Integrated single cell analysis of human lung fibrosis resolves cellular origins of predictive protein signatures in body fluids

Mayr, Christoph H.; Simon, Lukas M.; Leuschner, Gabriela; Ansari, Meshal; Geyer, Philipp; Angelidis, Ilias; Strunz, Maximilian; Singh, Pawandeep; Kneidinger, Nikolaus; Reichenberger, Frank; Silbernagel, Edith; Boehm, Stephan; Adler, Heiko; Hilgendorff, Anne; Lindner, Michael; Prasse, Antje; Behr, Jüergen; Mann, Matthias; Eickelberg, Oliver; Theis, Fabian J.; Schiller, Herbert B.

doi:10.1101/2020.01.21.20018358

Cited by 8 publications

(7 citation statements)

References 60 publications

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“…However, in a recent study an Ebf1 high fibroblast population was identified as a distinct cluster in a scRNA-seq analysis of wound fibroblasts 61 . Recent pre-print publications identified an “adventitial fibroblast” subtype with a similar transcriptomic signature to the Ebf1/EBF1 population in our study 11,12 . The in-situ hybridization localization of SFRP2, SERPINF1, PI16 prominent genes in the Ebf1/EBF1 cluster we identify in a recent study are compatible with the results of our Ebf1 immunofluorescence localizing a proportion of Ebf1 + fibroblasts to the adventitia 12 .…”

Section: Discussionsupporting

confidence: 66%

Definition and Signatures of Lung Fibroblast Populations in Development and Fibrosis in Mice and Men

Rowan

Liang

et al. 2020

Preprint

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The heterogeneity of fibroblasts in the murine and human lung during homeostasis and disease is increasingly recognized. It remains unclear if the different phenotypes identified to date are characteristic of unique subpopulations with unique progenitors or whether they arise by a process of differentiation from common precursors. Our understanding of this ubiquitous cell type is limited by an absence of well validated, specific, markers with which to identify each cell type and a clear consensus on the distinct populations present in the lung. Here we describe single cell RNA sequencing (scRNA-seq) analysis on mesenchymal cells from the murine lung throughout embryonic (E) development (E9.5 – 17.5), at post-natal day (P1 – 15), as well as in the adult and the aged murine lungs before and after bleomycin-induced fibrosis. We carried out complementary scRNA-seq on human lung tissue from a P1 lung, a month 21 lung and lung tissue from healthy donors and patients with idiopathic pulmonary fibrosis (IPF). The murine and human data were supplemented with publicly available scRNA-seq datasets. We consistently identified lipofibroblasts, myofibroblasts, pericytes, mesothelial cells and smooth muscle cells. In addition, we identified a novel population delineated by Ebf1 (early B-cell factor 1) expression and an intermediate subtype. Comparative analysis with human mesenchymal cells revealed homologous mesenchymal subpopulations with remarkably conserved transcriptomic signatures. Comparative analysis of changes in gene expression in the fibroblast subpopulations from age matched non-fibrotic and fibrotic lungs in the mouse and human demonstrates that many of these subsets contribute to matrix gene expression in fibrotic conditions. Subtype selective transcription factors were identified and putative divergence of the clusters during development were delineated. Prospective isolation of these fibroblast subpopulations, localization of signature gene markers, and lineage-tracing each cluster are under way in the laboratory. This analysis will enhance our understanding of fibroblast heterogeneity in homeostasis and fibrotic disease conditions.

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Section: Discussionsupporting

confidence: 66%

Definition and Signatures of Lung Fibroblast Populations in Development and Fibrosis in Mice and Men

Rowan

Liang

et al. 2020

Preprint

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“…We here identified and characterized the transient appearance of Krt8 + ADI cells during lung regeneration. Single cell analysis of human lung fibrosis recently identified a disease specific cell state that was termed aberrant basaloid cell (KRT17+/KRT5-) based on some similarities to airway basal cells 42,43 . It is currently unclear if these cells are indeed airway derived or could represent stem cells undergoing alveolar repair.…”

Section: Involvement Of Airway Stem Cells In Alveolar Regenerationmentioning

confidence: 99%

Alveolar regeneration through a Krt8+ transitional stem cell state that persists in human lung fibrosis

et al. 2020

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The cell type specific sequences of transcriptional programs during lung regeneration have remained elusive. Using time-series single cell RNA-seq of the bleomycin lung injury model, we resolved transcriptional dynamics for 28 cell types. Trajectory modeling together with lineage tracing revealed that airway and alveolar stem cells converge on a unique Krt8 + transitional stem cell state during alveolar regeneration. These cells have squamous morphology, feature p53 and NFkB activation and display transcriptional features of cellular senescence. The Krt8+ state appears in several independent models of lung injury and persists in human lung fibrosis, creating a distinct cell-cell communication network with mesenchyme and macrophages during repair. We generated a model of gene regulatory programs leading to Krt8+ transitional cells and their terminal differentiation to alveolar type-1 cells. We propose that in lung fibrosis, perturbed molecular checkpoints on the way to terminal differentiation can cause aberrant persistence of regenerative intermediate stem cell states.

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“…Markers of several pro-fibrogenic cell types were strongly increased in protein measurements of patients that were diagnosed with different forms of interstitial lung diseases (ILD) including patients with IPF when compared to non-ILD controls. These protein signatures correlated with diagnosis, lung function, smoking and injury status (27). scRNAseq data also offer the opportunity to identify transcription factors, signaling interactions and hormone/immune targets linked to disease onset and progression (20).…”

Section: New Insights From Scrnaseq Studies Into Disease Progression mentioning

confidence: 98%

“…scRNAseq studies have vastly increased our understanding of cell types resident in the distal lung and also highlighted differences in cell populations/states between healthy and IPF tissue (12-15, 20, 25, 28, 44, 45). Recent efforts aimed at extending these insights to a better understanding of mechanisms for the local and temporal disease progression or leveraging scRNAseq analysis to progress disease biomarker identification (27,46). Subclassifying IPF tissue samples into three pathological defined groups of no fibrosis detectable, mild fibrosis and advanced fibrosis a recent study detected large changes in IPF patients' samples even before histological changes in the respective regions were detectable.…”

Section: New Insights From Scrnaseq Studies Into Disease Progression mentioning

confidence: 99%

“…In addition, the abundance of data generated in scRNAseq studies are currently lacking realistic methods of validation on the protein level due to the lack of methods for proteomics at this scale. Albeit, first attempts are under way link scRNAseq data and proteomic analysis to identify disease specific changes on the cellular level and their corresponding reflection in body fluid proteomes (27). Animal studies, despite their limitations, will also benefit in validating data from scRNAseq studies in experimental models.…”

Section: Challenges and Future Perspectivesmentioning

confidence: 99%

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Insights Into Development and Progression of Idiopathic Pulmonary Fibrosis From Single Cell RNA Studies

2020

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Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disease with limited therapeutic options. The current model suggests that chronic or repetitive “micro-injuries” of the alveolar epithelium lead to activation and proliferation of fibroblasts and excessive extracellular matrix (ECM) deposition. Disruption of alveolar type II (ATII) epithelial cell homeostasis and the characteristics of mesenchymal cell populations in IPF have received particular attention in recent years. Emerging data from single cell RNA sequencing (scRNAseq) analysis shed novel light on alterations in ATII cell progenitor dysfunction and the diversity of mesenchymal cells within the fibrotic lung. Within this minireview, we summarize the data from most recent human scRNAseq studies. We aim to collate the current knowledge on cellular plasticity and heterogeneity in the development and progression of IPF, effects of drug treatment on transcriptional changes. Finally, we provide a brief outlook on future challenges and promises for large scale sequencing studies in the development of novel therapeutics for IPF.

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Integrated single cell analysis of human lung fibrosis resolves cellular origins of predictive protein signatures in body fluids

Cited by 8 publications

References 60 publications

Definition and Signatures of Lung Fibroblast Populations in Development and Fibrosis in Mice and Men

Definition and Signatures of Lung Fibroblast Populations in Development and Fibrosis in Mice and Men

Alveolar regeneration through a Krt8+ transitional stem cell state that persists in human lung fibrosis

Insights Into Development and Progression of Idiopathic Pulmonary Fibrosis From Single Cell RNA Studies

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