SUMMARYA comprehensive cellular anatomy of normal human prostate is essential for solving the cellular origins of benign prostatic hyperplasia and prostate cancer. The tools used to analyze the contribution of individual cell types are not robust. We provide a cellular atlas of the young adult human prostate and prostatic urethra using an iterative process of single-cell RNA sequencing (scRNA-seq) and flow cytometry on ~98,000 cells taken from different anatomical regions. Immunohistochemistry with newly derived cell type-specific markers revealed the distribution of each epithelial and stromal cell type on whole mounts, revising our understanding of zonal anatomy. Based on discovered cell surface markers, flow cytometry antibody panels were designed to improve the purification of each cell type, with each gate confirmed by scRNA-seq. The molecular classification, anatomical distribution, and purification tools for each cell type in the human prostate create a powerful resource for experimental design in human prostate disease.

A cellular anatomy of normal human organs is essential for solving the cellular origins of disease. We report the first comprehensive cellular atlas of the young adult human prostate and prostatic urethra using an iterative process of single cell RNA sequencing and flow cytometry on ~98,000 cells taken from different anatomical regions. Two previously unrecognized epithelial cell types were identified by KRT13 and SCGB1A1 expression and found to be highly similar to hillock and club cells of the proximal lung. It was demonstrated by immunohistochemistry that prostate club and hillock cells are similarly concentrated in the proximal prostate. We also optimized a new flow cytometry antibody panel to improve cell type-specific purification based on newly established cellular markers. The molecular classification, anatomical distribution, and purification methods for each cell type in the human prostate create a powerful new resource for experimental design in human prostate disease.

Background: Castration-insensitive epithelial progenitors capable of regenerating the prostate have been proposed to be concentrated in the proximal region based on facultative assays. Functional characterization of prostate epithelial populations isolated with individual cell surface markers has failed to provide a consensus on the anatomical and transcriptional identity of proximal prostate progenitors.Methods: Here, we use single-cell RNA sequencing to obtain a complete transcriptomic profile of all epithelial cells in the mouse prostate and urethra to objectively identify cellular subtypes. Pan-transcriptomic comparison to human prostate cell types identified a mouse equivalent of human urethral luminal cells, which highly expressed putative prostate progenitor markers. Validation of the urethral luminal cell cluster was performed using immunostaining and flow cytometry.Results: Our data reveal that previously identified facultative progenitors marked by Trop2, Sca-1, KRT4, and PSCA are actually luminal epithelial cells of the urethra that extend into the proximal region of the prostate, and are resistant to castration-induced androgen deprivation. Mouse urethral luminal cells were identified to be the equivalent of previously identified human club and hillock cells that similarly extend into proximal prostate ducts. Benign prostatic hyperplasia (BPH) has long been considered an "embryonic reawakening," but the cellular origin

Summary Prostate inflammation has been suggested as an etiology for benign prostatic hyperplasia (BPH). We show that decreased expression of the androgen receptor (AR) in luminal cells of human BPH specimens correlates with a higher degree of regional prostatic inflammation. However, the cause-and-effect relationship between the two events remains unclear. We investigated specifically whether attenuating AR activity in prostate luminal cells induces inflammation. Disrupting luminal cell AR signaling in mouse models promotes cytokine production cell-autonomously, impairs epithelial barrier function, and induces immune cell infiltration, which further augments local production of cytokines and chemokines including Il-1 and Ccl2. This inflammatory microenvironment promotes AR-independent prostatic epithelial proliferation, which can be abolished by ablating IL-1 signaling or depleting its major cellular source, the macrophages. This study demonstrates that disrupting luminal AR signaling promotes prostate inflammation, which may serve as a mechanism for resistance to androgen-targeted therapy for BPH.

Stromal–epithelial interactions are critical to the morphogenesis, differentiation, and homeostasis of the prostate, but the molecular identity and anatomy of discrete stromal cell types is poorly understood. Using single‐cell RNA sequencing, we identified and validated the in situ localization of three smooth muscle subtypes (prostate smooth muscle, pericytes, and vascular smooth muscle) and two novel fibroblast subtypes in human prostate. Peri‐epithelial fibroblasts (APOD+) wrap around epithelial structures, whereas interstitial fibroblasts (C7+) are interspersed in extracellular matrix. In contrast, the mouse displayed three fibroblast subtypes with distinct proximal–distal and lobe‐specific distribution patterns. Statistical analysis of mouse and human fibroblasts showed transcriptional correlation between mouse prostate (C3+) and urethral (Lgr5+) fibroblasts and the human interstitial fibroblast subtype. Both urethral fibroblasts (Lgr5+) and ductal fibroblasts (Wnt2+) in the mouse contribute to a proximal Wnt/Tgfb signaling niche that is absent in human prostate. Instead, human peri‐epithelial fibroblasts express secreted WNT inhibitors SFRPs and DKK1, which could serve as a buffer against stromal WNT ligands by creating a localized signaling niche around individual prostate glands. We also identified proximal–distal fibroblast density differences in human prostate that could amplify stromal signaling around proximal prostate ducts. In human benign prostatic hyperplasia, fibroblast subtypes upregulate critical immunoregulatory pathways and show distinct distributions in stromal and glandular phenotypes. A detailed taxonomy of leukocytes in benign prostatic hyperplasia reveals an influx of myeloid dendritic cells, T cells and B cells, resembling a mucosal inflammatory disorder. A receptor–ligand interaction analysis of all cell types revealed a central role for fibroblasts in growth factor, morphogen, and chemokine signaling to endothelia, epithelia, and leukocytes. These data are foundational to the development of new therapeutic targets in benign prostatic hyperplasia. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.