Activation of NAD(P)H oxidase by lipid hydroperoxides: mechanism of oxidant-mediated smooth muscle cytotoxicity

Oxidized lipids, such as 13-hydroperoxyoctadecadienoic acid (13-HPODE), have been implicated in the pathogenesis of atherosclerosis. 13-HPODE, a constituent of oxidized low-density lipoproteins, can induce cytotoxicity of vascular smooth muscle cells (SMC), which may facilitate plaque destabilization and/or rupture. 13-HPODE-induced cytotoxicity has been linked to oxidative stress, although the mechanisms by which this occurs are unknown. In the present study, we show that 13-HPODE and 9-HPODE (10–30 μM) increased superoxide (O2•−) production and induced cytotoxicity in SMC. The 13-HPODE-induced increase in O2•− was blocked by transfecting the cells with antisense oligonucleotides against p22phox, suggesting that the O2•− was produced by NAD(P)H oxidase. Similar concentrations of the corresponding HPODE reduction products, 13-hydroxyoctadecadienoic acid (13-HODE) and 9-HODE, neither increased O2•− production nor induced cytotoxicity, while 4-hydroxy nonenal (4-HNE), an unsaturated aldehyde lipid peroxidation product, induced cytotoxicity without increasing O2•− production. Treatment with superoxide dismutase or Tiron to scavenge O2•−, or transfection with p22phox antisense oligonucleotides to inhibit O2•− production, attenuated 13-HPODE-induced cytotoxicity, but not that induced by 4-HNE. These findings suggest that activation of NAD(P)H oxidase, and production of O2•−, play an important role in lipid hydroperoxide-induced smooth muscle cytotoxicity.