Cystic Fibrosis Clinical Isolates of Aspergillus fumigatus Induce Similar Muco-inflammatory Responses in Primary Airway Epithelial Cells.
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SUBMITTER: McLean SA
PROVIDER: S-EPMC8399118 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Aspergillus fumigatus is the most commonly isolated fungus in chronic lung diseases, with a prevalence of up to 60% in cystic fibrosis patients. Despite this, the impact of A. fumigatus colonisation on lung epithelia has not been thoroughly explored. We investigated the influence of A. fumigatus supernatants and the secondary metabolite, gliotoxin, on human bronchial epithelial cells (HBE) and CF bronchial epithelial (CFBE) cells. CFBE (F508del CFBE41o-) and HBE (16HBE14o-) trans-epithelial electrical resistance (TEER) was measured following exposure to A. fumigatus reference and clinical isolates, a gliotoxin-deficient mutant (ΔgliG) and pure gliotoxin. The impact on tight junction (TJ) proteins, zonula occludens-1 (ZO-1) and junctional adhesion molecule-A (JAM-A) were determined by western blot analysis and confocal microscopy. A. fumigatus conidia and supernatants caused significant disruption to CFBE and HBE TJs within 24 h. Supernatants from later cultures (72 h) caused the greatest disruption while ΔgliG mutant supernatants caused no disruption to TJ integrity. The ZO-1 and JAM-A distribution in epithelial monolayers were altered by A. fumigatus supernatants but not by ΔgliG supernatants, suggesting that gliotoxin is involved in this process. The fact that ΔgliG conidia were still capable of disrupting epithelial monolayers indicates that direct cell-cell contact also plays a role, independently of gliotoxin production. Gliotoxin is capable of disrupting TJ integrity which has the potential to contribute to airway damage, and enhance microbial invasion and sensitisation in CF.
| S-EPMC10141846 | biostudies-literature
Project description:Aspergillus fumigatus is a ubiquitous fungal pathogen capable of causing multiple pulmonary diseases, including invasive aspergillosis, chronic necrotizing aspergillosis, fungal colonization, and allergic bronchopulmonary aspergillosis. Intact mucociliary barrier function and early airway neutrophil responses are critical for clearing fungal conidia from the host airways prior to establishing disease. Following inhalation, Aspergillus conidia deposit in the small airways, where they are likely to make their initial host encounter with epithelial cells. Challenges in airway infection models have limited the ability to explore early steps in the interactions between A. fumigatus and the human airway epithelium. Here, we use inverted air-liquid interface cultures to demonstrate that the human airway epithelium responds to apical stimulation by A. fumigatus to promote the transepithelial migration of neutrophils from the basolateral membrane surface to the apical airway surface. Promoting epithelial transmigration with Aspergillus required prolonged exposure with live resting conidia. Swollen conidia did not expedite epithelial transmigration. Using A. fumigatus strains containing deletions of genes for cell wall components, we identified that deletion of the hydrophobic rodlet layer or dihydroxynaphthalene-melanin in the conidial cell wall amplified the epithelial transmigration of neutrophils, using primary human airway epithelium. Ultimately, we show that an as-yet-unidentified nonsecreted cell wall protein is required to promote the early epithelial transmigration of human neutrophils into the airspace in response to A. fumigatus Together, these data provide critical insight into the initial epithelial host response to Aspergillus.
| S-EPMC6977131 | biostudies-literature
Project description:Our laboratory has held a long interest in the glycosylation changes seen on the surface of airway epithelia of patients with the disease cystic fibrosis (CF). Experiments from our laboratory have detailed a CF glycosylation phenotype of increased Fuca1,3/4 and decreased Fuca1,2 and sialic acid on the surfaces of immortalized and primary CF cells compared to non-CF cells. Further, we have shown that gene transfer and subsequent expression of a wild type CF plasmid in CF airway cells results in correction or reversal of this glycosylation phenotype. We hypothesize that the changes in glycosylation seen in CF cells are key in the pathophysiology of the cystic fibrosis airway disease. For example, it has been shown that Pseudomonas aeruginosa, a bacterium that has a predilection for colonizing CF airways, adheres to asialylated glycolipids and glycoconjugates with terminal Fuca1,3/4. One focus of our laboratory is to elucidate the etiology of the glycosylation changes seen in CF cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer. We propose to study the gene expression of immortalized and primary CF and non-CF airway epithelial cells: 1. CF/T43 vs. BEAS-2B cells. These are two widely used immortalized airway cell lines that we have used extensively in the past. 2. C38 cells; C38 cells are IB3 cells expressing wtCFTR. The experimental focus is to elucidate the etiology of the glycosylation changes seen in Cystic Fibrosis (CF) cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer. To do so, the gene expression of immortalized and primary CF and non-CF airway epithelial cells were compared and studied. Cell lines used were CF/T43 and BEAS-2B, both widely used immortalized airway cell lines. Other cell lines studied included C38 cell lines (clonal derivatives of IB3 cells expressing wtCFTR).
2011-01-07 | E-GEOD-26482 | biostudies-arrayexpress
Project description:Given the complex nature of the responses that can occur in host-pathogen interactions, dual transcriptomics offers a powerful method of elucidating these interactions during infection. The gene expression patterns of Aspergillus fumigatus conidia or host cells have been reported in a number of previous studies, but each focused on only one of the interacting organisms. In the present study, we profiled simultaneously the transcriptional response of both A. fumigatus and human airway epithelial cells (AECs).16HBE14o- transformed bronchial epithelial cells were incubated with A. fumigatus conidia at 37°C for 6 hours, followed by genome-wide transcriptome analysis using human and fungal microarrays. Differentially expressed gene lists were generated from the microarrays, from which biologically relevant themes were identified. Human and fungal candidate genes were selected for validation, using RT-qPCR, in both 16HBE14o- cells and primary AECs co-cultured with conidia.We report that ontologies related to the innate immune response are activated by co-incubation with A. fumigatus condia, and interleukin-6 (IL-6) was confirmed to be up-regulated in primary AECs via RT-qPCR. Concomitantly, A. fumigatus was found to up-regulate fungal pathways involved in iron acquisition, vacuolar acidification, and formate dehydrogenase activity.To our knowledge, this is the first study to apply a dual organism transcriptomics approach to interactions of A. fumigatus conidia and human airway epithelial cells. The up-regulation of IL-6 by epithelia and simultaneous activation of several pathways by fungal conidia warrants further investigation as we seek to better understand this interaction in both health and disease. The cellular response of the airway epithelium to A. fumigatus is important to understand if we are to improve host-pathogen outcomes.
| S-EPMC3105077 | biostudies-literature
Project description:Aspergillus fumigatus is an opportunistic, ubiquitous, saprophytic mould that can cause severe allergic responses in atopic individuals as well as life-threatening infections in immunocompromised patients. A critical step in the establishment of infection is the invasion of airway epithelial cells by the inhaled fungi. Understanding how A. fumigatus senses and responds to airway cells is important to understand the pathogenesis of invasive pulmonary aspergillosis. Here, we analysed the transcriptomes of two commonly used clinical isolates, Af293 and CEA10, during infection of the A549 type II pneumocyte cell line in vitro. We focused our RNA-seq analysis on the core set of genes that are present in the genomes of the two strains. Our results suggest that: (a) A. fumigatus does not mount a conserved transcriptional response to airway epithelial cells in our in vitro model and (b) strain background and time spent in the tissue culture media have a greater impact on the transcriptome than the presence of host cells. Our analyses reveal both common and strain-specific transcriptional programmes that allow for the generation of hypotheses about gene function as it pertains to pathogenesis and the significant phenotypic heterogeneity that is observed among A. fumigatus isolates.
| S-EPMC5857381 | biostudies-literature
Project description:Aspergillus fumigatus is commonly isolated from CF airways. However, the impact on CF lung progression is not completely understood. In this study, using a 16-year retrospective observational cohort study (2000-2015) that included 132 patients, we determined the annual lung function, measured as percent predicted forced expiratory volume in the first second (ppFEV1), decline before and after the first colonization with A. fumigatus. Further, in the same individual, the ratios of lung function when patients were colonized with A. fumigatus and when they were not were calculated. The impact of eradication, with antifungal treatment or spontaneously, was assessed. The annual ppFEV1 was significantly lower after the first colonization with A. fumigatus. Furthermore, within the same individual, colonization with A. fumigatus for two and three years in a row was associated with 4.3% and 7.9% lower ppFEV1, respectively, compared to when not colonized. Finally, patients who eradicated A. fumigatus the following two years after colonization exhibited 9.9% and 14.5% higher ppFEV1 compared to patients who continued to produce cultures with A. fumigatus for two and three years. Our study demonstrated that A. fumigatus colonization was associated with a negative impact on lung function in the long term and eradication, spontaneously or with treatment, was associated with a better pulmonary outcome.
| S-EPMC8618016 | biostudies-literature
Project description:Pathological remodeling of the airway epithelium is commonly observed in Cystic Fibrosis (CF). The different cell types that constitute the airway epithelium are regenerated upon injury to restore integrity and maintenance of the epithelium barrier function. The molecular signature of tissue repair in CF airway epithelial cells has, however, not well been investigated in primary cultures. We therefore collected RNA-seq data from well-differentiated primary cultures of bronchial human airway epithelial cells (HAECs) of CF (F508del/F508del) and non-CF (NCF) origins before and after mechanical wounding, exposed or not to flagellin. We identified the expression changes with time of repair of genes, the products of which are markers of the different cell types that constitute the airway epithelium (basal, suprabasal, intermediate, secretory, goblet and ciliated cells as well as ionocytes). Researchers in the CF field may benefit from this transcriptomic profile, which covers the initial steps of wound repair and revealed differences in this process between CF and NCF cultures.
| S-EPMC6820749 | biostudies-literature
Project description:BACKGROUND:Defective chloride transport in airway epithelial cells (AECs) and the associated lung disease are the main causes of morbidity and early mortality in cystic fibrosis (CF). Abnormal airway iron homeostasis and the presence of lipid peroxidation products, indicative of oxidative stress, are features of CF lung disease. RESULTS:Here, we report that CF AECs (IB3-1) are susceptible to ferroptosis, a type of cell death associated with iron accumulation and lipid peroxidation. Compared to isogenic CFTR corrected cells (C38), the IB3-1 cells showed increased susceptibility to cell death upon exposure to iron in the form of ferric ammonium citrate (FAC) and the ferroptosis inducer, erastin. This phenotype was accompanied by accumulation of intracellular ferrous iron and lipid peroxides and the extracellular release of malondialdehyde, all indicative of redox stress, and increased levels of lactate dehydrogenase in the culture supernatant, indicating enhanced cell injury. The ferric iron chelator deferoxamine (DFO) and the lipophilic antioxidant ferrostatin-1 inhibited FAC and erastin induced ferroptosis in IB3-1 cells. Glutathione peroxidase 4 (GPX4) expression was decreased in IB3-1 cells treated with FAC and erastin, but was unchanged in C38 AECs. Necroptosis appeared to be involved in the enhanced susceptibility of IB3-1 AECs to ferroptosis, as evidenced by partial cell death rescue with necroptosis inhibitors and enhanced mixed lineage kinase domain-like (MLKL) localisation to the plasma membrane. CONCLUSION:These studies suggest that the increased susceptibility of CF AECs to ferroptosis is linked to abnormal intracellular ferrous iron accumulation and reduced antioxidant defences. In addition, the process of ferroptotic cell death in CF AECs does not appear to be a single entity and for the first time we describe necroptosis as a potential contributory factor. Iron chelation and antioxidant treatments may be promising therapeutic interventions in cystic fibrosis.
| S-EPMC8670191 | biostudies-literature
Project description:Background and purposeThe present study aimed to evaluate the effect of cyproconazole, the most used fungicide in Iranian wheat farms, on the induction of voriconazole resistance in Aspergillus fumigatus isolates.Materials and methodsA collection of 20 clinical and environmental isolates were selected for investigation of the in vitro activity of fungicides. The minimum inhibitory concentrations (MICs) were determined by the documented broth microdilution method M38-A2 (CLSI, 2008). Induction experiments were performed and the possibly induced isolate(s) were subjected to antifungal susceptibility testing, sequencing of the CYP51A promoter, and full coding gene. Furthermore, CYP51-protein homology modeling and docking modes were evaluated using SWISS-MODEL (https://swissmodel.expasy.org/) and SEESAR software (version 9.1).ResultsAmong 10 susceptible isolates, only one strain showed a high MIC value against voriconazole (MIC=4µg/ml) after 25 passages. Nevertheless, sequencing of the CYP51A promoter and full coding gene did not reveal any mutations. Cyproconazole, which has three nitrogen atoms in the aromatic ring, coordinated to the iron atom of heme through a hydrogen bond contact to residue Lys147 present in the active site of the A. fumigates Cyp51 homology model.ConclusionCyproconazole is being applied extensively in wheat farms in Iran. According to the results, cyproconazole may not play a key role in the induction of azole resistance in the isolates through the environmental route. However, the potential ability of the fungicide to induce medically triazole-resistant strains over a long period of application should not be neglected.
| S-EPMC8226051 | biostudies-literature
Project description:Our laboratory has held a long interest in the glycosylation changes seen on the surface of airway epithelia of patients with the disease cystic fibrosis (CF). Experiments from our laboratory have detailed a CF glycosylation phenotype of increased Fuca1,3/4 and decreased Fuca1,2 and sialic acid on the surfaces of immortalized and primary CF cells compared to non-CF cells. Further, we have shown that gene transfer and subsequent expression of a wild type CF plasmid in CF airway cells results in correction or reversal of this glycosylation phenotype. We hypothesize that the changes in glycosylation seen in CF cells are key in the pathophysiology of the cystic fibrosis airway disease. For example, it has been shown that Pseudomonas aeruginosa, a bacterium that has a predilection for colonizing CF airways, adheres to asialylated glycolipids and glycoconjugates with terminal Fuca1,3/4. One focus of our laboratory is to elucidate the etiology of the glycosylation changes seen in CF cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer.
2011-01-07 | GSE26482 | GEO