Fig. 2.

Effect of H₂O₂ on AMPK phosphorylation in HeLa cells. A and B: similar to the findings with H4IIEC3 cells, the levels of AMPK phosphorylation in HeLa cells were increased after H₂O₂ (1 mM) treatment to 1.2-fold over control at 10 min after treatment with 1 mM H₂O₂ (P < 0.05). C and D: AMPK phosphorylation in HeLa cells was affected by Ca²⁺/calmodulin-dependent protein kinase kinase (CaMKK) inhibitor, STO-609 at a concentration of 1 μg/ml. At baseline, STO-609 significantly inhibits AMPK phosphorylation by 75%, and it also significantly prevents AMPK activation in response to H₂O₂. To further examine the major pathway for the activation of AMPK by H₂O₂, similar experiments were performed in H4IIEC3 cells in the presence of STO-609 (E). Contrary to the experiments in the HeLa cells, STO-609 did not significantly inhibit AMPK phosphorylation at baseline. The activation of AMPK by H₂O₂ persists despite the presence of STO-609. Means ± SE (the blot is a representative of 3 blots from 3 individual experiments) are shown. *Significant difference vs. control; P < 0.05, by one-way ANOVA; $significant difference compared with cells that were not treated with STO-609.