Optical and Thermal Control of Pore Architecture in Collagen Hydrogels for Vascular-like Tissue Engineering Scaffolds

Vascularization remains a central challenge in thick tissue engineering. Building on our prior demonstration that carbonate buffer concentration governs multi-channel collagen gel (MCCG) architecture and perfusion culture performance, this study aimed to establish non-contact, orthogonal control of pore size and density in riboflavin-sensitized Type I collagen hydrogels via UV irradiation intensity and preparation temperature. UV intensity was modulated by varying the source-to-sample distance (25–52 mm); preparation temperature was set at 5, 25, or 40 °C; gelation kinetics were quantified using a vial-tilt assay. Pore area fraction ranged from 0.9% to 8.6% and Young’s modulus from 16 to 49 kPa depending on UV dose. Higher preparation temperatures accelerated gelation and produced smaller, more densely distributed pores, consistent with kinetically arrested phase separation. NIH/3T3 fibroblasts cultured on intermediate- and low-intensity UV scaffolds achieved >80% confluency by Day 7, with three-dimensional tissue-like organization and directionally aligned cellular bundles within large pores; cell metabolic activity, assessed by CCK-8 assay, remained consistently high throughout the culture period. These results demonstrate that UV irradiation intensity and preparation temperature are independently tunable, non-contact parameters for reproducible fabrication of collagen scaffolds with tunable vascular-like pore networks, complementing and extending the chemical (buffer concentration) design space of MCCG-based perfusion culture systems.