Senescence intensifies bleomycin-induced injury in IPF-lung epithelial cells

Introduction Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease involving injury and a dysregulated repair response of an ageing epithelium. Here we investigate the contribution of senescence and telomere shortening to bleomycin-induced injury in alveolar epithelial cells (EpCs) derived from lung parenchymal tissue of patients with pulmonary fibrosis (PF), predominantly IPF, in vitro . Methods Injury responses of EpCs in air liquid interface (ALI) culture were assessed by ELISA, qPCR and/or immunofluorescence at 48 and 96 h post-bleomycin (30 μg·mL −1 ) treatment. Telomere length was evaluated by qPCR and corroborated by fluorescence in situ hybridisation (FISH). Results Bleomycin treatment increased markers of inflammation ( i.e. interleukin-6 and −36γ) and senescence ( i.e. p16 and p21) in EpCs from PF patients (PF-EpCs) ( p <0.05); responses markedly higher than control EpCs (Ctrl-EpCs). Conversely, the effects of bleomycin on the expression of type I and type II alveolar EpC markers were supressed or down-regulated in cultures of PF-EpCs (when compared to Ctrl-EpCs), whereas the percentage of keratin 17 positive type I and II intermediate EpCs were increased ( p <0.05). Telomere length was also significantly shorter in PF- than Ctrl-EpCs at baseline and further reduced post-bleomycin treatment ( p <0.05). Induction of telomere shortening by cell passaging augmented bleomycin-induced PF-EpC injury, whereas pharmacological telomerase activation and selected senotherapeutics were protective. Conclusion This study provides evidence that senescence-associated telomere shortening increases the susceptibility of IPF-lung EpCs to injury. Our cell-based lung epithelial injury model has utility as a pre-clinical tool for drug discovery and the evaluation of potential senotherapeutics for IPF treatment.