Facile Fabrication of Cartilage-Bone Interface-Mimicking Ductile Scaffolds for Zone-Specific Stem Cell Differentiation and Angiogenesis

The regeneration of osteochondral tissue remains a significant clinical challenge due to the structural and biochemical complexity of the cartilage-bone interface. Current osteochondral polymeric scaffold implants face limitations, including biologically inert surface chemistries, insufficient mechanical properties, and simplistic architectures that fail to mimic native tissue heterogeneity. To address these challenges, we developed a biomimetic osteochondral scaffold using an equipment-free, stepwise thermally induced phase separation (TIPS) with porogen leaching, yielding gradient-interconnected porosity and zonal bioactivity without the need for sophisticated fabrication methods. The scaffold exhibited a hierarchical pore distribution, transitioning from nanofibrous, low-porosity superficial regions (<50 μm pores) to highly porous deep/subchondral (Sc) zones (700-1000 μm pores), closely mimicking the natural osteochondral interface. Bioactive glass nanoparticles (BGNs) were synthesized and incorporated at varying concentrations across the scaffold, with the highest content in the deep/Sc zone, imparting osteoconductive and osteoinductive properties. Despite high BGN content (70-90 wt %), the scaffold retained mechanical ductility through the addition of polyethylene glycol (PEG), facilitating intraoperative handling and defect conformation. <i>In vitro</i> biochemical and immunofluorescence assays showed zone-specific mesenchymal stem cell (MSC) differentiation: sulfated glycosaminoglycan (sGAG), type II collagen, and lubricin predominated in superficial/middle zones, whereas alkaline phosphatase (ALP), osteocalcin, and type X collagen were elevated in the deep/Sc zone, confirming osteoinduction. <i>In vivo</i>, the chick chorioallantoic membrane (CAM) model revealed angiogenesis predominantly in the deep/Sc zone, highlighting the scaffold's ability to promote vascularization, a critical factor for bone regeneration. These findings demonstrate a facile, scalable, and cost-effective route to clinically viable osteochondral scaffolds. With biomimetic architecture, zonal bioactivity, and ease of fabrication, this scaffold presents a promising candidate for osteochondral regeneration with future investigations focusing on refining its clinical application and long-term <i>in vivo</i> performance.