海藻酸钠-去铁胺/壳聚糖微球对大鼠骨髓间充质干细胞成骨分化的影响

Effect of sodium alginate-g-deferoxamine/chitosan microspheres on osteogenic differentiation of rat bone mesenchymal stem cells

目的 :探讨海藻酸钠(sodium alginate,SA)-去铁胺(deferoxamine,DFO)/壳聚糖(chitosan,CS)微球对大鼠骨髓间充质干细胞(bone mesenchymal stem cells,BMSCs)增殖及成骨分化的影响。方法:将化学接枝DFO的SA与CS多孔微球通过静电吸附作用制备SA-g-DFO/CS微球,检测其形貌、孔隙率、孔径及DFO体外释放情况。体外分离、培养大鼠BMSCs,与SA-g-DFO/CS微球共培养、成骨诱导分化,以MTT法检测细胞增殖状态,钙黄绿素(calcein acetoxymethyl ester,Calcein-AM)/碘化丙啶(propidium,PI)细胞双染观察细胞活性,检测微球促细胞分化过程中碱性磷酸酶(alkaline phosphatase, ALP)、成血管及成骨相关基因的表达。采用SPSS 22.0软件包对数据进行统计学分析。结果:成功制备SA-g-DFO/CS多孔微球,可实现DFO缓释。与SA/CS微球相比,SA-g-DFO/CS多孔微球有利于细胞增殖、分化。ALP和成血管相关基因缺氧诱导因子1α(hypoxia inducible factor 1-alpha,HIF-1α)、血管内皮生长因子(vascular endothelial growth factor,VEGF)及成骨相关基因Ⅰ型胶原(CollagenⅠ,COLI)、骨钙素(osteocalcin,OCN)表达增高。结论:SA-g-DFO/CS微球可为牙槽骨缺损修复提供新的植入载体选择。

PURPOSE:To explore the effect of sodium alginate-g-deferoxamine/chitosan(SA-g-DFO/CS)microspheres on proliferation and osteogenic differentiation of rat bone mesenchymal stem cells(BMSCs).METHODS:A kind of SA-g-DFO/CS microsphere was developed through electrostatic interaction between porous chitosan microspheres and sodium alginate chemically grafted on the surface of DFO.Its morphology,porosity rate,pore size and sustained release of DFO in vitro were examined.Rat BMSCs were isolated and co-cultured with microspheres in osteogenic differentiation medium.MTT assay was used to study the influence of cell proliferation,and Calcein-AM/PI staining was used to observe the cell viability.Alkaline phosphatase(ALP)activity assay was conducted.PCR was used to detect the expression of genes related to angiogenesis and osteogenesis.Statistical analysis was performed using SPSS 22.0 software package.RESULTS:The SA-g-DFO/CS porous microspheres were successfully prepared with a sustained release of DFO.Compared with SA/CS microspheres,the SA-g-DFO/CS microspheres were conducive to cell proliferation and differentiation,with the increases in expression level of ALP,related angiogenesis genes HIF-1α,VEGF and osteogenesis genes COLI,OCN.CONCLUSIONS:The SA-g-DFO/CS porous microspheres can provide a new choice for the development of alveolar bone regeneration.