3D Printed Bioactive Calcium Phosphate Scaffolds Regulate Macrophage Polarization To Enhance Angiogenesis And Promote Bone Repair

Vascularization of bone repair materials is one of the most critical and urgent problems in the process of bone defect repair.Changes in macrophage phenotype and function play an important role in the process of vascularization.Endowing bone repair materials with immunomodulatory properties to enhance angiogenesis is undoubtedly a new strategy to improve bone defect repair.In this study,β-tricalcium phosphate(β-TCP)scaffolds with different pore sizes were designed and constructed,and the mechanism of β-TCP scaffolds with different pore sizes regulate macrophage polarization,thereby enhance angiogenesis and ultimately promote bone repair was studied from the perspective of immune regulation.Based on the excellent bone repair effect of β-TCP scaffolds,the repair effect of 3D printed β-TCP bone repair on clinical small-segment bone defects was preliminarily explored.Finally,strontium(Sr)with immunomodulatory effects was doped into 3D printed calcium phosphate scaffolds(Sr TCP),and whether the introduction of Sr can enhance early angiogenesis,form mature vascular network and promote the rapid repair of bone defects by regulating the phenotypic polarization and function of macrophages was investigated.Main research work and conclusions are stated as follows:(1)β-TCP powders were prepared by solid-phase sintering method,and β-TCP scaffolds with different pore size were prepared by extrusion 3D technology.The results of macrophage polarization showed that the large pore size β-TCP scaffold could inhibit the expression of proinflammatory genes and the secretion of cytokines in macrophages,and could promote the polarization of macrophages from M1 type to M2 type.β-TCP scaffold with 400 μm pore size can regulate macrophages to secrete high concentration of PDGF-bb and show excellent potential to promote angiogenesis.The paracrine secretion after co-culture of large pore size scaffold and macrophage can promote the rapid healing of scratches of HUVECs,up-regulate the expression of angiogenesis-related genes and induce the formation of tubular structure of HUVECs.(2)The effect of 3D printed β-TCP scaffolds on the repair of tibial defects was evaluated in vivo,and the mechanism by which the pore size of the scaffolds could regulate the phenotypic polarization of macrophages,enhance vascularization and promote bone regeneration was explored.It was found that compared with small pore size scaffolds,large pore size scaffolds promoted the polarization of macrophages from M1 type to M2 type in the process of bone defect repair.In addition,β-TCP scaffold with 400 μm can promote the expression and secretion of PDGF-bb during the process of bone defect repair,which ultimately leads to higher microvessel density in the bone defect area.The research results of bone regeneration showed that TCP400 scaffolds formed new bone earlier and the rate of new bone formation was faster,and finally formed dense new bone.(3)The combination of computer-aided technology and 3D printing technology realizes the personalized and precise repair of small-segment bone defects after deformity osteotomy and orthopedic surgery,and the 3D-printed β-TCP bone implant completely matches the smallsegment bone defect.The patient recovered well,and the appearance and function of the wrist were well recovered.After six months of the operation,the β-TCP bone implant was gradually absorbed,and better bone defect repair effect was achieved.(4)The performance of 3D printed Sr TCP scaffolds with different Sr-doping amounts in regulating macrophages to enhance in vitro angiogenesis was investigated.The results of the interaction between scaffolds and macrophages showed that Sr TCP scaffolds could promote the polarization of macrophages from M1 type to M2 type,and it was dependent on the doping amount of Sr.Sr TCP scaffolds could regulate macrophages to secrete high concentrations of VEGF and PDGF-bb,and exhibit excellent angiogenesis potential.The results of scratch healing,angiogenesis-related gene expression,and tube formation in HUVECs confirmed that Sr TCP scaffolds could promote early angiogenesis by regulating macrophage polarization.(5)The effect of Sr TCP scaffolds on the repair of tibial defects was evaluated in vivo,and the mechanism of scaffolds regulating macrophage polarization to enhance vascularization and then promote bone regeneration was explored.The results showed that Sr TCP scaffolds could regulate macrophages to secrete and express VEGF and PDGF-bb related to vascular sprouting,formation and maturation,and formed a vascular network with high density and quantity in the area of bone defect under the combined action of VEGF and PDGF-bb.The results of MicroCT,histology and immunohistochemistry staining showed that Sr TCP scaffold could advance the period of new bone formation,promote the transformation of cartilage to new bone and increase the rate of new bone formation,and finally achieved better effect of bone defect repair.This study found the cascade effect and regularity of 3D printed β-TCP bioactive ceramics regulating macrophage polarization to promote vascular sprouting,formation and maturation,and then to promote bone defect repair,which provides a theoretical basis for the design of bone repair materials with immunomodulatory functions.