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H Esterbauer

Publications and source records attributed to H Esterbauer.

At least 163 records · Page 9Linked to original sources
PubMed1987

Autoxidation of human low density lipoprotein: loss of polyunsaturated fatty acids and vitamin E and generation of aldehydes.

The alteration of structural and biological properties of human plasma low density lipoprotein (LDL) exposed to oxidative conditions is in part ascribed to lipid peroxidation. The objective of this investigation was to measure quantitatively several parameters in oxidizing LDL indicative for lipid peroxidation. Exposure of freshly prepared EDTA-free LDL to an oxygen-saturated buffer led to a complete depletion of alpha- and gamma-tocopherol within 6 hr, thereafter lipid peroxidation commenced as indicated by the kinetics of the loss of linoleic (18:2) and arachidonic (20:4) acids, the formation of aldehydic lipid peroxidation products and fluorescent apoB. Within 24 hr of oxidation, on average 79 nmol of 18:2 (initial 345) and 12.8 nmol of 20.4 (initial 25.6) were oxidized per mg of LDL and the sample contained in total 7.1 nmol of aldehydes with the following molar distribution: 36.6% malonaldehyde, 25% hexanal, 8.9% propanal, 8.2% 4-hydroxynonenal, 7.6% butanal, 4.1% 2.4-heptadienal, 3.4% pentanal, 3.4% 4-hydroxyhexenal, and 2.5% 4-hydroxyoctenal. Malonaldehyde was predominantly (93%) in the aqueous phase, whereas the other aldehydes remained mostly (34-98%) within the LDL particle, where the total aldehyde concentration was in the range of 12 mM. Oxidized LDL exhibited a 1.6-fold enhanced electrophoretic mobility. Similarily, native LDL incubated for 5 hr with aldehydes showed increased electrophoretic mobility. At equal concentrations (5 mM) 4-hydroxynonenal was most effective, followed by 2,4-heptadienal, hexanal, and malonaldehyde. This study reports for the first time the rate and extent of the change of LDL constituents occurring during lipid peroxidation.

Aldehydes↗
PubMed1987

Possible role of aldehydic lipid peroxidation products as chemoattractants.

Previous studies showed that the lipid peroxidation product 4-hydroxy-trans-2-nonenal (HNE) stimulates the rat neutrophil oriented migration in vitro within a micromolar range. Its effect is weak with respect to other known chemoattractants, but highly significant on the basis of a two-way analysis of variance. Other hydroxyalkenals were found to be chemotactic within a pico-micromolar range, and their chemotactic power was not correlated to their lipophilicity. The structural requirements for the chemotactic activity of the hydroxyalkenals were studied by testing the influence of alkanals and 2-alkenals on oriented migration. Alkanals are lacking both the trans double-bond and the hydroxy group, while 2-alkenals are lacking only the hydroxy group. The 2-alkenals (2-octenal, 2-nonenal) were found to be chemotactic, whereas alkanals (hexanal, octanal, nonanal) were ineffective. Therefore the chemotactic activity of the aliphatic aldehydes is dependent on the -C = C-CHO part of their molecule. The possibility that unsaturated aliphatic aldehydes are present in an inflammatory site at a concentration at which they are chemotactic in vitro was also investigated. Carbonyls in pleural exudates were analysed at different times after a pleurisy induction and HNE was detected both in the cells and in the cell-free supernant of the exudate at increasing concentrations during the 4 hours of the experiment. The exact source of HNE is unknown, but since HNE has been identified among the degradation products of peroxidized lipids it is likely that this aldehyde is formed consequent on lipid peroxidation reactions which occur at the phlogistic site. The possibility that HNE is involved, at least in part, in the recruitment of neutrophils in the inflammatory area is suggested.

Aldehydes↗
PubMed1986

Possible involvement of the lipid-peroxidation product 4-hydroxynonenal in the formation of fluorescent chromolipids.

The effects of the lipid-peroxidation product 4-hydroxynonenal on the formation of fluorescent chromolipids from microsomes, mitochondria and phospholipids were studied. Incubation of freshly prepared rat liver microsomes or mitochondria with 4-hydroxynonenal results in a slow formation of a fluorophore with an excitation maximum at 360 nm and an emission maximum at 430 nm. The rate and extent of the development of the 430 nm fluorescence can be significantly enhanced by ADP-iron (Fe3+). With microsomes, yet not with mitochondria. NADPH has a catalytic effect similar to that of ADP-iron. Fluorescent chromolipids with maximum excitation and emission at 360/430 nm are also formed during the NADPH-linked ADP-iron-stimulated lipid peroxidation. Phosphatidylethanolamine and phosphatidylserine react with 4-hydroxynonenal revealing a fluorophore with the same spectral characteristics as that obtained in the microsomal and mitochondrial system. The findings suggest that the fluorescent chromolipids formed by lipid peroxidation are not derived from malonaldehyde, but are formed from 4-hydroxynonenal or similar reactive aldehydes via a NADPH and/or ADP-iron-catalysed reaction with phosphatidylethanolamine and phosphatidylserine contained in the membrane.

Adenosine Diphosphate↗
PubMed1986

Resistance to oxidative stress by hyperplastic and neoplastic rat liver tissue monitored in terms of production of unpolar and medium polar carbonyls.

The susceptibility of rat liver tissue to oxidative stress during its neoplastic transformation was analyzed by both qualitative and quantitative measurements of the carbonyl products of lipid peroxidation. Diethylnitrosamine was used as initiating agent of hepatocarcinogenesis and lipid peroxidation levels were monitored in the homogenates from normal liver, hyperplastic nodules and tumour, incubated in the presence or in the absence of ascorbate or adenosine diphosphate-iron complex. While the basal levels of lipid peroxidation in the three experimental conditions were found to be quite similar, in the presence of the pro-oxidant stimulus a remarkable reduction in aldehyde production was shown not only by the hepatoma tissue but also by the preneoplastic nodules.

Aldehydes↗
PubMed1986

Investigation of human plasma low density lipoprotein by three-dimensional fluorescence spectroscopy.

Human plasma LDL exhibits a diffuse fluorescence (excitation 360 nm) in the 400-600 nm range. Application of three-dimensional fluorescence spectroscopy shows the presence of 7 fluorophores in the lipid and 6 fluorophores in the protein domain. The 430 nm fluorescence in freshly prepared LDL and its apo-B is most likely indicative for remnants of in vivo lipid peroxidation.

Apolipoproteins B↗
PubMed1986

Studies on the mechanism of formation of 4-hydroxynonenal during microsomal lipid peroxidation.

The mechanism of the formation of 4-hydroxynonenal through the NADPH-linked microsomal lipid peroxidation was investigated. The results were as follows: 4-hydroxynonenal arises exclusively from arachidonic acid contained in the polar phospholipids, neither arachidonic acid of the neutral lipids nor linoleic acid of the polar or neutral lipids are substrates for 4-hydroxynonenal generation. This finding results from the estimation of the specific radioactivity of 4-hydroxynonenal produced by microsomes prelabelled in vivo with [U-14C]arachidonic acid. Phospholipid-bound 15-hydroperoxyarachidonic acid would have the structural requirements needed for 4-hydroxynonenal (CH3-(CH2)4-CH(OH)-CH=CH-CHO). Microsomes supplemented with 15-hydroperoxyarachidonic acid and NADPH, ADP/iron converted only minimal amounts (0.6 mol%) of 15-hydroperoxyarachidonic acid into 4-hydroxynonenal; similarly, 15-hydroperoxyarachidonic acid incubated at pH 7.4 in the presence of ascorbate/iron yielded only small amounts of 4-hydroxynonenal with a rate orders of magnitude below that observed with microsomes. Phospholipid-bound 15-hydroperoxyarachidonic acid is therefore not a likely intermediate in the reaction pathway leading to 4-hydroxynonenal. The rate of 4-hydroxynonenal formation is highest during the very initial phase of its formation and the onset does not show a lag phase, suggesting a transient intermediate predominantly formed during the early phase of microsomal lipid peroxidation. After 60 min of incubation, 204 nmol polyunsaturated fatty acids (20 nmol 18:2, 143 nmol 20:4, 41 nmol 22:6) were lost per mg microsomal protein and the incubation mixture contained 206 nmol lipid peroxides, 71.6 nmol malonic dialdehyde and 4.6 nmol 4-hydroxynonenal per mg protein. Under artificial conditions (pH 1.0, ascorbate/iron, 20 h of incubation) not comparable to the microsomal peroxidation system, 15-hydroperoxyarachidonic acid can be decomposed in good yields (15 mol%) into 4-hydroxynonenal. Autoxidation of arachidonic acid in the presence of ascorbate/iron gave after 25 h of incubation 2.8 mol% (pH 7.4) and 1.5 mol% (pH 1.0) 4-hydroxynonenal. The most remarkable difference between the non-enzymic system and the enzymic microsomal system is that the latter forms 4-hydroxynonenal at a much higher rate.

Aldehydes↗
PubMed1986

Role of cardiac glutathione transferase and of the glutathione S-conjugate export system in biotransformation of 4-hydroxynonenal in the heart.

There is a remarkable difference in the isozyme pattern between cardiac and hepatic glutathione S-transferases in rat (Ishikawa, T., and Sies, H. (1984) FEBS Lett. 169, 156-160), and one near-neutral isozyme (pI = 6.9) of the cardiac glutathione S-transferases was found to have a significantly high activity toward 4-hydroxynonenal. The isozyme was inhibited by the resulting glutathione S-conjugate of 4-hydroxynonenal competitively with GSH and noncompetitively with 4-hydroxynonenal. The kinetic parameters estimated for the isozyme were: kcat = 460 mol X min-1 X mol enzyme-1, Km = 50 microM for 4-hydroxynonenal, Ki = 85 microM. When the heart was perfused with 4-hydroxynonenal, a marked decrease was observed in the intracellular GSH level, accompanied by an increase of glutathione S-conjugate of 4-hydroxynonenal in the heart. The rate of the conjugation reaction was more than 30 times the rate of the spontaneous reaction, the half-life of 4-hydroxynonenal in the heart being less than 4 s. The glutathione S-conjugate of 4-hydroxynonenal was released from the heart into the perfusion medium. Saturation kinetics were observed for the release with respect to the intracellular level of the S-conjugate (Vmax = 12 nmol X min-1 X g heart-1), and there was a competition by the S-conjugate for GSSG release. The release of the glutathione S-conjugate is considered as a carrier-mediated process and to be important not only in interorgan glutathione metabolism but also in diminishing the inhibitory effect of the S-conjugate on glutathione S-transferases and glutathione reductase.

Aldehydes↗
PubMed1986

Modification of human low-density lipoprotein by the lipid peroxidation product 4-hydroxynonenal.

The effects of the lipid peroxidation product 4-hydroxynonenal on freshly prepared human low-density lipoprotein (LDL) were studied. At a fixed LDL concentration (5.7 mg/ml) the amount of 4-hydroxynonenal incorporated into the LDL increased with increasing aldehyde concentration from 28-30 (0.2 mM) to 140 (1 mM) mol per mol LDL, whereas at a fixed aldehyde concentration (0.2 mM) its incorporation into LDL decreased with increasing LDL concentration from 48 (1 mg LDL/ml) to 26 (12 mg LDL/ml) mol 4-hydroxynonenal bound per mol LDL. Of the total hydroxynonenal taken up 78% was bound to the protein and 21% to the lipid moiety; the remaining 1% was dissolved as free aldehyde in the lipid fraction. Amino acid analysis of the apolipoprotein B revealed that 4-hydroxynonenal attacks mainly the lysine and tyrosine residues and to a lesser extent also serine, histidine and cysteine. Treatment of LDL with 4-hydroxynonenal results in a concentration-dependent increase of the negative charge of the LDL particle as evidenced by its increased electrophoretic mobility. Moreover, 4-hydroxynonenal treatment leads to a partial conversion of the apolipoprotein B-100 into higher molecular weight forms most probably apolipoproteins B-126 and B-151. Compared to malonaldehyde, 4-hydroxynonenal exhibits a much higher capacity to modify LDL and it is therefore believed that this aldehyde is a more likely candidate for being responsible for LDL modification under in vivo lipid peroxidation conditions.

Aldehydes↗
PubMed1986

Preparation of 18O-labelled standards of hydroxy-eicosatetraenoic acids and thromboxanes for quantitative measurement by gas chromatography/mass spectrometry.

A study on the extent of 18O incorporation into various hydroxy-eicosatetraenoic acids, thromboxane B2 and 2,3-dinor-thromboxane B2 by base-catalysed hydrolysis of their methyl ester derivatives is presented. The amount as well as the position of the inbuilt 18O atoms were determined by gas chromatography/mass spectrometry, utilizing electron impact and positive and negative ion chemical ionization.

Gas Chromatography-Mass Spectrometry↗
PubMed1986

Relationships between respiratory activity and water-soluble sulfhydryl content of Norway spruce needles.

The rate of respiration of spruce needles was affected by the level of foliar glutathione. At 37 degrees C the rate of respiration increased with increasing water-soluble sulfhydryl content up to 1.3 micromol g(-1) fresh weight. Higher thiol concentrations inhibited the rate of respiration. At 25 degrees C no relationship was observed between glutathione content and rate of respiration. It was concluded that the mechanism by which SO(2) affects plants is temperature dependent.

Journal Article↗
PubMed1986

New data on kinetics of lipid peroxidation in experimental hepatomas and preneoplastic nodules.

Lipid peroxidation has been found decreased in several hepatomas. The decline has been shown already at the level of preneoplastic nodules obtained after DEN treatment of rats. A substantial exception is represented by the hepatoma cell line MH1C1, deriving from a slightly deviated Morris tumor. Most of the described experiments estimated lipid peroxidation levels in terms of malonaldehyde production by the thiobarbituric acid test. It is now clear that this test does not account for several other aldehydes produced during lipid peroxidation. We now investigated by high performance liquid chromatography (HPLC) the whole range of non-polar aldehydes produced by tumor homogenates and by preneoplastic nodules both in basal conditions and after stimulation with ADP-iron or ascorbate. It was reduced in the preneoplastic nodules as well as in the DEN-induced hepatoma. The susceptibility to the prooxidant effect of ADP-iron or ascorbate was strongly decreased in all hepatomas as well as in preneoplastic nodules. It has been recently published that hepatoma cells are more susceptible than normal liver to the toxic action of aldehydes. This was attributed at least in part to the decreased activity of aldehyde dehydrogenases, as well as to their different distribution in tumor cells. A deeper study on aldehyde metabolism in hepatomas has shown that alcohol dehydrogenase and NADPH-aldehyde reductase also are markedly decreased in Yoshida hepatoma cells and the MH1C1 cell line. However, glutathione transferase, that can use hydroxynonenal as a substrate, is strongly decreased in Yoshida hepatoma cells but not in MH1C1 cells.

Alcohol Dehydrogenase↗
PubMed1986

Chemotactic activity of the lipid peroxidation product 4-hydroxynonenal and homologous hydroxyalkenals.

The effect of the lipid peroxidation product 4-hydroxynonenal and homologous aldehydes (4-hydroxyoctenal, 4-hydroxyundecenal, 4-hydroxytetradecenal and 4-hydroxypentadecenal) on migration and polarization of rat neutrophils was examined. The most effective aldehydes were 4-hydroxyoctenal and 4-hydroxypentadecenal, which stimulated oriented migration at ED50 = 1.4 X 10(-12) M and 1.3 X 10(-12) M, resp., whereas the other aldehydes had ED50 between 1 X 10(-7) and 6 X 10(-11) M. The peptides fMet-Phe and fMet-Leu-Phe used as positive controls had ED50 values of 4.2 X 10(-7) M and 4.5 X 10(-10) M resp. The 4-hydroxyalkenals induced only a small increase of the percentage of polarized cell and did not enhance the random migration. The effects of 4-hydroxyalkenals were only observed when the incubation buffer contained bovine serum albumin (BSA), in the absence of BSA neither the aldehydes nor the peptides exhibited chemotactic properties. Since the aldehydes easily react with the sulfhydryl groups of the BSA to form the S-alkylated BSA in an equilibrium reaction, the chemotactic substance could either be the free aldehyde or the BSA-aldehyde adduct. The adduct prepared from BSA and 4-hydroxynonenal was chemotactic at doses of 0.65 to 0.0065 mg/ml, when tested in the presence of unmodified BSA. Since the adduct released free 4-hydroxyalkenal during the assay in the reverse reaction, it can not be decided whether the active principle is the aldehyde itself or the aldehyde attached to the BSA. From the effective doses of the aldehydes (10(-7) to 10(-12)M) and the BSA-aldehyde adduct it appears very unlikely that the BSA itself gained chemotactic properties through the alkylation of its sulfhydryl groups by the aldehyde.

Aldehydes↗
PubMed1986

Comparison of the inactivation of microsomal glucose-6-phosphatase by in situ lipid peroxidation-derived 4-hydroxynonenal and exogenous 4-hydroxynonenal.

1) The effect of 4-hydroxynonenal and lipid peroxidation on the activities of glucose-6-phosphatase and palmitoyl CoA hydrolase were studied. 2) 4-Hydroxynonenal inactivates glucose-6-phosphatase but has no effect on palmitoyl-CoA hydrolase. These effects are similar with those observed during lipid peroxidation of microsomes. 3) The inhibition of glucose-6-phosphatase by 4-hydroxynonenal can be prevented by glutathione but not by vitamin E. The inactivation of glucose-6-phosphatase during lipid peroxidation is prevented by glutathione and delayed by vitamin E. 4) The formation of 4-hydroxynonenal during lipid peroxidation was followed in relation to the inactivation of glucose-6-phosphatase. At 50% inactivation of glucose-6-phosphatase the 4-hydroxynonenal concentration was 1.5 microM. To obtain 50% inactivation of glucose-6-phosphatase by added 4-hydroxynonenal a concentration of 150 microM or 300 microM was needed with a preincubation time of 30 and 60 min, respectively. 5) It is concluded that the glucose-6-phosphatase inactivation during lipid peroxidation can be due to the formation of 4-hydroxynonenal. The formed 4-hydroxynonenal which inactivates glucose-6-phosphatase is located in the membrane. If this mechanism is valid it implies that a functional SH group of glucose-6-phosphatase is layered in the membrane. However, an inactivation of glucose-6-phosphatase by desintegration of the membrane by lipid peroxidation cannot be ruled out.

Aldehydes↗
PubMed1985

Qualitative and quantitative measurement of hydroxy fatty acids, thromboxanes and prostaglandins using stable isotope dilutions and detection by gas chromatography-mass spectrometry.

Methods for measurement of the metabolites of arachidonic acid (AA), namely prostaglandins (PGs), thromboxanes (TXs) and hydroxy fatty acids, using stable isotope dilution gas chromatography--mass spectrometry are described. With a few exceptions, labelled species of the various AA metabolites are not commercially available and were therefore synthesized in our laboratory. [2H8]AA, produced by deuteration of eicosatetraynoic acid, was used for comparing the metabolism of exogenously added and endogenously present AA in fibroblast cultures. After derivatization and catalytic hydrogenation, structure elucidation and quantification of the different hydroxy fatty acids was carried out by determination of the fragment ions resulting from alpha-cleavage at the site of the hydroxy function. During catalytic hydrogenation a significant hydrogen--deuterium exchange was observed. To eliminate this problem, 18O-labelled standards were prepared by exchanging the oxygen of the carboxylic acid group. The preparation and the use of hydroxy fatty acids, PGs and TXs labelled with 18O is described.

Fatty Acids↗
PubMed1985

Quantitative determination of the lipid peroxidation product 4-hydroxynonenal by high-performance liquid chromatography.

4-Hydroxynonenal is a product formed in tissue and tissue fractions from polyunsaturated membrane lipids through a free radical-induced lipid peroxidation process. The biological properties of this aldehyde have been studied in many respects. This article describes for the first time a sensitive and reproducible method for quantitative analysis of 4-hydroxynonenal in biological samples as well as in lipid-containing foodstuffs. The method involves extraction of the aldehyde by dichloromethane from cells or microsomes trapped on an Extrelut column. Oils and foodstuffs are extracted with excess water. After additional sample cleanup by solid-phase extraction on a disposable octadecyl silica gel (ODS) extraction column, the sample is analyzed by high-performance liquid chromatography using an ODS column and methanol/water 65/35 (v/v) or acetonitrile/water 40/60 (v/v) as eluant; the detection wavelength is 220 nm. The method developed has a high precision with coefficients of variation of 1.4% (microsomes) to 3.5% (olive oil). The recovery depends on the sample type and lies between 45% (control microsomes) and 96% (solution of hydroxynonenal in water). The method has been used for the determination of 4-hydroxynonenal in microsomes, platelets, and various foodstuffs.

Aldehydes↗
PubMed1985

Effects of the lipidperoxidation product 4-hydroxynonenal and related aldehydes on proliferation and viability of cultured Ehrlich ascites tumor cells.

The mechanism by which the lipid peroxidation product 4-hydroxynonenal and several other homologous, yet non biogenic aldehydes inhibit proliferation of cultured Ehrlich ascites tumor cells has been studied. Incubation of cells (5 X 10(-4)/ml) in a minimum essential medium supplemented with 10 or 20 microM 4-hydroxynonenal reduces the 36-hr cell count to 65 and 30% of the control value. The reduced growth rate is most likely due to a blockage of the DNA synthesis. Cells labelled by a [3H]-thymidine pulse prior to exposure to 4-hydroxynonenal (20 microM, 8 hr) showed no change of the specific radioactivity of the DNA, indicating that no de novo synthesis occurred in the presence of the aldehyde. In the absence of the aldehyde the specific radioactivity of the DNA decreased by 25%. A 2-hr incubation in the presence of 10 or 20 microM of 4-hydroxynonenal reduced [3H]-thymidine incorporation into the HClO4 insoluble fraction to 85 and 50% of the controls, but had no effect of the [3H]-thymidine and 86Rb uptake. Moreover, examination of the cell cultures by the Trypan Blue exclusion technique revealed that 20 microM 4-hydroxynonenal does not cause cell death. The high reactivity of 4-hydroxynonenal towards sulfhydryl groups suggests that the aldehyde inhibits DNA synthesis by interacting with a functional SH group of DNA polymerase. The specific action on DNA synthesis is abolished at an aldehyde concentration of 50 microM, which leads to 30% (6 hr exposure) and 95% (36 hr exposure) of dead cells. The cytostatic index (CI), i.e. concentration at 50% Trypan Blue positive cells/concentration at 50% inhibition of cell growth deducted from the dose effect curves is 3.0 for 4-hydroxynonenal. The other homologous 4-hydroxyalkenals with chain length of 5, 6, 7, 8, 10 and 11 carbon atoms also inhibit cell growth. The CI varied from 1.20 to 1.94, indicating that these non biogenic 4-hydroxyalkenals have a distinctively lower specific effect on proliferation than the biogenic 4-hydroxynonenal. The Michael adducts of 4-hydroxynonenal with glutathione and cysteine were nearly one order of magnitude less toxic than the free aldehyde, the CI (2.41 cysteine adduct, 2.06 glutathione adduct), however, were not improved since the growth inhibitory action was also reduced.

Aldehydes↗