Figure 5. VEGF-induced annexin A2 expression was PKC-dependent. A: Serum-starved RF/6A cells were pretreated with 1 µM calphostin C, 3 µM GF109203X, 3 µM LY333531, or 5 µM rottlerin for 30 min and then treated with 25 ng/ml VEGF for 6 h. Annexin A2 protein was detected by western blotting. B: After overnight serum starvation, RF/6A cells were pretreated with 1 µM calphostin C or 2 µM LY333531 for 30 min and then were treated with 25 ng/ml VEGF for 2 h. Total RNA was isolated, and quantitative real-time PCR was used to detect the expression of annexin A2 mRNA. Double asterisk (**) indicates p<0.01 compared with control samples. C: Serum-starved RF/6A cells were pretreated with 10 µM U0126 or 50 µM PD98059 for 30 min and then were treated with 25 ng/ml VEGF for 6 h. Total cell lysates were subjected to western blot analysis for annexin A2. The same membrane was blotted with anti–GAPDH antibody for normalization.