Copper-induced LDL peroxidation: interrelated dependencies of the kinetics on the concentrations of copper, hydroperoxides and tocopherol (original) (raw)
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Prooxidant role of vitamin E in copper induced lipid peroxidation
FEBS Letters, 1993
When exposed to Ct?', a-tocopherol, in detergent dispersion, is rapidly oxidised. Moreover, if phospholipids and traces of their hydroperoxide derivatives are included in these dispersions, Cu" initiates lipid peroxidation, the rate of which is dramatically stimulated by a-tocopherol. The observation that the rate of a-tocopherol consumption is identical in the absence and in the presence of lipids undergoing peroxidation, apparently rules out any antioxidant effect. These results are consistent with a prooxidant effect of vitamin E, mediated by its capability to reduce Cu*+ to Cu' which, in turn, produces, from lipid hydroperoxides, the highly reactive alkoxyl radicals. Present data highlight the risk of misleading results in interpreting the significance of lags in peroxidation of LDL challenged with Cu*'. Vitamin E; a-Tocopherol; Copper; Phospholipid hydroperoxide; Low density lipoprotein; Atherosclerosis Published by Elsevier Science Publishers B. F!
Archives of Biochemistry and Biophysics, 2001
The mechanisms by which low-density lipoprotein (LDL) particles undergo oxidative modification to an atherogenic form that is taken up by the macrophage scavenger-receptor pathway have been the subject of extensive research for almost two decades. The most common method for the initiation of LDL oxidation in vitro involves incubation with Cu(II) ions. Although various mechanisms have been proposed to explain the ability of Cu(II) to promote LDL modification, the precise reactions involved in initiating the process remain a matter of contention in the literature. This review provides a critical overview and evaluation of the current theories describing the interactions of copper with the LDL particle. Following discussion of the thermodynamics of reactions dependent upon the decomposition of preexisting lipid hydroperoxides, which are present in all crude LDL preparations, attention is turned to the more difficult (but perhaps more physiologically-relevant) system of the hydroperoxide-free LDL particle. In both systems, the key role of ␣-tocopherol is discussed. In addition to its protective, radical-scavenging action, ␣-tocopherol can also behave as a prooxidant via its reduction of Cu(II) to Cu(I). Generation of Cu(I) greatly facilitates the decomposition of lipid hydroperoxides to chaincarrying radicals, but the mechanisms by which the vitamin promotes LDL oxidation in the absence of preformed hydroperoxides remain more speculative. In addition to the so-called tocopherol-mediated peroxidation model, in which polyunsaturated fatty acid oxidation is initiated by the ␣-tocopheroxyl radical (generated during the reduction of Cu(II) by ␣-tocopherol), an evaluation of the role of the hydroxyl radical is provided. Important interactions between copper ions and thiols are also discussed, particularly in the context of cell-mediated LDL oxidation. Finally, the mechanisms by which ceruloplasmin, a copper-containing plasma protein, can bring about LDL modification are discussed. Improved understanding of the mechanisms of LDL oxidation by copper ions should facilitate the establishment of any physiological role of the metal in LDL modification. It will also assist in the interpretation of studies in which copper systems of LDL oxidation are used in vitro to evaluate potential antioxidants.
Lipids, 1999
As a first step in evaluating the significance of our recently developed method of monitoring the kinetics of copper-induced oxidation in unfractionated serum, we recorded the kinetics of lipid oxidation in the sera of 62 hyperlipidemic patients and analyzed the correlation between oxidation and lipid composition of the sera [high density lipoprotein (HDL) cholesterol, low density lipoprotein (LDL) cholesterol, and triglycerides]. We used six factors to characterize the kinetics of oxidation, namely, the maximal absorbance of oxidation products (OD max ), the maximal rate of their production (V max ), and the time at which the rate was maximal (t max ) at two wavelengths (245 nm, where 7-ketocholesterol and conjugated dienic hydroperoxides absorb intensely, and 268 nm, where the absorbance is mostly due to dienals). The major conclusions of our analyses are that: (i) Both OD max and V max correlate positively with the sum of concentrations of the major oxidizable lipids, cholesterol, and cholesteryl esters. (ii). The value of t max , which is a measure of the lag preceding oxidation and therefore reflects the resistance of the serum lipids to copper-induced oxidation, exhibits a negative correlation with HDL cholesterol. Although this finding accords with the observation of shorter lags for HDL than for LDL, it is apparently inconsistent with the role of HDL as an antirisk factor in coronary heart diseases. Abbreviations: HDL, high density lipoprotein; LDL, low density lipoprotein; OD max , maximal absorbance of oxidation products; PAF-AH, platelet-activating factor-acetyl hydrolase; PUFA, polyunsaturated fatty acids; t max , time at which the rate of oxidation was maximal; TG, triglycerides; V max , maximal rate of oxidation products accumulation.
Kinetic analysis of copper-induced peroxidation of LDL
Biochimica et biophysica acta, 1998
We have employed our recently developed spectroscopic method of continuous monitoring of lipid oxidation to study the formation and decomposition of hydroperoxides in the time course of LDL oxidation. The results show satisfactory agreement with simulated time courses based on the following assumptions: (a) Both the rates of formation and decomposition of hydroperoxides depend on the ratio of bound copper to LDL as computed under the assumption that each LDL particle has 17 equivalent copper binding sites characterized by a dissociation constant K = 1 microM. (b) Peroxidation is initiated by copper-catalyzed decomposition of hydroperoxides (LOOH) into peroxy radicals (LOO.) and other products, including dienals. Under these assumptions, the rate of accumulation of LOOH can be computed from the equation (equation in text). The agreement between the simulated and experimentally-observed kinetics supports the assumptions used for simulations. The close agreement between the values of l...
Copper-induced LDL peroxidation investigated by 1H-NMR spectroscopy
1995
Oxidatively modified LDL (oLDL) is thought to play a key role in the pathogenesis of atherosclerosis. We have studied Cu(2+)-induced peroxidation reactions of LDL and have elucidated the sequence of events which subsequently occur within LDL particles by 1H-NMR spectroscopy. Studies of chloroform/methanol extracts show that LDL arachidonate is oxidised by Cu2+ at a higher rate and to a greater extent than linoleate, giving isomeric hydroperoxides with predominantly trans,trans double-bonds, whilst only cis,trans isomers were detected as intrinsic hydroperoxides in control LDL samples. These intrinsic hydroperoxides were not degraded during peroxidation, suggesting that they are not involved in the initiation of Cu(2+)-induced peroxidation. Aldehydes arising from the decomposition of hydroperoxides were also detected, as well as saturated fatty acids which were released into the external aqueous medium. Decomposition pathways of the two major isomeric hydroperoxides are discussed. Cu(2+)-induced oxidation of LDL cholesterol appears to occur only after hydroperoxide breakdown, with esterified cholesterol being oxidised to a greater extent than free cholesterol. Phospholipid hydrolysis appeared to parallel the peroxidation of arachidonic acid, and the released lysophosphatidylcholine may become associated with apoB. These results suggest that hydroperoxide breakdown (probably in phospholipids) may be a key event in the peroxidation process, leading to the oxidation of cholesterol and propagation into the core of LDL.
Journal of Biological Chemistry
Initiation of lipid peroxidation by Cu(II) requires reduction of Cu(II) to Cu(I) as a first step. It is unclear, however, whether this reaction occurs in the course of lipoprotein oxidation. It is also unknown which reductant, if any, can drive the reduction of Cu(II) in this case. We found that Cu(II) was rapidly reduced to Cu(I) by all major human lipoproteins (high, low, and very low density lipoproteins (HDL, LDL, and VLDL), and chylomicrons). Cu(II)-reducing activity was associated with a lipid moiety of the lipoproteins. The rates of Cu(II) reduction by different lipoproteins were similar when the lipoproteins were adjusted to similar ␣-tocopherol concentrations. Enriching lipoproteins with ␣-tocopherol considerably increased the rate of Cu(II) reduction. Cu(II) reduction by ␣-tocopherol-deficient LDL isolated from a patient with familial inherited vitamin E deficiency was found to occur much slower in comparison with LDL isolated from a donor with a normal plasma level of ␣-tocopherol. Initial rate of Cu(II) reduction by ␣-tocopherol-deficient LDL was found to be zero. Enriching LDL with ubiquinol-10 to concentrations close to those of ␣-tocopherol did not influence the reaction rate. When LDL was treated with ebselen to eliminate preformed lipid hydroperoxides, the reaction rate was also not changed significantly. Cu(II) reduction was accompanied by a consumption of lipoprotein ␣-tocopherol and accumulation of conjugated dienes in the samples. Increasing ␣-tocopherol content in lipoproteins slightly decreased the rate of conjugated diene accumulation in LDL and HDL and considerably increased it in VLDL. The results suggest that ␣-tocopherol plays a triggering role in the lipoprotein oxidation by Cu(II), providing its initial step as follows: ␣TocH ؉ Cu(II) 3 ␣Toc ⅐ ؉ Cu(I) ؉ H ؉ . This reaction appears to diminish or totally eliminate the antioxidative activity of ␣-tocopherol in the course of lipoprotein oxidation.
The Role of Copper Reduction by α-Tocopherol in Low-Density Lipoprotein Oxidation
Free Radical Biology and Medicine, 1997
The oxidation of lipoproteins is thought to be an important early step in atherogenesis. The measurement of lipid peroxidation in low-density lipoprotein (LDL) challenged with Cu 2/ has become a widespread test to determine the ''susceptibility'' of LDL to oxidation. The determination of lag time to oxidation is thought to be a measure of the total antioxidant capacity of the LDL. However, we and others have failed to observe any correlation between lag time and the LDL content of its major lipid antioxidant, a-tocopherol. In fact, several studies now suggest a pro-oxidant role for tocopherol under some conditions of LDL oxidation. In the present study we sought to determine if there was a relationship between Cu 2/ reduction by LDL and kinetic parameters of LDL oxidation. LDL (0.3 mmol/l cholesterol, Ç0.1 mg protein/ml) was incubated at 30ЊC with 2 mM Cu 2/ and the formation of conjugated dienes measured over a 4-h period. Using neocuproine, an indicator molecule that specifically complexes Cu / but not Cu 2/ , the reduction of Cu 2/ by LDL was monitored. The final Cu concentration in these assays was 100 mM and neocuproine 750 mM. Cu / formation was measured by absorbance at 454 nm. A strong negative correlation was observed between copper reduction by LDL and lag time to oxidation (r Å 00.66, p õ .005, n Å 16). Further experiments showed that (1) LDL was able to reduce Cu 2/ to Cu / in a time and concentration-dependent manner; (2) blocking of free -SH groups on LDL apoprotein B by preincubation with dithionitrobenzoic acid (DTNB) had no significant effect on the rate and extent of Cu 2/ reduction; (3) consumption of tocopherol in LDL undergoing oxidation with Cu was very rapid (rate Å 6 1 10 010 M s 01 ). When Cu / formed during incubation with LDL was complexed with neocuproine, there was significant inhibition of LDL oxidation, as indicated by lipid peroxide formation and mobility on agarose gel electrophoresis. Surprisingly, tocopherol consumption was even more rapid in the presence of neocuproine, consistent with a shift in Cu 2/ / Cu / equilibrium and faster reduction of Cu 2/ by a-tocopherol. These results indicate that under these conditions tocopherol is a major reducing agent in LDL, converting Cu 2/ to Cu / , and therefore, may play an important role in promoting LDL oxidation. However, there was no correlation between LDL tocopherol content and reduction of Cu 2/ . Examination of the time course of Cu 2/ reduction in tocopherol enriched and depleted LDL indicates that tocopherol may determine Cu reduction at early time points but that the eventual capacity of LDL to reduce Cu may depend on more complex interactions between tocopherol and other LDL components. ᭧ 1997 Elsevier Science Inc.
1997
To determine whether lipid peroxidation is required (LDL) 2 are likely to be the main source of cholesterol for apolipoprotein B (apoB) carbonyl formation of huthat accumulates in arteriosclerotic plaques (1-3). man low-density lipoproteins (LDL) during copper-Therefore, the mechanisms that result in LDL oxidamediated oxidation, we investigated oxidation of nation and protein carbonyl formation are of interest. tive and probucol-preloaded LDL by measuring thio-Trace amounts of copper can induce LDL oxidation barbituric acid-reactive substances (TBARS) and apoB (4), resulting in highly reproducible LDL oxidative carbonyls. Probucol was used because it is known to damage (5, 6). This process leads to oxidized LDL that inhibit lipid peroxidation, but not protein modificashares many structural and functional properties with tion. During copper-mediated oxidation, apoB carbon-LDL oxidized by cells or LDL extracted from arterial yls formed in a time-dependent manner; high copper atherosclerotic plaques (8). Moreover, this process can concentrations ( §30 mM) resulted in saturation of mimic the in vivo situation because copper ions are apoB carbonyl content. ApoB carbonyl formation and present in the arterial wall and plaque extracts have lipid peroxidation were linearly related during incubeen shown to oxidize LDL in vitro (9). bation of LDL with copper for 3 h. During Cu 2/ -medi-Probucol, 4,4-(isopropylidenedithio)bis[2,6-di-tertated LDL oxidation of probucol-LDL, TBARS producbutylphenol], being structurally similar to butylated tion was very low, nonetheless apoB carbonyls inhydroxytoluene (10), is a lipid-soluble antioxidant. It creased significantly, and vitamin E was depleted.