Integrated trilayered silk fibroin scaffold for osteochondral differentiation of adipose-derived stem cells.

Repairing osteochondral defects (OCD) remains a formidable challenge due to the high complexity of native osteochondral tissue and the limited self-repair capability of cartilage. Osteochondral tissue engineering is a promising strategy for the treatment of OCD. In this study, we fabricated a novel integrated trilayered scaffold using silk fibroin and hydroxyapatite by combining paraffin-sphere leaching with a modified temperature gradient-guided thermal-induced phase separation (TIPS) technique. This biomimetic scaffold is characterized by three layers: a chondral layer with a longitudinally oriented microtubular structure, a bony layer with a 3D porous structure and an intermediate layer with a dense structure. Live/dead and CCK-8 tests indicated that this scaffold possesses good biocompatibility for supporting the growth, proliferation, and infiltration of adipose-derived stem cells (ADSCs). Histological and immunohistochemical stainings and real-time polymerase chain reaction (RT-PCR) confirmed that the ADSCs could be induced to differentiate toward chondrocytes or osteoblasts in vitro at chondral and bony layers in the presence of chondrogenic- or osteogenic-induced culture medium, respectively. Moreover, the intermediate layer could play an isolating role for preventing the cells within the chondral and bony layers from mixing with each other. In conclusion, the trilayered and integrated osteochondral scaffolds can effectively support cartilage and bone tissue generation in vitro and are potentially applicable for OC tissue engineering in vivo.