Abstract:

BACKGROUND: Studies have found that cord blood contains a large member of hematopoietic stem cells and abundant mesenchymal stem cells.
OBJECTIVE: To establish an isolating, culturing and proliferating method for human umbilical cord blood mesenchymal stem cells (HUMSCs) and then to observe cell biological characteristics in vitro.
METHODS: HUMSCs were isolated and purified by density gradient centrifugation and adhering to the culture plastic. After successive subculture and amplification, the morphology was observed under microscope and the growth curve was drawn. Cell cycle was determined by flow cytometry; The expression of CD29, CD44 and CD34 in HUMSCs were detected by immunohistochemistry (SP); HUMSCs were induced to differentiate into adipocytes in DMEM with 20% horse serum and identified by oil red O staining; HUMSCs were induced to differentiate into osteoblasts by 0.1 μmol/L dexamethasone, 10 mmol/L β-glycerol phosphate sodium ascorbate and 50 μmol/L ascorbic acid and identified by alkaline phosphatase staining.  
RESULTS AND CONCLUSION: High purity HUMSCs were isolated from umbilical cord blood. HUMSCs were spindle cells and their living behavior was quite stable. In vitro subcultured nine generations had no morphological changes and no signs of aging; Passage 3 of HUMSCs could differentiate into adipocytes and osteoblasts; The results showed that density gradient centrifugation, adherent culture and digestion time control can obtain an access to high purity HUMSCs with strong proliferation and passage ability in vitro.