Adipose-derived stem cells：isolation, culture and differentiation into endothelial progenitor cells / 中国组织工程研究

ABSTRACT

BACKGROUND:Adipose-derived stem cels are regarded as the potential seed cels for tissue engineering. Colagenase digestion is used to isolate adipose-derived stem cels from fat pads currently. However, there are some problems, such as cumbersome operation and high cost. OBJECTIVE: To study the basic biological characteristics of adipose-derived stem cels by tissue explants culture and to explore the differentiation potential into osteoblasts, adipocytes and endothelial progenitor cels in vitro. METHODS:Adipose-derived stem cels were isolated by tissue explants technique from the bilateral groin fat pads of rats under aseptic conditions, and cultured in vitro. Cel counting kit-8 was used to detect the proliferative activity, and flow cytometry was employed to analyze the expression of cel surface markers. Passage 4 adipose-derived stem cels were cultured in osteogenic medium, adipogenic medium and endothelial progenitor cel medium for 2-3 weeks, and then the cels were identified. RESULTS AND CONCLUSION:Adipose-derived stem cels that were isolated by tissue explants culture were easily cultured, and after subculture, cels were mainly spindle-shaped and grew in clone-like manner with swirling arrangement. Cels that experienced repeated subcultures stil kept stronger proliferative ability and the cel growth curve was shaped as a parabola. Immunochemical staining analysis revealed that adipose-derived stem cels were positive for CD44, CD90 and CD29, but negative for CD31, CD45. After adipogenic/osteogenic induction, the cels were respectively positive for oil red O staining and alizarin red staining. Induced endothelial progenitor cels were identified with CD34 and the ability to uptake Dil-ac-LDL and FITC-UEA. These findings indicate using the using tissue explants culture, high-purity adipose-derived stem cels easy to proliferate can be harvested, highly express stem cels-related antigens, and have the ability to differentiate into osteoblasts, adipocytes and endothelial progenitor cels, which meet the needs of seed cels in tissue engineering research.