Abstract
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/β-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.
Highlights
Vaginal epithelium (VE) is a keratinized stratified squamous epithelium consisting of three layers: basal layer, suprabasal layer, and apical cornified layer (Bragulla and Homberger, 2009)
In this study, using genetic cell lineage tracing, vaginal organoid culture, targeted gene deletion, and single-cell sequencing approaches, we revealed a unique population of VE stem cells and provided evidence that canonical Wnt/b-catenin signaling is required for the proliferation and differentiation of these stem cells
A total of 4,940 cells, of the combined duplicates, that passed QC filtering were clustered into six cell clusters using the k-means method and visualized using t-Distributed Stochastic Neighbor Embedding projection
Summary
Vaginal epithelium (VE) is a keratinized stratified squamous epithelium consisting of three layers: basal layer, suprabasal layer, and apical cornified layer (Bragulla and Homberger, 2009). Experimental or age-related decline in E2 levels in mice and humans leads to regression of VE, which is reversed upon the external supplementation of E2 (Edgren, 1959). This exceptional regenerative capacity of VE suggests the existence of a population of epithelial stem/progenitor cells. Wnt signaling is an important regulator of various stages of female reproductive tract development and is a known master controller of adult stem cell maintenance in many organ systems (Kobayashi and Behringer, 2003; Nusse, 2008). Very little is known about the role of Wnt signaling in VE homeostasis
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