Cellular glutathione peroxidase deficiency and endothelial dysfunction

Abstract

Cellular glutathione peroxidase (GPx-1) is the most abundant intracellular isoform of the GPx antioxidant enzyme family. In this study, we hypothesized that GPx-1 deficiency directly induces an increase in vascular oxidant stress, with resulting endothelial dysfunction. We studied vascular function in a murine model of homozygous deficiency of GPx-1 (GPx-1 Ϫ/Ϫ ). Mesenteric arterioles of GPx-1 Ϫ/Ϫ mice demonstrated paradoxical vasoconstriction to ␤-methacholine and bradykinin, whereas wildtype (WT) mice showed dose-dependent vasodilation in response to both agonists. One week of treatment of GPx-1 Ϫ/Ϫ mice with L-2-oxothiazolidine-4-carboxylic acid (OTC), which increases intracellular thiol pools, resulted in restoration of normal vascular reactivity in the mesenteric bed of GPx-1 Ϫ/Ϫ mice. We observed an increase of the isoprostane iPF 2␣-III, a marker of oxidant stress, in the plasma and aortas of GPx-1 Ϫ/Ϫ mice compared with WT mice, which returned toward normal after OTC treatment. Aortic sections from GPx-1 Ϫ/Ϫ mice showed increased binding of an anti-3-nitrotyrosine antibody in the absence of frank vascular lesions. These findings demonstrate that homozygous deficiency of GPx-1 leads to impaired endothelium-dependent vasodilator function presumably due to a decrease in bioavailable nitric oxide and to increased vascular oxidant stress. These vascular abnormalities can be attenuated by increasing bioavailable intracellular thiol pools. nitric oxide; peroxynitrite; oxidant stress NITRIC OXIDE (NO) synthesized by the endothelium contributes to vascular tone (23), preserves endothelial integrity (13), inhibits smooth muscle cell migration and proliferation (16), and acts as an antioxidant (30). An increase in reactive oxygen species (ROS), leading to increased oxidant stress in the vasculature, promotes endothelial dysfunction (28) associated with NO insufficiency (15).The enzyme glutathione peroxidase (GPx) is a selenocysteine-containing protein that serves an important role in the cellular defense against oxidant stress (24) by utilizing reduced glutathione (GSH) to reduce hydrogen peroxide (H 2 O 2 ) and lipid peroxides to their corresponding alcohols (37). GPx exists in several isoforms, and the most abundant intracellular isoform is cellular GPx, or GPx-1. We (7, 36, 39) have previously shown that elevated homocysteine concentrations suppress GPx-1 expression in endothelial cells in vitro and in mildly hyperhomocysteinemic mice in vivo and suggested that this effect may account, in part, for the vascular oxidant stress of hyperhomocysteinemic states. Hydrogen peroxide forms the toxic oxygen species hydroxyl radical (⅐OH), which is highly reactive and causes lipid peroxidation, and hydroxide anion (OHϪ), which promotes alkaline tissue damage, a process that is offset in part by catalase and GPx-1-dependent reduction to H 2 O. Elevated levels of lipid peroxides are accompanied by an increase in peroxyl radicals, which can inactivate NO through the formation of lipid peroxynitrites (19,30), although the prec...

Deficiency of Glutathione Peroxidase-1 Sensitizes Hyperhomocysteinemic Mice to Endothelial Dysfunction

“…GPx-1 may also provide protection from the toxic effects of peroxynitrite through its peroxynitrite reductase activity. 48 Our results confirm the findings of Forgione et al 26 that deficiency of GPx-1 leads to impairment of endotheliumdependent vasodilator function in mice with normal plasma levels of tHcy. Forgione et al observed endothelial dysfunction with paradoxical vasoconstriction to endotheliumdependent agonists in mesenteric arterioles of Gpx1 Ϫ/Ϫ mice fed standard chow.…”

“…To minimize the potential influence of differences in genetic background, GPx-1-deficient mice were crossbred to C57BL6 mice (The Jackson Laboratory, Bar Harbor, Me) for at least 7 generations and then interbred to generate littermates that were wild type (Gpx1 ϩ/ϩ ), heterozygous (Gpx1 ϩ/Ϫ ), or homozygous (Gpx1 Ϫ/Ϫ ) for the mutated Gpx1 allele. Genotyping for the wild-type and mutated Gpx1 alleles was performed by polymerase chain reaction as described by Forgione et al 26 At the time of weaning, mice were placed on either a control diet (LM-485 chow, Harlan Teklad) or a high methionine diet (LM-485 chow and drinking water supplemented with 0.5% L-methionine). After 17 weeks of the experimental diet, mice were euthanized with sodium pentobarbital (75 mg IP).…”

Influence of Hyperhomocysteinemia on the Cellular Redox State – Impact on Homocysteine-Induced Endothelial Dysfunction

“…Thus a deficiency of GPx-1 may lead to a decrease in bioavailable nitric oxide via at least two mechanisms, an increase in reactive oxygen species and an increase in lipid hydroperoxides. In support of this hypothesis we have shown previously that mice deficient in GPx-1 have endothelial dysfunction owing to decreased bioavailable endothelium-derived nitric oxide; increased oxidative stress and increased lipid hydroperoxide generation as measured by increased tissue isoprostane F 2α -III and hepatic phospholipid hydroperoxide levels; and increased nitrosative stress, as indicated by increased immunostaining for 3-nitrotyrosine in aortic tissue of GPx-1 knockout mice compared to wild-type mice (36).…”

Plasma antioxidants in subjects before hematopoietic stem cell transplantation

“…41 In animals with targeted gene disruption of the GPX-1 gene, endothelial function was impaired. 42 Therefore, it is conceivable that subjects with a reduction in the plasma concentrations of circulating GPX-3 undergoing HSCT may be at increased risk of organ damage beyond day 100 following transplant. In the small cohort of subjects in this study we were unable to demonstrate an increased incidence of organ damage in those with concentrations of GPX-3 below 100 mg/ml (data not shown).…”