Cultured alveolar basal cells differ in their transcriptome, but display similar characteristics and functional properties to airway basal cells

<bold>Rationale:</bold> Basal cells (BC) ectopically appear within the alveolar space of the lungs of interstitial lung disease (ILD) patients. The role, function and origin of these ectopic alveolar BC remains poorly understood. By comparing ectopic alveolar to “classical” airway BC, we aimed to get a better understanding of the origin and disease-specific characteristics of alveolar BC in ILD. <bold>Methods:</bold> Alveolar and airway BC were isolated from transbronchial- and airway mucosal biopsies of the same ILD patient and were cultured on plastic, an air liquid interface (ALI), or in a 3D organoid model. Additionally, BC were intratracheally injected into bleomycin-challenged mice. Samples were analyzed by single cell RNA sequencing (scRNA-seq), TaqMan RT-PCR, immunocytochemistry/immunofluorescence (IF), or immunohistochemistry/IF. <bold>Results:</bold> scRNA-seq analysis revealed several differentially expressed genes in alveolar compared to airway BC. However, BC marker expression and the capacity for wound repair, proliferation, organoid formation and differentiation was similar in alveolar and airway BC. Furthermore, there was no difference in engraftment and differentiation potential after instillation into bleomycin-injured mice between the two human BC types. <bold>Conclusion:</bold> Close similarities between alveolar and airway BC may indicate that alveolar BC derive from migrating airway BC. Transcriptomic differences may arise through imprinting of alveolar and airway BC by their specific in vivo microenvironment. If transcriptomic differences result in any functional differences or may serve as potential drug targets needs to be determined in future studies.