Intrinsic Phenotypic Differences of Asthmatic Epithelium and Its Inflammatory Responses to Respiratory Syncytial Virus and Air Pollution

Abstract

A substantial proportion of healthcare cost associated with asthma is attributable to exacerbations of the disease. Within the airway, the epithelium forms the mucosal immune barrier, the first structural cell defense against common environmental insults such as respiratory syncytial virus (RSV) and particulate matter. We sought to characterize the phenotype of differentiated asthmatic-derived airway epithelial cultures and their intrinsic inflammatory responses to environmental challenges. Air-liquid interface (ALI) cultures were generated from asthmatic (n = 6) and nonasthmatic (n = 6) airway epithelial cells. Airway tissue and ALI cultures were analyzed by immunohistochemistry for cytokeratin-5, E-cadherin, Ki67, Muc5AC, NF-κB, the activation of p38, and apoptosis. ALI cultures were exposed to RSV (4 × 10(6) plaque forming unit/ml), particulate matter collected by Environmental Health Canada (EHC-93, 100 μg/ml), or mechanically wounded for 24, 48, and 96 hours and basolateral supernatants analyzed for inflammatory cytokines, using Luminex and ELISA. The airway epithelium in airway sections of patients with asthma as well as in vitro ALI cultures demonstrated a less differentiated epithelium, characterized by elevated numbers of basal cells marked by the expression of cytokeratin-5, increased phosphorylation of p38 mitogen-activated protein kinase, and less adherens junction protein E-cadherin. Transepithelial resistance was not different between asthmatic and nonasthmatic cultures. In response to infection with RSV, exposure to EHC-93, or mechanical wounding, asthmatic ALI cultures released greater concentrations of IL-6, IL-8, and granulocyte macrophage colony-stimulating factor, compared with nonasthmatic cultures (P < 0.05). This parallel ex vivo and in vitro study of the asthmatic epithelium demonstrates an intrinsically altered phenotype and aberrant inflammatory response to common environmental challenges, compared with nonasthmatic epithelium.

A Heterotopic Xenograft Model of Human Airways for Investigating Fibrosis in Asthma

“…Both the asthmatic grafted epithelium and the asthmatic human lung biopsy tissue show decreased E-cadherin expression. This is all consistent with our previous work showing that, when differentiated at air-liquid interface culture, asthmatic-derived epithelial cells are less differentiated in comparison to nonasthmatic-derived cells, as demonstrated by increased basal cell number and lack of E-cadherin (5,6,11,19).…”

“…An important feature of asthma is the loss of junction proteins in the airway epithelium (18), such as E-cadherin (19). This is demonstrated in asthmatic human airway tissue by a 35% lower level of E-cadherin compared with nonasthmatic tissues ( Figure 2C).…”

Non-animal models of epithelial barriers (skin, intestine and lung) in research, industrial applications and regulatory toxicology

“…Such changes have been seen to correspond to an increase in ionic and macromolecular permeability (Xiao et al, 2011;Bai et al, 2015;Freishtat et al, 2011;Watson et al, 2010). It has also been shown that the permeability of the bronchial epithelial cell layer is only increased in asthmatic subjects after exposure to ozone or nitrogen dioxide (Bayram et al, 2002), and that mediator release is altered after exposure of ALI cultures from asthmatic donors to respiratory syncytial virus (RSV) or particulate matter (Hackett et al, 2011b). Using cells derived from patients with COPD, ALI cultures show higher baseline levels of cytokine expression and increased susceptibility to RSV infection, despite an increased IFN response tem is specially designed for direct contact between cells and components of the test atmosphere at the air/liquid interface.…”

Transforming Growth Factor β1 Function in Airway Remodeling and Hyperresponsiveness. The Missing Link?

“…Epithelial cell-to-cell contacts, however, may become compromised in the airways of subjects with asthma, allowing epithelial cell mediators to reach the underlying structural cells. Several studies show that many adhesion proteins necessary for epithelial cell-to-cell contacts are downregulated in the airway epithelium of patients with asthma (131)(132)(133). Because of this uncertainty, determining the circumstances under which airway epithelium and subepithelium modify HASM contraction and relaxation responses will be essential for future studies investigating the effect of the surrounding milieu on HASM contraction and AHR.…”