Cobalt-induced oxidant stress in cultured endothelial cells: Prevention by ascorbate in relation to HIF-1α

Endothelial cells respond to hypoxia by decreased degradation of hypoxia-inducible factor 1 alpha (HIF-1 alpha), accumulation of which leads to increased transcription of numerous proteins involved in cell growth and survival. Ascorbic acid prevents HIF-1 alpha stabilization in many cell types, but the physiologic relevance of such effects is uncertain. Given their relevance for angiogenesis, endothelial cells in culture were used to evaluate the effects of ascorbate on HIF-1 alpha expression induced by hypoxia and the hypoxia mimic cobalt. Although EA.hy926 cells in culture under oxygenated conditions did not contain ascorbate, HIF-1 alpha expression was very low, showing that the vitamin is not necessary to suppress HIF-1 alpha. On the other hand, hypoxiaor cobalt-induced HIF-1 alpha expression/stabilization was almost completely suppressed by what are likely physiologic intracellular ascorbate concentrations. Increased HIF-1 alpha expression was not associated with significant changes in expression of the SVCT2, the major transporter for ascorbate in these cells. Cobalt at concentrations sufficient to stabilize HIF-1 alpha both oxidized intracellular ascorbate and induced an oxidant stress in the cells that was prevented by ascorbate. Whereas the interaction of ascorbate and cobalt is complex, the presence of physiologic low millimolar concentrations of ascorbate in endothelial cells effectively decreases HIF-1 alpha expression and protects against cobalt-induced oxidant stress. (C) 2009 International Union of Biochemistry and Molecular Biology, Inc. Volume 35, Number 3, May/June 2009, Pages 306-313 E-mail: james.may@vanderbilt.edu