One-Pot Crosslinking Enables 3D Printing of Shear-Recovery Fish Collagen VEGF-Functionalized Inks

Fish collagen is gaining attention as a sustainable biomaterial for three-dimensional (3D) scaffold fabrication in tissue engineering. In this study, a biomimetic, one-pot crosslinking strategy for native fish skin collagen was developed and compared with a conventional periodate oxidation-Schiff base approach using oxidized maltose. Both approaches increased viscosity and thermal stability while preserving native structural features. The oxidized maltose-crosslinked collagen demonstrated Schiff-base crosslinking, and the one-pot crosslinking method produced a covalent bond that was not a Schiff base. Precrosslinked collagens were processed into microgels and incorporated into calcium alginate to yield a shear-recovering, extrudable ink suitable for 3D extrusion printing. The printed scaffolds maintained structural resilience under physiological conditions, exhibited shear-recovery behavior confirmed by rheological analysis, and supported high cell viability. To enhance biofunctionality, vascular endothelial growth factor was conjugated to the 3D scaffolds, which were subsequently seeded with human bone marrow-derived mesenchymal stem cells. Immunofluorescence staining indicated endothelial lineage differentiation, suggesting that this platform may support the development of vascularized 3D tissue constructs.