Evidence for carbon tetrachloride-induced lipid peroxidation in vivo by 23Na relaxation times t1

We report here the results of a study performed on the livers of rats intoxicated with CCl4 in order to investigate the dependence of 23Na n.m.r. parameters on specific cell alterations. In this connection, CCl4 represents a useful probe, since the hepatocellular damaging effects induced by this aliphatic halocarbon have been extensively investigated. The t1relaxationtimes of intracellular sodium ion under physiological conditions in the presence of the paramagnetic shift reagent dysprosium tripolyphosphate were investigated. The significant increase of t1 in the livers of rats intoxicated with CCl4 with respect to the t1relaxationtimes of normal rats was studied. Evidence is given that neither liver cell necrosis nor fat accumulation nor proliferative processes affected the observed t1 lengthening. However, when t1relaxationtimes were measured in the livers of vitamin E-treated rats subsequently intoxicated with CCl4 a significant shortening of t1 with respect to the times measured in intoxicated rats was observed. We report the results of the present study, taking into consideration that peroxidation of microsomal lipids is the key factor in the CCl4-induced liver injury process and that vitamin E exerts an antioxidant action against CCl4-induced microsomal lipidperoxidation

NMR determination of site specific deuterium distribution in squalene from different sources

The H-2 NMR spectra of squalene from shark liver oil and olive oil sources were investigated by the SNIF-NMR technique. The areas of the absorption signals of the olefinic, methylene and methyl sites were carefully determined, and the deuterium distribution among the various sites was shown to depart from the statistical distribution in a way that is different for squalene from different sources. The results are discussed in connection with the different biosynthetic pathways of squalene involved in plants and animals

Oxidative stability of polyunsaturated fatty acids: effect of squalene

The propensity of polyunsaturated fatty acids (PUFAs) to undergo oxidation plays an important role in the integrity of biological membrane and lipid containing foods, The ability of squalene (SQ), a naturally occurring dehydrotriterpene present in animal and plant tissues, to protect linoleic, linolenic, arachidonic and docosahexaenoic acids against temperature-dependent autoxidation and UVA (ultraviolet A, 320-880 nm) mediated oxidation was assessed. The oxidation of PUFAs was protected in varying degrees, with highest protection observed for linolenic, arachidonic and docosahexaenoic acids. Linoleic acid was less protected. At a molar ratio of 7:1 (PUFA:SQ) the inhibition of the oxidation process was 22% in the presence of linoleic acid and about 50% in presence of the other PUFAs tested. The different protection exerted by SQ against PUFAs with different degrees of unsaturation may be accounted for by the higher stability of octadecadienoic acid hydroperoxide isomers compared with respective PUFA hydroperoxides. Observing mild UVA-mediated oxidation and the temperature-dependent autoxidation reactions we found similarities in the oxidation pattern and the protection exerted by SQ. These findings suggest that the reaction of autoxidation is predominant and SQ acts mainly as peroxyl radical scavenger

Methyl mercury enhances [3H]diazepam binding in different areas of the rat brain.

Three days after the acute oral administration of methyl mercury (MeHg), a 27-60% increase in the total number of binding sites for [3H]diazepam was seen in the retina and different areas of the rat brain, with no change, except in the retina, in the apparent dissociation constant for its ligand. In contrast, MeHg failed to change [3H]spiroperidol and [3H]GABA binding in the same areas. Moreover, MeHg decreased cyclic GMP content in the cerebellar cortex. The various possible mechanisms involved in the action of MeHg on benzodiazepine binding are discussed

Non-random peroxidation of different classes of membrane phospholipids in live cells detected by metabolically integrated cis-parinaric acid

Quantitative assays of lipid peroxidation in intact, living cells are essential for evaluating oxidative damage from various sources and for testing the efficacy of antioxidant interventions. We report a novel method based on the use of cis-parinaric acid (PnA) as a reporter molecule for membrane lipid peroxidation in intact mammalian cells. Using four different cell lines (human leukemia HL-60, K562 and K/VP.5 cells, and Chinese hamster ovary (CHO) fibroblasts), we developed a technique to metabolically integrate PnA into all major classes of membrane phospholipids, i.e., phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol and cardiolipin, that can be quantified by HPLC with fluorescence detection. Integrated PnA constituted less than 1% of lipid fatty acid residues, suggesting that membrane structure and characteristics were not significantly altered. Low concentrations (20-40 mu M) of tert-butyl hydroperoxide (t-BuOOH) caused selective oxidation of PnA residues in phosphatidylserine and phosphatidylethanolamine of K562 cells and K/VP.5 cells while cell viability was unaffected. At higher t-BuOOH concentrations (exceeding 100 mu M), however, a progressive, random oxidation of all major phospholipid classes occurred and was accompanied by significant cell death. In HL-60 cells, phosphatidylethanolamine, phosphatidylserine and cardiolipin were sensitive to low concentrations of t-BuOOH, while phosphatidylcholine and phosphatidylinositol were not affected. Phosphatidylinositol was the only phospholipid that responded to the low concentrations of t-BuOOH in CHO cells, At high t-BuOOH concentrations, again, all phospholipid classes underwent extensive oxidation, All phospholipids were nearly equally affected by peroxidation induced by a initiator of peroxyl radicals, 2,2'-azobis-(2,4-dimethylvaleronitrile) (AMVN), in K562 cells. In gamma-irradiated (4-128 Gy) CHO cells, phosphatidylserine was the most affected phospholipid class (34% peroxidation) followed by phosphatidylinositol (24% peroxidation) while the other three phospholipid classes were apparently unaffected, Since loss of PnA fluorescence is a direct result of irreparable oxidative loss of its conjugated double bond system, the method described allows for selective and sensitive monitoring of oxidative stress in live cells without interference from cell repair mechanisms