Chitosan-based composite bilayer scaffold as an in vitro osteochondral defect regeneration model.

Prolonged osteochondral tissue damage can result in osteoarthritis and decreased quality of life. Multiphasic scaffolds, where different layers model different microenvironments, are a promising treatment approach, yet stable joining between layers during fabrication remains challenging. Here, a bilayer scaffold for osteochondral tissue regeneration was fabricated using thermally-induced phase separation (TIPS). Two distinct polymer solutions were layered before TIPS, and the resulting porous, bilayer scaffold was characterized by seamless interfacial integration and a mechanical stiffness gradient reflecting the native osteochondral microenvironment. Chitosan is a critical component of both scaffold layers to facilitate cell attachment and the formation of polyelectrolyte complexes with other biologically relevant natural polymers. The articular cartilage region was optimized for hyaluronic acid content and stiffness, while the subchondral bone region was defined by higher stiffness and osteoconductive hydroxyapatite content. Following co-culture with chondrocyte-like (SW-1353 or mesenchymal stem cells) and osteoblast-like cells (MG63), cell proliferation and migration to the interface along with increased gene expression associated with relevant markers of osteogenesis and chondrogenesis indicates the potential of this bilayer scaffold for osteochondral tissue regeneration.