue. HIF-1α, a key regulator of glycolysis, nonetheless appeared to play no critical role in proinflammatory activation of ATMs during early stages of obesity. CONCLUSIONS/INTERPRETATION: Our results reveal unique metabolic activation of ATMs in obesity that promotes inflammatory cytokine release. Further understanding of metabolic programming in ATMs will most likely lead to novel therapeutic targets to curtail inflammatory responses in obesity. DATA AVAILABILITY: Microarray data of ATMs isolated from obese or lean mice have been submitted to the Gene Expression Omnibus (accession no. GSE84000).
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Tejido Adiposo/citología , Macrófagos/metabolismo , Obesidad/metabolismo , Activación Metabólica , Adipocitos/metabolismo , Animales , Citocinas/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Glucólisis , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Inflamación , Resistencia a la Insulina , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células Mieloides/citología , Oxígeno/química , FenotipoRESUMEN
Recent estimates suggest that more than 3 million people have chronic or invasive fungal infections, causing more than 600,000 deaths every year. Aspergillus fumigatus causes invasive pulmonary aspergillosis (IPA) in patients with compromised immune systems and is a primary contributor to increases in human fungal infections. Thus, the development of new clinical modalities as stand-alone or adjunctive therapy for improving IPA patient outcomes is critically needed. Here we tested the in vitro and in vivo impacts of hyperbaric oxygen (HBO) (100% oxygen, >1 atmosphere absolute [ATA]) on A. fumigatus proliferation and murine IPA outcomes. Our findings indicate that HBO reduces established fungal biofilm proliferation in vitro by over 50%. The effect of HBO under the treatment conditions was transient and fungistatic, with A. fumigatus metabolic activity rebounding within 6 h of HBO treatment being removed. In vivo, daily HBO provides a dose-dependent but modest improvement in murine IPA disease outcomes as measured by survival analysis. Intriguingly, no synergy was observed between subtherapeutic voriconazole or amphotericin B and HBO in vitro or in vivo with daily HBO dosing, though the loss of fungal superoxide dismutase genes enhanced HBO antifungal activity. Further studies are needed to optimize the HBO treatment regimen and better understand the effects of HBO on both the host and the pathogen during a pulmonary invasive fungal infection.
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Antifúngicos/farmacología , Aspergillus fumigatus/patogenicidad , Oxigenoterapia Hiperbárica/métodos , Animales , Antifúngicos/uso terapéutico , Aspergilosis/tratamiento farmacológico , Aspergilosis/microbiología , Aspergillus fumigatus/efectos de los fármacos , Biopelículas/efectos de los fármacos , Modelos Animales de Enfermedad , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Balance of physiological levels of iron is essential for every organism. In Aspergillus fumigatus and other fungal pathogens, the transcription factor HapX mediates adaptation to iron limitation and consequently virulence by repressing iron consumption and activating iron uptake. Here, we demonstrate that HapX is also essential for iron resistance via activating vacuolar iron storage. We identified HapX protein domains that are essential for HapX functions during either iron starvation or high-iron conditions. The evolutionary conservation of these domains indicates their wide-spread role in iron sensing. We further demonstrate that a HapX homodimer and the CCAAT-binding complex (CBC) cooperatively bind an evolutionary conserved DNA motif in a target promoter. The latter reveals the mode of discrimination between general CBC and specific HapX/CBC target genes. Collectively, our study uncovers a novel regulatory mechanism mediating both iron resistance and adaptation to iron starvation by the same transcription factor complex with activating and repressing functions depending on ambient iron availability.
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Adaptación Fisiológica , Aspergilosis/metabolismo , Aspergillus fumigatus/patogenicidad , Proteínas Fúngicas/metabolismo , Regulación Fúngica de la Expresión Génica , Hierro/metabolismo , Factores de Transcripción/metabolismo , Aspergilosis/genética , Aspergilosis/virología , Western Blotting , Inmunoprecipitación de Cromatina , Proteínas Fúngicas/genética , Homeostasis , Inmunoprecipitación , Inanición , Resonancia por Plasmón de Superficie , Factores de Transcripción/genética , Vacuolas/metabolismo , VirulenciaRESUMEN
ß-Glucan is a naturally occurring glucose polysaccharide with immunostimulatory activity in both infection and malignancy. ß-Glucan's antitumor effects have been attributed to the enhancement of complement receptor 3-dependent cellular cytotoxicity, as well as modulation of suppressive and stimulatory myeloid subsets, which in turn enhances antitumor T cell immunity. In the present study, we demonstrate antitumor efficacy of yeast-derived ß-glucan particles (YGP) in a model of metastatic-like melanoma in the lung, through a mechanism that is independent of previously reported ß-glucan-mediated antitumor pathways. Notably, efficacy is independent of adaptive immunity, but requires inflammatory monocytes. YGP-activated monocytes mediated direct cytotoxicity against tumor cells in vitro, and systemic YGP treatment upregulated inflammatory mediators, including TNFα, M-CSF, and CCL2, in the lungs. Collectively, these studies identify a novel role for inflammatory monocytes in ß-glucan-mediated antitumor efficacy, and expand the understanding of how this immunomodulator can be used to generate beneficial immune responses against metastatic disease.
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Subunidad alfa del Factor 1 Inducible por Hipoxia/fisiología , Mediadores de Inflamación/inmunología , Neoplasias Pulmonares/tratamiento farmacológico , Melanoma Experimental/tratamiento farmacológico , Monocitos/inmunología , Receptores CCR2/fisiología , beta-Glucanos/farmacología , Inmunidad Adaptativa/inmunología , Adyuvantes Inmunológicos , Animales , Mediadores de Inflamación/metabolismo , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundario , Melanoma Experimental/inmunología , Melanoma Experimental/metabolismo , Melanoma Experimental/patología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Linfocitos T/inmunología , Células Tumorales CultivadasRESUMEN
Heterogeneity among Aspergillus fumigatus isolates results in unique virulence potential and inflammatory responses. How these isolates drive specific immune responses and how this affects fungally induced lung damage and disease outcome are unresolved. We demonstrate that the highly virulent CEA10 strain is able to rapidly germinate within the immunocompetent lung environment, inducing greater lung damage, vascular leakage, and interleukin 1α (IL-1α) release than the low-virulence Af293 strain, which germinates with a lower frequency in this environment. Importantly, the clearance of CEA10 was consequently dependent on IL-1α, in contrast to Af293. The release of IL-1α occurred by a caspase 1/11- and P2XR7-independent mechanism but was dependent on calpain activity. Our finding that early fungal conidium germination drives greater lung damage and IL-1α-dependent inflammation is supported by three independent experimental lines. First, pregermination of Af293 prior to in vivo challenge drives greater lung damage and an IL-1α-dependent neutrophil response. Second, the more virulent EVOL20 strain, derived from Af293, is able to germinate in the airways, leading to enhanced lung damage and IL-1α-dependent inflammation and fungal clearance. Third, primary environmental A. fumigatus isolates that rapidly germinate under airway conditions follow the same trend toward IL-1α dependency. Our data support the hypothesis that A. fumigatus phenotypic variation significantly contributes to disease outcomes.
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Aspergilosis/inmunología , Aspergillus fumigatus/inmunología , Aspergillus fumigatus/patogenicidad , Interleucina-1alfa/inmunología , Pulmón/inmunología , Animales , Células Cultivadas , Inmunocompetencia , Inflamación , Pulmón/microbiología , Macrófagos/inmunología , Macrófagos/microbiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Fenotipo , Esporas Fúngicas/inmunología , Esporas Fúngicas/patogenicidad , VirulenciaAsunto(s)
Proteínas Fúngicas , Hongos , Regulación Fúngica de la Expresión Génica , Micosis , Factores de Virulencia , Animales , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Hongos/genética , Hongos/metabolismo , Hongos/patogenicidad , Hongos/ultraestructura , Humanos , Micosis/genética , Micosis/metabolismo , Micosis/patología , Factores de Virulencia/genética , Factores de Virulencia/metabolismoRESUMEN
Aspergillus fumigatus is a mold that causes severe pulmonary infections. Our knowledge of how A. fumigatus growth is controlled in the respiratory tract is developing, but still limited. Alveolar macrophages, lung resident macrophages, and airway epithelial cells constitute the first lines of defense against inhaled A. fumigatus conidia. Subsequently, neutrophils and inflammatory CCR2+ monocytes are recruited to the respiratory tract to prevent fungal growth. However, the mechanism of neutrophil and macrophage recruitment to the respiratory tract after A. fumigatus exposure remains an area of ongoing investigation. Here we show that A. fumigatus pulmonary challenge induces expression of the inflammasome-dependent cytokines IL-1ß and IL-18 within the first 12 hours, while IL-1α expression continually increases over at least the first 48 hours. Strikingly, Il1r1-deficient mice are highly susceptible to pulmonary A. fumigatus challenge exemplified by robust fungal proliferation in the lung parenchyma. Enhanced susceptibility of Il1r1-deficient mice correlated with defects in leukocyte recruitment and anti-fungal activity. Importantly, IL-1α rather than IL-1ß was crucial for optimal leukocyte recruitment. IL-1α signaling enhanced the production of CXCL1. Moreover, CCR2+ monocytes are required for optimal early IL-1α and CXCL1 expression in the lungs, as selective depletion of these cells resulted in their diminished expression, which in turn regulated the early accumulation of neutrophils in the lung after A. fumigatus challenge. Enhancement of pulmonary neutrophil recruitment and anti-fungal activity by CXCL1 treatment could limit fungal growth in the absence of IL-1α signaling. In contrast to the role of IL-1α in neutrophil recruitment, the inflammasome and IL-1ß were only essential for optimal activation of anti-fungal activity of macrophages. As such, Pycard-deficient mice are mildly susceptible to A. fumigatus infection. Taken together, our data reveal central, non-redundant roles for IL-1α and IL-1ß in controlling A. fumigatus infection in the murine lung.
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Aspergillus fumigatus/inmunología , Quimiotaxis de Leucocito , Interleucina-1alfa/fisiología , Aspergilosis Pulmonar/inmunología , Animales , Pruebas de Provocación Bronquial , Células Cultivadas , Quimiotaxis de Leucocito/genética , Quimiotaxis de Leucocito/inmunología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Aspergilosis Pulmonar/genética , Transducción de Señal/genética , Transducción de Señal/inmunologíaRESUMEN
Aspergillus fumigatus forms ubiquitous airborne conidia that humans inhale on a daily basis. Although respiratory fungal infection activates the adaptor proteins CARD9 and MyD88 via C-type lectin, Toll-like, and interleukin-1 family receptor signals, defining the temporal and spatial pattern of MyD88- and CARD9-coupled signals in immune activation and fungal clearance has been difficult to achieve. Herein, we demonstrate that MyD88 and CARD9 act in two discrete phases and in two cellular compartments to direct chemokine- and neutrophil-dependent host defense. The first phase depends on MyD88 signaling because genetic deletion of MyD88 leads to delayed induction of the neutrophil chemokines CXCL1 and CXCL5, delayed neutrophil lung trafficking, and fatal pulmonary damage at the onset of respiratory fungal infection. MyD88 expression in lung epithelial cells restores rapid chemokine induction and neutrophil recruitment via interleukin-1 receptor signaling. Exogenous CXCL1 administration reverses murine mortality in MyD88-deficient mice. The second phase depends predominately on CARD9 signaling because genetic deletion of CARD9 in radiosensitive hematopoietic cells interrupts CXCL1 and CXCL2 production and lung neutrophil recruitment beyond the initial MyD88-dependent phase. Using a CXCL2 reporter mouse, we show that lung-infiltrating neutrophils represent the major cellular source of CXCL2 during CARD9-dependent recruitment. Although neutrophil-intrinsic MyD88 and CARD9 function are dispensable for neutrophil conidial uptake and killing in the lung, global deletion of both adaptor proteins triggers rapidly progressive invasive disease when mice are challenged with an inoculum that is sub-lethal for single adapter protein knockout mice. Our findings demonstrate that distinct signal transduction pathways in the respiratory epithelium and hematopoietic compartment partially overlap to ensure optimal chemokine induction, neutrophil recruitment, and fungal clearance within the respiratory tract.
Asunto(s)
Aspergillus fumigatus/fisiología , Proteínas Adaptadoras de Señalización CARD/metabolismo , Quimiocinas/metabolismo , Factor 88 de Diferenciación Mieloide/metabolismo , Aspergilosis Pulmonar/inmunología , Transducción de Señal , Animales , Humanos , Inmunidad Innata , Pulmón/inmunología , Ratones , Ratones Noqueados , Infiltración Neutrófila/inmunología , Neutrófilos/inmunología , Aspergilosis Pulmonar/microbiología , Receptores de Interleucina-1/metabolismoRESUMEN
Invasive aspergillosis (IA), most commonly caused by the filamentous fungus Aspergillus fumigatus, occurs in immune compromised individuals. The ability of A. fumigatus to proliferate in a multitude of environments is hypothesized to contribute to its pathogenicity and virulence. Transcription factors (TF) have long been recognized as critical proteins for fungal pathogenicity, as many are known to play important roles in the transcriptional regulation of pathways implicated in virulence. Such pathways include regulation of conidiation and development, adhesion, nutrient acquisition, adaptation to environmental stress, and interactions with the host immune system among others. In both murine and insect models of IA, TF loss of function in A. fumigatus results in cases of hyper- and hypovirulence as determined through host survival, fungal burden, and immune response analyses. Consequently, the study of specific TFs in A. fumigatus has revealed important insights into mechanisms of pathogenicity and virulence. Although in vitro studies have identified virulence-related functions of specific TFs, the full picture of their in vivo functions remain largely enigmatic and an exciting area of current research. Moreover, the vast majority of TFs remain to be characterized and studied in this important human pathogen. Here in this mini-review we provide an overview of selected TFs in A. fumigatus and their contribution to our understanding of this important human pathogen's pathogenicity and virulence.