The challenge of long-term cultivation of human precision-cut lung slices.

Abstract Human precision-cut lung slices (PCLS) have proven to be an invaluable tool for numerous toxicological, pharmacological and immunological studies. For most studies, a cultivation period of less than one week is sufficient, but modelling of complex disease mechanisms and investigating effects of long-term exposure to certain substances require much longer cultivation periods. So far, data regarding tissue integrity of long-term cultivated PCLS is incomplete. Over 1500 human PCLS from 16 different donors were cultivated under standardized, serum-free conditions for up to 28 days and assessed the preservation of viability, tissue integrity and the transcriptome to an unprecedented extent. Even though viability of PCLS was well preserved during long term cultivation, a continuous loss of cells could be observed. While the bronchial epithelium was well preserved over the entire course of cultivation, the alveolar integrity was mostly preserved for 2 weeks and the vasculatory system suffered from significant loss in integrity within the first week. Furthermore, ciliary beat in the small airways gradually decreased after one week. Interestingly, keratinizing squamous metaplasia of the alveolar epithelium with significantly increasing manifestation over time were found. Transcriptome analysis revealed a significantly increased immune response and significantly decreased metabolic activity within the first 24 hours after PCLS generation. Overall, this study provides a comprehensive overview of histomorphological and –pathological changes during long-term cultivation of PCLS.