骨髓基质干细胞与负载血管内皮生长因子/骨形态发生蛋白2聚乳酸-羟基乙酸共聚物/明胶纳米纤维支架的生物相容性

Biocompatibility of bone marrow stromal stem cells with vascular endothelial growth factor/bone morphogenetic protein 2-poly(lactic-co-glycolic acid) copolymer/gelatin nanofibrous scaffold

背景:应用骨组织工程学方法进行骨修复重建是治疗骨缺损的理想方法,负载生长因子的生物支架与种子细胞的结合是关键。目的:将大鼠骨髓基质干细胞种植于负载血管内皮生长因子(vascular endothelial growth factor,VEGF)/骨形态发生蛋白2(bone morphogenetic protein-2,BMP-2)的聚乳酸-羟基乙酸共聚物(poly(lactic-co-glycolic acid),PLGA)/明胶纳米纤维支架,观察二者的生物相容性。方法:实验分为5组:空白支架组、BMP-2组、VEGF组、VEGF/BMP-2组(1.2∶1)、VEGF/BMP-2组(0.8∶1)、VEGF/BMP-2组(0.4∶1),将各组纳米纤维支架放入细胞培养孔板底部,然后将大鼠骨髓基质干细胞接种于培养孔内,扫描电镜观察细胞在各组支架上的黏附情况,CCK-8法检测细胞增殖能力,RT-PCR检测成骨分化基因RUNX-2,ALP,OCN的表达。结果与结论:(1)在扫描电镜下可见细胞与支架黏附,并进入支架内部,VEGF/BMP-2(0.4︰1)组的细胞黏附生长状态最好,细胞与支架紧密结合成为支架细胞复合体;(2)负载生长因子的纳米纤维支架上的细胞增殖及成骨分化标记物ALP、RUNX-2和OCN表达量高于空白支架组,并且VEGF/BMP-2浓度比例为0.4︰1时成骨分化标记物表达量最高;(3)结果表明,负载VEGF/BMP-2的PLGA/明胶纳米纤维支架可促进大鼠骨髓基质干细胞的黏附、增殖及成骨分化,具备良好的细胞相容性,且VEGF/BMP-2浓度比例0.4∶1为最佳比例。

BACKGROUND:Using bone tissue engineering methods for reconstruction of bone repair is an ideal treatment for bone defects,and it is crucial to combine biological scaffolds carrying growth factors with seed cells.OBJECTIVE:To observe the biocompatibility of rat bone marrow stromal stem cells(BMSCs)seeded onto vascular endothelial growth factor(VEGF)/bone morphogenetic protein2(BMP-2)-poly(lactic-co-glycolic acid)(PLGA)copolymer/gelatin nanofibrous scaffolds.METHODS:Rat BMSCs were divided into five groups:blank scaffold group,BMP-2group,VEGF group,VEGF/BMP-2groups(1.2:1,0.8:1,0.4:1).In each group,the nanofibrous scaffold was placed at the bottom of the cell culture plate,and then rat BMSCs were seeded into the culture plate.Cell adhesion was observed under scanning electron microscope;cell proliferation was detected by cell counting kit-8;and ALP,RUNX-2and OCN expression was determined by RT-PCR.RESULTS AND CONCLUSION:Under the scanning electron microscope,the BMSCs adhered to the scaffold and further grew into the scaffold.The adherent cells grew best in the0.4:1group,in which the cells were tightly integrated with the scaffold to form a cell-scaffold complex.Compared with the blank scaffold group,the cell proliferation and expression of ALP,RUNX-2and OCN were both higher in the other groups,which was highest in the0.4:1group.To conclude,the VEGF/BMP-2-loaded PLGA copolymer/gelatin nanofibrous scaffold can promote cell adhesion,proliferation,osteogenic differentiation of rat BMSCs because of a good cytocompatibility.Moreover,the concentration ratio of VEGF/BMP-2is optimized at0.4:1.