Unknown,Transcriptomics,Genomics,Proteomics

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The response of human primary airway epithelial cells to Influenza or RSV infection [hAECs_Illumina]

ABSTRACT: To investigate how human airway epithelial cells respond to Influenza or RSV infection, we harvested airway epithelial cells from the mainstream bronchi of human donors and cultured them as previously described (Pickles et al,1998) in a polarized system that resembles the in vivo mucociliary pseudostratified epithelium. Quadruplicate hAEC cultures were infected with 2X105 PFUs Influenza A (Udorn) or with 1x106 PFUs RSV for 2h or mock inoculated and harvested 24h after Influenza infection and 48h after RSV infection. Quadruplicate polarized airway epithelial cell cultures were infected with 2x10^5 PFUs of Influenza A (Udorn) for 2h or infected with 1x10^6 PFUs RSV and harvested 24 h post infection for Influenza or 48h post infection for RSV.Duplicate cultures were used as controls for each condition (Two cultures were mock treated mor 2h and harvested after 24h for the Influenza infection and 2 cultures were mock treated for 2h and harvested after 48 hours for the RSV infection.Total RNA was harvested and gene expression was studied using Genespring GX v7.3.1.

ORGANISM(S): Homo sapiens

SUBMITTER: IOANNIS IOANNIDIS

PROVIDER: E-GEOD-32139 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Lower respiratory tract infections (LRTI) from human Respiratory Syncytial Virus (RSV) are a significant cause of morbidity in children. A component of LRTI pathogenesis is due to signals generated by infected lower airway cells that alter lymphocyte populations and trigger airway remodeling. To obtain insights into this process, we applied an unbiased quantitative proteomics analysis of the RSV-induced epithelial secretory response in cells representative of the trachea (hBECs) vs small airway bronchiolar cells (hSAECs). A workflow was standardized initially using telomerase immortalized human epithelial cells that showed high reproducibility and cell-type differences in proteomic signatures of both secreted proteins and isolated nanoparticles (exosomes). Over half of secretome proteins were contained within exosomal, with the remainder originating from lysosomal and vaculolar compartments. We applied this workflow to three independently derived primary human cultures. 577 differentially expressed proteins from control supernatants and 966 differentially expressed proteins from RSV-infected cell supernatants were identified at a 1% false discovery rate (FDR). 15 proteins were unique to RSV-infected hBECs regulated by epithelial-specific ets homology factor (EHF). 106 proteins were unique to RSV-infected hSAECs enriched in proteins regulated by the NFB transcription factor. In this latter group, we independently validated the differential expression of Chemokine (C-C Motif) Ligand 20 (CCL20)/macrophage inducible protein (MIP)3, Thymic Stromal Lymphopoietin (TSLP) and chemokine (CC) ligand 3-like 1(CCL3-L1). CCL20/MIP3 is the most active mucin inducing factor in the RSV infected hSAEC secretome, and is differentially expressed in smaller airways in a mouse model of RSV infection. These studies provide insight into role of exosomal production in innate responses and regional differences in epithelial secretomes that participate in pathogenesis of RSV LRTI-induced airway remodeling.

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The response of human primary airway epithelial cells to Influenza or RSV infection [hAECs_Illumina]

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