Fig. 3

Activation of PI3K/AKT signaling pathway is involved in EGF-suppressed KISS1 expression and secretion. a–c, HTR-8/SVneo cells were treated with 50 ng/mL EGF for 10, 30, and 60 min (a). HTR-8/SVneo cells were pretreated with vehicle control (DMSO) or 5 µM AG1478 for 1 h, and then treated with 50 ng/mL EGF for 10 min (b). HTR-8/SVneo cells were pretreated with vehicle control (DMSO), 5 µM LY294002, or 5 µM U0126 for 1 h, and then treated with 50 ng/mL EGF for 10 min (c). The levels of phosphorylated and total forms of ERK1/2 and AKT were determined by western blot. d, e HTR-8/SVneo cells were pretreated with vehicle control (DMSO), 5 µM LY294002, or 5 µM U0126 for 1 h, and then treated with 50 ng/mL EGF for 24 h. The KISS1 mRNA levels (d) and protein levels (e) were examined by RT-qPCR and western blot, respectively. f HTR-8/SVneo cells were pretreated with vehicle control (DMSO) or 5 µM LY294002 for 1 h, and then treated with 50 ng/mL EGF every 24 h for 48 h. The protein levels of KISS1 in culture media were examined by ELISA. Numbers under the western blots represent the densitometry quantifications. The results are expressed as the mean ± SEM of at least three independent experiments. Values without a common letter are significantly different (p < 0.05)