The Study On The Preparation And Osteogenic Induction Potential In Vitro Of 3D Printed Polyetheretherketone/Hydroxyapatite Whisker Composites

Background:PEEK has attracted the attention of researchers because of its suitable elastic modulus,good chemical stability,biocompatibility,X-ray permeability,thermal stability and self-lubrication,and good antibacterial properties.At present,PEEK has been applied in clinical implant such as interbody fusion cage,dental implant,anchor screw and skull prosthesis.However,the clinical application of PEEK is limited by the poor bone integration in vivo due to its biological inertia.In previous studies,nanohydroxyapatite（n HA）has been successfully added into PEEK and improved its biological activity.Micron-grade hydroxyapatite whisker（HAW）has the same chemical composition and calcium-phosphorus ratio as n HA and it could bind to bone by forming a hydroxyapatite layer on its surface.HAW have the same chemical composition and calciumphosphorus ratio as n HA,can bind to bone by forming a hydroxyapatite layer on its surface,and enhance the mechanical properties of the polymer.Objective:This research was aimed to investigate the effects of hydroxyapatite whisker incorporation and 3D printing process on the biocompatibility of polyetheretherketone and the osteogenic differentiation of rat bone marrow mesenchymal stem cells induced in vitro.It provides theoretical basis for the modification of polyetheretherketone and its application in orthopedics field.Methods:In this study,PEEK composites with different proportions of hydroxyapatite whisker was prepared by injection molding method.In order to select the best proportion of hydroxyapatite whisker,we tested its tensile properties and bending properties.The samples were printed into porous scaffolds by fused deposition modeling（FDM）and compared with pure PEEK scaffolds and solid discs.The surface morphology and hydrophilicity of the samples were tested by scanning electron microscope and droplet angle tester.Bone marrow mesenchymal stem cells（BMSCs）were obtained from SD rats by bone marrow rinsing and whole bone marrow culture.According to the requirements of different experiments,BMSCs were inoculated on the sample surface in a certain number at different time points for biological characterization research.The cytotoxicity of the materials was evaluated by Live/Dead staining and CCK-8 cell counting assay.The cytoskeleton and nuclear morphology of BMSCs stained by phalloidine were observed by confocal laser scanning microscope.Scanning electron microscope（SEM）was used to observe the adhesion morphology of cells on the material surface.Qualitative and quantitative alkaline phosphatase staining and alizarin red staining were used to analyze the effect of the samples on osteogenesis induction.Results:In this study,the mechanical properties of the composites were enhanced by adding hydroxyapatite whisker into the PEEK matrix,and the optimal proportion was selected as 20%,and the elastic modulus of the composites was closer to human cortical bone.HAW/PEEK scaffold with pore size of460μm was successfully fabricated by 3D printing technology of fused deposition modeling.Biocompatibility test proved that the addition of hydroxyapatite whiskers and 3D printing process did not bring cytotoxicity to the material.Rat BMSCs adhered to the surface of the material and prolifated well,and obvious cell pseudopodia were observed by scanning electron microscopy.Qualitative and quantitative results of alkaline phosphatase staining and alizarin red staining showed that the addition of HAW and three-dimensional porous structure promoted the osteogenic differentiation of BMSCs.Conclusion:In this study,different proportions of PEEK hydroxyapatite whisker composite were prepared.The addition of hydroxyapatite whisker enhanced the mechanical properties of the composite,and 20% proportion of hydroxyapatite whisker polyetheretherketone composite had the best mechanical properties.Hydroxyapatite whisker polyetheretherketone composites can be prepared by fused deposition modeling and constructed porous scaffolds.The addition of hydroxyapatite whisker increases the hydrophilicity of the surface of PEEK.The addition of hydroxyapatite whiskers and the porous structure promoted the adhesion,proliferation and osteogenic differentiation of BMSCs.We provides a theoretical basis for the study of in vivo implantation modification of PEEK.