Xerostomia or dry mouth are commonly diagnosed in head and neck cancer patients due to salivary gland (SG) epithelial injury after radiotherapy. Regenerative medicine has fetched the opportunity to replace or regenerate the SG epithelia and restore its secretory function. Early adult stem cell transplantation strategies in rodents have recently shown to improve clinical outcomes in radiotherapy-induced xerostomia in Phase 1/2 human trials. Mesenchymal stem cells from adipose tissue are the most promising, although the ones from the labial mucosa, bone marrow, or dental pulp have an attractive therapeutic value after successful findings in ex vivo and in vivo mouse models of SG injury. Emerging approaches using cell-free therapy with cell ''extracts'', ''soups'' or secretome components also exhibit favorable outcomes in the same rodent models. When compared to cell-based approaches, extracellular vesicles (EV) from the secretome (i.e., exosomes) can be easily extracted, quantified, and are more stable for long-term storage and use in SG tissue engineering. Additive manufacturing and threedimensional bioprinting or bioassembly have an important role on generating spheroids or organoids for cell transplantation to ameliorate SG injury. Moreover, organoids can secrete EV, which may have a therapeutic potential worth to explore in future studies. In this review, we will describe the technological advancements and challenges of these different cell-based and cell-free strategies in SG tissue engineering and regeneration.

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Antioxidant agents are promising pharmaceuticals to prevent salivary gland (SG) epithelial injury from radiotherapy and their associated irreversible dry mouth symptoms. Epigallocatechin-3-gallate (EGCG) is a well-known antioxidant that can exert growth or inhibitory biological effects in normal or pathological tissues leading to disease prevention. The effects of EGCG in the various SG epithelial compartments are poorly understood during homeostasis and upon radiation (IR) injury. This study aims to: (1) determine whether EGCG can support epithelial proliferation during homeostasis; and (2) investigate what epithelial cells are protected by EGCG from IR injury. Ex vivo mouse SG were treated with EGCG from 7.5–30 µg/mL for up to 72 h. Next, SG epithelial branching morphogenesis was evaluated by bright-field microscopy, immunofluorescence, and gene expression arrays. To establish IR injury models, linear accelerator (LINAC) technologies were utilized, and radiation doses optimized. EGCG epithelial effects in these injury models were assessed using light, confocal and electron microscopy, the Griess assay, immunohistochemistry, and gene arrays. SG pretreated with EGCG 7.5 µg/mL promoted epithelial proliferation and the development of pro-acinar buds and ducts in regular homeostasis. Furthermore, EGCG increased the populations of epithelial progenitors in buds and ducts and pro-acinar cells, most probably due to its observed antioxidant activity after IR injury, which prevented epithelial apoptosis. Future studies will assess the potential for nanocarriers to increase the oral bioavailability of EGCG.

Osteoblast differentiation requires the interaction of various cell signaling pathways to modulate cell responses. Notch and Wnt signaling are among the crucial pathways that control numerous biological processes, including osteo/odontogenic differentiation. The aim of the present study was to examine the involvement of Wnt signaling in the Jagged1-induced osteo/odontogenic differentiation in human dental pulp stem cells (hDPSCs). The Wnt-related gene expression was analyzed from publicly available data of Jagged1-treated human dental pulp cells. The mRNA expression of Wnt ligands (WNT2B, WNT5A, WNT5B, and WNT16) and Wnt inhibitors (DKK1, DKK2, and SOST) were confirmed using real-time polymerase chain reaction. Among the Wnt ligands, WNT2B and WNT5A mRNA levels were upregulated after Jagged1 treatment. In contrast, the Wnt inhibitors DKK1, DKK2, and SOST mRNA levels were downregulated. Recombinant WNT5A, but not WNT2B, significantly promoted in vitro mineral deposition by hDPSCs. Wnt signaling inhibition using IWP-2, but not DKK1, inhibited Jagged1-induced alkaline phosphatase (ALP) activity, mineralization, and osteo/odontogenic marker gene expression in hDPSCs. In conclusion, Jagged1 promoted hDPSC osteo/odontogenic differentiation by modulating the non-canonical Wnt pathway.

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