Homocyst(e)ine decreases bioavailable nitric oxide by a mechanism involving glutathione peroxidase.
Hyperhomocyst(e)inemia is believed to injure endothelial cells in vivo through a number of mechanisms, including the generation of hydrogen peroxide (H2O2). Earlier in vitro studies demonstrated that homocyst(e)ine (Hcy) decreases the biological activity of endothelium-derived relaxing factor and that this decrease can be reversed by preventing the generation of hydrogen peroxide. Here we show that Hcy treatment of bovine aortic endothelial cells leads to a dose-dependent decrease in NOx (p = 0.001 by one-way analysis of variance) independent of endothelial nitric-oxide synthase activity or protein levels and nos3 transcription, suggesting that Hcy affects the bioavailability of NO, not its production. We hypothesized that, in addition to increasing the generation of H2O2, Hcy decreases the cell's ability to detoxify H2O2 by impairing intracellular antioxidant enzymes, specifically the intracellular isoform of glutathione peroxidase (GPx). To test this hypothesis, confluent bovine aortic endothelial cells were treated with a range of concentrations of Hcy, and intracellular GPx activity was determined. Compared with control cells, cells treated with Hcy showed a significant reduction in GPx activity (up to 81% at 250 microM Hcy). In parallel with the decrease in GPx activity, steady-state GPx mRNA levels were also significantly decreased compared with control levels after exposure to Hcy, which appeared not to be a consequence of message destabilization. These data suggest a novel mechanism by which Hcy, in addition to increasing the generation of hydrogen peroxide, may selectively impair the endothelial cell's ability to detoxify H2O2, thus rendering NO more susceptible to oxidative inactivation.
Upchurch
G R
GR
Whitaker Cardiovascular Institute and the Evans Department of Medicine, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
Welch
G N
GN
Fabian
A J
AJ
Freedman
J E
JE
Johnson
J L
JL
Keaney
J F
JF
Loscalzo
J
J
D000818:Animals; D001682:Biological Availability; D002417:Cattle; D004730:Endothelium, Vascular / Q000378:metabolism; D004790:Enzyme Induction; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism*; D006710:Homocysteine / Q000494:pharmacology*; D006861:Hydrogen Peroxide / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D014158:Transcription, Genetic
D012333:RNA, Messenger; D006710:Homocysteine; D009569:Nitric Oxide; D006861:Hydrogen Peroxide; D007770:L-Lactate Dehydrogenase; D005979:Glutathione Peroxidase; D019001:Nitric Oxide Synthase; D005978:Glutathione