title: Induction of endothelial cell apoptosis by lipid hydroperoxide-derived bifunctional electrophiles.
abstract: Endothelial dysfunction is considered to be the earliest event in atherogenesis. Oxidative stress, inflammation, and apoptosis play critical roles in its progression and onset. Lipid peroxidation, which occurs during oxidative stress, results in the formation of lipid hydroperoxide-derived bifunctional electrophiles such as 4-hydroxy-2(E)-nonenal that induce apoptosis. In this study, recently identified lipid hydroperoxide-derived bifunctional electrophiles 4-oxo-2(E)-nonenal (ONE; 5-30 microm) and 4,5-epoxy-2(E)-decenal (EDE; 10-20 microM) were shown to cause a dose- and time-dependent apoptosis in EA.hy 926 endothelial cells. This was manifest by morphological changes, caspase-3 activation, and poly(ADP-ribose) polymerase cleavage. Bifunctional electrophiles caused cytochrome c release from mitochondria into the cytosol, implicating a mitochondrial pathway of apoptosis in the endothelial cells. The novel carboxylate-containing lipid hydroperoxide-derived bifunctional electrophile 9,12-dioxo-10(E)-dodecenoic acid was inactive because it could not translocate across the plasma membrane. However, its less polar methyl ester derivative (2-10 microM) was the most potent inducer of apoptosis of any bifunctional electrophile that has been tested. An acute decrease in intracellular glutathione (GSH) preceded the onset of apoptosis in bifunctional electrophile-treated cells. The ability of ONE and EDE to deplete GSH was directly correlated with their predicted reactivity toward nucleophilic amino acids. Liquid chromatography/mass spectrometry methodology was developed in order to examine the intracellular and extracellular concentrations of bifunctional electrophile-derived GSH adducts. Relative intracellular/extracellular ratios of the GSH adducts were identical with the rank order of potency for inducing caspase 3 activation. This suggests that there may be a role for the bifunctional electrophile-derived GSH adducts in the apoptotic response. N-Acetylcysteine rescued bifunctional electrophile-treated cells from apoptosis, whereas the GSH biosynthesis inhibitor d,l-buthionine-(R,S)-sulfoximine sensitized the cells to apoptosis. These data suggest that lipid hydroperoxide-derived bifunctional electrophiles may play an important role in cardiovascular pathology through their ability to induce endothelial cell apoptosis.
authors.affiliation: Center for Cancer Pharmacology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6160, USA.
authors.forename: Wenying
authors.initials: W
authors.lastname: Jian
authors.forename: Jasbir S
authors.initials: JS
authors.lastname: Arora
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Oe
authors.forename: Vladimir V
authors.initials: VV
authors.lastname: Shuvaev
authors.forename: Ian A
authors.initials: IA
authors.lastname: Blair
mesh_terms: D000447:Aldehydes / Q000737:chemistry / Q000494:pharmacology*; D017209:Apoptosis; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D004789:Enzyme Activation; D004852:Epoxy Compounds / Q000737:chemistry / Q000494:pharmacology*; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D015227:Lipid Peroxidation; D008054:Lipid Peroxides / Q000737:chemistry* / Q000378:metabolism; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism
chemical_list: C096140:4,5-epoxy-2-decenal; C403894:4-oxo-2-nonenal; C418762:9,12-dioxo-10-dodecenoic acid; D000447:Aldehydes; D004852:Epoxy Compounds; D005229:Fatty Acids, Monounsaturated; D008054:Lipid Peroxides; D045304:Cytochromes c; D011065:Poly(ADP-ribose) Polymerases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D005978:Glutathione