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目的:通过观察以人脐带血血清为主体的培养体系对脐带间充质干细胞(Mesnchymal Stem Cells,MSCs)体外培养扩增的影响,探讨人脐带血血清替代动物血清用于培养临床组织工程用干细胞的可行性。建立从足月脐带分离间充质干细胞和体外传代培养技术,并对其生物学特性进行研究。方法:采用双酶法分离脐带间充质干细胞,并通过传代进行纯化和扩增培养,绘制生长曲线:用流式细胞仪检测脐带表面抗原及细胞周期。结果:成功建立了以人脐带血血清为主体的培养体系的脐带间充质干细胞培养扩增的方法:流式细胞仪检测结果显示,贴壁细胞均表达CD44、CD29,低表达CD106,不表达造血细胞表型CD14、CD34、CD45和内皮细胞表型CD31,也不表达HLADR:细胞倍增时间为30h,细胞周期分析表明,G0～G1期和+G2+M期所占比例分别为78.84%和11.16%。结论:应用灭活脐带血清培养体系可成功扩增人脐带间充质干细胞,培养的细胞具有间充质干细胞的基本特性,为建立间充质干细胞库和临床应用提供了理论依据。
OBJECTIVE: To investigate the effects of human umbilical cord blood serum-based culture system on the growth and expansion of umbilical cord mesenchymal stem cells (MSCs) in vitro and to explore the feasibility of using human umbilical cord blood serum instead of animal serum for culturing clinical tissue engineering stem cells Feasibility. Establishment of isolation of mesenchymal stem cells from full-term umbilical cord and in vitro culture techniques, and to study its biological characteristics. Methods: Umbilical cord mesenchymal stem cells were isolated by double enzymatic method and purified by subculturing. The growth curves were drawn. The surface antigens and cell cycle of umbilical cord were detected by flow cytometry. Results: The method of culture and expansion of umbilical cord mesenchymal stem cells was established successfully. The results of flow cytometry showed that the adherent cells expressed CD44, CD29, but not CD106 The hematopoietic cell phenotype CD14, CD34, CD45 and endothelial cell phenotype CD31, also did not express HLADR: The cell doubling time was 30h, the cell cycle analysis showed that the proportion of G0 ~ G1 phase and + G2 + M phase were 78.84% 11.16%. Conclusion: Human umbilical cord mesenchymal stem cells can be successfully expanded by using inactivated umbilical cord blood serum culture system. The cultured cells have the basic characteristics of mesenchymal stem cells, which provides a theoretical basis for establishing the mesenchymal stem cell bank and clinical application.