Flow cytometry analysis of plasma membranes with annexin V-FITC / PI double staining. A-D: representative photos of different groups. A: control. Cells were incubated in serum-free medium for 24 h following overnight starvation. The numbers in upper left, upper right, lower left and lower right quadrants represent the percentage of necrotic cells (annexin V positive, PI positive), advanced apoptotic cells (annexin V negative, PI positive), viable cells (annexin V negative, PI negative) and early apoptotic cells (annexin V positive, PI negative) respectively; B: VEGF. Cells were incubated in serum-free medium containing 100 ng/ml VEGF for 24 h following starvation; C: VEGF+150 μM H-RN. Cells were pre-incubated with 150 μM H-RN for 30 min and VEGF (100 ng/ml) was added and incubated for 24 h; D: VEGF+1.5 mM H-RN. Cells were pre-incubated in 1.5 mM H-RN for 30 min and VEGF (100 ng/ml) was added for 24 h; E: Quantitative analysis of apoptotic cells under different experimental conditions. The values represent the mean percentages of apoptotic cells in different groups. (*p<0.05, each condition versus VEGF control).