Figure 8

HIMF-induced NF-κB activation and upregulation of VEGF is PI-3K/Akt pathway dependent. MLE-12 cells were pretreated with signal transduction inhibitors or co-transfected with luciferase constructs and PI-3K dominant-negative mutant, then stimulated with HIMF (40 nmol/L) for various periods as indicated. (8A) HIMF strongly induced the phosphorylation of Akt at Ser473 and Thr308. The Akt phosphorylation started at 30 minutes and sustained for 360 min. HIMF also induced phosphorylation of ERK1/2 and p38 MAPK, but not JNK MAPK in MLE-12 cells. (8B) The PI-3K inhibitor LY294002 (10 μmol/L), but not SB203580 (5 μmol/L), PD098059 (5 μmol/L) or U0126 (5 μmol/L), abolished HIMF-induced Akt phosphorylation and upregulation of VEGF in MLE-12 cells. (8C) Transfection of Δp85, into MLE-12 cells abolished HIMF-induced phosphorylation of IKK and IκBα, NF-κB activation and production of VEGF. The symbol (*) indicates a significant increase from MLE-12 controls without HIMF treatment (P < 0.05). The symbol (#) indicates a significant decrease from MLE-12 cells treated with HIMF only (P < 0.05). Triplicate experiments were performed with essentially identical results.