Long-Term In Vitro Expansion of Salivary Gland Stem Cells Driven by Wnt Signals

• EpCAMhigh cells generate salivary gland organoids in the presence of Wnt proteins
• Wnt signaling is required for the maintenance of salivary gland stem cells in vitro
• Wnt agonists promote long-term salivary gland stem cell expansion in culture
• Expanded salivary gland stem cells regenerate damaged salivary glands in vivo

SummaryAdult stem cells are the ultimate source for replenishment of salivary gland (SG) tissue. Self-renewal ability of stem cells is dependent on extrinsic niche signals that have not been unraveled for the SG. The ductal compartment in SG has been identified as the location harboring stem cells. Here, we report that rare SG ductal EpCAM+ cells express nuclear β-catenin, indicating active Wnt signaling. In cell culture experiments, EpCAMhigh cells respond potently to Wnt signals stimulating self-renewal and long-term expansion of SG organoids, containing all differentiated SG cell types. Conversely, Wnt inhibition ablated long-term organoid cultures. Finally, transplantation of cells pre-treated with Wnt agonists into submandibular glands of irradiated mice successfully and robustly restored saliva secretion and increased the number of functional acini in vivo. Collectively, these results identify Wnt signaling as a key driver of adult SG stem cells, allowing extensive in vitro expansion and enabling restoration of SG function upon transplantation.

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