Abstract Rationale Interstitial lung diseases (ILDs) are a group of conditions involving inflammation and fibrosis of the lung parenchyma and are associated with a variety of causes and risk factors. Existing single-cell RNA sequencing (scRNA-seq) analyses of the human lung have characterized the healthy lung as well as various states of disease, including idiopathic pulmonary fibrosis (IPF), the most common type of ILD. We hypothesized that integrated analysis of scRNA-seq of explant lung tissue from different ILD phenotypes compared with healthy donors would uncover shared molecular programs across ILDs and establish candidate regulators of distinct ILD phenotypes. Methods Single-cell suspensions were generated from 68 declined donor and 126 ILD lung explants from two lung transplant centers. Suspensions underwent scRNA-seq library preparation using the 10X Genomics Chromium 5’ system and libraries were sequenced on an Illumina Novaseq6000. After ambient RNA correction and quality control filtering, libraries were integrated and batch corrected, and cell types were annotated. Subject-level pseudobulked expression matrices were generated for each cell type and used to detect differentially expressed genes between ILD diagnoses and control. Co-expression module analysis and corresponding transcription factor regulatory network analysis were performed for cell types of interest. Results Analyzing 1,155,549 cells from these 194 unique lung samples, we identified 63 distinct cell types and states across samples. Compositional and differential expression analyses between ILD phenotypes with samples from at least 5 subjects (IPF, immune-related ILD, chronic hypersensitivity pneumonitis, coal worker’s pneumoconiosis, and sarcoidosis) and controls revealed significant changes in cell type proportion and transcriptional programs in the distal epithelium across phenotypes. Largely overlapping differentially expressed gene sets indicated broadly similar patterns of epithelial dysfunction in IPF and immune-related ILD, confirmed by gene set enrichment analysis. Transcription factor regulatory network analysis performed on cell type-specific gene co-expression modules identified common transcription factor regulons across these phenotypes, including upregulation of SOX4, TCF3/4, TRPS1, and RARG in multiple phenotypes. In contrast, downregulated regulons differed between phenotypes, including a decrease in expression of regulons mediated by FOS, JUN, and EGR1 in immune-related ILDs. These analyses suggest shared patterns of epithelial dysfunction with some differences in transcriptional regulation between ILD phenotypes. Conclusions Integration of transcriptomic profiles of various ILD lungs with healthy lungs revealed compositional and expression-based differences between ILD phenotypes and controls. We identified widespread dysregulation of distal epithelial cells across ILD phenotypes and candidate transcription factors that may regulate changes in expression profiles across ILDs. This abstract is funded by: NIH/NHLBI, DOD
Dietrich et al. (Fri,) studied this question.