ABSTRACT
BACKGROUND:A great amount of mesenchymal stem cels can be successfuly derived from fat tissue and induced to differentiate into osteoblasts, chondrocytes, adipocytes and myocardial cels. OBJECTIVE:To establish the method of isolating, culturing and osteogenic differentiation of adipose-derived stem celsin vitro, and to explore the potential of adipose-derived stem cels as seed cels for bone tissue engineering. METHODS:Colagenase enzymatic digestion was used to isolate adipose-derived stem cels from human fat tissue which were then culturedin vitro. Flow cytometry was used to detect cellsurface markers. cellcounting kit-8 assay was performed to examine cellviability. Adipose-derived stem cels were induced by osteogenic induced reagent to differentiate into bone cels. In addition, we also performed BCIP/NBT method to detect alkaline phosphatase activity. Alizarin red staining was used to detect the formation of calcium node. RT-PCR was performed to examine alkaline phosphatase and osteopontin expression. RESULTS AND CONCLUSION:We successfuly obtained adipose-derived stem cels from fat aspirated liquid. Adipose-derived stem cels obtained could passage stably and proliferate highly. Flow cytometry data showed the expression of stem cellsurface markers. Adipose-derived stem cels showed typical osteoblast morphology after osteogenic differentiation. Alkaline phosphatase staining was positive and alizarin red staining displayed the formation of calcium node. Furthermore, we found that alkaline phosphatase and osteopontin mRNA was expressed after differentiation 0, 3, 7, 14, 21, 28 days. These findings indicate that adipose-derived stem cels can be obtained from fat tissue through enzymatic digestion, differentiate towards bone cels, and express alkaline phosphatase and osteopontin, which can become potential seed cels for bone tissue engineering.