The Effect Of Three-dimensional Printed Poly L-lactic Acid Scaffolds Combined With Bone Marrow Mesenchymal Stem Cells On Periosteal Distraction Osteogenesis

Objective:To investigate the feasibility of three-dimensional printed poly 1-lactic acid(3D printed PLLA)scaffolds for periosteal distraction osteogenesis(PDO).To observe the effect of distraction devices in different pore sizes combined with bone marrow mesenchymal stem cells(BMSCs)on PDO and provide experimental basis for repairing large bone defects in clinical practice.Methods:BMSCs of New Zealand white rabbits were obtained by the whole bone marrow adherent culture method,then amplified and identified by osteogenic,adipogenic and chondrogenic differentiation in vitro.According to the difference of filling rates,three kinds of PLLA distraction devices in different pore sizes were designed and printed by 3D printing technology.Scanning electron microscopy(SEM),cell counting kit 8(CCK 8)and mechanical testing instruments were then used to measure the structure,toxicity and mechanical properties of these scaffolds.The in vivo study was divided into three groups in terms of the pore sizes of the scaffolds.The periosteum of rabbits skull were distracted by the 3D printed PLLA scaffolds in different pore sizes,and each group was injected with autologous BMSCs.At the end of 8 weeks after distraction,the newly formed bone of the distracted skull area was sampled,detected and analyzed by micro-computed tomography(Micro-CT),hematoxyin and eosin(HE)and immunological histological chemistry(IHC)staining.Results:The rabbit BMSCs obtained by the whole bone marrow adherent culture method had good proliferation and differentiation abilities.The 3D printed biodegradable PLLA periosteal distraction devices had low toxicity and good biocompatibility.With the decrease of the filling rates,the pore sizes of the scaffolds gradually increased,however,the mechanical properties decreased.In vivo experiments showed that scaffolds in different pore sizes could meet the requirements of distraction of rabbit skull periosteum.The Micro-CT and HE staining results demonstrated that the quantity and quality of the newly formed bone were gradually improved with the increasing pore sizes of the scaffolds,and it is mainly reflected on the bone volume,the bone mineral density as well as the trabecular thickness.Moreover,the results of IHC showed that the expression of osteocalcin(OCN)and CD31 were also gradually increased with the increasing pore sizes of the materials,which displayed that the differentiation of new bone was closer to the cortical bone and the vascularization was stronger.Conclusion:It is feasible to use 3D printing technology to design and fabricate degradable PLLA scaffold as a device for PDO.Within the range of mechanical condition,increasing the pore size of distraction scaffold is beneficial to the promotion of PDO.