No permanent cure exists for salivary gland (SG) damage and consequent xerostomia (dry mouth) in patients undergoing radiotherapy for head and neck cancers. The lack of commercially available healthy human SG-derived cell lines has hindered in vitro studies of radiation-induced glandular injury. In this study, we successfully immortalized and characterized two novel human major SG-derived cell lines. Leveraging these cell lines and hyaluronic-acid hydrogels, we bioengineered distinct multicellular SG spheroids and microtissues expressing key acinar, ductal, myoepithelial, and mesenchymal cell markers in long-term cultures. Further, using this platform, we developed a proof-of-concept radiation injury model, demonstrating spheroid disruption characterized by actin depolymerization, DNA damage, apoptosis, and loss of SG-specific markers following radiation exposure. Notably, these detrimental effects were partially mitigated with a radioprotective agent. Our findings demonstrate that the bioengineered SG spheroids provide a scalable and versatile platform with significant potential for disease modeling and drug testing, thereby accelerating the development of targeted therapies for radiation-induced xerostomia.