| Beta-tricalcium phosphate(β-TCP)has been used as a clinical bone filling material for filling bone defects or gaps between bone implants and bone beds due to its biocompatibility,good osteoconductivity,and degradability.However,block-shaped β-TCP implants lack a permeable porous structure and have insufficient osteoinductivity,resulting in a new bone growth rate lower than the scaffold degradation rate,which is not conducive to promoting bone integration.Therefore,in order to improve the bioactivity of block-shaped β-TCP bone implants,promote their osteoconductive properties and bone integration rate,it is necessary to prepare porous β-TCP implants and modify their surface.Although there have been reports of using 3D printing technology to prepare porous β-TCP scaffolds,it is still an urgent problem to find appropriate micro/nanostructure and coating modification on their surface to improve the osteoconductive properties of β-TCP scaffolds.In this study,β-TCP scaffolds with interconnected holes were first prepared using direct writing technology,followed by the preparation of a micro/nanorod modified layer on the surface of the scaffold using a specific hydrothermal process,and then an appropriate content of chitosan(CS)/polydopamine(PDA)coating was prepared to further improve the osteoconductive properties of the scaffold surface.First,the optimized preparation process of β-TCP ceramic slurry and the sintering process of the scaffold were studied,and the influence of slurry optimization on the mechanical strength and biological activity of the scaffold was explored.The results indicated that a threedimensional porous β-TCP scaffold was successfully fabricated using a direct writing technique.The slurry was prepared by utilizing β-TCP powder and hydroxypropyl methylcellulose as the solid phase,and a deionized water-based viscous solution containing 1 wt%gelatin and 1.5wt%sodium alginate as the liquid phase.The printed scaffold dried at 40-45℃;and a pre-sintering temperature of 300℃ was set,and the scaffold was kept at this temperature for 1 hour.Furthermore,the mechanical strength and bone marrow mesenchymal stem cell proliferation ability of the scaffold were confirmed to be improved after slurry optimization through mechanical property testing and cell proliferation level detection.Then,the preparation,physicochemical properties and in vitro biological properties of the hydrothermal-coating composite surface modified β-TCP scaffold were studied.The effects of different hydrothermal agents,solvent concentrations,and reaction times on the proliferation of stem cells were studied.After obtaining the optimized parameters for the four groups of hydrothermal processes,compared with the β-TCP scaffold control group,the surface micro/nano morphology,Ca-P ratio,and hydrophilicity of the four groups of hydrothermal modified scaffolds were explored,and the compressive strength of each group of scaffolds was tested.The ability of the scaffolds to mineralize in vitro was investigated by soaking the scaffolds in simulated body fluid(SBF)for 7-28 days.The influence of surface micro/nano morphology on stem cell morphology and fate was studied by observing cell adhesion and morphology,and detecting cell proliferation and osteogenic differentiation performance.The results showed that the surface with protruding micro/nano morphology could significantly enhance the hydrophilicity of the scaffold surface.The scaffold treated with SBF containing glycine had the highest amount of mineralized hydroxyapatite blocks on the surface after soaking in SBF for 28 days.Moreover,different surface micro/nano morphologies had different effects on stem cell proliferation and osteogenic differentiation.The nano-pitted surface was conducive to promoting stem cell proliferation but had a relatively small effect on promoting stem cell osteogenic differentiation.The intricate distribution of micro/nano rod morphology could significantly promote stem cell osteogenic differentiation,although not as much as the pitted surface morphology could promote stem cell proliferation.Subsequently,the preparation,physicochemical properties,and in vitro biological performance of the hydrothermal-coated composite modified β-TCP scaffold were investigated.The effects of changes in solution concentration and coating time during the preparation of CS and PDA coatings on the proliferation performance of stem cells were studied.The synergistic effect of CS/PDA composite coating on promoting stem cell proliferation was validated,and a hydrothermal-coated composite modified β-TCP scaffold was prepared by coating the surface of the hydrothermally modified scaffold with CS/PDA coating.The surface morphology,elements,hydrophilicity,and mechanical properties of the composite modified scaffold were characterized to explore its in vitro mineralization ability.Furthermore,the effects of surface micro/nano-rod structure,CS/PDA coating,and micro/nanostructure-coating composites on stem cell adhesion,proliferation,and osteogenic differentiation were studied through in vitro cell experiments.The results showed that CS/PDA coating could improve the compressive strength of the β-TCP scaffold and hydrothermal modified scaffold,but only increase the surface hydrophilicity of the β-TCP scaffold.After soaking in SBF for 28 days,the surface of the composite modified scaffold deposited the most hydroxyapatite(HA)blocks,followed by the coated modified scaffold,and the unmodified β-TCP scaffold the least.Moreover,CS/PDA coating could promote the early proliferation and osteogenic differentiation of stem cells better than the surface micro/nano-rod structure,and hydrothermal-coating composite modification had a more excellent synergistic promotion effect.Finally,the in vivo osteogenic performance of the hydrothermal-coated composite surfacemodified β-TCP scaffold was investigated.Untreated β-TCP scaffold,hydrothermally modified scaffold,coating-modified scaffold,and hydrothermal-coated composite modified scaffold were implanted in the rat skull,and the visceral tissues of rats were sliced and stained at 2 and 6 weeks after implantation to assess the scaffold toxicity.In addition,the growth of new bone tissue around and inside the scaffold was evaluated by Micro-CT analysis,bone volume evaluation index detection,and hard tissue section staining observation.The results showed that hydrothermal and coating modifications did not produce toxicity to β-TCP materials.Both surface micro/nanorod structures and CS/PDA coatings promoted the growth of bone tissue in the early stage of implantation,and the micro/nanorod structures could promote the growth of bone trabeculae in the long term.The hydrothermal-coated composite modified scaffold had the advantages of both hydrothermal and coating modifications,and could promote the rapid growth of new bone in the early stage of implantation and long-term promotion of the growth and thickness of bone trabeculae,demonstrating synergistic promotion of bone integration. |
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