Decellularized tissue-specific hydrogels support an engineered salivary gland within a microfluidic platform

• dSMG hydrogel retains key ECM components, exhibits a nanofibrous structure, and possesses a thermo-responsive gelation. • dSMG hydrogel ensures consistent biological effects with minimal batch-to-batch variations. • dSMG hydrogel supports human SG MEC cell viability, proliferation, ductal phenotype, cholinergic-specific signaling and functional activity within microfluidic systems. Mucoepidermoid carcinoma (MEC) is a rare malignancy of the salivary gland (SG) that poses significant treatment challenges. This highlights the need for in vitro cancer modeling platforms towards anti-cancer drug screening applications. Emerging organ-on-a-chip (OoC) microfluidic technologies represent promising new approach methodologies (NAMS) and a real alternative to animal testing. While tissue-specific decellularized extracellular matrix (ECM) can recapitulate in vivo-like microenvironments, its application in SG-on-a-chip (SGoC) is still underexplored. This study developed an injectable porcine decellularized submandibular gland (dSMG) hydrogel for bioengineering an SG MEC tissue chip. dSMG was prepared using a chemical and enzymatic decellularization process with 0.1% or 1% sodium dodecyl sulfate (SDS). Both treatments effectively removed DNA content while preserving key ECM components, including collagens, glycoproteins, and mucins. Proteomic analysis revealed that 1% SDS-treated dSMG contained a greater abundance of ECM components involved in matrix assembly and cell-ECM interactions compared to the 0.1% group. The 1% SDS-treated dSMG was subsequently digested with a pepsin-based buffer to form hydrogels. At 5 mg/mL, dSMG hydrogel exhibited nanofibrous architecture, thermo-responsive gelation, injectability into microfluidic devices, and minimal batch-to-batch biological variations. In static conditions, dSMG hydrogel significantly enhanced SG cell viability and mitochondria-dependent proliferation compared to Matrigel. Under gravity-driven flow, dSMG hydrogel promoted a ductal phenotype on human SG MEC cells, unlike on Matrigel. Additionally, dSMG hydrogel supported cholinergic-specific signaling and functional activity. These findings demonstrate the potential of dSMG hydrogel as a physiologically relevant matrix for SG cancer modeling towards drug screening applications in SGoC microfluidic systems.