Perturbational single-cell genomics to decode trajectories and interactions of regenerative intermediate cell states during lung fibrogenesis and anti-fibrotic drug treatments

Lang, Niklas Jonathan (2025): Perturbational single-cell genomics to decode trajectories and interactions of regenerative intermediate cell states during lung fibrogenesis and anti-fibrotic drug treatments. Dissertation, LMU München: Medizinische Fakultät

Abstract

Pulmonary Fibrosis (PF) is the central pathomechanism shared across all fibrotic Interstitial Lung Diseases (fILD), a heterogeneous group of diseases with limited therapeutic options and poor outcomes. Fibrotic remodeling of the alveolar gas exchange unit due to impaired regeneration following injury is the common pathophysiologic feature of all fILDs. Recent studies probing the cellular underpinnings of these processes have highlighted the emergence of regenerative intermediate cell states (RICS) as transitional stages regulating whether injury leads to regeneration or fibrosis. Most of these studies are, however, focused on end stage disease, which by design provide only limited insights into the role and contributions of RICS during disease initiation and progression. This thesis aims to investigate the emergence, interactions, and cellular trajectories of RICS underlying lung fibrogenesis and their modulation in response to anti-fibrotic drug treatments. To this end, it leverages single-cell RNA sequencing (scRNA-seq) in combination with both lineage tracing in mice and human precision-cut lung slices (hPCLS) treated with a pro-fibrotic cytokine mix to induce de novo fibrotic remodeling and study the cellular responses to pharmacological perturbations. This thesis shows that hPCLS not only retain the full cellular repertoire of the human lung but also reveals that they can recapitulate the fibrogenic RICS observed in lungs from patients with PF upon stimulation with pro-fibrotic cytokines. During early disease alveolar type 2 cells differentiate into fibrogenic KRT17+/KRT5- basaloid cells, while capillary cells give rise to fibrogenic PLVAP/VWA1+ endothelial cells. Additionally, CTHRC1+ myofibroblasts arise from transcriptional convergence of activated stromal cell populations rather than from a single progenitor cell type. Micro-CT staged patient tissues and cell-cell communication analyses map the appearance and interactions of these fibrogenic RICS, validating their appearance during early disease stages. Finally, this work explores the potential of hPCLS coupled to scRNA-seq for phenotypic drug testing directly in human lung tissue by providing a computational framework that accurately confirms known and identifies previously unrecognized mechanisms of the anti-fibrotic drug nintedanib. In summary, this thesis introduces a novel framework to study lung fibrogenesis directly in human lung tissue, enabling spatiotemporal analysis at single-cell resolution. The results advance our understanding of the emergence, interactions, and lineage relationships of fibrogenic RICS during disease initiation, which could inform novel disease-modifying therapeutics. Ultimately, it showcases the potential of hPCLS for next generation, high-resolution drug testing directly in human lung tissue - the environment where effective drugs would have to unfold their antifibrotic properties - thereby opening new avenues for accelerated drug development and translation.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 18. Dezember 2025
• 1. Bericht­erstatter:in: Behr, Jürgen