linoleic-acid-hydroperoxide and Arteriosclerosis

We have previously prepared the polyclonal antibody to the 13-hydroperoxyoctadecadienoic acid-modified protein (13Ab) (Kato et al. 1997. J. Lipid Res. 38: 1334-1346), however, the epitopes have not yet been structurally identified. In this study, we identified a novel amide-type adduct as one of the major epitopes of 13Ab and characterized the endogenous formation. Upon incubation of the lysine derivative with peroxidized linoleic acid, the formation of N epsilon -(azelayl)lysine (AZL) was confirmed using liquid chromatography-mass spectrometry. The chemically synthesized azelayl protein was significantly recognized by 13Ab. The peroxidation products of different polyunsaturated fatty acids also generated several analogous carboxyalkylamide-type adducts to AZL by the reaction with the lysine derivative, whereas 13Ab specifically recognized AZL, suggesting that the AZL moiety may be one of the major epitopes of 13Ab. The immunoreactive materials of 13Ab were immunohistochemically detected in atherosclerotic lesions from hypercholesterolemic rabbits. More strikingly, the immunoreactivity was significantly enhanced when the sections were treated with alkali or phospholipase A2 for hydrolyzing the ester bonds prior to the staining. These results suggest that the lipid hydroperoxide-derived carboxylic adducts, such as AZL, and their esters linked with phospholipids may be generated in vivo and involved in the pathogenesis of atherosclerosis associated with oxidative stress.

Topics: Amides; Animals; Antibodies; Aorta; Arteriosclerosis; Cross Reactions; Dicarboxylic Acids; Epitopes; Fatty Acids, Unsaturated; Hypercholesterolemia; Immunohistochemistry; Linoleic Acids; Lipid Peroxidation; Lipid Peroxides; Lysine; Phospholipases A; Phospholipases A2; Phospholipids; Proteins; Rabbits; Time Factors

Human serum paraoxonase (PON1) exists in two polymorphic forms: one that differs in the amino acid at position 192 (glutamine and arginine, Q and R, respectively) and the second one that differs in the amino acid at position 55 (methionine and leucine, M and L, respectively). PON1 protects LDL from oxidation, and during LDL oxidation, PON1 is inactivated.. The present study compared PON1 isoforms Q and R for their effect on lipid peroxide content in human coronary and carotid lesions. After 24 hours of incubation with PON1Q or PON1R (10 arylesterase units/mL), lipid peroxides content in both coronary and carotid lesion homogenates (0.1 g/mL) was reduced up to 27% and 16%, respectively. The above incubation was associated with inactivation of PON1Q and PON1R by 15% and 45%, respectively. Lesion cholesteryl linoleate hydroperoxides and cholesteryl linoleate hydroxides were hydrolyzed by PON1 to yield linoleic acid hydroperoxides and linoleic acid hydroxides. Furthermore, lesion and pure linoleic acid hydroperoxides were reduced to yield linoleic acid hydroxides. These results thus indicate that PON1 demonstrates esterase-like and peroxidase-like activities. Recombinant PON1 mutants in which the PON1-free sulfhydryl group at cysteine-284 was replaced with either alanine or serine were no longer able to reduce lipid peroxide content in carotid lesions.. We conclude that PON1 may be antiatherogenic because it hydrolyzes lipid peroxides in human atherosclerotic lesions.

Topics: Arteriosclerosis; Aryldialkylphosphatase; Carboxylic Ester Hydrolases; Carotid Artery Diseases; Coronary Artery Disease; Esterases; Humans; In Vitro Techniques; Linoleic Acids; Lipid Peroxides; Peroxidase; Protein Isoforms

Effect of lipid peroxidation on the development of atherosclerotic lesions was studied in 10 diet-induced hyper cholesterolemic rabbits. By the end of the 3rd week of hypercholesterolemia, lipid peroxidative injury of the endothelial cells was induced by injecting linoleic acid hydroperoxide into a defined segment of the carotid artery. By the end of the 8th and 10th week light and electron microscopy revealed that atherosclerotic lesions mainly the fibrous plaques developed in certain segments of the artery described. The results indicate that under hypercholesterolemic condition, endothelial peroxidative injury may promote and intensify the process of atherogenesis. Furthermore, endothelial peroxidative injury which is characterised by the presence of plasma membrane blebbing can also be induced by hypercholesterolemia.

Topics: Animals; Arteriosclerosis; Carotid Arteries; Endothelium, Vascular; Hypercholesterolemia; Linoleic Acids; Lipid Peroxides; Male; Rabbits