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Transcriptomic and proteomic host response to Aspergillus fumigatus conidia in an air-liquid interface model of human bronchial epithelium.

ABSTRACT: Aspergillus fumigatus (A. fumigatus) is a wide-spread fungus that is a potent allergen in hypersensitive individuals but also an opportunistic pathogen in immunocompromised patients. It reproduces asexually by releasing airborne conidiospores (conidia). Upon inhalation, fungal conidia are capable of reaching the airway epithelial cells (AECs) in bronchial and alveolar tissues. Previous studies have predominantly used submerged monolayer cultures for studying this host-pathogen interaction; however, these cultures do not recapitulate the mucocililary differentiation phenotype of the in vivo epithelium in the respiratory tract. Thus, the aim of this study was to use well-differentiated primary human bronchial epithelial cells (HBECs) grown at the air-liquid interface (ALI) to determine their transcriptomic and proteomic responses following interaction with A. fumigatus conidia. We visualized conidial interaction with HBECs using confocal laser scanning microscopy (CLSM), and applied NanoString nCounter and shotgun proteomics to assess gene expression changes in the human cells upon interaction with A. fumigatus conidia. Western blot analysis was used to assess the expression of top three differentially expressed proteins, CALR, SET and NUCB2. CLSM showed that, unlike submerged monolayer cultures, well-differentiated ALI cultures of primary HBECs were estimated to internalize less than 1% of bound conidia. Nevertheless, transcriptomic and proteomic analyses revealed numerous differentially expressed host genes; these were enriched for pathways including apoptosis/autophagy, translation, unfolded protein response and cell cycle (up-regulated); complement and coagulation pathways, iron homeostasis, nonsense mediated decay and rRNA binding (down-regulated). CALR and SET were confirmed to be up-regulated in ALI cultures of primary HBECs upon exposure to A. fumigatus via western blot analysis. Therefore, using transcriptomics and proteomics approaches, ALI models recapitulating the bronchial epithelial barrier in the conductive zone of the respiratory tract can provide novel insights to the molecular response of bronchial epithelial cells upon exposure to A. fumigatus conidia.

SUBMITTER: Toor A

PROVIDER: S-EPMC6307744 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Transcriptomic and proteomic host response to Aspergillus fumigatus conidia in an air-liquid interface model of human bronchial epithelium.

Toor Amreen A Culibrk Luka L Singhera Gurpreet K GK Moon Kyung-Mee KM Prudova Anna A Foster Leonard J LJ Moore Margo M MM Dorscheid Delbert R DR Tebbutt Scott J SJ

PloS one 20181227 12

Aspergillus fumigatus (A. fumigatus) is a wide-spread fungus that is a potent allergen in hypersensitive individuals but also an opportunistic pathogen in immunocompromised patients. It reproduces asexually by releasing airborne conidiospores (conidia). Upon inhalation, fungal conidia are capable of reaching the airway epithelial cells (AECs) in bronchial and alveolar tissues. Previous studies have predominantly used submerged monolayer cultures for studying this host-pathogen interaction; howev ...[more]

PMID: 30589860

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Project description:Aspergillus fumigatus (A. fumigatus) is a ubiquitous fungus which reproduces asexually by releasing abundant airborne conidia (spores), which are easily respirable. In allergic and immunocompromised individuals A. fumigatus can cause a wide spectrum of diseases, including allergic bronchopulmonary aspergillosis, aspergilloma and invasive aspergillosis. Previous studies have demonstrated that A. fumigatus conidia are internalized by macrophages and lung epithelial cells; however the exact transcriptional responses of airway epithelial cells to conidia are currently unknown. Thus, the aim of this study was to determine the transcriptomic response of the human bronchial epithelial cell line (16HBE14o-) following interaction with A. fumigatus conidia. We used fluorescence-activated cell sorting (FACS) to separate 16HBE14o- cells having bound and/or internalized A. fumigatus conidia expressing green fluorescent protein from cells without spores. Total RNA was then isolated and the transcriptome of 16HBE14o- cells was evaluated using Agilent Whole Human Genome microarrays.Immunofluorescent staining and nystatin protection assays demonstrated that 16HBE14o- cells internalized 30-50% of bound conidia within six hrs of co-incubation. After FAC-sorting of the same cell culture to separate cells associated with conidia from those without conidia, genome-wide analysis revealed a set of 889 genes showing differential expression in cells with conidia. Specifically, these 16HBE14o- cells had increased levels of transcripts from genes associated with repair and inflammatory processes (e.g., matrix metalloproteinases, chemokines, and glutathione S-transferase). In addition, the differentially expressed genes were significantly enriched for Gene Ontology terms including: chromatin assembly, G-protein-coupled receptor binding, chemokine activity, and glutathione metabolic process (up-regulated); cell cycle phase, mitosis, and intracellular organelle (down-regulated).We demonstrate a methodology using FACs for analyzing the transcriptome of infected and uninfected cells from the same cell population that will provide a framework for future characterization of the specific interactions between pathogens such as A. fumigatus with human cells derived from individuals with or without underlying disease susceptibility.

Dataset Information

Transcriptomic and proteomic host response to Aspergillus fumigatus conidia in an air-liquid interface model of human bronchial epithelium.

Publications

Transcriptomic and proteomic host response to Aspergillus fumigatus conidia in an air-liquid interface model of human bronchial epithelium.

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