Background aims. Adipose-derived stem cells (ASCs) are seen as a promising candidate for the treatment of diabetic foot ulcers, and it was hypothesized that treating ASCs with hypoxia and trypsin would increase the pro-angiogenic effect of these cells.
Methods. The possible increased pro-angiogenic effect was investigated using ASCs treated with hypoxia, trypsin, and a combination of these. The pro-angiogenic effect of ASCs was measured in two ways. First, the expression and secretion of VEGF by the ASCs was assessed. The expression of vascular endothelial growth factor (VEGF) was determined by real-time RT-PCR normalized to the reference genes PPIA and YWHAZ. Moreover, the secretion of VEGF was assessed by ELISA. Second, the influence of ASCs on endothelial cell migration was investigated.
Results. The hypoxic treatment of the ASCs resulted in a significant increased expression of VEGF (P = 0.000) compared to untreated ASCs, as did the trypsin treatment (P = 0.001). The combinational treatment of hypoxia and trypsin resulted in a significant higher up regulation of the relative expression of VEGF (P = 0.000), indicating an additive effect. The secretion of VEGF was both after hypoxic treatment and after trypsin treatment significantly increased (P = 0.000, P = 0.005). The combination of hypoxic and trypsin treatment increased the secretion even further, which was significantly higher than both control, hypoxic and trypsin treated cells (P = 0.000, P = 0.002, and P = 0.000, respectively). No results were obtained from the endothelial migration assay.
Conclusion. In conclusion, the combination of hypoxic and trypsin treatment of ASCs increased the expression and secretion of VEGF in these in an additive manner. These findings indicate the combinational treatment of hypoxia and trypsin as a potentiating preconditioning of ASCs to be used for clinical application. In the area of endothelial migration the influence of ASCs has yet to be determined.