携载rhBMP-2微球的新型复合人工骨的制备及生物相容性研究

Study on preparation and biocompatibility of a novel bone substitude rhBMP-2 loaded microspheres

目的制备一种具有良好生物相容性、降解性和成骨活性、可注射的自凝固新型骨修复材料。方法采用复乳溶剂挥发方法制备携载rhBMP-2的聚乳酸与聚乙醇酸共聚物(PLGA)微球,并将其与rhBMP-2/磷酸钙骨水泥(CPC)复合,制备出rhBMP-2/PLGA微球/CPC复合人工骨。探讨材料特性包括形貌和体外rhBMP-2释放速度,采用体外细胞培养的方法测定复合材料的细胞黏附能力及其浸提液对于人骨髓基质干细胞(MSCs)增殖和成骨分化的影响。结果与单纯rhBMP-2/CPC材料相比较,复合材料rhBMP-2体外释药明显提高。材料与MSCs可良好黏附并使其增殖。体外培养时材料不同时间的浸提液对MSCs细胞的增殖具有促进作用,对于细胞成骨分化的影响与单纯CPC无明显差别。结论rhBMP-2/PLGA微球/磷酸钙骨水泥新型复合人工骨具有良好的生物相容性和活性因子缓释功能,是一种有良好应用前景的骨修复材料。

Objective To develop a novel bone repairing biomaterial with the characteristics of good biocompatibility, degradability and osteogenic activity. Methods The recombinant human bone morphogenetic protein-2 （rhBMP-2） loaded poly （lactide-co-glycolide） （PLGA） microspheres were prepared by double-emulsion and solvent -extraction technique, then compounded with rhBMP-2/CPC （calcium phosphate cement） during setting. The final product was rhBMP-2/PLGA microspher/CPC complex artificial bone. The fundamental properties including the microstructure and in vitro rhBMP-2 release of the composite material were investigated. The biocompatibility of composite samples was assessed by proliferation,alkaline phosphatase （ALP） ,and cell adhesion assays using a MSCs cell line. The interaction of cell-material on the surface of the composite was observed by scanning electron microscope （SEM）. The effect of PLGA/CPC on the behavior of marrow stromal stem cells （MSCs） was evaluated and compared with those of pure CPC and a negative control samples. Results The rhBMP- 2/PLGA/CPC composite, showing a 5 % initial release followed by a prolonged release for 28 days, released a greater amount of rhBMP-2 compared to the CPC delivery system. Morphology investigation of adhered cells by SEM indicated that MSCs cells could adhere and proliferate on the surface of the composite with a normal morphology. The MSCs cells, cultured in the presence of PLGA/CPC composite extracts samples, had a higher proliferation rate as compared to the control sample after 7 days of incubation. Compared with the control group,ALP production was higher in CPC and PLGA/CPC,but the difference was not significant. Conclusion The in vitro rhBMP-2 release can be upgraded by the intermixing of the biodegradable rhBMP-2 loaded PLGA microsphere and CPC during setting. The composite is biocompatible,nontoxic,and showing increase of cell proliferation rate and ALP production in some cases as compared to the control. Therefore,the combination of rhBMP-2 loaded PLGA microsphere and CPC forms a promising synthetic bone graft.