Repair of articular osteochondral defects with CS/HA bilayered scaffold in rabbits

[Objective]To evaluate feasibility for the treatment of osteochondral defects on the femoral condyles by chitosan/ hydroxyapatite bilayered scaffold in rabbit knee joint.[Method]Bilayered scaffolds were produced with chitosan and hydroxyapatite using a lyophilization and sintering method.The pore size and interpores of the scaffolds were observed by scanning electron microscopy(SEM)anhydrous ethanol substitution method to determinate its porosity .Rabbit bone mesenchymal stem cells(BMSC) inoculated onto the scaffold. By SEM scanning,the condition of the cells adhering onto the scaffold was observed.The cell-scaffold composite were implanted into an rabbit model of osteochondral defect 10 hour later.The result was observed 6 and 12 weeks later by HE staining and toluidine blue staining. [Result]Physicochemical characterization of CS/HA bilayered scaffold:HA possessed a porosity of 72%± 4.23% CS layer possessed a porosity of 76%± 5.0% .SEM micrograph showed CS pore at the typical transversal cross-section of CS layer with open pores and pore size with average 300 μm ranging from 200–400 μm and HA pore at the typical transversal cross-section of HA layer with open pores and pore size with average 350 μm ranging from 200–500 μm。The FTIR data showed typical peaks of the CS and HA peaks and no typical peaks of polyethylene glycol peaks. It was observed by SEM that BMSCs adhered on the chitosan scaffold and grew well but did not completely extend ,adhered on the HA scaffold and extended partially after BMSCs was implanted on the CS/HA scaffold 10 hours. The treatment of simple CS/HA scaffold improved defect filling compared with that left untreated, while the regenerated tissue was mainly fibrocartilage and showed little bone formation.When implanted with CS/HA scaffold combined with MSCs, most of the defects were filled with a well-established layer of cartilage tissue with a little bone formation observed by HE staining and toluidine blue staining. Modified Wakitani grading scale showed that A group was superior to B、C groups in 6、12weeks. There were significant differences (P0.05) between them.[Conclusion]The novel chitosan/ hydroxyapatite bilayered scaffold possesses porous structure and biocompatibility, and will possibly become a new biomaterial of osteochondral tissue engineering.