Endothelial nitric oxide synthase negatively regulates hydrogen peroxide-stimulated AMP-activated protein kinase in endothelial cells

Jin, B.Y.; Sartoretto, J.L.; Gladyshev, V.N.; Michel, T.

Авторы: Jin B.Y., Sartoretto J.L., Gladyshev V.N., Michel T.
Журнал: Proceedings of the National Academy of Sciences of the United States of America
Том: 106
Номер: 41
Год издания: 2009
Издательство: National Academy of Sciences
Местоположение издательства: United States
Первая страница: 17343
Последняя страница: 17348
DOI: 10.1073/pnas.0907409106
Аннотация: Hydrogen peroxide and other reactive oxygen species are intimately involved in endothelial cell signaling. In many cell types, the AMP-activated protein kinase (AMPK) has been implicated in the control of metabolic responses, but the role of endothelial cell redox signaling in the modulation of AMPK remains to be completely defined. We used RNA interference and pharmacological methods to establish that H2O2 is a critical activator of AMPK in cultured bovine aortic endothelial cells (BAECs). H2O2 treatment of BAECs rapidly and significantly increases the phosphorylation of AMPK. The EC50 for H2O2-promoted phosphorylation of AMPK is 65 ± 15 μM, within the physiological range of cellular H2O2 concentrations. The Ca2+/calmodulin-dependent protein kinase kinase-β (CaMKKβ) inhibitor STO-609 abolishes H2O2-dependent AMPK activation, whereas eNOS inhibitors enhance AMPK activation. Similarly, siRNA-mediated knockdown of CaMKKβ abrogates AMPK activation, whereas siRNA-mediated knockdown of eNOS leads to a striking increase in AMPK phosphorylation. Cellular imaging studies using the H2O2 biosensor HyPer show that siRNA-mediated eNOS knockdown leads to a marked increase in intracellular H2O2 generation, which is blocked by PEG-catalase. eNOS−/− mice show a marked increase in AMPK phosphorylation in liver and lung compared to wild-type mice. Lung endothelial cells from eNOS−/− mice also show a significant increase in AMPK phosphorylation. Taken together, these results establish that CaMKKβ is critically involved in mediating the phosphorylation of AMPK promoted by H2O2 in endothelial cells, and document that eNOS is an important negative regulator of AMPK phosphorylation and intracellular H2O2 generation in endothelial cells.
Добавил в систему: Гладышев Вадим Николаевич

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Endothelial nitric oxide synthase negatively regulates hydrogen peroxide-stimulated AMP-activated protein kinase in endothelial cellsстатья