Role of stromal cells during human iPSC differentiation into bronchial epithelium

Abstract

Background: Chronic airway diseases are a common medical condition worldwide including diseases such as cystic fibrosis, asthma or COPD. Induced pluripotent stem cells (iPSC) provide a new approach to study lung diseases. We and others have recently generated functional airway multiciliated epithelium in air-liquid interface culture conditions from human iPSC (iALI) (Ahmed et al., BioRxiv, 2020). During this differentiation process, a mesenchymal stromal component is consistently described but poorly studied.

Objective/Methods: To comprehensively define the cell types, mechanisms, and mediators driving iALI model, we performed single-cell mRNA sequencing. RT-qPCR and immunofluorescence assays were used to confirm the sequencing results.

Results: In addition to the epithelial cell subtypes, the iALI cultures also contain a previously poorly documented EPCAM-COL1A1+DCN+ mesenchymal stromal compartment. Several populations, that partially overlap can be identified: 1/ACTA2+ myofibroblasts including fibromyocytes characterized by high expression of contractile genes such as CNN1 and TAGLN, 2/ a transition population closer to epithelial cells expressing MAF and the long non coding mRNA EPB41L4A-AS1 and 3/ a highly proliferating PCNA+CDK1+ cells. Moreover, stromal cells expressed growth factors such as FGF10 while its receptor FGFR2 is expressed on the epithelial cells, suggesting that stromal cells could support iALI development.

Conclusion: These data provide high-resolution insights into the complexity and plasticity of the iALI suggesting that a crosstalk between epithelium and mesenchyme can contribute to iALI development. The functional role of the relevant pathways will be assessed in our iALI model.