Parasympathetic Innervation Maintains Epithelial Progenitor Cells During Salivary Organogenesis

Abstract

The maintenance of a progenitor cell population as a reservoir of undifferentiated cells is required for organ development and regeneration. However, the mechanisms by which epithelial progenitor cells are maintained during organogenesis are poorly understood. We report that removal of the parasympathetic ganglion in mouse explant organ culture decreased the number and morphogenesis of keratin 5-positive epithelial progenitor cells. These effects were rescued with an acetylcholine analog. We demonstrate that acetylcholine signaling, via the muscarinic M1 receptor and EGFR, increased epithelial morphogenesis and proliferation of the keratin 5-positive progenitor cells. Parasympathetic innervation maintained the epithelial progenitor cell population in an undifferentiated state, which was required for organogenesis. This mechanism for epithelial progenitor cell maintenance may be targeted for organ repair or regeneration.

Roles of innervation in developing and regenerating orofacial tissues

“…This concomitant axonal outgrowth from the submandibular ganglion regulates epithelial salivary gland branching, and thus gland morphogenesis. Indeed, isolated E11 mouse submandibular epithelia cultured in vitro in absence of parasympathetic ganglia develop glands with a reduced number of branches [36]. The development of the appropriate number of terminal epithelial buds is dependent on acetylcholine and its muscarinic receptor M1 that is located on the surface of epithelial cells.…”

“…The development of the appropriate number of terminal epithelial buds is dependent on acetylcholine and its muscarinic receptor M1 that is located on the surface of epithelial cells. This has been evidenced after addition of inhibitors of acetylcholine/M1 signalling in the medium of an in vitro culture system of salivary glands [36]. The reduced number of progenitor epithelial gland cells, assessed by quantification of cells expressing progenitor/stem cell markers, is mainly responsible for the diminished number of branches [36].…”

“…This has been evidenced after addition of inhibitors of acetylcholine/M1 signalling in the medium of an in vitro culture system of salivary glands [36]. The reduced number of progenitor epithelial gland cells, assessed by quantification of cells expressing progenitor/stem cell markers, is mainly responsible for the diminished number of branches [36]. In the absence of parasympathetic ganglia, the expression of salivary gland progenitor/stem cell markers such as keratin5, keratin15 and aquaporin3 decreases significantly.…”

“…Thus, the parasympathetic innervation affects salivary epithelial morphogenesis by regulating the pool of gland progenitor/stem cells. A potential therapeutic role of parasympathetic innervation in salivary gland regeneration could be suggested, since increased expression of the stemness markers keratin5 and keratin15 was observed in denervated adult salivary glands treated with carbachol, which is an equivalent of acetylcholine [36]. Salivary glands produce high quantities of NGF and express all neurotrophin receptors [37][38][39].…”

Expression of a Neural Stem/Progenitor Cell Marker Nestin in Salivary Glands

“…6 Because cytokeratin 5 has been reported as a marker of progenitor cells in salivary glands, 7 increase of cytokeratin 5 in response to tissue injury can be interpreted as an activation and proliferation of stem/progenitor cells to replace damaged cells. However, cytokeratin 5 is also expressed in basal cells of ducts and myoepithelial cells, and identification of more specific markers for progenitor cells is desirable.…”