Interactions between airway epithelial cells and the microenvironment in idiopathic pulmonary fibrosis
File(s)
Author(s)
Hewitt, Richard
Type
Thesis
Abstract
Idiopathic pulmonary fibrosis (IPF) is a progressive, scarring lung disease driven by alveolar epithelial cell injury, dysregulated fibroblast activity and excessive extracellular matrix (ECM) deposition. Through unknown mechanisms, basal cells, normally restricted to the airway epithelium, are expanded in remodeled fibrotic lung. The aim of this thesis was to understand how airway epithelial cell phenotype and function is shaped through interactions with the local immune and tissue microenvironment in IPF, and how this contributes to disease pathogenesis.
Functional characterization of primary bronchial epithelial cells from newly diagnosed IPF patients and healthy controls was performed in vitro. There were no significant differences in proliferation, wound healing capacity, barrier function or response to viral challenge. Human precision- cut lung slices demonstrated a close spatial relationship between airway epithelial cells and CD206+ airway macrophages (AMs), therefore their molecular phenotype was explored with bulk RNA-sequencing. AMs purified from bronchoalveolar lavage displayed a unique transcriptomic profile with upregulation of genes involved in cell movement and matrix remodelling. Bulk RNA-seq of bronchial brushings exposed transcripts indicative of immune cells, therefore single-cell RNA-seq was utilised to illuminate cellular heterogeneity in the airway mucosa. This revealed expansion in AM subsets in IPF and increased interactions between epithelial secretory cells and AMs.
In distal fibrotic lung tissue, second harmonic generation imaging demonstrated regional differences in collagen organisation. A positive association was identified between the number of KRT5+ basal cells and collagen density and fibre orientation. Basal cell migration was modulated by components of the ECM in vitro. Cell-derived matrices (CDMs) generated from IPF fibroblasts had a distinct matrisome determined by mass spectrometry- based proteomics and were more prohibitive to basal cell migration than control CDMs.
Collectively, these findings demonstrate that interactions between airway epithelial cells and the microenvironment in the airway and distal lung, are integral to IPF pathogenesis.
Functional characterization of primary bronchial epithelial cells from newly diagnosed IPF patients and healthy controls was performed in vitro. There were no significant differences in proliferation, wound healing capacity, barrier function or response to viral challenge. Human precision- cut lung slices demonstrated a close spatial relationship between airway epithelial cells and CD206+ airway macrophages (AMs), therefore their molecular phenotype was explored with bulk RNA-sequencing. AMs purified from bronchoalveolar lavage displayed a unique transcriptomic profile with upregulation of genes involved in cell movement and matrix remodelling. Bulk RNA-seq of bronchial brushings exposed transcripts indicative of immune cells, therefore single-cell RNA-seq was utilised to illuminate cellular heterogeneity in the airway mucosa. This revealed expansion in AM subsets in IPF and increased interactions between epithelial secretory cells and AMs.
In distal fibrotic lung tissue, second harmonic generation imaging demonstrated regional differences in collagen organisation. A positive association was identified between the number of KRT5+ basal cells and collagen density and fibre orientation. Basal cell migration was modulated by components of the ECM in vitro. Cell-derived matrices (CDMs) generated from IPF fibroblasts had a distinct matrisome determined by mass spectrometry- based proteomics and were more prohibitive to basal cell migration than control CDMs.
Collectively, these findings demonstrate that interactions between airway epithelial cells and the microenvironment in the airway and distal lung, are integral to IPF pathogenesis.
Version
Open Access
Date Issued
2021-08
Date Awarded
2022-01
Copyright Statement
Creative Commons Attribution NonCommercial NoDerivatives Licence
Advisor
Lloyd, Clare
Maher, Toby
Byrne, Adam
Sponsor
Imperial College London
Publisher Department
National Heart & Lung Institute
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)