Abstract:

BACKGROUND: There are different methods to isolate and culture human umbilical cord mesenchymal stem cells (hUC-MSCs), so how to fast and efficiently harvest hUC-MSCs have become a research hotspot.
OBJECTIVE: To optimize digestive enzymes components for the preparation of hUC-MSCs, in order to lay the foundation of preparation for hUC-MSCs.
METHODS: Human umbilical cords were collected from full term deliveries under aseptic conditions. According to different mixed enzyme concentration ratio, the samples were divided into three groups: mixed enzyme Ⅰ group, mixed enzyme Ⅱ group, mixed enzyme Ⅲ group. Then according to digest time each group into three subgroups: 1 hour, 2 hours, and 3 hours. Finally, suspended cell volume was decided as 4 mL to count cells. Dulbecco’s Modified Eagle Medium with fetal bovine serum was used for cell culture.
RESULTS AND CONCLUSION: Based on the three digestion enzyme concentration, at the duration of 1 hour, 2 hours and 3 hours, mixed enzyme Ⅲ group had the highest total cell number and the total cell rate was statistically different from the other groups (P < 0.05). At the duration of 3 hours, live cell rate was the lowest in the mixed enzyme Ⅲ group, and there was statistically significant differences among these groups (P < 0.01). The optimal isolation for hUC-MSCs is 0.3% collagenase Ⅱ, 0.1% trypsin, 0.02% EDTA, 0.1% hyaluronidase and 0.1% DNA enzymeⅠ digestion for 2 hours.