循环灌注接种方法在人脐带间充质干细胞骨组织工程中的应用

背景：课题前期设计了一套模块式三维灌注生物反应器系统,并初步将其应用于大鼠骨髓间充质干细胞接种于三维非纺型聚对苯二甲酸乙二醇酯（polyethylene terephthalate,PET）纤维片状载体的骨组织工程研究中。目的：应用自制的循环灌注接种系统将传代人脐带间充质干细胞接种至三维无纺布PET片状载体,并与传统的静态接种方法接种的载体进行比较。方法：培养传代人脐带间充质干细胞,流式细胞仪检测细胞表面标记,将细胞用循环灌注方法（高速率组和低速率组）和静态接种方法接种至三维聚PET无纺布片状载体。结果与结论：传代细胞形态稳定、活力好,高标达CD90,CD105,不表达CD14,CD45。循环灌注方法组的接种效率,细胞密度,增殖能力均优于静态接种组。循环灌注高速率组乳酸脱氢酶偏高,并且细胞的延迟期长于静态接种。循环灌注组的碱性磷酸酶活性高于静态接种组。结果表明,循环灌注接种方法更适合间充质干细胞骨组织工程的应用,进一步应用仍需对工程参数进行优化。

Application of cycling perfusion seeding method in bone tissue engineering of human umbilical cord derived mesenchymal stem cells

BACKGROUND： A set of modular three-dimensional perfusion bioreactor system was pre-designed in the previous studies, which has be initially applied for implantation of rat bone marrow mesenchymal stem cells into a three-dimensional non-woven polyethylene terephthalate （PET） fiber sheet carrier in bone tissue engineering. OBJECTIVE： To investigate the use of self-made cycling perfusion seeding method for human umbilical cord derived mesenchymal stem cells （hUCMSCs） seeded on PET sheet carrier. METHODS： Primary hUCMSCs were cultured and passaged. The fifth passage of cells were characterized by determining their surface antigen using a flow cytometry and then seeded onto three-dimentional PET disc scaffolds by static seeding method and cycling perfusion seeding method （low perfusion rate and high perfusion rate）. RESULTS AND CONCLUSION： The hUCMSCs of the fifth passage were steady in the cell shapes, with good viability and highly expressed CD90, CD105, while were negative for CD14, CD45. The cycling perfusion seeding method was superior to the static seeding method in the seeding efficiency, cell density and proliferative ability. Dehydrogenase activity and delayed period of cells seeded by using the cycling perfusion seeding method with high perfusion rate were better than those by using the static seeding method. Alkaline phosphatase activity was also higher in the cycling perfusion seeding method group than the static seeding method. The data suggested that the cycling perfusion seeding method is more suitable for hUCMSCs bone tissue engineering and further optimization of this method is needed.