人脐血来源MSCs的主要生物学特性的研究

Biological characters of mesenchymal stem cells of human umbilical cord blood

目的:探讨人脐血间充质干细胞(MSCs)分离、纯化、扩增、表面标志、免疫表型及其造血生长因子(HGFs)分泌等生物学特性。方法:(1)F icoll法分离、纯化人脐血、成人骨髓和胎儿骨髓MSCs,动态观察不同来源MSCs的生长状况。(2)流式细胞仪检测脐血和骨髓MSCs表面CD34、CD45、CD14、CD29、CD44、CD105、CD166、CD80、CD86、CD40和CD40L的表达。(3)ELISA法检测脐血和骨髓MSCs培养上清中SCF、TPO、FLT-3L和IL-6的分泌水平。结果:(1)所有骨髓标本中均可分离纯化出MSCs,而15份脐血中仅4份分离纯化出MSCs,脐血MSCs和骨髓MSCs的细胞形态无明显差异,脐血MSCs原代培养所需要的单个核细胞(MNC)量为骨髓MSCs的3倍,且其原代培养的增殖速度较慢,但脐血MSCs传代培养的增殖速度和骨髓相似。(2)脐血MSCs和骨髓MSCs表面标志无差异,均不表达造血细胞表面标志以及与免疫识别有关的表面抗原。(3)脐血和骨髓MSCs分泌SCF、TPO、FLT-3L和IL-6的水平相似。结论:(1)脐血和骨髓同样可以分离、纯化MSCs,但脐血中MSCs的含量少,且大部分脐血中不含MSCs。(2)纯化的脐血MSCs扩增能力、表面标志、免疫学表型及HGFs分泌水平和骨髓MSCs相似。

AIM： To study the isolation, expansion and purification of mesenchymal stem cells （MSCs） from human umbilical cord blood （ UCB）, and investigate some biological identities of MSCs. METHODS ： （ 1 ） MSCs of UCB, adult bone marrow （BM） and fetus BM were isolated by centrifugation with Ficoll, and the different kinds of MSCs were observed everyday. （2） Surface markers of MSCs were identified by flow cytometry. （ 3 ） The level of HGFs （ TPO, SCF, FLT -3L, IL-6） secreted by different sources of MSCs was checked by ELISA method. RESULTS： （1） No difference in morphology of the colonies between UCB MSCs and BM MSCs was observed. However, the mononuclear cells needed in culture of UCB MSCs was about 3 times more than that in culture of BM MSCs. The times of UCB MSCs colony formation and confluencing were longer than that of BM in primary culture. （ 2 ） After passaged, there was no significant difference in the proliferation rates of 3 kinds of MSCs. Only 4 of 15 UCB samples contained a homogeneous population of MSCs. （3） UCB MSCs shared the same markers with BM MSCs. Neither hematopoietic marker nor immunologic recognition antigens were expressed. （4） The level of hematopoietic growth factors （HGFs） secreted by 3 kinds of MSCs was similar. CONCLUSIONS： （ 1 ） MSCs were isolated from UCB, but the amount of MSCs in UCB was smaller than that in BM, and just seldom samples of UCB contained homogeneous MSCs. （2） MSCs from UCB and BM shared the same biological characteristics, such as proliferation ability, surface markers, immunophenotypes and HGFs secretion.