Functional genomics of the human epididymis: further characterization of efferent ducts and model systems by single cell RNA-seq analysis.
Objectives Here we used single cell RNA-seq technologies to elucidate the identity of cells within the human efferent ducts and compared them to caput epididymis cells. We also compared the cellularity of primary tissues with those of 2D and 3D (organoid) culture models used for functional studies.  Materials and Methods Human epididymis tissue was dissected to separate different anatomical regions and digested to release single cells for processing on the 10X Genomics Chromium platform. Primary human epididymis epithelial (HEE) cells and HEE organoids were grown as described previously and subjected to single cell (sc) RNA-seq. ScRNA-seq data were processed by standard bioinformatics pipelines and used for comparative analysis.  Results We define the cell types in the efferent ducts which include specialized epithelial cells, connective tissue stromal cells, vascular endothelial cells, smooth muscle cells and immune cells, but lack basal cells that are seen in the caput epididymis. Furthermore, we identify a sub-population of epithelial cells which have marker genes found in the bladder and urothelium. Comparative genomics analysis of the 2D and 3D culture models shows cellular identities adapted to the culture environment while still maintaining similarity to the primary tissue.  Discussion Our data suggest that efferent ducts are lined with a transitional epithelium, which like the urothelium is able to stretch and contract depending on luminal volume. This is consistent with its primary role in seminal fluid resorption and sperm concentration. Moreover, we describe the cellularity of models to study the human epididymis epithelium in vitro. Conclusion Single cell RNA-seq data from the human epididymis make a valuable contribution to our understanding of this highly specialized organ. Overall design: Single cells from human primary epididmis epithelial cells (HEE), human efferent duct tissue, and human caput tissue were isolated and analyzed using scRNA-seq.