Characterising the changes in alveolar stem cells and immune cells during development of chronic lung fibrosis

Thesis

In the thesis, I hypothesised that the alveolar epithelial landscape during chronic fibrosis is greatly altered and contains altered alveolar epithelial cells that interacted with immune cells to contribute to abnormal alveolar regeneration and persistent fibrosis. I used publicly deposited data and our own IPF lung lavage scRNA-seq data to examine and characterize damage associated alveolar epithelial cells and immune cells in IPF. I then generated an acute (resolving lung fibrosis) and chronic (non-resolving) fibrotic murine model to allow me to understand the regenerating alveolar epithelial landscape and identify the potential immune drivers of non-resolving, chronic fibrosis. My studies show that there are likely two subsets of alveolar intermediates [alveolar damage-associated intermediate cells (ADIs) in murine studies, and aberrant basaloid (AB) cells in human IPF lungs)]), one originating from club cells, and the other from AT2 cells. The former may be more relevant to chronic fibrosis as it was found more abundant in the later time points after lung injuryinjuries and in both human and murine lungs, a subset of alveolar intermediate cells expressed Pmepa1 and Tgfbr1, which are highly relevant to the pathogenesis of IPF. In the chronic fibrosis vs acute (resolving) fibrotic model, a new epithelial landscape is revealed by single nucleus RNA sequencing (snRNA-seq), not known previously due to inability of scRNA-seq methods to capture these cells. This landscape is dominated by emergence of an AT1 subset highly expressing Notch signalling genes which is stable in abundance and transcriptome after the ‘tipping point’ to chronic fibrosis, in contrast to AT1 subsets in acute fibrotic model. There was also an increased expression of Th2 cytokines in AT2 cells in chronic fibrotic mice, which may affect the differentiation of neighbouring macrophages.

By flow cytometry and snRNA-seq of murine lung digests from the chronic and acute fibrotic model and by scRNA-seq in bronchoalveolar lavage of IPF patients, macrophages subsets were the most abundant cell compared to naive and healthy control lungs. However comparing immune cell profile between chronic and acute fibrotic mice, only DCs were found to be statistically different between the two models by flow cytometry (lower in chronic fibrotic model) , and CD163^-CD9⁺ alveolar macrophages (AMs)IMs by snRNA-seq (higher in chronic fibrotic model).

These findings map out a more complete landscape for epithelial and immune cells in the fibrotic lung, and uncovers epithelial and immune cell types that may be involved in chronic fibrosis.

Authors: Hu, T

• DOI: 10.5287/ora-5ryoyy7eg
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford
• Language: English
• Keywords: repair and regeneration; chronic lung fibrosis; idiopathic pulmonary fibrosis; immune cells; alveolar epithelial cells
• Subjects: Immunology; Epithelial cells; Regeneration (biology); Respiratory organs; Pulmonary fibrosis