Figure 4

Leptin activate PI3-kinase/AKT signaling pathway. (A) MDAH2774 cells were serum starved for 24 hours in serum free medium, followed by stimulation with recombinant leptin (100 ng/ml) for various time periods as indicated. After cell lysis, 20 μg proteins were separated by SDS-PAGE, transferred to immobilon membrane, and immunoblotted with antibodies against p-AKT-Ser 473, FOXO1, and beta actin. (B) Inhibition of PI3-kinase/AKT pathway prevented leptin-induced activation of AKT pathway in EOC cells. SKOV3 cells were serum starved in the presence and absence of LY294002 as indicated for 48 hours and subsequently stimulated with 100 ng/ml of recombinant leptin for 3 hours, Cells were lysed and proteins were separated by SDS-PAGE, transferred to immobilon membrane, and immunoblotted with antibodies against p-AKT-Ser 473, FOXO1, and beta actin. (C) LY294002-inhibitor abrogate leptin-mediated cell proliferation and prevented leptin-induced anti-apoptotic effects in EOC cells. MADH2774 and SKOV3 cells were serum starved in the presence and absence of LY294002 as indicated and subsequently stimulated with 100 ng/ml of recombinant leptin for 48 hours. Cell proliferation was measured by MTT assays and (D) apoptosis was measured by Anenexin/PI staining. The graph displays the mean +/- SD of six independent experiments (* p < 0.05).