Li Guan1, Vignesh Viswanathan1, Yuyan Jiang1, Sivakamasundari Vijayakumar2, Hongbin Cao1, Junfei Zhao3, Deana Rae Crystal Colburg4, Patrick Neuhöfer5, Yiru Zhang6, Jinglong Wang1, Yu Xu1, Eyiwunmi E. Laseinde1, Rachel Hildebrand1, Mobeen Rahman4, Richard Frock1, Christina Kong4, Philip A. Beachy2, Steven Artandi5 and Quynh-Thu Le1 1Department of Radiation Oncology, Stanford University School of Medicine, Stanford, California 94305, USA; 2Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, California 94305, USA; 3Department of Pathology and Cell Biology, Columbia University, New York, New York 10032, USA; 4Department of Pathology, Stanford University School of Medicine, Stanford, California 94305, USA; 5Stanford Cancer Institute, Stanford University School of Medicine, Stanford, California 94305, USA; 6Department of Radiology, Sloan Kettering Institute, Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA Corresponding author: qlestanford.edu Abstract

Salivary gland homeostasis and regeneration after radiotherapy depend significantly on progenitor cells. However, the lineage of submandibular gland (SMG) progenitor cells remains less defined compared with other normal organs. Here, using a mouse strain expressing regulated CreERT2 recombinase from the endogenous Tert locus, we identify a distinct telomerase-expressing (TertHigh) cell population located in the ductal region of the adult SMG. These TertHigh cells contribute to ductal cell generation during SMG homeostasis and to both ductal and acinar cell renewal 1 year after radiotherapy. TertHigh cells maintain self-renewal capacity during in vitro culture, exhibit resistance to radiation damage, and demonstrate enhanced proliferative activity after radiation exposure. Similarly, primary human SMG cells with high Tert expression display enhanced cell survival after radiotherapy, and CRISPR-activated Tert in human SMG spheres increases proliferation after radiation. RNA sequencing reveals upregulation of “cell cycling” and “oxidative stress response” pathways in TertHigh cells following radiation. Mechanistically, Tert appears to modulate cell survival through ROS levels in SMG spheres following radiation damage. Our findings highlight the significance of TertHigh cells in salivary gland biology, providing insights into their response to radiotherapy and into their use as a potential target for enhancing salivary gland regeneration after radiotherapy.

Received February 6, 2024. Accepted June 25, 2024.