Immobilization of Nanocarriers Within a Porous Chitosan Scaffold for the Sustained Delivery of Growth Factors in Bone Tissue Engineering Applications

To guide the natural bone regeneration process, bone tissue engineering strategies rely on the development of a scaffold architecture that mimics the extracellular matrix and incorporates important extracellular signaling molecules, which promote fracture healing and bone formation pathways. Incorporation of growth factors into particles embedded within the scaffold can offer both protection of protein bioactivity and a sustained release profile. In this work, a novel method to immobilize carrier nanoparticles within scaffold pores is proposed. A biodegradable, osteoconductive, porous chitosan scaffold was fabricated via the "freeze-drying method," leading to scaffolds with a storage modulus of 8.5 kPa and 300 μm pores, in line with existing bone scaffold properties. Next, poly(methyl methacrylate-co-methacrylic acid) nanoparticles were synthesized and immobilized to the scaffold via carbodiimide-crosslinker chemistry. A fluorescent imaging study confirmed that the conventional methods of protein and nanocarrier incorporation into scaffolds can lead to over 60% diffusion out of the scaffold within the first 5 min of implantation, and total disappearance within 4 weeks. The novel method of nanocarrier immobilization to the scaffold backbone via carbodiimide-crosslinker chemistry allows full retention of particles for up to 4 weeks within the scaffold bulk, with no negative effects on the viability and proliferation of human umbilical vein endothelial cells.