Cell Lineage Tracing Identifies Hormone-Regulated and Wnt-Responsive Vaginal Epithelial Stem Cells

The intact vaginal epithelium is essential for women's reproductive health and provides protection against HIV and sexually transmitted infections. How this epithelium maintains itself remains poorly understood. Here, we used single-cell RNA sequencing (RNA-seq) to define the diverse cell populations in the vaginal epithelium. We show that vaginal epithelial cell proliferation is limited to the basal compartment without any obvious label-retaining cells. Furthermore, we developed vaginal organoids and show that the basal cells have increased organoid forming efficiency. Importantly, Axin2 marks a self-renewing subpopulation of basal cells that gives rise to differentiated cells over time. These cells are ovariectomy-resistant stem cells as they proliferate even in the absence of hormones. Upon hormone supplementation, these cells expand and reconstitute the entire vaginal epithelium. Wnt/beta-catenin is essential for the proliferation and differentiation of vaginal stem cells. Together, these data define heterogeneity in vaginal epithelium and identify vaginal epithelial stem cells.