缓释骨诱导因子的玉米醇溶蛋白/左旋聚乳酸纳米纤维材料对骨髓间充质干细胞成骨分化的影响

Effect of Bone Induction Factor-Loaded Zein/PLLA Scaffolds on Vitro Osteogenesis of Mesenchymal Stem Cells

目的:探讨缓释骨诱导因子的玉米醇溶蛋白(Zein)/左旋聚乳酸(PLLA)同轴静电纺丝纳米纤维支架对骨髓间充质干细胞(MSCs)成骨分化作用。方法:制备载有骨形态发生蛋白-2(BMP-2)和地塞米松(DEX)的壳-核型同轴静电纺丝支架,使用透射电子显微镜(TEM)验证纳米纤维的壳-核结构。分离原代骨髓间充质干细胞并培养,使用扫描电子显微镜(SEM)评估骨髓间充质干细胞的黏附、生长情况。检测碱性磷酸酶(ALP)活性并定量分析,茜素红染色(ARS)检测钙矿化沉积物并定量分析。结果:骨髓间充质干细胞经成骨诱导分化后,表现出成骨细胞的特性。透射电子显微镜观察到以玉米醇溶蛋白和左旋聚乳酸为原料,利用同轴静电纺丝技术制备的纳米纤维具有完整的壳-核结构。扫描电子显微镜观察到骨髓间充质干细胞在加载骨形态发生蛋白-2和地塞米松的纳米纤维支架显示出良好的细胞黏附、伸展能力。加载双重诱导因子的Zein-DEX/PLLA-BMP-2纳米纤维支架上骨髓间充质干细胞表现出更高的碱性磷酸酶活性和细胞外基质(ECM)矿物沉积,优于对照组。结论:以天然生物材料玉米醇溶蛋白和生物高分子材料左旋聚乳酸为原料制备了加载生物活性因子骨形态发生蛋白-2和地塞米松的壳-核结构的新型同轴静电纺丝纳米纤维支架,形成类似细胞外基质样结构,刺激骨髓间充质干细胞早期黏附、增殖、诱导成骨分化,适合骨组织工程的应用。

Objective:To study the effect of Zein/PLLA co-axial electrospun nanofiber scaffold with sustained release of bone induction factor on the osteogenic differentiation of bone mesenchymal stem cells(MSCs).Methods:The core-shell coaxial electrospun scaffolds containing bone morphogenetic protein-2(BMP-2)and dexamethasone(DEX)were prepared,and the core-shell structure of the nanofibers was verified by transmission electron microscopy(TEM).The primary MSCs were isolated and cultured,and the adhesion and growth of MSCs were evaluated by scanning electron microscopy(SEM).Osteoblast differentiation and mineralization were determined by Alkaline phosphatase(ALP)activity and quantitative assay and Alizarin Red S(ARS).Results:MSCs showed osteoblastic characteristics after osteogenic induction and differentiation.TEM observed that the nanofibers prepared by coaxial electrospinning with zein and PLLA as raw materials had a complete shell core structure.SEM observed that MSCs showed good cell adhesion and extension ability in the nanofiber scaffold loaded with BMP-2 and DEX.MSCs on Zein DEX/PLLA-BMP-2 nanofiber scaffold loaded with double inducers showed higher ALP activity and extracellular matrix(ECM)mineral deposition,and were superior to the control group.Conclusion:A novel coaxial electrospun nanofiber scaffold with core-shell structure loaded with bioactive factors BMP-2 and DEX was prepared from natural biomaterials(Zein)and biopolymers(PLLA).It shows ECM like structure,stimulates early adhesion and proliferation of MSCs,induces osteogenic differentiation,and is suitable for the application of bone tissue engineering.