Spatial transcriptome analysis at single-cell resolution for understanding IPF pathogenesis

Introduction: Recent advances in single-cell RNA sequencing (scRNA-seq) and spatial transcriptomic technologies have provided extensive insight into the cellular composition of lungs. Histopathological pattern of idiopathic pulmonary fibrosis (IPF) is corresponding to usual interstitial pneumonia (UIP), which is characterized by temporal and spatial heterogeneity of fibrosing process composed of densely collagenized fibrosis and immature fibroblast foci. To further understand the molecular mechanism of heterogeneity in fibrosis, we simultaneously examined scRNA-seq and spatial transcriptome analysis by using IPF lungs. Methods: We performed scRNA-seq on 3 IPF and 3 normal lung tissues. For spatial transcriptome analysis, we analyzed 9 lung tissue sections from 3 IPF patients. Results: For scRNA-seq, we profiled a total of 29,872 cells and the average number of reads was 97,207 reads/cell. For spatial transcriptome analysis, total 21,523 spots were analyzed and the average number of reads was 148,284 reads/spot. In Comparison to normal lungs, characteristic cell distribution was demonstrated in IPF lungs by scRNA-seq. Deconvolution of the single-cell transcriptome and spatial transcriptomic spots revealed cellular heterogeneity within the fibroblast foci. By comparing locally expressed genes between dense fibrosis and fibroblastic foci, we elucidated the difference in mesenchymal cell populations and identified novel profibrotic markers mainly expressed in fibroblast foci. Conclusions: We demonstrated cellular heterogeneity within the fibroblast foci and identified novel profibrotic markers expressed in fibroblast foci, which may have a role in fibrosis progression during IPF pathogenesis.