| Osteochondral defects caused by trauma,tumor and other causes remain a huge clinical problem.Although there are many methods for the treatment of osteochondral defects,there is no operation or treatment that could promote the complete healing of osteochondral defects.In recent years,osteochondral tissue engineering graft is considered as a potential therapeutic strategy for repairing osteochondral defects.Considering the different biological characteristics of cartilage and subchondral bone tissue,it is necessary to develop a novel,biomimetic and bioactive scaffolds to simulate the hierarchically mechanical structure and function of osteochondral tissue.Natural polymer such as collagen(COL)are commonly used in the fabrication of osteochondral biological scaffolds,but their mechanical properties are insufficient.Under the condition of lack of seeded cells,their inducing ability remains weak,which seriously limits its clinical application.Graphene oxide(GO)is a new type of carbon nanomaterial.The addition of GO could improve the mechanical properties of biomaterials and promote the proliferation and differentiation of stem cells.In addition,we found that rat knee joint-derived chondrocytes could form chondrogenic/Osteogenic extracellular matrix membrane(Cg ECM/Og ECM)with different biological characteristics through continuous culture/osteogenic culture,which may have good tissue inducing ability.Therefore,we hope to synergize the advantages of GO and Cg ECM/Og ECM to enhance the mechanical properties and tissue inducing ability of collagen scaffold.Through a novel ECM scaffold construction method,we construct a biphasic extracellular matrix combined with GO-COL scaffold,which could be used in patients with osteochondral defects in time.This study is divided into three parts:Part I: Graphene oxide-collagen(GO-COL)porous scaffolds with different GO concentrations(0%,0.05%,0.1%,0.2%)were prepared by freeze-drying and chemical cross-linking.Cell implantation and animal models were used to explore its biocompatibility,degradability and osteogenic activity in vivo and in vitro.The results show that the GO-COL porous scaffold has unique wrinkled structure,and its mechanical strength was enhanced with the GO concentration increased.For in vitro,0.1% GO-COL porous scaffolds showed better bioactivity and cytocompatibility.For in vivo,the bone repair effect of 0.1%/0.2% GO-COL porous scaffold was better than that of COL porous scaffold alone in rat cranial defect models.Therefore,0.1%GO-COL porous scaffolds show good biocompatibility and osteogenic ability in vivo and in vitro,and are expected to become a potential scaffold for bone tissue engineering and regenerative medicine.Part II: Given the different biological characteristics of cartilage and subchondral bone tissue,the Cg ECM/Og ECM containing chondrogenic and osteogenic proteins with certain thickness were prepared by continuous culture/osteogenic culture of rat chondrocytes.In the meantime,the integrated GO-COL bilayer scaffold was constructed by adjusting the concentration of GO and n HAP.The GO-COL bilayer scaffold was assembled with Cg ECM/Og ECM to construct biphasic ECM-scaffold.After material characterization and in vitro BMSCs seeding test,it was found that it had biomimetic mechanical properties and chondrogenic/osteogenic activity to BMSCs.Part III: In order to verify the repair effect of rat osteochondral defect of biphasic ECM assembled with GO-COL biomimetic composite scaffold in vivo,the full-thickness osteochondral defect of rat knee joint was constructed.Three months after implantation,the repair effect of osteochondral defect was examined by imaging and histology.The results show that the biphasic ECM-scaffold group has a better effect of repairing bone and cartilage defects in vivo than the blank group and the simple bilayer GO-COL scaffold group.It indicated that the biphasic ECM-scaffold is a potential new bioactive scaffold for osteochondral tissue engineering. |
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