Preparation And Performance Of Bioactive Biomimetic Scaffolds For Vascularized Bone Repair

The repair of large bone defects has always been an international clinical challenge to be solved.The formation of early blood vessels is very important in the process of bone tissue repair.Blood vessels can enhance the transport and exchange of active factors,cells and nutrients in bone defects,thus promoting osteogenesis.Generally speaking,the main methods to treat bone defects are autogenous bone transplantation,allogenic bone transplantation and artificial bone repair material implantation.Because of their limited sources and the risk of disease transmission,the application of autologous and allograft bone implants has been limited.Artificial bone repair materials in tissue engineering have been widely investigated because of their wide sources,no risk of disease transmission,and controllable chemical composition and structure.Among them,3D printed bioceramics scaffolds are widely used in biomedical field due to their precise and controllable pore structure and personalized customization.However,conventional 3D printed bioceramic scaffolds still have some shortcomings.Therefore,the purpose of this paper is via controlling the pore structure,porosity and specific surface area of bone repair materials,to award the materials with the ability of inducing angiogenesis in the early stage,and to improve their mechanical properties.From the perspective of bionics,biomimetic bone repair scaffolds with biological internal microstructure of natural organisms were prepared by combining a variety of methods.The in vitro/vivo osteogenesis and angiogenesis of the scaffolds and the effects of biomimetic structure on their mechanical strength and toughness were mainly investigated.The main research contents and conclusions are as follows:1.Inspired by the internal structure of lotus root,lotus root-like 3D scaffolds with parallel multi-channel structure of each strut were prepared by using a new 3D printing nozzle and a special 3D printing paste with appropriate rheological and mechanical properties.By applying the new 3D printing strategy,the physical and chemical properties of the lotus root-like biomimetic scaffolds could be flexibly adjusted.We could not only prepare akermanite（AKT,Ca₂Mg Si₂O₇）,Zr O₂,Al₂O₃,Fe and Alginate lotus root-like biomimetic scaffolds,but also modulate the macro-shape,strut size,number of channels,porosity,specific surface area,and mechanical properties of these scaffolds.The lotus root-like AKT scaffolds we prepared had a porosity of up to 80%and a mechanical strength of more than 30MPa,which could meet the needs of unload-bearing bone repair.Biological analysis in vitro showed that the biomimetic scaffolds could promote the attachment,delivery and proliferation of r BMSCs in vitro.In addition,the skull defect repair in rabbit model showed that the vascular tissue successfully grew into the channels,and more new bone tissue grew into the biomimetic scaffolds than conventional 3D materials.It can be seen that this parallel multi-channel structure can promote nutrient transmission between the scaffolds and the microenvironment of bone defect,thereby greatly promote the regeneration of bone tissue inside the scaffolds.At the same time,this lotus root-like structure can indeed induce blood vessel formation in vivo,thus promote the repair of bone defects.2.Based on the preliminary exploration of the 3D printed lotus root-like biomimetic scaffolds,in order to further offer bone repair materials better bone repair performance,we prepared the 3D printed AKT scaffolds with micro-nano structures and channel structures（M-C-S scaffolds）via a single channel nozzle and hydrothermal synthesis.Microscopic morphology and composition analysis results showed that the surface of the scaffold was modified by a calcium phosphate mineralized layer with micro-nano structure.The results of biological experiments showed that the micro-nano structures and channel structures of the M-C-S scaffolds promoted the adhesion and proliferation of r BMSCs in vitro.Because their channel structure provided more space for tissue cells,the biomimetic scaffolds could deliver more cells than conventional 3D scaffolds.The M-C-S scaffolds were implanted into the defect of rabbit femur for 12 weeks.The results of Micro-CT stereogram and V-G staining of hard tissue sections indicated that more new bone was generated in the pores and channels of M-C-S scaffolds.In addition,the results of histomorphological quantitative analysis further revealed that the surface micro-nano structures and channel structures in the scaffolds showed a synergic promoting effect on osteogenesis in vivo.3.Based on the above two researches on the 3D printed bioceramic scaffolds,the third part of this study is aim to solve the main issues that traditional 3D printed bioceramic scaffolds are brittle and their plasticity does not match the human bones.In this study,inspired by the layered structure of nacre and cortical bone,by simulating biomineralization（in situ crystallization of hydroxyapatite,HA）and combining with vacuum filtration self-assembly technology,a co-inspired scaffold with multi-layer coil structure and multi-layer structure of"brick/mud".The co-inspired scaffolds had high compressive strength（98.4 MPa）,flexural strength（171.6 MPa）and toughness（1.1 MJ/m³）similar to cortical bone.In vitro biological analysis showed that the biomimetic material had greater promotion effects on the adhesion and proliferation of r BMSCs and HUVECs than other control groups.In addition,it can promote the in vitro expression of osteogenesis-related genes（BSP,OCN,Runx2 and OPN）and angiogenesis-related genes（e Nos,HIF-1α,VEGF and KDR）.We implanted the co-inspired scaffolds in the rat the rabbit femoral defects for8 weeks,and the results showed that the co-inspired scaffolds presented better in vivo angiogenic activity and osteogenic performance than control groups.Through the construction of co-inspired systems,significant improvements in the strength and toughness of biomaterials were achieved.The co-inspired scaffolds in this study had excellent mechanical strength and toughness which close to human bones,as well as good angiogenic and osteogenic properties.This kind of bone repair material with good biological activity and mechanical properties is expected to be used for repairing load-bearing bone defects.In summary,the studies address two distinct issues of how to improve the biological activity and mechanical properties of 3D printed bioceramic scaffolds.We received inspiration from natural biological structures.By constructing the lotus root-like parallel multi-channel structure,the micro-nano structure of biomineralized layer,the co-inspired system of"brick/mud"multi-layer structure and coil structure,bone repair materials with good mechanical properties as well as good angiogenesis and osteogenesis activities in vivo are prepared,which provides feasible new strategy for load-bearing bone defect repair and large bone defect repair.