控释重组人骨形态发生蛋白-2和血管内皮生长因子的微囊支架对骨髓间充质干细胞向成骨细胞分化的影响  

Effects of microcapsule-based scaffolds for controlled-release of bone morphogenetic protein 2 and vascular endothelial growth factor on differentiation of bone marrow-mesenchymal stem cells to osteoblasts

作　　者：任前贵[1] 张坤 任威[1] 孙韬[1] 李永峰 马丽波[1] REN Qian-gui;ZHANG Kun;REN Wei;SUN Tao;LI Yong-feng;MA Li-bo(The Emergency Department,Second Affiliated Hospital of Inner Mongolia Medical University,Hohhot 010059,Inner Mongolia Autonomous Region,China)

机构地区：[1]内蒙古医科大学第二附属医院急诊科

出　　处：《中国生物制品学杂志》2019年第12期1357-1360,1376,共5页Chinese Journal of Biologicals

基　　金：内蒙古医科大学科技百万项目（YKD2015059)

摘　　要：目的 探讨控释重组人骨形态发生蛋白-2(recombinant huaman bone morphogenetic protein-2,rhBMP-2)及血管内皮生长因子(vascular endothelial growth factor,VEGF)微囊支架对骨髓间充质干细胞(bone marrow mesenchymal stem cells,b MSCs)向成骨细胞分化的影响。方法 以聚乳酸-聚乙二醇-聚乳酸三嵌段共聚物[polylactide-poly(ethylene glycol)-polylactide,PELA]为囊材,采用复乳溶剂挥发法制备外黏附rhBMP-2内包封VEGF的微囊支架。经ELSIA法检测微囊支架在PBS中释放rhBMP-2和VEGF的浓度。将微囊支架加入bMSCs,于培养后第3、7、14天,MTT法检测微囊支架对bMSCs活性的影响,Western blot法检测微囊支架对bMSCs向成骨细胞分化过程中MAPK通路相关蛋白及碱性磷酸酶(alkaline phosphatase,ALP)表达水平的影响。结果 微囊支架于PBS中培养第2天rhBMP-2释放约60%,VEGF释放约32%。随着培养时间的延长,微囊支架对bMSCs的细胞活性无明显影响(P>0. 05);培养后第14天磷酸化ERK1/2及ALP表达水平均显著高于第3和7天(P <0. 05),培养后第7天显著高于第3天(P <0. 05);培养后第3、7、14天磷酸化JNK及磷酸化p38表达水平变化差异无统计学意义(P> 0. 05)。结论 控释rhBMP-2及VEGF的微囊支架可诱导bMSCs向成骨细胞分化,可能是通过激活MAPK通路发挥作用。Objective To investigate the effects of microcapsule-based scaffolds for controlled-release of recombinant human bone morpho-genetic protein-2(rhBMP-2)and vascular endothelial growth factor(VEGF)on differentiation of bone marrow mesen-chymal stem cells(bMSCs)to osteoblasts.Methods The microcapsule-based scaffolds with rhBMP-2 on the outer surface and VEGF encapsulated were prepared with polylactide-poly(ethylene glycol)-polylactide(PELA)by emulsion solvent volatilization method.The concentrations of rhBMP-2 and VEGF released by microcapsule-based scaffolds were determined by ELISA.The activities of bMSCs on days 3,7 and 14 after culture with microcapsule-based scaffolds were determined by MTT assay.The effects of microcapsule-based scaffolds on the expression levels of MAPK pathway-associated proteins and alkaline phosphatase(ALP)during differentiation of bMSCs to osteoblasts were evaluated by Western blot.Results On the second day after culture in PBS,about 60%of rhBMP-2 and about 32%of VEGF were released from the microcapsule-based scaffolds.The microcapsule-based scaffolds showed no significant effect on the activity of bMSCs with the increasing culture time(P>0.05).The expression levels of phosphorylated ERK1/2 and ALP were significantly higher on day 14 after culture than on days 3 and 7(P<0.05),and significantly higher on day 7 than on day 3(P<0.05).However,the expression levels of phosphorylated JNK and phosphorylated p38 on days 3,7 and 14 showed no significant difference(P>0.05).Conclusion The microcapsule-based scaffolds for controlledrelease of rhBMP-2 and VEGF induced differentiation of bMSCs to osteoblasts by a possible mechanism of activating MAPK pathway.

关 键 词：人骨形态发生蛋白-2 血管内皮生长因子 微囊支架 骨髓间充质干细胞 成骨细胞 MAPK通路 

分 类 号：Q28[生物学—细胞生物学]