BACKGROUND: Adipose-derived mesenchymal stem cells (MSCs) are taken from fat tissue obtained by liposuction aspirates, which can be repeatedly drawn, and there are adequate sources of raw materials.
OBJECTIVE: To establish the isolation and culture method of MSCs derived from human adipose in vitro, and to explore their biological properties.
METHODS: Separation by collagen enzyme digestion was used to obtain MSCs from human adipose that were cultured in vitro until the cells reached 80% confluence and passaged. The P3, P7, P10 generation cells were collected to observe and analyze the cell growth curve. The surface markers of the P5 generation cells were detected by using flow cytometry. The P4 cells were induced into osteoblasts in vitro. The cryopreserved passage cells were recovered respectively in 2 and 6 months to detect cell survival rate after resuscitation.
RESULTS AND CONCLUSION: Primary cells adhered to the wall at 3 days, then started the swift growth and formed the colony after 6 days, and reached 80%-90% fusions at 11 days, showing the desmoid shape. After passage, the cells maintained desmoid shape. The passage cells growth curves had the common characteristics: latent period was 24-48 hours, logarithm multiplication period was 3-4 days, and the cells entered the platform period at 5-6 days after the logarithm multiplication period. The flow cytometry examination showed that the expressions of CD34, CD14, HLA-DR were negative, CD44, CD105, CD13 were positive, and HLA-ABC were weakly positive on the surface of MSCs. After the recovery, the cell survival percentage reached above 90%; compared with the uncryopreserved passage cells, the cryopreserved passage cells had the same growth characteristics.