3D Plotting And Characterization Of A Novel Osteochondral Integrated Gradiented Hybrid Scaffold

Articular cartilage plays an important physiological role in the body such as weight-bearing,shock absorption,and anti-slipping.However,there is no nerve,blood vessel,and lymphocyte in the cartilage,thus it is difficult for the articular cartilage to repair and regenerate by itself after injuried,and most of damaged articular cartilage would cause joint lesions of subchondral bone.Articular cartilage and subchondral bone are closely connected in anatomical structure and interact with each other in biological function,and they forms an interdependent functional unit.However,there are differences between them in terms of composition and mechanical properties,which fully shows the importance of their tissue interface.Therefore,the bone,cartilage and osteochondral transitional layer should considered simultaneously in the repair of osteochondral defects.At present,the difficulty of osteochondral tissue engineering is the design of the gradient structure of the seamless connection between the cartilage and the bone,which makes the osteocalcular integrated gradient structure scaffold become a hot issue in current research.The purpose of this study is achieve the construction of osteochondral integrated repair gradient hybrid materials.The 3D printing technology was used to prepare the hybrid materials with different components.By studying the physical and chemical properties as well as biocompatibility,the best components was selected as bone and cartilage repair materials.Further,an osteochondral integration scaffold with gradients of material and structure was fabricated by 3D printing technology and freeze-drying technology,and the structure and performance of the integrated scaffold were preliminarily discussed to provide basic experimental evidence for further studies in vivo.Firstly,the hydroxyapatite-chitosan-silica hybrid(HA-CSH)gel was prepared by sol-gel method and in situ precipitation method.The HA-CSH bone scaffold with controllable morphology and three-dimensional connectivity was constructed by 3D printing technology.Rheological tests showed that the introduction of inorganic nanoparticles greatly improved the printability of CS.The physicochemical properties of the scaffolds indicated that the introduction of n-HA improved the thermal stability of the hybrids and reduced the swelling rate of the 3D scaffolds.The formation of nano HA in situ improved the mechanical strength of the hybrid scaffolds and at the same time confered superior biological activity to the hybrid materials.Cell experiments showed that the HA-CSH scaffold had good cell compatibility,which not only supported adhesion and proliferation of m BMSCs,but also improved osteoinductivity.Secondly,using ?-(2,3-epoxypropyloxy)propyltrimethoxysilane(GPTMS)as a cross-linking agent,the gelatin was introduced into the hybrid network and successfully prepared gelatin-chitosan(Gel-CSH)cartilage scaffold with secondary three-dimensional porous structure by combining 3D printing technology and freeze-drying technology.SEM observation showed that the introduction of Gel had no significant effect on the macroscopic morphology of the three-dimensional scaffold.As increasing Gel content,internal pore number of the scaffolds increased and the pore size decreased.The cross-linking degree of the scaffolds increased,but the swelling rate and degradation rate in vitro of scaffolds decreased significantly.The compressive strength and tensile strength of the scaffolds increased with increasing Gel content.The results of dynamic cycle tests showed that the cartilage scaffolds had viscoelasticity similar to cartilage tissue,and the introduction of Gel further improved the elasticity of scaffolds.The co-culture in vitro showed that Gel-CSH scaffolds had good cell compatibility,and the introduction of Gel significantly improved adhesion and proliferation of chondrocytes.Finally,in accordance with the structural characteristics of articular osteochondral,the Gel-CSH was selected as the cartilage layer,the CSH as the cartilage calcified layer,and the HA-CSH as the subchondral bone layer,the osteochondral integration scaffolds with gradiented component and structural were prepared by 3D printing and freeze-drying techniques.The morphology observation showed that the integrated scaffold had complete structure and tight connection among layers,which not only had regular macroporous structure,but also showed porous structure with mutual penetration(10~100 m).Mechanical tests showed that the compressive strength of the integrated scaffold containing cartilage calcification layer reached 0.25 MPa,which was obviously higher than that of the osteochondral bilayer scaffold,indicating that the biomimetic structure of the osteochondral transitional layer can further improve the mechanical properties of the integrated scaffold.mBMSCs could adhere and grow well on integrated scaffolds and proliferate rapidly,indicating the good cell compatibility.The integrated scaffold is expected to be used as osteochondral tissue engineering scaffold for repair of osteochondral defects.