Role of CFTR in airway epithelial differentiation and regeneration

<bold>Background:</bold> The loss of functional CF transmembrane conductance regulator (CFTR) has been shown to compromise the regeneration process of the airway epithelium after injury. However, the mechanistic insights remain unclear. <bold>Objective:</bold> Exploring the mechanistic role of CFTR in airway epithelial cell differentiation/regeneration. <bold>Methods:</bold> Epithelial/mesenchymal markers were assessed by Western Blot (WB) and immunofluorescence in CF and non-CF cells. Wound-closure of differentiated human BCi-NS1.1 cells was analysed by live-cell microscopy and WB. Using CRISPR/Cas9, two novel basal CF cells were generated. <bold>Results:</bold> Data show an increased N-cadherin expression only in p.Phe508del-CFTR cells vs p.Gly551Asp- and wt-CFTR cells, albeit both CF cells present a lower TEER vs wt-CFTR cells. Fully differentiated BCi-NS1.1 cells are able to close a wound within 72 h with CFTR protein levels increasing over time. Furthermore, two novel isogenic basal cell lines, BCi-CF1.1 (p.Phe508del/p.Phe508del) and BCi-CF2.1 (p.Gly551Asp/p.Gly551Asp) were successfully generated to study the mechanistic role of CFTR. Results show that BCi-CF1.1 and BCi-CF2.1 cells differentiate into the various airway cell types, including ionocytes. Ongoing studies are characterizing both CF cell models exploring the impact of CFTR variants on airway epithelial differentiation and regeneration. <bold>Conclusion:</bold> Plasma membrane localized p.Gly551Asp-CFTR, albeit not functional, improves epithelial junction organization vs ER-retained p.Phe508del-CFTR. Moreover, wound healing studies suggest that CFTR plays a role in these processes. Novel CF cells with differentiation capacity offer unique models to study CFTR's role in epithelial differentiation and regeneration.