Infrequently Dividing Oral Epithelial Cells Reside in Posterior Palatal Niches

Objectives: The oral cavity is lined by highly proliferative epithelia; how these epithelia utilize stem/progenitor populations to heal with minimal scarring is unknown. Oral epithelial stem/progenitor cells (OESCs) are theoretically located within the proliferative basal layer, which is assumed to be homogenous. However, it is believed in the gastrointestinal tract there are two stem cell pools: 1) those active during maintenance and 2) a reserve population, which infrequently divides and is not involved in epithelial homeostasis. The goal of this study was to assess whether heterogeneous OESC populations exist, specifically looking for evidence of reserve OESCs.
Methods: We adopted genetic label retention approaches to identify label-retaining cells (LRCs) using two distinct promoters (Krt14 and Krt5) to drive expression of a doxycycline-regulable histone-GFP, which is diluted as cells divide during a variable chase period. Additionally, we complemented these assays with 1) in vivo immunofluorescence-based proliferation assays, 2) FACS/qPCR, 3) lineage tracing (K5-; K14-; Sox9-CreER), and 4) diet modification assays.
Results: Both LRC strategies demonstrated rapid GFP dilution in 14-days chases; however, slight differences between the two approaches were observed. We identified LRCs (GFP^hi) in posterior palate (rugae/interrugae transitions) at later chases. We observed GFP^hi/GFP^lo cells show different division orientation—the former characterized by more planar/symmetric divisions. We isolated GFP^hi by FACS and compared gene expression to GFP^lo. GFP^hi are enriched for Sox9 and TAp63, while the ΔNp63 isoform is enriched in GFP^lo cells. This was confirmed by immunofluorescence microscopy in vivo. Lineage tracing using K5-, K14-, and Sox9-CreER revealed clonal populations with distinct sizes and morphologies reminiscent of stem and transit-amplifying cells. With normal “hard” diets, GFP^hi cells were rapidly depleted in highly innervated rugae peaks; however, soft diets restored GFP^hi cells in innervated regions.
Conclusions: Collectively, these data suggest that posterior palate may harbor IDCs reminiscent of reserve stem cells.