Figure 4.
Hes-1 suppresses VEGF-induced angiogenesis via downregulation of OPN expression in vitro and in vivo. (A) HUVECs (1 × 105) treated with siRNA/ pcDNA3.1of Hes-1 or OPN and then incubated with or without 10 ng/mL of VEGF for 24 h were seeded on matrigel-coated plates. After 6 h of incubation, endothelial cell tubular structure formation was photographed and analyzed. The number of tubular-like structures was counted and indicated as the number of branches per field. Averages of tubules were calculated from four fields in each sample. Pictures were taken at 20x magnification. *p < 0.05. Bar: 400 μm. (B) The CAM assay was done to assess angiogenesis in vivo, HUVECs (1 × 105) treated with siRNA/ pcDNA3.1 of Hes-1 or OPN and then incubated with or without 10 ng/mL of VEGF for 24 h were pipeted directly into the center of the silicone ring resting on the CAM. Sealed the window and incubated the egg for 48 h to observe the result of neovascularization. The CAM surface was photographed at the same position with a digital camera, and the area of newly formed vessels was calculated. The data are presented as mean ± SD of three independent experiments. *p < 0.05.