Single cell transcriptomes of mouse bladder urothelium uncover novel cell type markers and urothelial differentiation characteristics.

Objectives: Much of the information to date in terms of subtypes and function of bladder urothelial cells were derived from anatomical location or by the expression of a small number of marker genes. To have a comprehensive map of the cellular anatomy of bladder urothelial cells, we performed single-cell RNA-sequencing to thoroughly characterize mouse bladder urothelium. Materials and methods: A total of 18,917 single cells from mouse bladder urothelium was analyzed by unbiased single-cell RNA sequencing. The expression of the novel cell marker was confirmed by immunofluorescence using urinary tract infections models. Results: Unsupervised clustering analysis identified 8 transcriptionally distinct cell subpopulations from mouse bladder urothelial cells. We discovered a novel type of bladder urothelial cells marked by Plxna4 that may be involved with host response and wound healing. We also found a group of basal-like cells labeled by ASPM that could be the progenitor cells of adult bladder urothelium. ASPM+ urothelial cells are significantly increased after injury by UPEC. In addition, specific transcription factors were found to be associated with urothelial cell differentiation. At the last, a number of interstitial cystitis/bladder pain syndrome-regulating genes were found differentially expressed among different urothelial cell subpopulations. Conclusions: Our study provides a comprehensive characterization of bladder urothelial cells, which is fundamental to understanding the biology of bladder urothelium and associated bladder disease. Overall design: One sample from murine urothelium. A total of 18,917 single cells from mouse bladder urothelium was analyzed by unbiased single-cell RNA sequencing.