人脐带间充质干细胞原代培养方法的改良

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Abstract Objective: To improve the tissue mass primary culture protocol of human umbilical cord-derived mesenchymal stem cells （hUC-MSCs），so as to explore their morphology, cell surface markers，osteogenic and adipogenic differentiation potential and observe biological properties. Methods: Human umbilical cords were obtained from healthy full-term delivered newborns. After stripping off blood vessels,the remaining connective tissues were cut into small fragments，and primary hUC-MSCs were separated by culture of traditional and improved tissue explants methods. The hUC-MSCs were tested with MTT method and the growth curves of them were drawn. Cell surface markers were detected by flow cytometry. After osteogenic and adipogenic differentiation，the potentiality of their differentiation was detected by chemistry staining. Results: After 3-4 days of incubation, flat or spindle-shaped fibroblast-like primary cells were presented,and cells reached 80% confluence after 5-6 days of incubation. Culture period was shortened by more than half. Flowing water-like or spiral growth morphology was presented after cell passage. MTT revealed high capability of proliferation. Flow cytometry analysis showed CD90(+), CD105(+), CD73(+), CD166(+), CD44(+), CD29(+), positive rate of 95% or more ;CD45(-),HLA-DR(-), negative rate of not more than 2%.Its differentiation potential was proved by osteogenic and adipogenic differentiation，which was proved by chemistry staining. Conclusion: An improved primary culture method of hUC-MSCs has been developed. Primary cells can be obtained from improved tissue explants method combined enzyme digestion technique. Primary cell culture time was shortened,and the efficiency of cell culture was improved. Cells isolated by the new method from the human umbilical cords had the following biological characteristics:adherent growth, high proliferation, mesenchymal stem cell phenotype，and multipotent differentiation. They could be used as the source of seed cells in scientific research and clinical applications.

Received: 14 June 2014

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	Articles by authors
	LI Xiao-Zhan
	Ma-Hong
	Xu-Hui
	Li-Shu
	Li-Yu-Mei

Cite this article:

LI Xiao-Zhan,Ma-Hong,Xu-Hui等. Improved primary cultivation of human umbilical cord mesenchymal stem cells[J]. Journal of Jiangsu University(Medicine Edition), 2014, 24(05): 375-379.

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http://zzs.ujs.edu.cn/xbyxb/EN/ OR http://zzs.ujs.edu.cn/xbyxb/EN/Y2014/V24/I05/375

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