Dynamic process enhancement on chitosan/gelatin/nano-hydroxyapatite-bone derived multilayer scaffold for...

Accurate repair of osteochondral defects is a great challenge due to the complex structure of osteochondral defects. The current study aims to prepare a biomimetic osteochondral gradient scaffold based on chitosan, gelatin and nano-hydroxyapatite and bone-derived materials for repair cartilage defects. Hereon, the chitosan/gelatin/nano-hydroxyapatite multilayer scaffold with complex hierarchical structure using iterative hierarchical method is prepared to simulate the structure of natural cartilage. And porcine femur in distal metaphysis was treated by chemical decellularization, deproteinization, decalcification, and degreasing to obtain the bone-derived scaffold simulating subchondral bone layer. We also used a spinner bioreactor for the in vitro modeling of the microenvironment which can promote adipose mesenchymal stem cell (ADSCs) recruitment into the chitosan/gelatin/nano-hydroxyapatite-bone derived multilayer scaffold under physiological flow conditions. Biological experiments have shown that osteochondral layered materials can induce ADSCs to differentiate into chondrocytes and osteoblasts and exhibit chondrogenic and osteogenic phenotypes similar to natural tissues, respectively. Furthermore, the expression both of the chondrogenic gene (Col II, ACAN, and Sox9) and osteogenic gene (Runx2, OCN and Col I) of ADSCs differentiated by mechanical stimulation were increased. These results indicate that osteochondral materials and dynamic environment are the key factors to improve cell regulation or signal molecule transfer function, and provide a feasible plan for osteochondral regeneration for future medical services.