17 beta-estradiol inhibition of NADPH oxidase expression in human endothelial cells

We investigated the hypothesis that the antiatherosclerotic effect of 17 beta -estradiol (E-2) is due to a shift in the nitric oxide (NO)/superoxide (O-2(-)) balance in the vessel wall, thereby increasing the bioavailability of NO. In human umbilical vein cultured endothelial cells, E-2 (1-100 nmol/l), but not 17 alpha -estradiol, caused a time- and concentration-dependent decrease in expression of the NADPH oxidase subunit gp91phox (up to 60% inhibition at both the mRNA and protein level). This effect was prevented by coincubation with the estrogen receptor antagonists tamoxifen and ICI 182,780 (1 mu mol/l each). Within the same concentration range, E-2 also up-regulated endothelial nitric oxide synthase expression (similar to twofold). Moreover, preincubation of the cells with E-2 or a gp91phox antisense oligonucleotide significantly decreased their capacity to generate O-2(-) on phorbol ester stimulation (i.e., assembly of the active NADPH oxidase complex). Blockade of NO synthase activity, on the other hand, had no effect on phorbol ester-stimulated O-2(-) formation. In addition, E-2 (100 nmol/l) inhibited the increase in adhesion molecule and chemokine expression in cells exposed to cyclic strain. Cyclic strain enhanced endothelial O-2(-) formation, thereby offsetting the inhibitory effect of NO on the expression of these gene products. E-2 thus seems to act as an antioxidant at the genomic level which by improving the NO/O-2(-) balance normalizes expression of proatherosclerotic gene products in endothelial cells.