Umbilical cord-derived mesenchymal stem cells were isolated from fresh umbilical cord cells (UCs) that were obtained from informed, healthy mothers after normal deliveries and were processed as quickly as possible.
The cords were rinsed twice in PBS containing penicillin and streptomycin, and the cord blood was removed during this process. The washed cords were cut into 1-2mm pieces and floated in low-glucose DMEM containing 10% FBS. The pieces of cord were subsequently incubated at 37°C in a humidified atmosphere consisting of 5% CO2. Nonadherent cells were removed by washing. The medium was replaced every 3 days after the initial plating. When well-developed colonies of fibroblast-like cells appeared after 10 days, the cultures were trypsinized and passaged into a new flask for further expansion. All of the infused UC MSCs were derived from passages 2–5, with rigorous purification and quality control. Cell viability of purified MSCs was 92%.