Alginate-based hydrogels are widely used for the development of biomedical scaffolds in regenerative medicine. The use of sugar glass as a sacrificial template for fluidic channels fabrication within alginate scaffolds remains a challenge because of the premature dissolution of sugar by the water contained in the alginate as well as the relatively slow internal gelation rate of the alginate. Here, a new and simple method, based on a sugar glass fugitive ink loaded with calcium chloride to build sacrificial molds, is presented. We used a dual calcium cross-linking process by adding this highly soluble calcium source in the printed sugar, thus allowing the rapid gelation of a thin membrane of alginate around the sugar construct, followed by the addition of calcium carbonate and gluconic acid δ-lactone to complete the process. This innovative technique results in the rapid formation of "on-demand" alginate hydrogel with complex fluidic channels that could be used in biomedical applications such as highly vascularized scaffolds promoting pathways for nutrients and oxygen to the cells.

Transplantation of hydrogel-encapsulated pancreatic islets is a promising long-term treatment for type 1 diabetes that restores blood glucose regulation while providing graft immunoprotection. Most human-scale islet encapsulation devices that rely solely on diffusion fail to provide sufficient surface area to meet islet oxygen demands. Perfused macroencapsulation devices use blood flow to mitigate oxygen limitations but increase the complexity of blood-device interactions. Here we describe a human-scale in vitro perfusion system to study hemocompatibility and performance of islet-like cell clusters (ILCs) in alginate hydrogel. A cylindrical perfusion device was designed for multi-day culture without leakage, contamination, or flow occlusion. Rat blood perfusion was assessed for prothrombin time and international normalized ratio and demonstrated no significant change in clotting time. Ex vivo perfusion performed with rats showed patency of the device for over 100 min using Doppler ultrasound imaging. PET-CT imaging of the device successfully visualized metabolically active mouse insulinoma 6 ILCs. ILCs cultured for 7 days under static conditions exhibited abnormal morphology and increased activated caspase-3 staining when compared with the perfused device. These findings reinforce the need for convective transport in macroencapsulation strategies and offer a robust and versatile in vitro system to better inform preclinical design.

A prominent obstacle in scaling up tissue engineering technologies for human applications is engineering an adequate supply of oxygen and nutrients throughout artificial tissues. Sugar glass has emerged as a promising 3D-printable, sacrificial material that can be used to embed perfusable networks within cell-laden matrices to improve mass transfer. To characterize and optimize a previously published sugar ink, we investigated the effects of sucrose, glucose, and dextran concentration on the glass transition temperature ( T g ), printability, and stability of 3D-printed sugar glass constructs. We identified a sucrose ink formulation with a significantly higher T g (40.0 ± 0.9°C) than the original formulation (sucrose-glucose blend, T g = 26.2 ± 0.4°C), which demonstrated a pronounced improvement in printability, resistance to bending, and final print stability, all without changing dissolution kinetics and decomposition temperature. This formulation allowed printing of 10-cm-long horizontal cantilever filaments, which can enable the printing of complex vascular segments along the x-, y-, and z-axes without the need for supporting structures. Vascular templates with a single inlet and outlet branching into nine channels were 3D printed using the improved formulation and subsequently used to generate perfusable alginate constructs. The printed lattice showed high fidelity with respect to the input geometry, although with some channel deformation after alginate casting and gelation—likely due to alginate swelling. Compared with avascular controls, no significant acute cytotoxicity was noted when casting pancreatic beta cell-laden alginate constructs around improved ink filaments, whereas a significant decrease in cell viability was observed with the original ink. The improved formulation lends more flexibility to sugar glass 3D printing by facilitating the fabrication of larger, more complex, and more stable sacrificial networks. Rigorous characterization and optimization methods for improving sacrificial inks may facilitate the fabrication of functional cellular constructs for tissue engineering, cellular biology, and other biomedical applications.