Visible Light Cross-Linked Methacrylated Silk Fibroin Enables Enhanced Osteogenic Response in Bioprinted Dual-Layer
Soham Ghosh1, Fathima Shajahan1, Jaideep Adhikari1
1BioFabTE Lab, Department of Biomedical Engineering, Indian Institute of Technology Hyderabad, Kandi, Sangareddy 502285, Telangana, India.
Abstract:
Precise design and fabrication of photo-cross-linked hydrogels with controlled network architecture and tailored mechanical properties are essential for advancing complex tissue engineering applications. In this study, a visible-light-activated type II photo-cross-linking system was developed using eosin Y/triethanolamine/N-vinylpyrrolidone (VE) to fabricate methacrylated silk fibroin (SFM) hydrogels through oxygen-mediated controlled photolysis. In comparison to conventional UV-initiated lithium phenyl-2,4,6-trimethylbenzoylphosphinate cross-linking, which rapidly yields mechanically stiffer networks with pronounced β-structures, the VE system (0.02 mM eosin Y, 100 mM triethanolamine, and 50 mM N-vinylpyrrolidone) enabled the fabrication of gradually formed compliant homogeneous networks. This tunability allowed using VE cross-linked SFM in light-assisted extrusion bioprinting to fabricate a dual-layer guided bone regeneration membrane incorporating a collagen I-rich ECM layer. Under calcium-supplemented conditions, the dual-layer membrane exhibited robust osteogenic potential, evidenced by significantly elevated ALP activity and distinctive nodular mineralization patterns compared with single-layer controls. Gene expression profiles revealed coordinated regulation of early (RUNX2, COL1A1), mid-to-late (SPP1, SPARC), and late-stage (BGLAP) markers, indicating successful progression through the osteogenic program. The heterogeneous design achieved a desired balance between its barrier function and tissue integration with an interconnected porous architecture that limits soft tissue downgrowth while supporting matrix organization conducive to bone regeneration. These findings establish critical structure-function relationships in photo-cross-linked biomaterials and highlight how mechanistic understanding of cross-linking chemistry can guide the rational design of functional scaffolds for biomedical applications.
