A urine-dependent human urothelial organoid offers a viable alternative to rodent models of infection

Murine models describe a defined host/pathogen interaction for urinary tract infection, but human cell studies are scant. Although recent human urothelial organoid models are promising, none tolerate urine, the natural substrate of the tissue and of the uropathogens that live there. We developed a novel human organoid from progenitor cells which demonstrates the key structural hallmarks and biomarkers of the urothelium. After three weeks of transwell culture with 100% urine at the apical interface, the surface of the three-dimensional organoid differentiated into enlarged and flattened umbrella cells bearing characteristic tight junctions, asymmetric until membrane plaques, fusiform vesicles and a glycosaminoglycan layer. The umbrella cells also expressed apical cytokeratin-20, a spatial feature of the mammalian urothelium. Further experiments showed that urine itself is necessary for full development, and that undifferentiated cells are urine-tolerant despite the lack of membrane plaques and a glycosaminoglycan layer. Infection with Enterococcus faecalis revealed the expected invasive outcome, including urothelial sloughing and the formation of intracellular colonies similar to those previously observed in patient cells. This new biomimetic model could help illuminate invasive behaviours of uropathogens in the human system, and serve as a reproducible test bed for disease formation, treatment and resolution in patients. . CC-BY-NC-ND 4.0 International license not peer-reviewed) is the author/funder. It is made available under a The copyright holder for this preprint (which was . http://dx.doi.org/10.1101/152033 doi: bioRxiv preprint first posted online Jun. 20, 2017;