An all-silk-derived functional nanosphere matrix for sequential biomolecule delivery and in situ osteochondral regeneration.

Endogenous repair of osteochondral defect is usually limited by the insufficient number of cells in the early stage and incomplete cell differentiation in the later stage. The development of drug delivery systems for sequential release of pro-migratory and pro-chondrogenic molecules to induce endogenous bone marrow-derived mesenchymal stem cells (BMSCs) recruitment and chondrogenic differentiation is highly desirable for in situ osteochondral regeneration. In this study, a novel, all-silk-derived sequential delivery system was fabricated by incorporating the tunable drug-loaded silk fibroin (SF) nanospheres into a SF porous matrix. The loading efficiency and release kinetics of biomolecules depended on the initial SF/polyvinyl alcohol (PVA) concentrations (0.2%, 1% and 5%) of the nanospheres, as well as the hydrophobicity of the loaded molecules, resulting in controllable and programmed delivery profiles. Our findings indicated that the 5% nanosphere-incorporated matrix showed a rapid release of E7 peptide during the first 120 h, whereas the 0.2% nanosphere-incorporated matrix provided a slow and sustained release of Kartogenin (KGN) longer than 30 days. During in vitro culture of BMSCs, this functional SF matrix incorporated with E7/KGN nanospheres showed good biocompatibility, as well as enhanced BMSCs migration and chondrogenic differentiation through the release of E7 and KGN. Furthermore, when implanted into rabbit osteochondral defect, the SF nanosphere matrix with sequential E7/KGN release promoted the regeneration of both cartilage and subchondral bone. This work not only provided a novel all-silk-derived drug delivery system for sequential release of molecules, but also a functional tissue-engineered scaffold for osteochondral regeneration.