A study on isolation,cultivation and biological characterisitics of human adipose derived mesenchymal stem cells in vitro

Objective To explore a practical method for effective isolation and culture of human adipose derived mesenchymal stem cells (ADMSCs) and observe its biological characteristics.Methods Subcutaneous adipose tissue were collected. Primary ADMSCs were isolated subsequently by the method of stirring type Ⅰ collagenase digestion and adhering to culture flask. The morphological changes of ADMSCs were observed under an inserted microscope. The expressions of CD29, CD44, CD105,CD31, CD34, and CD106 were detected by flow cytometry (FCM). Ultrastructure of ADMSCs were observed via electron microscopy. Cell cycle was analyzed by flow cytometry. Results ADSCs in both primary and passage culture were fusiform shapes, and stable in growth with active proliferation. The cells were in latency for 2 days, converted into growth period on day 3 and entered the stable phase on day 5. FCM results indicated that the positive rates of CD29, CD44, CD105, CD31, CD34, and CDl06were respectively 95.3%, 98.6%, 86.5%, 3.5%, 2.6%, and 1.3%. Cell cycle analysis showed that 84.8% of the cells were in G0/G1 phase and they revealed the structural characteristics of stem cells under an election microscope.Conclusions Stirring type Ⅰ collagenase digestion and adhering to culture flask can be effectively used to isolate and culture human ADSCs from the subcutaneous adipose tissue. The cultured ADSCs are stable in growth with active proliferation, which provides a much simple and effective collagenase digestion method. Key words: Adipose derived mesenchymal stem cells;  Cell isolation and culture;  Stem cells