Porosity Tunable Silk Fibroin Microspheres Enhance Cartilage Repair in Rabbit Model

Repairing articular cartilage defects is particularly challenging due to the tissue's hypovascularity and limited intrinsic healing capacity, which pose significant clinical challenges. Biomaterial-based scaffolds have shown promising outcomes in terms of repair capacity; however, finding a robust scaffold system remains challenging in cartilage tissue engineering. In this study, silk fibroin porous microspheres were fabricated using the co-axial electrospraying technique, followed by ethanol treatment to induce crosslinking and enhance microsphere stability. Interestingly, we observed that while changing the ethanol concentration from 100% to 75% to crosslink the silk microsphere resulted in the formation of tunable micropores, with pore sizes up to 87 μm in nonporous microspheres and ranging from 80-200 μm in porous microspheres. Then, we seeded human adipose-derived mesenchymal stem cells (hAdMSCs) onto microspheres to evaluate cell adhesion and proliferation over time. The microspheres exhibited enhanced cell adhesion during in vitro cytocompatibility assessments. Notably, porous microspheres displayed superior cell migration compared to nonporous microspheres, attributed to their interconnected pore architecture. Additionally, hAdMSCs-laden microspheres were further studied for their chondrogenic potential under induction conditions, demonstrating cartilage-specific marker expression. Moreover, macroscopic observations and histological staining images showed cartilage regeneration in a rabbit cartilage-defect model, in which porous microspheres outperformed nonporous counterparts due to their enhanced nutrient transfer and host-scaffold integration. Together, these results suggest that silk fibroin microspheres can mimic the extracellular matrix microenvironment, support cell growth, and promote matrix accumulation for effective cartilage repair.