Simple, and effective isolation and differentiation of endothelial cells and their progenitor cells from human fat tissue

Recently, it has been documented that human adipose tissue (AT) has mesenchymal stem cells which can be differentiated into multiple cell lineage including endothelia cells, and there are many attempts to differentiate these cells into endothelial cells for the experimental and clinical study about angiogenesis. However, it seems to be that endothelial cells and their precursor cells in AT could be discarded in the process of extracting adipose derived stromal cells (ADSCs). In this study, we evaluated that endothelial differentiation of discardable cells. After the isolation of ADSCs, 6 hour and 24 hour non adherent floating cells were collected and characterized by the flow cytometry analysis using CD29, CD31, CD34 and CD90 cell surface markers. Also, we cultured these cells in the endothelial medium and RT-PCR for endothelial-specific mRNA was performed. Lastly, CD31 staining and Dillabeled ac-LDL uptake study were performed to confirm endothelial differentiation. In vitro experiment indicated that ADSCs showed a spindle like shape, but 6 hour and 24 hour non adherent floating cells showed a cobblestone like shape. When 6 hour and 24 hour non adherent floating cells were incubated for up to 1 weeks in endothelial medium, they detected to express a variety of endothelial-specific mRNA. Differentiated cells were also found to be able to uptake ac-LDL and stain CD 31. These results suggest that a subset of 6 hour and 24 hour non adherent floating cells derived from AT can give rise to cells with an endothelial cell-like phenotype, in vitro, at high percentages, which could be applied to in vivo vasculogenesis.