Generation of oxidant stress in cultured endothelial cells by methylene blue: protective effects of glucose and ascorbic acid

被引:38
作者
May, JM
Qu, ZC
Whitesell, RR
机构
[1] Vanderbilt Univ, Sch Med, Dept Med, Nashville, TN 37232 USA
[2] Vanderbilt Univ, Sch Med, Dept Mol Physiol & Biophys, Nashville, TN 37232 USA
关键词
methylene blue; ascorbic acid; ferricyanide; dihydrofluorescein; oxidant stress; EA.hy926 endothelial cells;
D O I
10.1016/S0006-2952(03)00408-8
中图分类号
R9 [药学];
学科分类号
1007 ;
摘要
The thiazine dye methylene blue has long been used to stimulate cellular redox metabolism. To determine the extent to which it also generates oxidant stress in cells, its effects in cultured human-derived endothelial cells were studied. As expected, low concentrations of the dye (2-20 muM) activated the pentose phosphate pathway and oxidized both NADPH and NADH. Methylene blue enhanced extracellular ferricyanide reduction, indicating that the reduced form of the dye was present outside the cells. This reduction was greater when ferricyanide was added just before rather than 15 min after methylene blue, confirming that the dye is at least initially reduced at the cell surface. In the absence of glucose, methylene blue at concentrations above 5 muM increased intracellular oxidant stress, as manifest by oxidation of dihydrofluorescein and cellular GSH. Inclusion of glucose protected against these effects. In cells that had been loaded with ascorbate, the dye caused progressive oxidation of ascorbate, even in the presence of D-glucose. Loading cells with ascorbate also partially prevented oxidation of dihydrofluorescein by methylene blue. These results suggest that concentrations of the dye above 5 muM generated intracellular reactive oxygen species that were scavenged by ascorbate and GSH. Further, although D-glucose enhanced reduction of methylene blue, it ameliorated the oxidant stress generated by the dye. (C) 2003 Elsevier Inc. All rights reserved.
引用
收藏
页码:777 / 784
页数:8
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