Metformin inhibits IGF-I-stimulated IGF-IR and Akt activation in normal glucose. (A-C) BxPC-3 and (D-F) MIAPaCa-2 cells were treated in normal (5 mM) or high (25 mM) glucose-containing SFM, with or without 10 mM metformin for 24 h. Cells were spiked with IGF-I (100 ng/ml) as indicated for the final 15 min prior to lysis. pIGF-IRβ/pIRβ, IGF-IRβ, pAkt, and Akt were assessed by Western immunoblotting. GAPDH is shown as loading control. One representative blot is shown. Graphs display densitometry measurements of relative levels at 5 mM (black bars) or 25 mM (striped bars) glucose conditions and represent means ± SE of three independent experiments. P-values are based on two-way ANOVA with Bonferroni post hoc test. N.S. not significant.