骨髓间充质干细胞膜片和脐静脉内皮细胞共培养模型的构建及鉴定  

Co-culture model of bone marrow mesenchymal stem cell sheet and umbilical vein endothelial cells: establishment and identification

作　　者：周震 王亚敏 任飞 张兆强 曾曙光 杨熙 Zhou Zhen;Wang Yamin;Ren Fei;Zhang Zhaoqiang;Zeng Shuguang;Yang Xi(Stomatology Hospital, Southern Medical University,Guangzhou 510280, Guangdong Province, China)

机构地区：[1]南方医科大学口腔医院

出　　处：《中国组织工程研究》2019年第25期3951-3955,共5页Chinese Journal of Tissue Engineering Research

基　　金：广东省自筹经费类科技计划项目(2017ZC0124),项目负责人:周震;广东省医学科学技术研究基金项目(C2017061),项目负责人:任飞;广东省自然科学基金项目(2018A030313759),项目负责人:张兆强;广州市科技计划项目(201707010193),项目负责人:曾曙光~~

摘　　要：背景:细胞膜片技术是近年来热门的组织工程学技术,但是膜片中间部位的营养供给和移植后血供问题易导致失败,所以预血管化是提高组织工程材料移植后存活率的关键。目的:比较在不同氧含量下,人骨髓间充质干细胞和脐静脉内皮细胞共培养形成预血管化骨髓间充质干细胞膜片的生物学性能。方法:实验分2组:常氧共培养组即常氧(体积分数20%O2)条件下形成人骨髓间充质干细胞膜片,常氧条件下加入人脐静脉内皮细胞共培养;低氧共培养组,即低氧(体积分数2%O2)条件下形成人骨髓间充质干细胞膜片,常氧条件下加入人脐静脉内皮细胞共培养。采用免疫荧光染色技术观察并比较共培养7d两组的微血管生成情况,并通过ELISA法比较不同氧含量下骨髓间充质干细胞膜片内血管内皮生长因子的水平。结果与结论:①与常氧共培养组比较,低氧共培养组的血管更长、血管密度更高、管间交互连接更多(P <0.05);②低氧共培养组的血管内皮生长因子水平高于常氧共培养组(P <0.05);③结果证实,低氧共培养组较常氧共培养组膜片更利于血管生长,对于建立优化的预血管化膜片具有可行性。BACKGROUND: Cell sheet technology is a recent popular technique for tissue engineering. However, a failure is prone to occur because of poor nutrient supply in the middle part of the membrane and post-transplantation blood supply, so pre-vascularization determines the survival rate of tissue engineering materials after transplantation.OBJECTIVE: To investigate the pre-vascularization of bone marrow mesenchymal stem cell sheets co-cultured with human umbilical vein endothelial cells under different oxygen tensions.METHODS: Bone marrow mesenchymal stem cell sheets were prepared under physiological hypoxia(2% O2) and normoxia(20% O2), and then co-cultured with human umbilical vein endothelial cells under normoxia. At 7 days of co-culture the angiogenesis was observed and compared by immunofluorescence staining. The level of vascular endothelial growth factor was compared under different oxygen tensions using ELISA.RESULTS AND CONCLUSION: Compared with the normoxia co-culture group, there were longer microvessels, higher vessel density and more microvessel networks in the hypoxia co-culture group(P < 0.05). The expression of vascular endothelial growth factor in the hypoxia co-culture group was higher than that in the normoxia co-culture group(P < 0.05). Therefore, the hypoxia co-culture is more beneficial to the growth of microvessels than the normoxia co-culture, and it is feasible to construct an optimized prevascularized cell sheets.

关 键 词：低氧 常氧 细胞膜片 骨髓间充质干细胞 免疫荧光染色 脐静脉内皮细胞 血管内皮生长因子 干细胞 血管化 组织工程 

分 类 号：R453[医药卫生—治疗学] R364[医药卫生—临床医学]