Vascular endothelial growth factor A (VEGF-A), a glycosylated mitogen, is known to be a vascular permeability factor and an endothelial cell growth factor secreted by the smooth muscle and endothelial cells. It has been reported that VEGF-A induces vascular permeability and growth, promotes monocyte/macrophage migration, and inhibits cell apoptosis and so on. To test the effect of VEGF-A on cell proliferation of ECV304 endothelium cell line, cells were seeded into triplicate wells of 96-well plates at a density of 2,000 cells/well and allowed to attach overnight, then the medium was replaced with serum-free standard DMEM prior to the addition of various concentrations of VEGFA. After incubated for 72h, cells were observed by inverted microscope and cell proliferation was measured by Cell Counting Kit-8 (CCK-8). Briefly, 10µL of CCK-8 solution was added to each well of the plate, then measure the absorbance at 450nm using a microplate reader after incubating the plate for 1-4 hours at 37oC .
Cell proliferation of ECV304 cells after incubation with VEGFA for 72h observed by inverted microscope was shown in Figure 1.

The dose-effect curve of VEGFA was shown in Figure 2. It was obvious that VEGFA significantly promoted cell proliferation of ECV304 cells. The ED50 for this effect is typically 5.58 to 9.98 ng/mL.