Fig. 1.

H₂O₂ stimulates Akt phosphorylation and activity in human cardiac microvascular endothelial cells (HCMVEC). HCMVEC were treated with increasing concentrations of H₂O₂ for 10 min, and phosphorylation status of Akt at Ser-473 (A) and Thr-308 (B) were determined by Western blot analysis. C and D: to further determine the time course of Akt activation, HCMVEC were stimulated with 500 μM H₂O₂ for 1.5, 10, and 30 min and Akt Ser-473 and Thr-308 phosphorylation was determined by Western blotting. In some cells, H₂O₂-containing medium was replaced with a fresh medium 10 min after stimulation. E: HCMVEC were treated with 500 μM H₂O₂ and Akt-related phosphorylation of GSK3 was measured. Blots are representative of 3 independent experiments. Densitometric analyses represent means ± SE of 3 independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001. au, Arbitrary units; p, phospho.