Figure 2.

Norcantharidin (NCTD) inhibits vascular endothelial growth factor (VEGF)‐induced migration, invasion, and tubular structure formation of endothelial cells. (a) Norcantharidin inhibits human umbilical vein endothelial cell (HUVEC) migration. Human umbilical vein endothelial cells were allowed to grow into full confluence in six‐well plates, and inactivated with 10 μg/mL mitomycin C for 2 h. Cells were wounded with pipette and treated with or without 4 ng/mL VEGF and different concentration of NCTD in endothelial cell medium supplemented with 0.5% fetal bovine serum (FBS). After incubation, the migrated cells were quantified by manual counting. (b) Norcantharidin inhibits the invasion of HUVEC. Human umbilical vein endothelial cells were seeded in the upper chamber of transwell and treated with different concentrations of NCTD. The bottom chamber was filled with ECGM supplemented with VEGF. After about 8–10 h, the invasive HUVECs passed through the membrane and were quantified by counting the cells that migrated onto the membrane. (c) Norcantharidin inhibits VEGF‐induced tube formation of HUVECs. Human umbilical vein endothelial cells were placed in the 24‐well plates coated with Matrigel at the density of 4 × 10⁴ cells/well. After 4–6 h, cells were fixed and tubular structure was quantified by manual counting of high power fields (200×). Column, mean from three different experiments with duplicates; bar, SE (*P < 0.05, **P < 0.01 versus VEGF alone).