With no further endothelial cells remaining to be stimulated to form additional capillary-like structures. In case of a higher cell seeding number, there are more capillary-forming endothelial cells in a cobblestone pattern which can be stimulated via VEGF-A/PDGFBB to reorganize into capillary-like structures. In this case, the difference is measureable. Regarding the 3rd donor, the positive effect by adding VEGF to PDGF-BB was only statistically different at concentrations of 40 ng/ml VEGF and 10 ng/ml PDGF-BB. In the course of this study, it was observed that the presence of VEGFR-2 receptors on HUVECs correlates with the number of capillary-like structures obtained in the endothelial co-culture assay. A lower expression of VEGFR-2 on HUVEC cells compared to higher levels of expression in donor 1 and donor 2 correlated to a decrease in length, number of junctions and area of the capillary-like structures, and it is clearly evident that the cells with low concentrations of VEGFR2 receptors are less responsive to stimulation by VEGF-A. With regard to VEGFR-2 expression and its activity in combination with different supportive cell numbers, the increases in length and number of junctions correlated with increasing numbers of supportive cells for all donors. With regards to the 3rd donor, however, the increase in both length and number of junctions in the presence of increasing numbers of supportive cells was lower compared to the other two donors. This limitation may be explained by the lower concentrations of VEGFR-2, and by the fact that more supportive cells lead to improved ECM development resulting in enhanced endothelial cell attachment to the supportive layer. VEGFR-2 is a critical VEGF receptor during development and adult angiogenesis. It is well accepted that VEGFR-2 is the most important mediator for mitogenic, angiogenic and permeabilityenhancing GDC-0879 effects of VEGF on endothelial cells. In the literature, it has been shown that VEGFR-2 knockdown completely inhibits network formation in a 2D co-culture system. The inhibition of VEGFR-2 led to similar results in a 3D coculture model. With respect to the testing of an anti-angiogenic drug in the presented co-culture system, the negative effect of Bevacizumab on the formation of capillary-like structures could be demonstrated. The effect was investigated for donor 3 with the addition of both 40 ng/ml VEGF and 10 ng/ml PDGF-BB. In this case, the amount of capillary-like structures decreased. Bevacizumab is an FDA-approved VEGF antibody which binds VEGF-A and proteolytic fragments, thereby inhibiting the development of blood vessels. In co-culture as well as in in vivo models, this effect has also been demonstrated by other groups. In other co-culture experiments, suramin or curcumin have been successfully used as anti-angiogenic drugs. The presently introduced co-culture system of ovine carotidderived supportive cells and HUVECs demonstrates that the effect of pro- and anti-angiogenic effects on capillary network development can be measured and quantified in an easy and reliable manner. However, donor variability as a possible limitation in the endothelial co-culture assay needs to be taken into account.