Early changes of liver microsomal lipids in acute thiobenzamide poisoning

The administration to rats of Thiobenzamide, a new liver toxin, induces the early appearance of conjugated dienes among microsomal lipids. Along with an increased malonic dialdehyde "in vitro", the finding suggests lipid peroxidation to occur.     

Liver lipid peroxidation in the ontogeny of rats during perinatal exposure to polychlorinated biphenyls

We studied the activity of NADPH-cytochrome P-450 reductase, NADPH- and ascorbate-dependent systems of lipid peroxidation in liver microsomes, the activity of superoxide dismutase in the supernatant and the level of malonic acid dialdehyde in liver tissue of rats of various age. The activity of lipid peroxidation system and the malonic dialdehyde content in the early postnatal period increased to the adult level. The NADPH-cytochrome P-450 reductase activity increased during the first four months of animals life while that of superoxide dismutase increased until the animals were seven months old. A single administration of polychlorinated diphenyls at a dose of 500 mg/kg (1/10 LD50) to pregnant rats drastically stimulated and changed the pattern of the studied activities in their offspring. The role of lipid peroxidation in modification of microsomal membranes after the monooxygenase system induction by polychlorinated diphenyls in early ontogenesis is discussed.     

Functioning of the monooxygenase system of the liver in experimental myocardial infarct

The experiments on rats have shown that coronary artery ligation reduces the content of microsomal cytochromes P-450 and b5 and causes amidopyrine-N-demethylation and aniline-p-hydroxylation disturbances that persist throughout a 3-week period of myocardial infarction. The investigation of spontaneous lipid peroxidation of microsomal membranes in myocardial infarction has shown that concentration of malonic dialdehyde in microsomal fraction significantly increased by the 7th day after coronary artery ligation, as compared to sham-operated rats.     

Kinetics of lipid peroxidation in isolated surviving rat livers

The kinetics of accumulation of lipid peroxidation products (hydroperoxides as primary products and malonic dialdehyde and "fluorescent pigments" as secondary ones) was investigated in an isolated non-perfused and preliminarily perfused liver during aerobic incubation. In the course of surviving there takes place an intensive accumulation of primary, secondary and final products of lipid peroxidation whose kinetics is of an extreme character. The rate of this process in a non-perfused liver is considerably higher than in a preliminarily perfused liver.     

Cytochrome P450 inactivation in microsomal membranes damaged by Fe2+-ascorbate-dependent lipid peroxidation

It has been established that Fe2+-ascorbate-dependent lipid peroxidation in rat liver microsome membranes is followed by the decrease of microsome cytochrome P450 content and the increase of the reduced haemoprotein inactivation rate. These changes are proportional to the amount of lipid peroxidation products (malonic dialdehyde) accumulating in the membranes.     

The reaction of 2-thiobarbituric acid with biologically active alpha,beta-unsaturated aldehydes

The standard assay for lipid peroxidation is the measurement of the pink, 532 n, absorbing chromogen which is formed upon reaction of 2-thiobarbituric acid (TBA) with the lipid peroxidation product malonaldehyde (MDA). The present studies indicate that the toxic lipid peroxidation product trans-4-hydroxynonenal and its dehydration product trans, trans-nonadienal react with TBA to form chromogens which absorb maximally at 530 and 532 nm, respectively. Other biologically active alpha, beta-unsaturated aldehydes, such as acrolein and crotonaldehyde, short-chain homologs of alkenals formed during lipid peroxidation, and trans,trans-muconaldehyde, a novel diene dialdehyde, react with TBA to form products which absorb maximally at 495 nm. The molar extinction coefficients of the aldehyde: TBA chromogens formed were found to vary widely, suggesting that only small contributions to the 532 nm absorption by TBA adducts of reactive aldehydes other than MDA may be encountered during the use of the TBA assay.     

Oxidation of aldehydic products of lipid peroxidation by rat liver microsomal aldehyde dehydrogenase

Lipid peroxidation in microsomal membranes produces a large number of aldehydes, alcohols, and ketones, some of which have been shown to be cytotoxic. This study has determined the kinetic parameters for the oxidation of aldehyde lipid peroxidation products by purified rat hepatic microsomal aldehyde dehydrogenase (ALDH). Livers were obtained from male Sprague-Dawley rats for preparation of microsomal ALDH which was purified 400-fold. Kinetic parameters, Vmax and V/K, were determined for saturated and unsaturated aldehydes of three to nine carbons in length in the presence of NAD+. Of the aldehydes examined, only acrolein and 4-hydroxynonenal were not oxidized by ALDH. The Vmax values (mumol NADH produced/min/mg protein) increased linearly with carbon chain length and ranged from 6.5 to 23 for the saturated series and 4.0 to 9.0 for the unsaturated aldehydes. The affinity constant V/K (nmol NADH produced/min/mg protein/nmol aldehyde/liter) also increased with carbon chain length and ranged from 12 to 9000 for the saturated aldehydes and 13 to 5300 for the unsaturated aldehydes. These results suggest that microsomal ALDH may serve a biological role for detoxification of reactive aldehydes produced by lipid peroxidation of microsomal membranes.     

Activation of chemiluminescence accompanying lipid peroxidation in monolayer liposome suspension

In the presence of rhodamine J the chemiluminescence intensity of monolayer liposomes induced by ferrous ions is increased by three orders. There is no accumulation of malonic dialdehyde. It is suggested that chemiluminescence activation is related to rhodamine J interaction with the products of lipid peroxidation whose molecules are in the excited state.     

Inhibition of Class-3 aldehyde dehydrogenase and cell growth by restored lipid peroxidation in hepatoma cell lines

Hepatoma cells have a below-normal content of polyunsaturated fatty acids; this reduces lipid peroxidation and the production of cytotoxic and cytostatic aldehydes within the cells. In proportion to the degree of deviation, hepatoma cells also show an increase in the activity of Class-3 aldehyde dehydrogenase, an enzyme important in the metabolism of lipid peroxidation products and also in that of several drugs. When hepatoma cells with different degrees of deviation were enriched with arachidonic acid and stimulated to peroxidize by ascorbate/iron sulphate, their growth rate was reduced in proportion to the quantity of aldehydes produced and to the activity of aldehyde dehydrogenase. Therefore, 7777 cells, less deviated and with low Class-3 aldehyde dehydrogenase activity, were more susceptible to lipid peroxidation products than JM2 cells. It is noteworthy that repeated treatments with prooxidant also caused a decrease in mRNA and activity of Class-3 aldehyde dehydrogenase, contributing to the decreased growth and viability. Thus, Class-3 aldehyde dehydrogenase could be considered relevant for the growth of hepatoma cells, since it defends them against cell growth inhibiting aldehydes derived from lipid peroxidation.     

The role of aldehyde dehydrogenases in the malonic dialdehyde metabolism in the rat liver

The enzymes catalyzing the NAD-dependent oxidation of malonic dialdehyde (MDA) were isolated from rat liver extracts. Upon 5'-AMP-Sepharose chromatography MDA dehydrogenase was separated into two isoforms, I and II. Isoform I was eluted from the affinity carrier with a 0.1 M phosphate buffer pH 8.0. This isoform had a broad substrate specificity towards aliphatic and aromatic aldehydes. Kinetic studies showed that short- and medium-chain aliphatic aldehydes (C2-C6) were characterized by the lowest Km values and the highest Vmax values. The Km' values for MDA and acetaldehyde were 2.8 microM and 0.69 microM, respectively. Isoform II was eluted with a 0.1 M phosphate buffer pH 8.0 containing 0.5 mM NAD, was the most active with medium- and long-chain aliphatic aldehydes (C6-C11) and had Km values for MDA and acetaldehyde equal to 37 microM and 52 microM, respectively. Isoform I was much more sensitive towards disulfiram inhibition than isoform II. Both isoforms had an identical molecular mass (93 kD) upon gel filtration. It is concluded that MDA dehydrogenase isoform I is identical to mitochondrial aldehyde dehydrogenase having a low Km for acetaldehyde, whereas isoform II may be localized in liver cytosol. The role of aldehyde dehydrogenases in the metabolism of aldehydes derived from lipid peroxidation is discussed.     

A New Role of Phosphoglucose Isomerase. Involvement of the Glycolytic Enzyme in Aldehyde Metabolism

Lipid peroxidation in biological membranes is accompanied by malonic dialdehyde (MDA) formation, but the problem of its further metabolism in cytoplasm remains unsolved. The experimental data obtained in this work showed that the liver fraction prepared by centrifugation at 10,000g contained phosphoglucose isomerase and enzymes of the glyoxalase system. In this fraction in the presence of GSH there is an aggregate of reactions taking place both in membranes (lipid peroxidation) and outside membranes (MDA conversion to methylglyoxal and further to neutral D-lactate). This means that MDA is slowly accumulated because it is a substrate of aldehyde isomerase (MDA <--> methylglyoxal). Most probably, phosphoglucose isomerase serves as this enzyme. We concluded that D-lactate should be regarded as the end product of two different parametabolic reactions: lipid peroxidation or protein glycation.     

Effect of genistein and daidzein obtained by acid hydrolysis of their glycosides on phospholipid peroxidation

A mixture of isoflavones was obtained by acid hydrolysis of isoflavone glycosides isolated from the products of soybean processing by a successive extraction with aqueous acetone and methanol. Homogeneous isoflavones genistein and daidzein were isolated from the aglycone mixture by adsorption chromatography and identified by spectral and chromatographic methods. The effect of both isoflavones on lipid peroxidation of soy phospholipids in multilamellar vesicles was studied at various concentrations. These aglycones were found to inhibit the formation of lipid hydroperoxides and malonic dialdehyde at the concentrations as low as 1 mM.     

Isoflavones Daidzein and Genistein: Preparation by Acid Hydrolysis of Their Glycosides and the Effect on Phospholipid Peroxidation

A mixture of isoflavones was obtained by acid hydrolysis of isoflavone glycosides isolated from the products of soybean processing by successive extraction with aqueous acetone and methanol. The homogeneous isoflavones daidzein and genistein were isolated from the aglycone mixture by adsorption chromatography and identified by spectral and chromatographic methods. The effect of both isoflavones on lipid peroxidation of soy phospholipids in multilamellar vesicles was studied at various concentrations. These aglycones were found to inhibit the formation of lipid hydroperoxides and malonic dialdehyde at concentrations as low as 1 mM.     

Inhibition of phospholipid hydrolysis by phospholipase A2 in microsomal and mitochondrial membranes subjected to lipid peroxidation

The rate of phospholipid hydrolysis in rat liver microsomal and mitochondrial membranes catalyzed by phospholipase A2 was shown to decrease after ascorbate + Fe2+-induced lipid peroxidation. The degree of inhibition was linearly dependent on the amount of lipid peroxidation products (malonyl dialdehyde) accumulated in the membrane. The decreased phospholipid hydrolysis rate in membranes after lipid peroxidation was registered using phospholipases A2 from two sources: porcine pancreas and bee venom. It was established that the inhibitory action of phospholipid peroxidation products was not linked with a direct effect on the enzyme and was not caused by depletion of phospholipase reaction substrates (as a result of lipid peroxidation). A possible role of lateral separation of oxidized and non-oxidized lipid phases in the mechanisms of inhibition of phospholipid hydrolysis by phospholipase A2 is discussed.     

Lipid peroxidation in experimental Salmonella infection

The results of investigation of lipid peroxidation in experimental Salmonella infection in 21-day-old rabbits are analyzed. Salmonella infection was accompanied by activation of lipid peroxidation not only in enterocytes, but also in blood serum. An increase in the level of malonic dialdehyde leads to a decrease in the antioxidation capacity and in the peroxidation resistance of erythrocytic membranes. The severity of the pathological process was found related to the activity of lipid peroxidation.     

Correction of lipid peroxidation disorders in experimental traumatic shock using the antioxidant ionol

The level of certain parameters of lipid peroxidation and the activity of lysosome hydrolases were studied on the shock model in rats. It was established that the traumatic shock in the experiment is accompanied by the growth of the level of over-oxidation products (malonic dialdehyde, diene conjugates), the rate of erythrocyte hemolysis as well as by an increase in the hydrolase activity. Administration of ionol (60 mg/kg) inhibits the higher activity of radical-free lipid oxidation, decreases the damage of membrane structure and the metabolism disturbance.     

Influence of whole-body gamma-irradiation upon rat erythrocyte: lipid peroxidation and osmotic fragility

The effects of whole-body gamma-irradiation of rats (8 Gy) on erythrocyte enzymes and biochemical components involved in lipid peroxidation were studied. Decreased superoxide dismutase and glutathione reductase activities, and lowered concentrations of reduced glutathione, were found to be the main factors responsible for the observed increase in lipid peroxidation in the erythrocytes of irradiated rats. This increased lipid peroxidation did not result in a greater tendency to hemolysis in hypotonic media; on the contrary, the mean osmotic fragility was decreased at days D + 1 and D + 3 after irradiation. The behavior of the erythrocyte populations towards hemolysis in hypotonic media appeared to be most homogeneous at days D + 4 and D + 8 after irradiation, which correspond to maxima of malonic dialdehyde concentrations in erythrocytes. Such a synchrony of variations suggests that crosslinking of primary amino groups of proteins or phospholipids by malonic dialdehyde might produce a rigidification in erythrocyte membranes, possibly leading to a more homogeneous behavior of the erythrocyte populations towards hemolysis in hypotonic media.     

Target enzymes on hepatic dysfunction caused by dietary products of lipid peroxidation

Dietary products of lipid peroxidation cause hepatic dysfunction due to decreases in the activities of some hepatic enzymes and to depletion of CoA. An idea about the decreases and depletion is that the enzymes and CoA could be injured directly by the incorporated products in the liver. Their inactivations in vitro were then examined using a reasonable amount of peroxidation products. The hepatic cytosol, microsomes, and mitochondria were incubated with 10, 15, and 20 micrograms/mg protein of peroxidation products, respectively, and changes in the enzymatic activities were monitored. Glucose-6-phosphate dehydrogenase, mitochondrial NAD-dependent aldehyde dehydrogenase, glucokinase, and glyceradehyde phosphate dehydrogenase were inactivated, and the CoA level was decreased, but the other hepatic enzymes were not. Although glyceraldehyde phosphate dehydrogenase was most sensitive to peroxidation products in vitro, the decrease in activity was not detected by the oral dose of secondary products. On the other hand, among the components of peroxidation products, hydroperoxides and polymers are not incorporated in the liver, but decomposed products of low molecular weight are incorporated. Glucokinase among the above enzymes was not inactivated by the low-molecular-weight products. It was therefore concluded that glucose-6-phosphate dehydrogenase, mitochondrial NAD-dependent aldehyde dehydrogenase, and CoA were targets of the direct attack by incorporated components of peroxidation products in the liver.     

Changes in energy metabolism and lesion of nerve cells in rats after repeated administrations of high doses of insulin

In the brain of rats exposed to 5–7 hypoglycemic comas, at the 2nd day after the last coma, an increase of NADP-isocitrate dehydrogenase activity and acceleration of catabolism of adenyl nucleotides as well as a decrease of activities of NADH-dehydrogenase, mitochondrial NADP-isocitrate dehydrogenase, glucose-6-phosphate dehydrogenase, glutathione reductase, and superoxide dismutase were found, whereas no changes of the rate of glycolysis were revealed. After placing sections of brain large hemispheres from experimental animals into hypoosmotic medium supplied with Fe²⁺ and ascorbate, the release of lactate dehydrogenase was increased. A considerable increase of concentration of malonic dialdehyde is observed in brain sections of experimental rats. The obtained results indicate that disturbances of energy metabolism and activation of processes of lipid peroxidation are involved in pathogenesis of post-hypoglycemic encephalopathy.     

Evidence for mitochondrial DNA damage by lipid peroxidation

When mitochondria of rat liver were incubated in an in vitro system containing NADPH and ferrous chloride, marked lipid peroxidation occurred, as evidenced by the evolution of malonic dialdehyde. DNA isolated from these peroxidized mitochondrial preparations had completely different electrophoretic mobility than DNA isolated from mitochondria protected from peroxidation. Scavengers of superoxide anion, hydrogen peroxide and hydroxyl radicals offered no protection against either lipid peroxidation or DNA damage. However, alpha-tocopherol protected against both lipid peroxidation and damage to the mitochondrial genome. These results support the hypothesis that lipid peroxidation can mediate DNA damage.