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H Esterbauer

Publications and source records attributed to H Esterbauer.

At least 181 records · Page 10Linked to original sources
PubMed1985

Metabolism of the lipid peroxidation product 4-hydroxynonenal by isolated hepatocytes and by liver cytosolic fractions.

The metabolism of the lipid peroxidation product 4-hydroxynonenal and of several other related aldehydes by isolated hepatocytes and rat liver subcellular fractions has been investigated. Hepatocytes rapidly metabolize 4-hydroxynonenal in an oxygen-independent process with a maximum rate (depending on cell preparation) ranging from 130 to 230 nmol/min per 10(6) cells (average 193 +/- 50). The aldehyde is also rapidly utilized by whole rat liver homogenate and the cytosolic fraction (140 000 g supernatant) supplemented with NADH, whereas purified nuclei, mitochondria and microsomes supplemented with NADH show no noteworthy consumption of the aldehyde. In cytosol, the NADH-mediated metabolism of the aldehyde exhibits a 1:1 stoichiometry, i.e. 1 mol of NADH oxidized/mol of hydroxynonenal consumed, and the apparent Km value for the aldehyde is 0.1 mM. Addition of pyrazole (10 mM) or heat inactivation of the cytosol completely abolishes aldehyde metabolism. The various findings strongly suggest that hepatocytes and rat liver cytosol respectively convert 4-hydroxynonenal enzymically is the corresponding alcohol, non-2-ene-1,4-diol, according to the equation: CH3-[CH2]4-CH(OH)-CH = CH-CHO + NADH + H+----CH3-[CH2]4-CH(OH)-CH = CH-CH2OH + NAD+. The alcohol non-2-ene-1,4-diol has not yet been isolated from incubations with hepatocytes and liver cytosolic fractions, but was isolated in pure form from an incubation mixture containing 4-hydroxynonenal, isolated liver alcohol dehydrogenase and NADH and its chemical structure was confirmed by mass spectroscopy. Compared with liver, all other tissues possess only little ability to metabolize 4-hydroxynonenal, ranging from 0% (fat pads) to a maximal 10% (kidney) of the activity present in liver. The structure of the aldehyde has a strong influence on the rate and extent of its enzymic NADH-dependent reduction to the alcohol. The saturated analogue nonanal is a poor substrate and only a small proportion of it is converted to the alcohol. Similarly, nonenal is much less readily utilized as compared with 4-hydroxynonenal. The effective conversion of the cytotoxic 4-hydroxynonenal and other reactive aldehydes to alcohols, which are probably less toxic, could play a role in the general defence system of the liver against toxic products arising from radical-induced lipid peroxidation.

Alcohol Dehydrogenase↗
PubMed1985

Separation and characterization of the aldehydic products of lipid peroxidation stimulated by carbon tetrachloride or ADP-iron in isolated rat hepatocytes and rat liver microsomal suspensions.

Carbonyl products were separated and identified in suspensions of rat liver microsomal fractions and in isolated hepatocytes, after stimulation of lipid peroxidation by incubation with the pro-oxidants CCl4 and ADP-iron. The carbonyl products were allowed to react with 2,4-dinitrophenylhydrazine, and the derivatives were extracted and separated by t.l.c. into three zones of non-polar materials, and one fraction of polar derivatives that remained at the origin. Separation of the individual non-polar hydrazones in each zone by h.p.l.c. demonstrated that zone I prepared from microsomal fraction or hepatocytes incubated with CCl4 or ADP-iron contained mainly 4-hydroxyhex-2-enal, 4-hydroxynon-2-enal and 4-hydroxynona-2,5-dienal. Zone III consisted mainly of the alkanals propanal, pentanal and hexanal, the 2-alkenals propenal, pent-2-enal, hex-2-enal, hept-2-enal, oct-2-enal and non-2-enal, the ketones butanone, pentan-2-one and pentan-3-one, and deca-2,4-dienal. Incubation of a microsomal fraction with ADP-iron was much more effective in producing malonaldehyde and other carbonyl products than an incubation with CCl4. Despite such quantitative differences, there were no obvious qualitative differences in the h.p.l.c. spectra obtained from zones I and III. However, the stoichiometric evaluation of fatty acid loss and the production of malonaldehyde and other carbonyls suggests that the pathways of lipid peroxidation triggered by CCl4 and ADP-iron are different. The accumulation of carbonyl products of lipid peroxidation in isolated hepatocytes is strongly affected by their metabolism; in particular, 4-hydroxyalkenals were found to be metabolized very rapidly. Nonetheless, both CCl4 and ADP-iron produced stimulation in the production of malonaldehyde and non-polar carbonyl production. After incubation of rat hepatocytes with CCl4 or ADP-iron it was found that approx. 50% of the total amount of non-polar carbonyls produced during incubation escaped into the external medium. This was not leakage from dead cells, as 90-95% of the hepatocytes had retained their integrity at the end of the incubation. Release of carbonyl products from cells stimulated to undergo lipid peroxidation may be a mechanism for spreading an initial intracellular disturbance to affect critical targets outside the parent cell.

Adenosine Diphosphate↗
PubMed1985

Cytotoxicity, thiol depletion and inhibition of O6-methylguanine-DNA methyltransferase by various aldehydes in cultured human bronchial fibroblasts.

Lipid peroxidation aldehydes of the 4-hydroxy-alpha, beta-unsaturated type, as well as the tobacco-smoke related alpha, beta-unsaturated aldehyde, acrolein, were highly cytotoxic and decreased the intracellular thiol content in cultured human bronchial fibroblasts after treatment with micromolar concentrations. In comparison, formaldehyde and acetaldehyde were less toxic and 100- to 300-fold higher doses were required to affect cell survival or thiol levels. The unsaturated aldehydes also markedly inhibited the DNA repair enzyme O6-methylguanine-DNA methyltransferase known to have a cysteine residue in its active site, but had no effect on the activity of uracil-DNA glycosylase. Our results indicate that reactive aldehydes of either exogenous or endogenous origin have direct cytotoxic effects and may also make cells more susceptible to other toxic chemicals due to an impairment in cellular defense mechanisms, e.g., DNA repair and detoxification by systems requiring glutathione.

Aldehydes↗
PubMed1985

Chemotactic activity of hydroxyalkenals on rat neutrophils.

Previous results from our laboratory showed that in vitro 4-hydroxyoctenal, 4-hydroxynonenal (HNE) and 4-hydroxytetradecenal elicit a neutrophil chemotactic response in the presence of albumin. Data presented here indicate that other hydroxyalkenals (4-hydroxyundecenal and 4-hydroxy-pentadecenal) also exert a chemotactic attraction on rat pleural neutrophils. The different aldehydes tested show a wide activity-concentration range, the most powerful being 4-hydroxyoctenal. Unlike most chemoattractants, the hydroxyalkenals seem deprived of chemokinetic properties. The mechanism by which these compounds may act on granulocytes is analysed. Experiments investigating the deactivation phenomenon induced by HNE indicate that this aldehyde acts as a chemoattractant by triggering mechanisms different from N-formylmethionyl-leucyl-phenylalanine (fMLP). In fact, cells pre-incubated with this peptide fail to respond chemotactically to the same compound, but are still able to respond to HNE. Similarly, the preventive contact of neutrophils with HNE is unable to influence the chemotactic effect of fMLP.

Aldehydes↗
PubMed1984

Detection of 4-hydroxynonenal as a product of lipid peroxidation in native Ehrlich ascites tumor cells.

4-Hydroxynonenal, which is a major product of lipid peroxidation in rat liver microsomes, was detected in native Ehrlich ascites tumor cells. Its formation was stimulated either by ferrous ions or by Fe(II)-histidinate. The identification was based on chromatographic (TLC/HPLC) and ultraviolet-spectroscopic evidence using synthetic 4-hydroxynonenal as reference. Highest values of 4-hydroxynonenal concentration (about 0.1 microM in the cell suspension) after 30 min of incubation were observed with Fe(II)-histidinate as stimulant. Saturation was already reached after an incubation period of 10 min. The results confirm the expectation by Schauenstein and Esterbauer (in Submolecular Biology and Cancer, Ciba Foundation Series 67 (1979) pp. 225-244, Excerpta Medica, Amsterdam) that endogenous lipid peroxidation gives rise to a distinct intracellular level of alpha, beta-unsaturated aldehydes. A simple hypothetical mechanism for the formation of 4-hydroxynonenal from n-6-polyunsaturated fatty acids is presented.

Aldehydes↗
PubMed1984

Characterization of lipoxygenase metabolites of arachidonic acid in cultured human skin fibroblasts.

Cultured human skin fibroblasts were incubated in the presence of [14C]arachidonic acid (50 microM; 1 m Ci/mmol) and the divalent cation ionophore A23187 at 37 degrees C for 60 min. The metabolites formed were extracted from the cell-free medium in diethyl ether, separated by thin layer chromatography and identified unequivocally by GC-MS. The distribution of the arachidonic acid metabolites as estimated from the recovered radioactivity showed as major product prostaglandin E2 (26%). Minor amounts of other prostaglandins, i.e., 6-oxo-prostaglandin F1 alpha (1%), prostaglandin F2 alpha (1%), prostaglandin D2 (0.5%) and prostaglandin A2 (1%) were also present. In addition to the prostaglandins, monohydroxy fatty acids (4.5%) were also detected. This fraction contained 33% 12-hydroxy-5,8,10-heptadecatrienoic acid (HHT), 22% 11-hydroxy-5,8,11,14-eicosatetraenoic acid (11-HETE) and 31% 15-hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE). Lipid extracts of the cells did not show any detectable amount of the monohydroxy fatty acids, indicating that they are not incorporated metabolically in the cellular lipids. The monohydroxy fatty acids originate mainly from the exogenously added arachidonic acid as evidenced by the 2H/H ratio (30:1) from experiments with octadeuterated arachidonic acid [( 2H8]arachidonic acid). Indomethacin inhibited the formation of all prostaglandins, HHT and 11-HETE; moreover, eicosatetraynoic acid (also blocked the formation of 15-HETE. From these results, it can be concluded that in human skin fibroblasts prostaglandin E2 is the major product of the cycloxygenase pathway, while 15-HETE is the main lipoxygenase product.

Arachidonic Acid↗
PubMed1984

Cytotoxic aldehydes originating from the peroxidation of liver microsomal lipids. Identification of 4,5-dihydroxydecenal.

During the NADPH-Fe-induced peroxidation of liver microsomal lipids products are formed which are provided with cytopathological activities. In a previous study one of the major products was identified as an aldehyde of the 4-hydroxyalkenal class, namely 4-hydroxynonenal. In the present study another cytotoxic product has been isolated and identified as 4,5-dihydroxy-2,3-decenal. The isolation was performed by means of thin-layer chromatography and high-pressure liquid chromatography and the structure was ascertained mainly by means of mass spectroscopy of the free aldehyde and of its derivatives. In the absence of NADPH-Fe liver microsomes produced no 4,5-dihydroxydecenal. The inhibitory activity of 4,5-dihydroxydecenal on microsomal glucose-6-phosphatase is somewhat lower than that exhibited by 4-hydroxynonenal. This lower inhibitory activity correlates with the lower capacity to bind to the microsomal protein of 4,5-dihydroxydecenal as compared to 4-hydroxynonenal. The reactivities of the two aldehydes with cysteine were comparable. The production of toxic aldehydes may represent a mechanism by which lipid peroxidation causes deleterious effects on cellular functions.

Aldehydes↗
PubMed1983

Effects of 4-hydroxynonenal on isolated hepatocytes. Studies on chemiluminescence response, alkane production and glutathione status.

The effect of 4-hydroxy-2,3-trans-nonenal, a diffusible product of lipid peroxidation, on isolated hepatocytes was evaluated with two non-invasive techniques measuring low-level chemiluminescence and alkane evolution. Oxygen-induced low-level chemiluminescence and ethane and n-pentane formation by hepatocytes is enhanced over 7-fold in the presence of 4-hydroxynonenal (2 mM). Glutathione-depleted hepatocytes show a higher increase than controls in both low-level chemiluminescence and alkane formation upon supplementation with 4-hydroxynonenal. The effects on both parameters are diminished by vitamin E pretreatment of rats and are absent under anaerobiosis. At variance with chemiluminescence and alkane formation, 4-hydroxynonenal does not elicit a concomitant increase in malonaldehyde or diene-conjugate formation. Addition of 4-hydroxynonenal to a suspension of hepatocytes causes a rapid loss of cellular glutathione in the form of a glutathione conjugate with the alkenal as observed with high-pressure liquid-chromatographic analysis. The reaction between glutathione and 4-hydroxynonenal proceeds also spontaneously in vitro at 1:1 stoichiometry. The cellular effects of 4-hydroxynonenal evaluated by low-level chemiluminescence and alkane formation are independent of the formation of a glutathione conjugate and seem to rely on the remaining not-bound 4-hydroxynonenal. The sensitivity of 4-hydroxynonenal-enhanced chemiluminescence and alkane formation to free-radical quenchers suggests the participation of a free-radical propagation process.

Aldehydes↗
PubMed1982

Separation and characterization of the aldehydic products of lipid peroxidation stimulated by ADP-Fe2+ in rat liver microsomes.

1. Methods using t.l.c. and high-pressure liquid chromatography (h.p.l.c.) have been used to separate the complex variety of substances possessing a carbonyl function that are produced during lipid peroxidation. 2. The major type of lipid peroxidation studied was the ADP-Fe2+-stimulated peroxidation of rat liver microsomal phospholipids. Preliminary separation of the polar and non-polar products was achieved by t.l.c.: further separation and identification of individual components was performed by h.p.l.c. Estimations were performed on microsomal pellets and the supernatant mixture after incubation of microsomes for 30 min at 37 degrees C. 3. The polar fraction was larger than the non-polar fraction when expressed as nmol of carbonyl groups/g of liver. In the non-polar supernatant fraction the major contributors were n-alkanals (31% of the total), alpha-dicarbonyl compounds (22%) and 4-hydroxyalkenals (37%) with the extraction method used. 4. Major individual contributors to the non-polar fraction were found to be propanal, 4-hydroxynonenal, hexanal and oct-2-enal. Other components identified include butanal, pent-2-enal, hex-2-enal, hept-2-enal, 4-hydroxyoctenal and 4-hydroxyundecenal. The polar carbonyl fraction was less complex than the non-polar fraction, although the identities of the individual components have not yet been established. 5. Since these carbonyl compounds do not react significantly in the thiobarbituric acid reaction, which largely demonstrates the presence of malonaldehyde, it is concluded that considerable amounts of biologically reactive carbonyl derivatives are released in lipid peroxidation and yet may not be picked up by the thiobarbituric acid reaction.

Adenosine Diphosphate↗
PubMed1982

Detection of carbonyl functions in phospholipids of liver microsomes in CCl4- and BrCCl3-poisoned rats.

Since the peroxidative cleavage of unsaturated fatty acids can result in either the release of carbonyl compounds or the formation of carbonyl functions in the acyl residues, evidence for the presence of carbonyl groups in liver microsomal phospholipids was searched for in in vivo conditions (CCl4 and BrCCl3 intoxications) in which peroxidation of lipids of hepatic endoplasmic reticulum had been previously demonstrated. The spectrophotometric examination of 2,4-dinitrophenylhydrazine-treated phospholipids of liver microsomes from the intoxicated animals showed absorption spectra similar to those observed for the dinitrophenylhydrazones of various carbonyls. Similar spectra, although magnified from a quantitative point of view, were also observed with 2,4-dinitrophenylhydrazine-treated phospholipids of liver microsomes peroxidized in the NADPH-Fe-dependent system. A time-course study of microsomal lipid peroxidation showed that the amount of 2,4-dinitrophenylhydrazine-reacting groups (carbonyl functions) in phospholipids of liver microsomes increases with the incubation time and is correlated to the amount of malonic dialdehyde formed in the incubation mixture. The kinetics of the production of 4-hydroxynonenal was somewhat similar to that of malonic dialdehyde formation. In both the in vivo conditions (CCl4 and BrCCl3 intoxications) the amount of carbonyl functions in microsomal phospholipids, which was higher in the BrCCl3-intoxicated animals as compared to the CCl4-poisoned ones, was close to that found in the vitro condition in which lipid peroxidation is induced by 6 microM Fe2+. The possible pathological significance of formation of carbonyl functions in membrane phospholipids is discussed.

Animals↗
PubMed1982

Evidence for aldehydes bound to liver microsomal protein following CCl4 or BrCCl3 poisoning.

Since it has been demonstrated in previous studies that peroxidation of liver microsomal lipids leads to the production of aldehydes provided with cytopathological activities--namely 4-hydroxyalkenals--evidence was searched for aldehydes bound to microsomal protein in in vivo conditions (CCl4 and BrCCl3 intoxications) in which peroxidation of lipids of hepatic endoplasmic reticulum had been demonstrated previously. The spectrophotometric analysis of 2,4-dinitrophenylhydrazine-treated non-lipoidal residues of liver microsomes from the intoxicated rats shows absorption spectra similar to those observed for the dinitrophenylhydrazones formed in the reaction of alkenals with -SH groups of proteins or low molecular weight thiols. Similar spectra, although magnified from a quantitative point of view, were obtained either with liver microsomes allowed to react with synthetic 4-hydroxynonenal or with liver microsomes peroxidized in the NADPH-Fe-dependent system. A time-course study of microsomal lipid peroxidation shows that the amount of 2,4-dinitrophenylhydrazine-reacting groups in the non-lipoidal residue of liver microsomes increases with the incubation time and is correlated to the amount of thiobarbituric acid-reacting products formed in the incubation mixture. In both the in vivo conditions (CCl4 and BrCCl3 intoxications) the amount of 2,4-dinitrophenylhydrazine-reacting groups in the non-lipoidal residue of liver microsomes increases from 15 min up to 2 h after poisoning and is higher, in every instance, in the BrCCl3-intoxicated animals compared to the CCl4-poisoned ones. Experiments carried out to ascertain the reliability of the spectrophotometric detection of protein-bound alkenals showed that in the in vitro system in which liver microsomes are allowed to react with 4-hydroxynonenal there is a good agreement between the binding value that can be calculated from the absorption spectrum and the binding value obtained by using labelled 4-hydroxynonenal.

Aldehydes↗
PubMed1982

Neutrophil chemotactic responses to aldehydes.

The influence of 4-hydroxynonenal (HNE), 4-hydroxytetradecenal (HTDE) and methylglyoxal (MGL) on rat polymorphonuclear leukocyte locomotion has been studied by means of boyden chambers. The two alkenals have been found to exert a chemotactic activity in a range of concentrations between 10-9 and 10-5 M, the most potent chemical being HTDE. The chemotactic effect, however, appeared influenced by the concentration of serum albumin in the medium. In addition, chemotactic doses of the same aldehydes were unable to affect the cell random migration. Higher concentrations of these alkenals depressed both the directed and random locomotion. Methylglyoxal appeared deprived of chemotactic power when assayed at the doses that were active with the above mentioned alkenals and it did not interfere with random migration. 5 X 10-4 M Methylglyoxal exerted a slight, but significant, inhibition of the locomotion events. These results suggest that aldehydes, present at the inflammatory sites, as final products of lipid peroxidation, might play a role in modulating the granulocyte cell locomotion, therefore interfering with the development of phlogosis.

Aldehydes↗
PubMed1981

[Preliminary data on the influence of 4-hydroxyalkenals on the phagocytic activity polymorphonuclear leukocytes in the rat].

The effect of low concentrations of 4-hydroxy-2-trans-pentenal, 4-hydroxy-2-trans-nonenal and 4-hydroxy-2-trans-tetradecenal (4-hydroxyalkenals produced during lipid peroxidation) was evaluated on the phagocytic activity of polymorphonuclear cells obtained from rat peritoneum. Our results show that the above compounds can influence, at different degrees, the endocytic powers, as measured "in vitro", of leukocytes through an inhibition that appears, to some extent, dose-related. Since lipoperoxidative processes can take place at the inflammed area, aldehydes originating from the peroxidative derangement of fatty acids may play a role in interfering with the cellular reactions at the inflammatory site.

Aldehydes↗
PubMed1980

Identification of 4-hydroxynonenal as a cytotoxic product originating from the peroxidation of liver microsomal lipids.

During the NADPH-Fe induced peroxidation of liver microsomal lipids, products are formed which show various cytopathological effects including inhibition of microsomal glucose-6-phosphatase. The major cytotoxic substance has been isolated and identified as 4-hydroxy-2,3-trans-nonenal. The structure was ascertained by means of ultraviolet, infrared and mass spectrometry and high-pressure liquid chromatographic analysis. Moreover, 4-hydroxynonenal, prepared by chemical synthesis, was found to reproduce the biological effects brought about by the biogenic aldehyde. Preliminary investigations suggest that as compared to 4-hydroxynonenal very low amounts of other 4-hydroxyalkenals, namely 4-hydroxyoctenal, 4-hydroxydecenal and 4-hydroxyundecenal are also formed by actively peroxidizing liver microsomes. In the absence of NADPH-Fe liver microsomes produced only minute amounts of 4-hydroxyalkenals. The biochemical and biological effects of synthetic 4-hydroxyalkenals have been studied in great detail in the past. The results of these investigations together with the finding that 4-hydroxyalkenals, in particular 4-hydroxynonenal, are formed during NADPH-Fe stimulated peroxidation of liver microsomal lipids, may help to elucidate the mechanism by which lipid peroxidation causes deleterious effects on cells and cell constituents.

Aldehydes↗
PubMed1979

[Spin-lattice-relaxationtime T1 measurements of nucleosides (author's transl)].

Nucleosides and nucleotides have been investigated by means of the spin-lattice-relaxationtime T1, the measurements carried out in CDCl3 on the nucleosides adenosine (A), uridine (U) and inosine (I) indicate that the strength of H-bonding is highest in the basepair (A)--(U). The systems (A)--(A), (U)--(U), (I)--(A) and (I)--(U) however, show only a low tendency to associate. Plotting the spin-lattice-relaxationtime, of the nucleotide protons in fragments (mol weight below 40,000) originated from DNA or RNA, against increasing temperature gives a curve with a sigmoide shape and a Tm value of 80 degrees C. The average of the activation energy for the H-bonding is 18.02 KJ/mol. The investigations clearly show that the spin-lattice-relaxationtime measurements open first of all a new method for research on H-bonding, secondly it is even in the case of nucleosides and oligonucleotides, where most of the common methods fail, a way to get information about H-bonding effects.

Animals↗
PubMed1978

Formation and properties of reactive aldehydes.

4-Hydroxyalkenals react easily with SH groups in enzymes and structural proteins to give stable adducts in which the aldehyde is bound through a thioether linkage to the protein. This reaction represents the mechanism by which these compounds block certain metabolic processes (DNA, RNA and protein synthesis, respiration, glycolysis) and exert their antitumour activity. A highly significant inverse correlation exists between the number of hydroxypentenal-sensitive SH groups in the soluble proteins and the doubling time of 12 transplantable animal tumours. Hydroxyalkenals applied peritumourally completely prevent or significantly delay growth of the solid forms of four animal tumours. The almost non-toxic adducts of 4-hydroxypentenal-cysteine 1:1 and crotonal-cysteine 1:2 applied intraperitoneally prevent or inhibit growth of Ehrlich ascites tumour in mice. The possible formation of unsaturated aldehydes in vivo from lipids, carbohydrates and diamines is discussed.

Aldehydes↗