Dataset Information

Human Neutrophils Are Primed by Chemoattractant Gradients for Blocking the Growth of Aspergillus fumigatus.

ABSTRACT: The contribution of human neutrophils to the protection against fungal infections by Aspergillus fumigatus is essential but not fully understood. Whereas healthy people can inhale spores of A. fumigatus without developing disease, neutropenic patients and those receiving immunosuppressive drugs have a higher incidence of invasive fungal infections. To study the role of neutrophils in protection against A. fumigatus infections, we developed an in vitro assay in which the interactions between human neutrophils and A. fumigatus were observed in real time, at single-cell resolution, in precisely controlled conditions. We measured the outcomes of neutrophil-fungus interactions and found that human neutrophils have a limited ability to migrate toward A. fumigatus and block the growth of A. fumigatus conidia (proportion with growth blocked, 69%). The blocking ability of human neutrophils increased to 85.1% when they were stimulated by uniform concentrations of fMLP and was enhanced further, to 99.4%, in the presence of chemoattractant gradients. Neutrophils from patients receiving immunosuppressive treatment after transplantation were less effective against the fungus than those from healthy donors, and broader heterogeneity exists between patients, compared with healthy individuals. Further studies using this microfluidic platform will help understand the relevance of innate immune deficiencies responsible for the higher risk of fungal infections in patients with immunosuppressive disease.

SUBMITTER: Jones CN

PROVIDER: S-EPMC4704668 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Human Neutrophils Are Primed by Chemoattractant Gradients for Blocking the Growth of Aspergillus fumigatus.

Jones Caroline N CN Dimisko Laurie L Forrest Kevin K Judice Kevin K Poznansky Mark C MC Markmann James F JF Vyas Jatin M JM Irimia Daniel D

The Journal of infectious diseases 20150812 3

The contribution of human neutrophils to the protection against fungal infections by Aspergillus fumigatus is essential but not fully understood. Whereas healthy people can inhale spores of A. fumigatus without developing disease, neutropenic patients and those receiving immunosuppressive drugs have a higher incidence of invasive fungal infections. To study the role of neutrophils in protection against A. fumigatus infections, we developed an in vitro assay in which the interactions between huma ...[more]

PMID: 26272935

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Project description:Polymorphonuclear granulocytes (PMNs) are indispensable for controlling life-threatening fungal infections. In addition to various effector mechanisms, PMNs also produce extracellular vesicles (EVs). Their contribution to antifungal defense has remained unexplored. We reveal that the clinically important human-pathogenic fungus Aspergillus fumigatus triggers PMNs to release a distinct set of antifungal EVs (afEVs). Proteome analyses indicated that afEVs are enriched in antimicrobial proteins. The cargo and the release kinetics of EVs are modulated by the fungal strain confronted. Tracking of afEVs indicated that they associated with fungal cells and even entered fungal hyphae, resulting in alterations in the morphology of the fungal cell wall and dose-dependent antifungal effects. To assess as a proof of concept whether the antimicrobial proteins found in afEVs might contribute to growth inhibition of hyphae when present in the fungal cytoplasm, two human proteins enriched in afEVs, cathepsin G and azurocidin, were heterologously expressed in fungal hyphae. This led to reduced fungal growth relative to that of a control strain producing the human retinol binding protein 7. In conclusion, extracellular vesicles produced by neutrophils in response to A. fumigatus infection are able to associate with the fungus, limit growth, and elicit cell damage by delivering antifungal cargo. This finding offers an intriguing, previously overlooked mechanism of antifungal defense against A. fumigatus IMPORTANCE Invasive fungal infections caused by the mold Aspergillus fumigatus are a growing concern in the clinic due to the increasing use of immunosuppressive therapies and increasing antifungal drug resistance. These infections result in high rates of mortality, as treatment and diagnostic options remain limited. In healthy individuals, neutrophilic granulocytes are critical for elimination of A. fumigatus from the host; however, the exact extracellular mechanism of neutrophil-mediated antifungal activity remains unresolved. Here, we present a mode of antifungal defense employed by human neutrophils against A. fumigatus not previously described. We found that extracellular vesicles produced by neutrophils in response to A. fumigatus infection are able to associate with the fungus, limit growth, and elicit cell damage by delivering antifungal cargo. In the end, antifungal extracellular vesicle biology provides a significant step forward in our understanding of A. fumigatus host pathogenesis and opens up novel diagnostic and therapeutic possibilities.

Project description:BackgroundAspergillus fumigatus is the most common etiologic agent of invasive aspergillosis in immunocompromised patients. Several studies have addressed the mechanism involved in host defense but only few have investigated the pathogen's response to attack by the host cells. To our knowledge, this is the first study that investigates the genes differentially expressed in conidia vs hyphae of A. fumigatus in response to neutrophils from healthy donors as well as from those with chronic granulomatous disease (CGD) which are defective in the production of reactive oxygen species.Methodology/principal findingsTranscriptional profiles of conidia and hyphae exposed to neutrophils, either from normal donors or from CGD patients, were obtained by using the genome-wide microarray. Upon exposure to either normal or CGD neutrophils, 244 genes were up-regulated in conidia but not in hyphae. Several of these genes are involved in the degradation of fatty acids, peroxisome function and the glyoxylate cycle which suggests that conidia exposed to neutrophils reprogram their metabolism to adjust to the host environment. In addition, the mRNA levels of four genes encoding proteins putatively involved in iron/copper assimilation were found to be higher in conidia and hyphae exposed to normal neutrophils compared to those exposed to CGD neutrophils. Deletants in several of the differentially expressed genes showed phenotypes related to the proposed functions, i.e. deletants of genes involved in fatty acid catabolism showed defective growth on fatty acids and the deletants of iron/copper assimilation showed higher sensitivity to the oxidative agent menadione. None of these deletants, however, showed reduced resistance to neutrophil attack.ConclusionThis work reveals the complex response of the fungus to leukocytes, one of the major host factors involved in antifungal defense, and identifies fungal genes that may be involved in establishing or prolonging infections in humans.

Dataset Information

Human Neutrophils Are Primed by Chemoattractant Gradients for Blocking the Growth of Aspergillus fumigatus.

Publications

Human Neutrophils Are Primed by Chemoattractant Gradients for Blocking the Growth of Aspergillus fumigatus.

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