Lipid peroxidation in low density lipoproteins from human plasma and egg yolk promotes accumulation of 1-acyl analogues of platelet-activating factor-like lipids

Oxidative modification of low density lipoprotein (LDL) is known to be a key event for induction of atherosclerosis. However, there has been little progress in structural elucidation of oxidized lipids, especially oxidatively fragmented phospholipids retaining a glycerol backbone. In this study, we found that LDL derived from egg yolk has no platelet-activating factor (PAF) acetylhydrolase activity, and that prolonged incubation of egg yolk LDL with Cu2+ resulted in the formation of various PAF-like lipids: 1-acyl type phosphatidylcholines with an sn-2-short-chain dicarboxylate or monocarboxylate group. Only a very small amount of the PAF-like lipid having an sn-2-short-chain monocarboxylate group was detected by gas chromatography-mass spectrometry in Cu(2+)-oxidized LDL from human plasma with high PAF-acetylhydrolase activity, which has been reported to hydrolyze PAF-like lipids to lysophosphatidyl-cholines. Preincubation of plasma LDL with diisopropyl fluorophosphate dose-dependently inhibited PAF-acetylhydrolase activity, resulting in accumulation of the PAF-like lipids when the LDL was oxidized with Cu2+. As well as PAF and lysophosphatidylcholines, several PAF-like lipids were found to inhibit [3H]thymidine incorporation into cultured vascular smooth muscle cells derived from rat aorta. The possible formation of PAF-like lipids by lipid peroxidation in LDL is discussed as well as its possible significance for induction of atherosclerosis.     

Effect of L-carnitine on lipid peroxidation and lipid composition in blood serum in hemic hypoxia]

Sodium nitrite has been studied for its effect on lipid metabolism of Wistar line rats. It is shown that a single administration of sodium nitrite in account 5 mg per 100 g of the body weight results in the intensification of lipids peroxidation, hyperbetalipoproteinemia, hypertriglyceridemia, hypercholesterolemia and to the decrease of the coefficient phospholipids/cholesterin/. Prophylactic administration of carnitine chloride in account of 20 mg per 100 g of the body weight stabilizes the level of lipids peroxidation, decreases concentration of total lipids, triglycerides, total cholesterin, phospholipids, lipoproteids of low and very low density, in the rat blood serum, normalizes the coefficient phospholipids/cholesterin.     

Fine structure of the olfactory organ of elasmobranchs (Elasmobranchii)]

The study acetylene amines were capable of inhibiting the processes of lipids peroxidation both in the enzymatic and non-enzymatic peroxidation system. The inhibitory effect of the study compounds depended upon the degree of the compound unsaturation. An elevation of the unsaturation degree to a definite extent is attended also by an increased inhibitory action on the processes of the lipids peroxidation.     

Possible mechanism of inhibition by polyamines of lipid peroxidation in rat liver microsomes

The rate of malondialdehyde formation in rat liver microsomes decreased as concentration of spermine added to the reaction mixture was increased. Inhibitory effect on lipid peroxidation was observed even when spermine was added at 15 min after the onset of reaction. Microsomal lipid peroxidation was markedly increased when extracted lipids were added to the reaction mixture. It was, moreover, found that in the presence of exogeneous microsomal lipids, spermine inhibition was weakened. The character of spermine inhibition with respect to the amounts of exogeneous and endogeneous lipids was shown to be competitive. From these results, it was suggested that spermine interaction with microsomal lipids may be responsible for the inhibition of lipid peroxidation.     

Antioxidant activity of the drug distinol in vitro

Distinol, ionol and dimethylsulphoxide were investigated in model reaction of thermal oxidation of methyl oleate for their antioxidant activity and for their influence on the rate of malonic dialdehyde formation in the liver tissue of chicken with E-hypovitaminosis in vitro. It has been found that the level of antioxidant activity in the homogeneous system of methyl oleate oxidation permits arranging the preparations studied in a series: ionol > distinol > dimethylsulphoxide. Distinol inhibits peroxidation of lipids in the liver homogenates of chicken most efficiently. Its inhibitory effect is 4.6 times higher than that of dimethylsulphoxide and 1.3 times higher than that of ionol. No correlation was found between antioxidant activity of distinol in the homogeneous model system and its influence on peroxidation of lipids in biomembranes.     

Copper ions promote peroxidation of low density lipoprotein lipid by binding to histidine residues of apolipoprotein B100, but they are reduced at other sites on LDL

Oxidized LDL is implicated in the pathogenesis of atherosclerosis. A widely studied model for oxidation of the lipid in LDL involves Cu2+. Recent studies suggest that Cu2+ may be reduced to Cu1+ by alpha-tocopherol to initiate LDL lipid peroxidation. LDL demonstrates binding sites for Cu2-, but the nature of these binding sites, as well their role in promoting Cu2+ reduction and lipid peroxidation, has not been established. In the current studies, we used diethylpyrocarbonate (DEPC) to modify the histidine residues of apolipoprotein B100, the major protein in LDL. First, we demonstrated that histidine residues were preferentially modified by DEPC under our experimental conditions. Then we monitored the kinetics of Cu(2+)-promoted oxidation of LDL and DEPC-modified LDL. In both cases, the progress curve of lipid peroxidation exhibited a lag phase and a propagation phase. However, when LDL was modified with DEPC, the length of the lag phase was prolonged whereas the rate of lipid peroxidation during the propagation phase was lower. Studies with LDL oxidized by 2,2'-azobis (2-amidinopropane) hydrochloride and phosphatidylcholine liposomes oxidized with hydroxyl radical established that DEPC was not acting simply as a nonspecific inhibitor of lipid peroxidation. DEPC treatment of LDL almost completely inhibited its ability to bind Cu2+. These observations suggest that peroxidation of the lipids in LDL can proceed with normal kinetics only when Cu2+ binds preferentially to sites on apolipoprotein B100 that contain histidine residues. We also compared the kinetics of Cu2+ reduction in the absence and presence of DEPC. There was no effect of DEPC modification on either the rate or extent of Cu2+ reduction by LDL. Therefore LDL is likely to contain a second class of binding sites for Cu2+ that does not involve histidine residues. Thus, LDL appears to contain at least two classes of Cu(2+)-binding sites: histidine containing sites, which are responsible in part for promoting lipid peroxidation during the propagation phase, and sites at which Cu2+ is reduced without binding to histidine.     

The dependence of thrombocyte aggregation on temperature

The effect of temperature (range of 4-44 degrees C) upon induced aggregation of platelets from normal subjects was studied. It was shown that temperature dependences of aggregation degree and rate have maxima. A relationship between aggregation agonist kind and some features of the temperature dependence curve of the aggregation parameters is established. It was shown that the limitation of platelet membrane lipids mobility by means of lipids peroxidation leads to the shift of maxima of the temperature dependence curve of platelet aggregation degree and rate. It was concluded that character of aggregation temperature dependence was governed by phase transition in platelet membrane lipids at room temperature.     

Inhibitory effect of ethanolamine plasmalogen on iron- and copper-dependent lipid peroxidation

The effect of ethanolamine plasmalogen (EtnPm) on lipid peroxidation was investigated in liposomal suspension of egg yolk phosphatidylcholine. EtnPm inhibited iron- and copper-dependent peroxidation in the presence of preformed hydroperoxides, although it was not effective for radical initiator mediated lipid peroxidation. EtnPm resulted in complete binding of iron to liposomal lipids, suggesting that EtnPm exerts its inhibitory effect on lipid peroxidation through inhibiting preformed peroxide decomposition by trapping transition metal ions.     

First direct evidence for lipid/protein conjugation in oxidized human low density lipoprotein

It has been postulated that lipids incorporated in atherosclerotic plaques are derived from the uptake of oxidized low density lipoprotein (LDL) by a macrophage-bound receptor. In vitro studies of LDL oxidation have established that reactive lipids are formed and that the exposure of native LDL to these products leads to modified protein with physical properties similar to oxidized LDL. Here we describe the application of highly specific tandem mass spectrometric techniques to the first characterization of lipid-modified LDL by demonstrating the addition of 4-hydroxy-2-nonenal to histidine residues of apolipoprotein B-100, following oxidation of LDL. The modified residues have been assigned to specific locations that have been previously shown to reside on the surface of the LDL particle.     

Lipid peroxidation: a review of causes, consequences, measurement and dietary influences

In this review the process of lipid peroxidation and the atherogenicity of peroxidied lipids are discussed. Recent findings with regard to the effect of selected dietary factors on susceptibility of lipids to oxidative stress and on antioxidant defences are analysed with particular reference to their potential use in the prevention and treatment of atherogenesis and, by extension, coronary heart disease. Laboratory methods of assessing antioxidant defences, lipid peroxidation and the effects of lipid peroxidation are also reviewed and discussed with particular reference to their ability to assess in vivo oxidative stress and lipid peroxidation status. A range of oxidative stress indices are presented and their limitations discussed, but the main focus is on the most commonly used laboratory test for lipid peroxidation, the thiobarbituric acid reacting substances (TBARS) test. Finally, the influence of selected dietary factors on measured peroxidation status is discussed, with particular reference to the antioxidant vitamins C (ascorbic acid) and E (alpha tocopherol) and the type of fatty acids (mono- and poly-unsaturated) in the diet.     

British Inflammation Research Association meeting: metalloproteinases and inflammatory diseases, the Fielder Centre, Hatfield, UK, 18 April 1997

Nitric oxide released by macrophages during inflammation reacts with active oxygen to form peroxynitrite. Peroxynitrite nitrates protein and peroxidizes lipids. gamma-Tocopherol traps peroxynitrite and is more effective than alpha-tocopherol in protecting lipids against such peroxidation.     

Low doses of eicosapentaenoic acid, docosahexaenoic acid, and hypolipidemic eicosapentaenoic acid derivatives have no effect on lipid peroxidation in plasma

It was of interest to investigate the influence of both high doses of eicosapentaenoic acid (EPA) and low doses of 2- or 3-methylated EPA on the antioxidant status, as they all cause hypolipidemia, but the dose required is quite different. We fed low doses (250 mg/d/kg body wt) of different EPA derivatives or high doses (1500 mg/d/kg body wt) of EPA and DHA to rats for 5 and 7 d, respectively. The most potent hypolipidemic EPA derivative, 2,2-dimethyl-EPA, did not change the malondialdehyde content in liver or plasma. Plasma vitamin E decreased only after supplementation of those EPA derivatives that caused the greatest increase in the fatty acyl-CoA oxidase activity. Fatty acyl-CoA oxidase activity increased after administration of both EPA and DHA at high doses. High doses of EPA and DHA decreased plasma vitamin E content, whereas only DHA elevated lipid peroxidation. In liver, however, both EPA and DHA increased lipid peroxidation, but the hepatic level of vitamin E was unchanged. The glutathione-requiring enzymes and the glutathione level were unaffected, and no significant changes in the activities of xanthine oxidase and superoxide dismutase were observed in either low- or high-dose experiments. In conclusion, increased peroxisomal beta-oxidation in combination with high amounts of polyunsaturated fatty acids caused elevated lipid peroxidation. At low doses of polyunsaturated fatty acids, lipid peroxidation was unchanged, in spite of increased peroxisomal beta-oxidation, indicating that polyunsaturation is the most important factor for lipid peroxidation.     

Lipoproteins accumulate in immune deposits and are modified by lipid peroxidation in passive Heymann nephritis

Proteinuria in passive Heymann nephritis is primarily caused by reactive oxygen species that are produced by glomerular cells. Reactive oxygen species apparently exert their damaging effects on the glomerular filter by lipid peroxidation and subsequent adduct formation on matrix proteins of glomerular basement membranes. This raised the question as to the source of polyunsaturated fatty acids required as substrates for lipid peroxidation. Here we have localized by immunocytochemistry rat apolipoprotein E and apolipoprotein B within subepithelial immune deposits. Moreover, apolipoprotein B extracted from isolated glomeruli of proteinuric passive Heymann nephritis rats shows degradation and lipid peroxidation adduct formation, similar to apoproteins of oxidized lipoproteins in atherosclerotic lesions. These data provide evidence that lipoproteins accumulate within immune deposits and suggest that their lipids generate lipid-peroxidation-derived reactive compounds.     

Linoleic acid intake and susceptibility of very-low-density and low density lipoproteins to oxidation in men

Lipoprotein peroxidation is thought to play an important role in atherogenesis. In the Kuopio Atherosclerosis Prevention Study (KAPS) the intake of fat and fatty acids, the oxidation susceptibility of the plasma very-low-density + low-density lipoprotein (VLDL+LDL) fraction (by induction with copper or hemin and hydrogen peroxide), and concentrations of plasma antioxidants, serum lipids, and lipoproteins were measured in 393 men. In the multivariate-regression model dietary linoleic acid was the most important determinant of the maximal oxidation velocity for the hemin assay (standardized regression coefficient = 0.294, P<0.0001). In the copper assay the association of dietary linoleic acid and maximal oxidation velocity was second in order of strength (standardized regression coefficient = 0.324, P< 0.0001). We conclude that high linoleic acid intake is associated with increased oxidation susceptibility of atherogenic lipoproteins in men.     

Oxidative stress and the development of diabetic complications--antioxidants and lipid peroxidation in erythrocytes and cell membrane

Erythrocytes isolated from 131 cases of Non-Insulin Dependent Diabetes Mellitus (NIDDM) were studied for lipid peroxidation, antioxidant defences, and the maximum peroxidisable substrate in the cell membrane. Antioxidant defences are lowered in NIDDM, followed by significant rise in lipid peroxidation products. However, in the erythrocyte membrane, the total polyunsaturated peroxidisable lipids are lower than in normal erythrocytes which may be a causative factor affecting the survival of the cells.     

Thermal stability of apolipoprotein B100 in low-density lipoprotein is disrupted at early stages of oxidation while neutral lipid core organization is conserved

The time course of the unfolding characteristics of the protein moiety and of the thermotropic behavior of the core-located apolar lipids of highly homogeneous low-density lipoprotein (LDL) subspecies (d 1.030-1.040 g/mL) have been evaluated during transition metal- and azo radical-induced oxidation using differential scanning calorimetry. Apolipoprotein B100 (apo-B100) structure was highly sensitive to oxidative modification; indeed, a significant loss of thermal stability was observed at initial stages irrespective of whether oxidation was mediated by site-specific binding of copper ions or by free radicals generated during decomposition of azo compounds. Subsequently, thermal protein integrity was destroyed, as a result of potentially irreversible protein unfolding, cross-linking reactions, and aggregation. Our results suggest that even minimal oxidative modification of apo-B100 has a major impact on the stability of this large monomeric protein. By contrast, the core lipids, which consist primarily of cholesteryl esters and triglycerides and play a determinant role in the thermal transition occurring near physiological temperature, preserved features of an ordered arrangement even during propagation of lipid peroxidation.     

gamma-Glutamyl transpeptidase-dependent lipid peroxidation in isolated hepatocytes and HepG2 hepatoma cells

Gamma-glutamyltranspeptidase (GGT), a plasma membrane-bound enzyme, provides the only activity capable to effect the hydrolysis of extracellular glutathione (GSH), thus favoring the cellular utilization of its constituent amino acids. Recent studies have shown however that in the presence of chelated iron prooxidant species can be originated during GGT-mediated metabolism of GSH, and that a process of lipid peroxidation can be started eventually in suitable lipid substrates. The present study was undertaken to verify if a GGT-dependent lipid peroxidation process can be induced in the lipids of biological membranes, including living cells, and if this effect can be sustained by the GGT highly expressed at the surface of HepG2 human hepatoma cells. In rat liver microsomes (chosen as model membrane lipid substrate) exposed to GSH and ADP-chelated iron, the addition of GGT caused a marked stimulation of lipid peroxidation, which was further enhanced by the addition of the GGT co-substrate glycyl-glycine. The same was observed in primary cultures of isolated rat hepatocytes, where the lipid peroxidation process did not induce acute toxic effects. GGT-stimulation of lipid peroxidation was dependent both on the concentration of GSH and of ADP-chelated iron. In GGT-rich HepG2 human hepatoma cells, the exposure to GSH, glycyl-glycine, and ADP-chelated iron resulted in a nontoxic lipid peroxidation process, which could be prevented by means of GGT inhibitors such as acivicin and the serine-boric acid complex. In addition, by co-incubation of HepG2 cells with rat liver microsomes, it was observed that the GGT owned by HepG2 cells can act extracellularly, as a stimulant on the GSH- and iron-dependent lipid peroxidation of microsomes. The data reported indicate that the lipid peroxidation of liver microsomes and of living cells can be stimulated by the GGT-mediated metabolism of GSH. Due to the well established interactions of lipid peroxidation products with cell proliferation, the phenomenon may bear particular significance in the carcinogenic process, where a relationship between the expression of GGT and tumor progression has been envisaged.     

Hypersensitivity following low one-time irradiation dosage and induced radio resistance

There is now little doubt of the existence of radioprotective mechanisms, or stress responses, that are upregulated in response to exposure with small doses of ionizing radiation and other DNA-damaging agents. Phenomenologically, there are two ways in which these induced mechanisms operate. First, a small conditioning dose (generally below 30 cGy) may protect against a subsequent, separate irradiation. This has been termed the adaptive response. Second, the response to single doses may itself be dose-dependent so that small acute radiation exposures are more effective per unit dose than larger exposures above the threshold where the induced radioprotection is triggered. This combination has been termed low dose hypersensitivity (HRS) and induced radioresistance (IRR) as the dose increases. Both the adaptive response and HRS/IRR have been well documented in studies with yeast, bacteria, protozoa, algae, higher plant cells, insect cells, mammalian and human cells in vitro, and in studies on animal models in vivo. There is indirect evidence that the adaptive response and the IRR phenomenon in response to single doses is a manifestation of the same underlying mechanisms, namely an increase of the amount and rate of DNA repair induced by low radiation doses.     

Lipid hydroperoxide generation, turnover, and effector action in biological systems

Lipid peroxidation is a well known example of oxidative damage in cell membranes, lipoproteins, and other lipid-containing structures. Peroxidative modification of unsaturated phospholipids, glycolipids, and cholesterol can occur in reactions triggered by i) free radical species such as oxyl radicals, peroxyl radicals, and hydroxyl radicals derived from iron-mediated reduction of hydrogen peroxide or ii) non-radical species such as singlet oxygen, ozone, and peroxynitrite generated by the reaction of superoxide with nitric oxide. Lipid hydroperoxides (LOOHs) are prominent non-radical intermediates of lipid peroxidation whose identification can often provide valuable mechanistic information, e.g., whether a primary reaction is mediated by singlet oxygen or oxyradicals. Certain cholesterol-derived hydroperoxides (ChOOHs) have been used very effectively in this regard, both in model systems and cells. Being more polar than parent lipids, LOOHs perturb membrane structure/function and can be deleterious to cells on this basis alone. However, LOOHs can also participate in redox reactions, the nature and magnitude of which often determines whether peroxidative injury is exacerbated or prevented. Exacerbation may reflect iron-catalyzed one-electron reduction of LOOHs, resulting in free radical-mediated chain peroxidation, whereas prevention may reflect selenoperoxidase-catalyzed two-electron reduction of LOOHs to relatively non-toxic alcohols. LOOH partitioning between these two pathways in an oxidatively stressed cell is still poorly understood, but recent cell studies involving various ChOOHs have begun to shed light on this important question. An aspect of related interest that is under intensive investigation is lipid peroxidation/LOOH-mediated stress signaling, which may evoke a variety of cellular responses, ranging from induction of antioxidant enzymes to apoptotic death. Ongoing exploration of these processes will have important bearing on our understanding of disease states associated with peroxidative stress.     

Acrolein is a product of lipid peroxidation reaction. Formation of free acrolein and its conjugate with lysine residues in oxidized low density lipoproteins

Lipoprotein peroxidation, especially the modification of apolipoprotein B-100, has been implicated to play an important role in the pathogenesis of atherosclerosis. However, there have been few detailed insights into the chemical mechanism of derivatization of apolipoproteins during oxidation. In the present study, we provide evidence that the formation of the toxic pollutant acrolein (CH2=CH-CHO) and its conjugate with lysine residues is involved in the oxidative modification of human low density lipoprotein (LDL). Upon incubation with LDL, acrolein preferentially reacted with lysine residues. To determine the structure of acrolein-lysine adduct in protein, the reaction of acrolein with a lysine derivative was carried out. Employing Nalpha-acetyllysine, we detected a single product, which was identified to be a novel acrolein-lysine adduct, Nalpha-acetyl-Nepsilon-(3-formyl-3,4-dehydropiperidino )lysine. The acid hydrolysis of the adduct led to the derivative that was detectable with amino acid analysis. It was revealed that, upon in vitro incubation of LDL with acrolein, the lysine residues that had disappeared were partially recovered by Nepsilon-(3-formyl-3, 4-dehydropiperidino)lysine. In addition, we found that the same derivative was detected in the oxidatively modified LDL with Cu2+ and that the adduct formation was correlated with LDL peroxidation assessed by the consumption of alpha-tocopherol and cholesteryl ester and the concomitant formation of cholesteryl ester hydroperoxide. Enzyme-linked immunosorbent assay that measures free acrolein revealed that a considerable amount of acrolein was released from the Cu2+-oxidized LDL. Furthermore, metal-catalyzed oxidation of arachidonate was associated with the formation of acrolein, indicating that polyunsaturated fatty acids including arachidonate represent potential sources of acrolein generated during the peroxidation of LDL. These results indicate that acrolein is not just a pollutant but also a lipid peroxidation product that could be ubiquitously generated in biological systems.