Habibie, H.; Vlasma, J. R.; Putri, K.; Song, S.; Jager, M. d.; Petersen, A.; Boorsma, C. E.; Cool, R. H.; Quax, W.; Nawijn, M.; Timens, W.; Burgess, J. K.; Brandsma, C.-A.; Melgert, B. N.

Introduction: Chronic obstructive pulmonary disease (COPD) is the third leading cause of death globally, with progressive emphysema driven by repeated epithelial damage and impaired repair. Recently, we found that secretion of cytokine receptor agonist of nuclear factor {kappa}B signaling ligand (RANKL) is higher in lung fibroblasts from patients with COPD compared to control and that RANKL represses lung epithelial cell death. However, the underlying mechanism, its cross-species conservation and the involved epithelial cell types remain unclear. Methods: To investigate how RANKL affects lung epithelial cells we employed in vitro and in vivo models. Primary lung organoids from human and mouse epithelial cells assessed progenitor activation and expansion. Furthermore, we employed a murine model of lung damage to examine which epithelial cell types were affected by RANKL in vivo and whether this was altered upon lung damage. Results: RANKL supplementation increased organoid forming capacity in both murine and human epithelial cells compared to vehicle treated controls. Finally, in elastase-treated mice, RANKL was able to rescue the elastase-induced loss of epithelial cells compared to vehicle-treated controls and it augmented the proportion of epithelial cells in transitional states expressing Krt8 and MHCII. Conclusion: RANKL signaling in lung epithelial cells resulted in enhanced cell survival, primarily observed in alveolar epithelial cells in both mice and humans. In vivo, RANKL was able to expand transitional cell states after elastase-induced lung injury. These results suggest that RANKL is a novel factor in epithelial repair and that its signaling pathway could be explored for therapeutic applications.