Abstract: BACKGROUND: A large number of studies have confirmed that tissue engineering scaffolds can almost completely repair osteochondral defects. However, when osteochondral defects are complicated with infection, even after thorough debridement in the early stage, the repair effect of simple osteochondral tissue engineering scaffolds is often unsatisfactory.
OBJECTIVE: To prepare fibroin/chitosan/nano-hydroxyapatite scaffold loaded with vancomycin hydrochloride sustained release microspheres, and to investigate the repair effect on infected osteochondral defect in distal femur of rabbit. 
METHODS: (1) Vancomycin hydrochloride sustained release microspheres were prepared by emulsified solvent evaporation method. The sustained-release microspheres of different weights (7.5, 10, and 12.5 mg) were mixed with fibroin protein-chitosan nanohydroxyapatite solution, and the scaffolds of fibroin protein/chitosan/nano-hydroxyapatite were prepared by chemical crosslinking method. The porosity, water absorption and expansion rate, hot water loss rate of the scaffolds, and drug sustained-release in vitro were characterized. (2) Forty-five New Zealand white rabbits were randomly divided into blank group, control group, and experimental group, with 15 rabbits in each group. The osteochondral defect and infection model of the distal femur of the right hind limb was established in both groups. The blank group was not treated, and the control group was implanted with fibroin protein-chitosan-nano-hydroxyapatite scaffold. Vancomycin hydrochloride sustained-release microspheres (10 mg) of fibroin/chitosan/nano-hydroxyapatite scaffold were implanted in the defect of the experimental group. The levels of C-reactive protein and leukocytes in blood samples were detected 1 week after operation. At 4, 8 and 12 weeks after operation, the tissue of the operative area was taken for gross observation and pathological observation. 
RESULTS AND CONCLUSION: (1) With the increase of sustained-release microspheres content, the porosity of scaffolds decreased, and there was significant difference among groups (P < 0.05). There were no significant differences in the pore size, water absorption expansion rate and hot water loss rate among the three groups (P > 0.05). Vancomycin hydrochloride was released sustainably in vitro for more than 30 days in all three groups of scaffolds. (2) The levels of C-reactive protein and leukocytes in blood samples of the experimental group were lower than those of the blank group and control group (P < 0.05). The repair of gross cartilage in the experimental group was significantly better than that in the blank group and the control group. Hematoxylin-eosin, Masson, Alcian blue and type II collagen immunohistochemical stainings showed that the osteochondral repair effect of the experimental group was significantly better than that of the blank group and the control group at each time point. (3) The results showed that fibroin/chitosan/nano-hydroxyapatite scaffolds loaded with vancomycin hydrochloride sustained-release microspheres could effectively promote the repair of open osteochondral defects.

Key words: drug-loaded microsphere, vancomycin, osteochondral defect, tissue engineering, fibroin, chitosan, nano-hydroxyapatite, scaffold