Effect of AACOCF3 or PD98059 on FSH- or EGF stimulated follicular DNA synthesis, PLA2G4 and PKC activity. (A) Follicles were cultured with 60 μM AACOCF3 for 1h before the administration of FSH or EGF, and [3H]thymidine. After 6h of culture, DNA synthesis was determined. (B) Follicles were cultured with 60 μM AACOCF3 or 10 μM PD98059 for 1h before the administration of FSH or EGF. After 6h culture, PLA2G4 or PKC activity was measured. AACOCF3 was used as a positive control for PLA2G4 inhibition. (C) Cell-free determination of the specificity of kinase inhibitors at the tested dose level. Follicles were cultured without or with 25 ng/ml FSH, homogenate was mixed with an appropriate concentration of respective inhibitors for 30 min on ice and used in enzyme reaction. Each bar represents a mean ± SEM of three separate values. P > 0.05: bars with a same letter; P < 0.05: bars with a different letter.