脐带间充质干细胞分离培养方法的优化

Objective: To investigate the best method to isolate and cultivate umbilical cord mesenchymal stem cells. Methods: Type I collagenase plus trypsin enzyme, type II collagenase plus trypsin, type II collagenase plus trypsin and hyaluronidase and adherent method were used to isolate human umbilical cord mesenchymal stem cells, and DMEM/F12 containing 10% fetal bovine serum (FBS) was used to cultivate it. During the research, we changed the culture conditions to enable the cells digested of the dissociated from umbilical cord to adhere and grow. The serum concentration was adjusted to 15% or 20%; the medium was adjusted to low glucose DMEM or high glucose DMEM medium plus epidermal growth factor and recombinant basic fibroblast growth factor. Inverted microscope was used to check the cell. MTT method was used to depict growth curve of the cells. Results: Only type I collagenase plus trypsin method could efficiently and successfully isolate human umbilical cord mesenchymal stem cells; other separation methods were negative, even though changing the culture medium, serum concentrations or adding growth factors. Conclusion: Type I collagenase plus trypsin digestion method and DMEM/F12 containing 10% fetal bovine serum can be fast and efficiently separate the umbilical cord mesenchymal stem cells.