An improved method for isolation of human umbilical cord mesenchymal stem cells / 中国组织工程研究

ABSTRACT

BACKGROUND:Human umbilical cord mesenchymal stem cells with capabilities for self-renewal and multi-differentiation have attracted widespread attention. OBJECTIVE:To develop an efficient method for isolation and culture of human umbilical cord mesenchymal stem cells, and to analyze the cellbiological features. METHODS:Mesenchymal stem cells were isolated and cultured from human umbilical cord by improved tissue cultivation. Immunophenotype and cellcycle were analyzed by flow cytometry. Growth curve was determined by MTT assay, and differentiation ability was evaluated by in vitro osteogenic and adipogenic induction as wel . RESULTS AND CONCLUSION:Some fusiform cells crawled out from human umbilical cord tissues after cultivation for 5 days and formed colonies about 10 days later. When the removed tissues were further cultured, more cells appeared again within 2 days and formed colonies after 5 days. The isolated cells exhibited similar morphology of fibroblast-like shape after passage. Furthermore, the cells expressed CD90, CD105, but were negative for the markers of CD34, CD45, HLA-DR. Population doubling time of the cells calculated from the result of MTT was about 50 hours and cellcycle analysis showed that 41.24%cells were in the G 2/S phrase. Therefore, the isolated cells had a high prolification ability. In addition, the isolated cells could be induced into osteoblasts and adipocytes in vitro. In a word, the results of this study demonstrated that the cells from the second tissues culture possessed the biological characteristics of mesenchymal stem cells and more primary umbilical cord mesenchymal stem cells were acquired through the improved method.