Isolation and culture of rat adipose-derived stem cells and the test of their differentiation capacity

Objective To isolate,culture and identify adipose-derived stem cells(ADSCs) from SD rats so as to provide a theoretical basis for later experiments which are to select suitable seed cells.Methods Adipose tissues were isolated from the inguinal fat of SD rats.By combining zymogen digestion with differential velocity adherent method to obtain cells,the morphology and proliferation characteristics of the cells were observed under the inverted phase contrast microscope.Their growth curves were detected;freezing and resuscitation were performed.The third-generation cells differentiated into the osteogenic direction by osteogenesis-inducing liquid,and then underwent type Ⅰ collagen staining,alkaline phosphatase dyeing,and alizarin red staining.Adipogenic-inducing liquid was used to induce them into the adipogenic direction,and oil red O staining was conducted.Results ADSCs were successfully obtained and cultured from the rat adipose tissues.They presented fibroblast-like growth.After being cultured by osteogenesis-inducing liquid,they showed osteogenic characteristics.Type Ⅰ collagen staining,alkaline phosphatase dyeing and alizarin red staining were all positive.After being cultured by adipogenic-inducing liquid,they displayed adipogenic properties.After oil red O staining,fat droplets were visible within the cytoplasm.The ADSCs showed no significant decrease in their proliferation activity and capability of differentiating into diverse cell types after being cryopreserved in liquid nitrogen for two months.Conclusion Rat ADSCs were successfully isolated and cultured.The ADSCs obtained could grow and proliferate rapidly,capable of differentiating into diverse cell types.It provides a theoretical basis for later experiments which use tissue engineering to repair tissue damage.