The role of covalent binding and lipid peroxidation in liver damage by carbon tetrachloride]

4-[N-sodium-N-(5-ethyl-1,3,4-thiadiazol-2-yl)]- sulphanylamido-5-methoxy-1,2-benzoquinone selectively inhibiting lipid peroxidation (LPO) was used to study the hepatotoxic effect of carbon tetrachloride in vivo. It was found that inactivation of the liver microsomal oxidation system during the first few hours after CCl4 injection is due to covalent binding rather than LPO.     

The role of phospholipase A activity in rat liver microsomal lipid peroxidation

The involvement of phospholipase(s) A in lipid peroxidation of rat liver microsomes was investigated by: (a) determining the effects of phospholipase A inhibitors (p-bromophenylacyl bromide, chlorpromazine, mepacrine) on the accumulation of thiobarbituric acid reactivity or on levels of oxidized phospholipids in response to selected oxidative stimuli and (b) measurement of phospholipase A activities in response to these agents. Lipid peroxidation in response to various peroxidation systems was inhibited completely by exposure of microsomes to p-bromophenylacyl bromide (250 microM). The effectiveness of p-bromophenylacyl bromide was dependent on the presence of glutathione (200 microM) in preincubation mixtures. Chlorpromazine (100 microM) and mepacrine (100 microM) also effectively inhibited peroxidation, and their potency was independent of glutathione. The accumulation of oxidized phospholipids in response to the potent peroxidation stimulus alloxan/ferrous ion was similarly inhibited by p-bromophenylacyl bromide, although the level of oxidized phospholipid in response to the initiator ADP/ferrous ion was not affected. Microsomal phospholipase A1 activity, assessed using a liposomal substrate, was substantially enhanced by promoters of lipid peroxidation. Phospholipase A2 activity was not detected using a liposomal substrate but was evident using radiolabeled microsomes as endogenous substrate and was enhanced by oxidative stimuli. We conclude that phospholipase A activity may play an integral role in the microsomal lipid peroxidation mechanism. Based on this study, we hypothesize a role for phospholipases in facilitating propagation reactions.     

The mechanism of liver microsomal lipid peroxidation

In the presence of Fe³⁺ and complexing anions, the peroxidation of unsaturated liver microsomal lipid in both intact microsomes and in a model system containing extracted microsomal lipid can be promoted by either NADPH and NADPH : cytochrome c reductase or by xanthine and xanthine oxidase. Erythrocuprein effectively inhibits the activity promoted by xanthine and xanthine oxidase but produces much less inhibition of NADPH-dependent peroxidation. The singlet-oxygen trapping agent, 1,3-diphenylisobenzofuran, had no effect on NADPH-dependent peroxidation but strongly inhibited the peroxidation promoted by xanthine and xanthine oxidase. NADPH-dependent lipid peroxidation was also shown to be unaffected by hydroxyl radical scavengers.. The addition of catalase had no effect on NADPH-dependent lipid peroxidation, but it significantly increased the rate of malondialdehyde formation in the reaction promoted by xanthine and xanthine oxidase. These results demonstrate that NADPH-dependent lipid peroxidation is promoted by a reaction mechanism which does not involve either superoxide, singlet oxygen, HOOH, or the hydroxyl radical. It is concluded that NADPH-dependent lipid peroxidation is initiated by the reduction of Fe³⁺ followed by the decomposition of hydroperoxides to generate alkoxyl radicals. The initiation reaction may involve some form of the perferryl ion or other metal ion species generated during oxidation of Fe²⁺ by oxygen.     

The role of lipid peroxidation in CCl4-induced damage to liver microsomal enzymes: comparative studies in vitro using microsomes and isolated liver cells

The question as to whether CCl4 decreases the activities of glucose-6-phosphatase and cytochrome P-450 in liver endoplasmic reticulum mainly through its action in stimulating lipid peroxidation has been investigated using Promethazine to block lipid peroxidation. The investigation, moreover, has compared the effects of CCl4, with and without Promethazine, on isolated rat hepatocytes with corresponding effects on rat liver microsomal suspensions. Our data give no support for the view that products of lipid peroxidation are the main cause of the decrease in cytochrome P-450 observed in CCl4-intoxication. However, our present results are consistent with lipid peroxidation being a major contributory factor to the decrease in glucose-6-phosphatase activity observed in CCl4-induced liver injury.     

Rabbit liver microsomal lipid peroxidation. The effect of lipid on the rate of peroxidation

Rat and rabbit liver microsomes catalyze an NADPH-cytochrome P-450 reductase-dependent peroxidation of endogenous lipid in the presence of the chelate, ADP-Fe3+. Although liver microsomes from both species contain comparable levels of NADPH-cytochrome P-450 reductase and cytochrome P-450, the rate of lipid peroxidation (assayed by malondialdehyde and lipid hydroperoxide formation) catalyzed by rabbit liver microsomes is only about 40% of that catalyzed by rat liver microsomes. Microsomal lipid peroxidation was reconstituted with liposomes made from extracted microsomal lipid and purified protease-solubilized NADPH-cytochrome P-450 reductase from both rat and rabbit liver microsomes. The results demonstrated that the lower rates of lipid peroxidation catalyzed by rabbit liver microsomes could not be attributed to the specific activity of the reductase. Microsomal lipid from rabbit liver was found to be much less susceptible to lipid peroxidation. This was due to the lower polyunsaturated fatty acid content rather than the presence of antioxidants in rabbit liver microsomal lipid. Gas-liquid chromatographic analysis of fatty acids lost during microsomal lipid peroxidation revealed that the degree of fatty acid unsaturation correlated well with rates of lipid peroxidation.     

The crucial role of hypoxia in halothane-induced lipid peroxidation

Halothane-induced lipid peroxidation in NADPH-reduced liver microsomes from phenobarbital-pretreated male rats was studied under defined steady state oxygen partial pressures (Po2). Under anaerobic conditions, as well as at a Po2 above 10 mm Hg no halothane-induced formation of malondialdehyde was detected. At a Po2 below 10 mm Hg, however, with a maximum near 1 mm Hg oxygen, significant halothane-induced malondialdehyde formation was found. This evidence supports the hypothesis that halothane can induce lipid peroxidation. The Po2 (i) must be low enough to permit the reductive formation of . CF3 CHCl-radicals but (ii), it must be high enough to promote formation of lipid peroxides.     

The protective role of plasmalogens in iron-induced lipid peroxidation

The role of plasmalogens in iron-induced lipid peroxidation was investigated in two liposomal systems. The first consisted of total brain phospholipids with and without plasmalogens, and the second of phosphatidylethanolamine/phosphatidylcholine liposomes with either diacyl- or alkenylacyl-phosphatidylethanolamine. By measuring thiobarbituric acid reactive substances, oxygen consumption, fatty acids and aldehydes, we show that plasmalogens effectively protect polyunsaturated fatty acids from oxidative damage, and that the vinyl ether function of plasmalogens is consumed simultaneously. Furthermore, the lack of lag phase, the increased antioxidant efficiency with time, and the experiments with lipid- and water-soluble azo compounds, indicate that plasmalogens probably interfere with the propagation rather than the initiation of lipid peroxidation, and that the antioxidative effect cannot be related to iron chelation.     

The mechanism of liver microsomal lipid peroxidation.

In the presence of Fe-3+ and complexing anions, the peroxidation of unsaturated liver microsomal lipid in both intact microsomes and in a model system containing extracted microsomal lipid can be promoted by either NADPH and NADPH : cytochrome c reductase or by xanthine and xanthine oxidase. Erythrocuprein effectively inhibits the activity promoted by xanthine and xanthine oxidase but produces much less inhibition of NADPH-dependent peroxidation. The singlet-oxygen trapping agent, 1, 3-diphenylisobenzofuran, had no effect on NADPH-dependent peroxidation but strongly inhibited the peroxidation promoted by xanthine and xanthine oxidase. NADPH-dependent lipid peroxidation was also shown to be unaffected by hydroxyl radical scavengers.. The addition of catalase had no effect on NADPH-dependent lipid peroxidation, but it significantly increased the rate of malondialdehyde formation in the reaction promoted by xanthine and xanthine oxidase. The results demonstrate that NADPH-dependent lipid peroxidation is promoted by a reaction mechanism which does not involve either superoxide, singlet oxygen, HOOH, or the hydroxyl radical. It is concluded that NADPH-dependent lipid peroxidation is initiated by the reduction of Fe-3+ followed by the decomposition of hydroperoxides to generate alkoxyl radicals. The initiation reaction may involve some form of the perferryl ion or other metal ion species generated during oxidation of Fe-2+ by oxygen.     

Protective role of L-carnitine on liver and heart lipid peroxidation in atherosclerotic rats

Lipid peroxides are considered to be the initiation factor for atherosclerosis. Present study depicts that L-carnitine treatment (300 mg/kg body weight/day) for 7 and 14 days caused significant reduction in the tissue lipid peroxidations. It also shows marked improvement in the antioxidant status. By this way carnitine maintain the normal function of the cells.     

The role of iron in the initiation of lipid peroxidation

Iron is required for the initiation of lipid peroxidation. Evidence is presented that lipid peroxidation requires both Fe3+ and Fe2+, perhaps with oxygen to form a Fe3+-dioxygen-Fe2+ complex. Other mechanisms of initiation, mostly involving the iron-catalyzed formation of hydroxyl radical, are described and discussed from both theoretical and experimental view points.     

The role of lipid peroxidation in the mechanism of stress

A concept on the mechanism of stress response is substantiated proceeding from the data available in literature and obtained from the author's research made on the radiation stress model. The conception envisages that products of lipid peroxidation (LPO) appear as primary (under direct effect of a stress factor on tissues) and secondary (as a consequence of high- and long-term catecholamine++) mediators. Mobilization of stress-realizing systems in that process is regarded as an adequate response of the auto-oxidative++ system to the primary activation of LPO. Transformation of catecholamines into the factor of LPO stimulation (secondary) is a result of an increase in the relative role of the quinoid way to transform catecholamines in the case of their high concentration. Radical intermediates of the quinoid metabolism appear as LPO initiators. An important pathogenetic role of LPO activation in the stress mechanism substantiates expedience to use antioxidants as agents for prophylaxis and early treatment of stress-factor injuries.     

The role of iron in oxygen radical mediated lipid peroxidation

The role of iron in the peroxidation of polyunsaturated fatty acids is reviewed, especially with respect to the involvement of oxygen radicals. The hydroxyl radical can be generated by a superoxide-driven Haber-Weiss reaction or by Fenton's reaction; and the hydroxyl radical can initiate lipid peroxidation. However, lipid peroxidation is frequently insensitive to hydroxyl radical scavengers or superoxide dismutase. We propose that the hydroxyl radical may not be involved in the peroxidation of membrane lipids, but instead lipid peroxidation requires both Fe2+ and Fe3+. The inability of superoxide dismutase to affect lipid peroxidation can be explained by the fact that the direct reduction of iron can occur, exemplified by rat liver microsomal NADPH-dependent lipid peroxidation. Catalase can be stimulatory, inhibitory or without affect because H2O2 may oxidize some Fe2+ to form the required Fe3+, or, alternatively, excess H2O2 may inhibit by excessive oxidation of the Fe2+. In an analogous manner reductants can form the initiating complex by reduction of Fe3+, but complete reduction would inhibit lipid peroxidation. All of these redox reactions would be influenced by iron chelation.     

The possible role of lipid peroxidation in breast cancer risk

Breast cancer remains the commonest cause of death from cancer in women in most of the Western world. There is considerable evidence that breast cancer risk is influenced by environmental factors and can therefore potentially be modified. In this paper we describe evidence suggesting a relationship of lipid peroxidation to breast cancer risk, and propose that the method used to generate this information might usefully be applied to other disease states, and make some suggestions for further work. We have compared the urinary excretion of the mutagen malonaldehyde (MDA) in premenopausal women at different risks for breast cancer as determined by the appearance of the breast parenchyma on mammography. MDA was measured in 24-h urine samples from both groups and excretion in 30 women with mammographic dysplasia (high risk) was found to be approximately double that of 16 women without these radiological changes (p less than 0.02). These results suggest that mammographic dysplasia may be associated with lipid peroxidation. Further study of environmental factors associated with states that precede the development of breast and other cancers may lead to the identification of factors that can be modified and that may prevent the development of malignant disease.     

Dehydroepiandrosterone-induced lipid peroxidation in rat liver mitochondria

Administration of dehydroepiandrosterone (DHEA), a steroid hormone of the adrenal cortex which acts as a peroxisome proliferator and hepatocarcinogen in the rat, caused an increase in NADPH-dependent lipid peroxidation in mitochondria isolated from the liver, kidney and heart, but not from the brain. The effect of DHEA on rat liver mitochondrial lipid peroxidation became discernible after feeding steroid-containing diet (0.6% w/w) for 3 days, and reached maximal levels between 1 and 2 weeks. DHEA in the concentration range 0.001–0.02% did not significantly increase lipid peroxidation compared to the control. Lipid peroxidation was significantly enhanced in animals given a diet containing ≥ 0.05% DHEA. The addition of DHEA in the concentration range 0.1–100 μM to mitochondria isolated from control rats had no effect on lipid peroxidation. It seems, therefore, that the steroid effect is mediated by an intracellular process. Our data indicate that induction of mitochondrial membrane lipid peroxidation is an early effect of DHEA administration at pharmacological doses.     

DNA damage caused by lipid peroxidation products

Lipid peroxidation is a process involving the oxidation of polyunsaturated fatty acids (PUFAs), which are basic components of biological membranes. Reactive electrophilic compounds are formed during lipid peroxidation, mainly alpha, beta-unsaturated aldehydes. These compounds yield a number of adducts with DNA. Among them, propeno and substituted propano adducts of deoxyguanosine with malondialdehyde (MDA), acrolein, crotonaldehyde and etheno adducts, resulting from the reactions of DNA bases with epoxy aldehydes, are a very important group of adducts. The epoxy aldehydes are more reactive towards DNA than the parent unsaturated aldehydes. The compounds resulting from lipid peroxidation mostly react with DNA showing both genotoxic and mutagenic action; among them, 4-hydroxynonenal is the most genotoxic, while MDA is the most mutagenic. DNA damage caused by the adducts of lipid peroxidation products with DNA can be removed by the repairing action of glycosylases. The formed adducts have been hitherto analyzed using the IPPA (Imunopurification-(32)P-postlabelling assay) method and via gas chromatography/electron capture negtive chemical ionization/mass spectrometry (GC/EC NCI/MS). A combination of liquid chromatography with electrospray tandem mass spectrometry (LC/ES-MSMS) with labelled inner standard has mainly been used in recent years.     

The role of cholesterol in activating lipid peroxidation in platelets]

The relationship between the cholesterol (Ch) content and the concentration of lipid peroxidation (LPO) products in activated platelets and the effect of these parameters on the structure-function characteristics of platelet membranes were studied. It was found that esterified Ch activates free radical processes occurring in platelets. Nonesterified Ch does not induce the production of primary products of LPO (dienoic conjugates) but promotes the accumulation of a secondary LPO metabolite, malonic dialdehyde, this reaction being mediated via indirect mechanisms. The higher (in comparison with normal) orderliness and orientation of membranes in platelets reflect the increase in the concentration of dienoic conjugates and nonesterified Ch. The observed differences in the aggregability of platelets are due to the changes in the Ch content as well as to the "rigidity" of blood platelets.     

Pathobiochemical mechanisms of hepatocellular damage following lipid peroxidation

In hepatocytes, cytotoxic events induced by haloalkanes or acute iron-overload exhibit neither a quantitative nor a temporal correlation to lipid peroxidation or covalent binding. Thus, secondary pathological mechanisms have been postulated linking initial focal reactions of free radicals and end stage pathological consequences. Due to the crucial role of plasma-membrane integrity in the cytotoxic process it has to be supposed that relevant secondary pathological mechanisms finally impair the physico-chemical and functional properties of this membrane. Based on recent developments a chain of causality is proposed as a two-step activation of phospholipase A2 producing cytolytic amounts of lysophosphatides. In this cascade, the initial activating step is a decrease of membrane lipid fluidity induced by lipid peroxidation and/or by calcium binding and intramembranous formation of 4-hydroxynonenal. This enzyme activation is further amplified by the early rise of cytosolic calcium. Consequently, increasing amounts of lysophosphatides progressively impair membrane configuration thus improving the substrate accessibility for phospholipase A2 in a second activation step. Finally, the lysophosphatides reach plasma membrane-lytic concentrations by this autocatalytic enzyme activation.     

The role of NADPH-cytochrome b 5 reductase in microsomal lipid peroxidation

Spectrophotometric changes in the extent of NADPH, but not NADH, reduction of microsomal cytochrome b₅ are correlated with the utilization of oxygen and the accumulation of lipid peroxidation products. The results suggest that NADPH-cytochrome b₅ reductase (NADPH-cytochrome c reductase) participates in the reduction of obligatory ferric chelates to their ferrous form prior to the initiation of lipid peroxidation. Further, an increased oxidation of cytochrome b₅ observed only in the presence of peroxidation products implicates a peroxidase activity associated with b₅ in the microsomal electron transport chain.     

Pregnancy-associated decrease in lipid peroxidation in rat liver

A significant decrease in the hepatic malonaldehyde content and lipid peroxidation, induced by ascorbate, NADPH and cumene hydroperoxide, was observed during gestation in the rat. Lipid peroxidation tends to reach normal levels 3 days post partum. While a significant decrease in the lipid peroxidation of hepatic mitochondria was observed with ascorbate and NADPH, that of microsomes was affected by ascorbate and cumene hydroperoxide. The observed decrease in lipid peroxidation during pregnancy seems to be due to lesser phospholipid content, a lower degree of unsaturation in lipids, and an increase in the level of antioxidants.     

The role of radical reactions in organomercurials impact on lipid peroxidation

The organomercury compounds RHgX and R(2)Hg are broad-spectrum biocidal agents acting via diverse mechanisms in biological systems. Despite the enormous amount of studies carried out in last decades to elucidate the detailed mechanisms of organomercurials toxicity their biomolecular mode of action is still under debate. Among various toxicity mechanisms the action of RHgX and R(2)Hg at the membrane level due to the lipophilic properties of their molecules is discussed. Organomercurials are supposed to induce membrane associated oxidative stress in living organism through different mechanisms including the enhancement of the lipid peroxidation and intracellular generation of reactive oxygen species (ROS), H(2)O(2), O(2)(-), HO(). The perturbation of antioxidative defense system and the peroxidation of unsaturated fatty acids in membrane lipid bilayer are consequences of this impact. On the other hand, the involvement of organomercurials in radical and redox biochemical processes is manifested in carbon to metal bond cleavage that leads to the generation of reactive organic radicals R(). This pathway is discussed as one of the multiple mechanisms of organomercurials toxicity. The goal of this review is to present recent results in the studies oriented towards the role of organomercurials in the xenobiotic-mediated enhancement of radical production and hence in the promotion of lipids peroxidation. The application of natural and synthetic antioxidants as detoxification agents is presented.