Biomedical subjects

G Waeg

Publications and source records attributed to G Waeg.

At least 19 recordsLinked to original sources

PubMed2002

Down-modulation of nuclear localisation and pro-fibrogenic effect of 4-hydroxy-2,3-nonenal by thiol- and carbonyl-reagents.

Among the oxidative breakdown products of omega-6 unsaturated fatty acids, the aldehyde 4-hydroxy-2,3-nonenal (HNE) is receiving increasing attention for its potential pathophysiological implication, which at least partly lies on the demonstrated ability to modulate gene expression of a number of genes. Here we show that a marked down-modulation of HNE nuclear localisation in cells of a macrophage line (J774-A1) can be afforded by treatment with sulfydryl and carbonyl reagents without significantly interfering with cell viability. As regards the addition of thiol-group reagents to the cell suspension, N-ethylmaleimide (NEM) led to a sustained decrease of HNE nuclear localisation, while 4-(chloromercuri)-benzene-sulfonic acid (PCMBS) gave a similar but more transient effect. Hydroxylamine (HYD), a carbonyl-group reagent, was also able to inhibit HNE nuclear localisation. The actual efficacy of the inhibitors used was then tested on the HNE-induced stimulation of transforming growth factor beta1 (TGFbeta1) production by J774-A1 cells. Indeed, the thiol reagents NEM and PCMBS, both markedly down-modulating HNE nuclear localisation, were able to inhibit HNE-induced increase of TGFbeta1 protein synthesis. The carbonyl reagent HYD was less effective on this respect, producing strong but incomplete protection against HNE-induced TGFbeta1 increase. Taken together, the results indicate that sulfydryl groups are involved in the process of HNE cellular internalisation, while both sulfydryl and carbonyl groups are involved in the process of HNE nuclear translocation, and consequently in the modulation of gene expression by the aldehyde. Further, an actual demonstration is provided that HNE-induced effect on gene regulation can be efficiently counteracted by suitable interference with HNE biochemistry.

Aldehydes↗

PubMed2001

4-Hydroxynonenal, a lipid peroxidation byproduct of spinal cord injury, is cytotoxic for oligodendrocyte progenitors and inhibits their responsiveness to PDGF.

Oligodendroglial reactions to compression injury of spinal cord include apoptosis, secondary demyelination, and remyelination failure. Within hours after contusion, the membrane lipid peroxidation (MLP) byproduct, 4-hydroxynonenal (HNE), increases rapidly in gray matter and thereafter in white matter tracts beyond the initial lesion level. Considering that HNE is a mediator and marker of neuronal MLP toxicity in various neurodegenerative conditions, the present study examined its effect on the regeneration potential of oligodendrocyte progenitors, as defined by their capacity to survive, proliferate and migrate in primary culture. Treatment of oligodendroblasts with HNE evoked a time- and dose-dependent cytotoxicity resembling apoptosis at aldehyde concentrations known to be produced by neurons and achieved in tissue undergoing peroxidative injury. In addition, sublethal concentrations of HNE inhibited the mitogenic and chemotactic responses of more immature progenitors to platelet-derived growth factor. These effects appear to be mediated in part by the formation of HNE adducts with progenitor proteins located within the plasma membrane and cytoplasmic compartments. Our data are the first to show that HNE can have direct, deleterious effects on oligodendrocyte precursors. The present study also suggests a mechanism by which the striking accumulation of HNE in white matter tracts surrounding the site of spinal cord compression injury and in other ischemic-hypoxic insults associated with MLP could suppress the potential regenerative response of endogenous oligodendrocyte progenitor cells.

Aldehydes↗

PubMed2000

Immunohistochemical evidence for the myeloperoxidase/H2O2/halide system in human atherosclerotic lesions: colocalization of myeloperoxidase and hypochlorite-modified proteins.

The 'oxidation theory' of atherosclerosis proposes that oxidation of low density lipoprotein (LDL) contributes to atherogenesis. Although the precise mechanisms of in vivo oxidation are widely unknown, increasing evidence suggests that myeloperoxidase (MPO, EC 1.11.1.7), a protein secreted by activated phagocytes, generates modified/oxidized (lipo)proteins via intermediate formation of hypochlorous acid (HOCl). In vitro generation of HOCl transforms lipoproteins into high uptake forms for macrophages giving rise to cholesterol-engorged foam cells. To identify HOCl-modified-epitopes in human plaque tissues we have raised monoclonal antibodies (directed against human HOCl-modified LDL) that do not cross-react with other LDL modifications, i.e. peroxynitrite-LDL, hemin-LDL, Cu2+-oxidized LDL, 4-hydroxynonenal-LDL, malondialdehyde-LDL, glycated-LDL, and acetylated-LDL. The antibodies recognized a specific epitope present on various proteins after treatment with OCl- added as reagent or generated by the MPO/H2O2/halide system. Immunohistochemical studies revealed pronounced staining for HOCl-modified-epitopes in fibroatheroma (type V) and complicated (type VI) lesions, while no staining was observed in aortae of lesion-prone location (type I). HOCl-oxidation-specific epitopes are detected in cells in the majority of atherosclerotic plaques but not in control segments. Staining was shown to be inside and outside monocytes/macrophages, endothelial cells, as well as in the extracellular matrix. A similar staining pattern using immunohistochemistry could be obtained for MPO. The colocalization of immunoreactive MPO and HOCl-modified-epitopes in serial sections of human atheroma (type IV), fibroatheroma (type V) and complicated (type VI) lesions provides further convincing evidence for MPO/H2O2/halide system-mediated oxidation of (lipo)proteins under in vivo conditions. We propose that MPO could act as an important link between the development of atherosclerotic plaque in the artery wall and chronic inflammatory events.

Aged↗

PubMed2000

Involvement of lipid peroxidation, oncogene expression and induction of apoptosis in the antitumorous activity of ferric-sorbitol-citrate.

We described before that iron-containing, anti-anaemic drug, ferric-sorbitol-citrate complex (FSC) inhibited proliferation of various murine cancer cells in vitro and caused tumour regression in vivo, but did not affect proliferation of the non-malignant cells. The aim of this study was to evaluate further the anticancer activity mechanism of FSC using human colon cancer cell line CaCo2. After treatment with FSC for 72 hours impaired proliferative ability and viability of CaCo2 cells as observed. Growth modification caused by FSC involved diminished expression of Bcl-2, and over-expression of mp53 proto-oncogenes, accompanied by increased incidence of apoptosis. Immunostaining the cells applying monoclonal antibodies for lipid peroxidation product 4-hydroxynonenal (HNE) showed that FSC-iron increased intracellular HNE, but did not induce severe HNE-mediated oxidative stress. Thus, antitumorous mechanism of FSC involves modulation of oncogene expression and induction of apoptosis apparently not triggered by lipid peroxidation-mediated oxidative stress, although FSC might restore endogenous HNE production in the CaCo2 cells to level resembling physiological for various non-malignant cells and tissues. Higher dose of FSC increased also number of intracellular ferritin positive CaCo2 cells.

Aldehydes↗

PubMed1999

4-Hydroxynonenal as a second messenger of free radicals and growth modifying factor.

Immunohistochemical analysis of the distribution of the lipid peroxidation product 4-hydroxynonenal (HNE) in the brain of baboons exposed to experimental hemorrhagic traumatic shock or sepsis showed that systemic oxidative stress and the thereby generated HNE affect the blood:brain barrier and the regulation of cerebral blood flow determining secondary brain damage. Similarly, HNE was determined during ischemia in the brain blood vessels of rats exposed to ischemia/reperfusion injury of the brain. After reperfusion, HNE disappeared from the blood vessels but remained in neurones and in glial cells. Since HNE modulates cell proliferation and differentiation (including proto-oncogene expression), it is postulated that HNE might have prominent local and systemic effects that are not only harmful but beneficial, too, determining the outcome of various pathophysiological conditions based on oxidative stress.

Aldehydes↗

PubMed1998

HNE interacts directly with JNK isoforms in human hepatic stellate cells.

4-Hydroxy-2,3-nonenal (HNE) is an aldehydic end product of lipid peroxidation which has been detected in vivo in clinical and experimental conditions of chronic liver damage. HNE has been shown to stimulate procollagen type I gene expression and synthesis in human hepatic stellate cells (hHSC) which are known to play a key role in liver fibrosis. In this study we investigated the molecular mechanisms underlying HNE actions in cultured hHSC. HNE, at doses compatible with those detected in vivo, lead to an early generation of nuclear HNE-protein adducts of 46, 54, and 66 kD, respectively, as revealed by using a monoclonal antibody specific for HNE-histidine adducts. This observation is related to the lack of crucial HNE-metabolizing enzymatic activities in hHSC. Kinetics of appearance of these nuclear adducts suggested translocation of cytosolic proteins. The p46 and p54 isoforms of c-Jun amino-terminal kinase (JNKs) were identified as HNE targets and were activated by this aldehyde. A biphasic increase in AP-1 DNA binding activity, associated with increased mRNA levels of c-jun, was also observed in response to HNE. HNE did not affect the Ras/ERK pathway, c-fos expression, DNA synthesis, or NF-kappaB binding. This study identifies a novel mechanism linking oxidative stress to nuclear signaling in hHSC. This mechanism is not based on redox sensors and is stimulated by concentrations of HNE compatible with those detected in vivo, and thus may be relevant during chronic liver diseases.

Aldehydes↗

PubMed1998

U-101033E (2,4-diaminopyrrolopyrimidine), a potent inhibitor of membrane lipid peroxidation as assessed by the production of 4-hydroxynonenal, malondialdehyde, and 4-hydroxynonenal--protein adducts.

4-Hydroxy-2-nonenal (4-HNE) and malondialdehyde (MDA) are major lipid peroxidation products generated by free radical attack on membranes and appear to contribute to the cytotoxic effects of oxidative stress by a mechanism involving adduct formation with cellular proteins. In the present studies, we investigated the relationship between lipid peroxidation and eventual inactivation of plasma membrane proteins using a model system consisting of purified red blood cell membranes and Fe2+/EDTA. Using this system, we also analyzed the ability of a novel antioxidant, U-101033E (2,4-diaminopyrrolopyrimidine), to inhibit lipid peroxidation and associated protein damage. Our results demonstrated that significant levels of MDA and 4-HNE are generated in this model system, and that both aldehydes are capable of cross-linking membrane proteins. In addition, we used a monoclonal antibody to demonstrate the presence of 4-HNE-protein adducts in this system. The generation of 4-HNE-protein adducts closely paralleled the time course of lipid peroxidation and membrane protein cross-linking, suggesting that 4-HNE may contribute to membrane protein cross-linking. Analysis of U-101033E in this system showed that this antioxidant inhibited lipid peroxidation, prevented the appearance of 4-HNE-protein adducts, and strongly reduced membrane protein cross-linking, with an EC50 of 0.5 microM. We also show that these antioxidant effects were not due to the scavenging of superoxide anion. Thus, these studies demonstrate the potential usefulness of U-101033E for treating certain disease processes where lipid peroxidation plays a role in disease pathogenesis.

Aldehydes↗

PubMed1998

Human low density lipoprotein as a target of hypochlorite generated by myeloperoxidase.

The aim of this study was to further clarify which part of human low density lipoprotein (LDL) is attacked by the MPO/H2O2/Cl- -system and which reactive oxygen species is responsible for the attack. Therefore the influence of this system on the modification of the lipid and protein moiety of LDL was studied in vitro. Using the monochlorodimedone assay it was found that HOCl is produced in micromolar quantities in the absence of LDL and is rapidly consumed by LDL in a concentration dependent manner. The consumption of HOCl was reflected in the formation of HOCl-specific epitopes on apo B-100 as determined by an antibody raised against HOCl-modified LDL. The absorbency at 234 nm was applied to measure continuously the extent of modification of LDL. The general kinetic pattern of the absorbency measurement consisted of a lag phase where no LDL modification was observed, followed by a rapid increase of absorbency and a plateau phase. Finally the absorbency decreased due to LDL precipitation. Time dependent absorption spectra indicated that this kinetic pattern is mainly caused by light scattering due to particle aggregation rather than by a specific absorption at 234 nm due to conjugated diene formation. In agreement with this finding a low rate of thiobarbituric acid reactive substances (TBArS) formation was observed after a lag phase. The aggregation of LDL occurs most likely by modification of apo B-100, which was determined fluorimetrically in terms of LDL-tryptophan destruction in presence of the MPO/H2O2/Cl(-)-system. The kinetic course of tryptophan fluorescence generally consisted of a rapid decrease leveling off into a low plateau phase. Gas chromatographic determinations of linoleic acid in LDL in presence of the MPO system showed that this polyunsaturated fatty acid (PUFA) is easily attacked by HOCl. Consistent with this finding NMR spectra of HOCl modified LDL indicated a complete disappearance of bis-allylic methylene groups. Since lipid peroxidation products only partially account for this loss of PUFAs, other reactions of HOCl with unsaturated lipids--probably chlorohydrin formation--must be involved. Summarizing, although the rate of lipid peroxidation is low, both the lipid and the protein moiety of LDL are readily modified by the MPO system. It appears that the immediate consequence of apo B-100 modification is its aggregation. It is concluded that MPO, which has been detected in atherosclerotic lesions, is able to contribute to the modification of LDL into a form recognizable for uncontrolled uptake by macrophages.

Adult↗

PubMed1998

The presence of 4-hydroxynonenal/protein complex as an indicator of oxidative stress after experimental spinal cord contusion in a rat model.

OBJECT: The authors tested the hypothesis that breach of the blood-spinal cord barrier (BSCB) will produce evidence of oxidative stress and that a similar staining pattern will be seen between 4-hydroxynonenal (HNE)/protein complexes and extravasated immunoglobulin G (IgG). METHODS: Adult female Fischer 344 rats, each weighing 200 to 225 g, were subjected to a spinal cord contusion at T-10 by means of a weight-drop device. Spinal cord tissue was assessed for oxidative stress by localizing extravasated plasma contents with a monoclonal antibody for rat IgG and protein conjugation with HNE, which is an aldehyde byproduct of lipid peroxidation. The animals were killed at 1 and 6 hours, and 1, 2, and 7 days after surgery. Maximum HNE/protein staining was observed at 2 days postinjury, and HNE/protein and IgG manifested similar staining patterns. Analysis revealed a graduated but asymmetrical rostral-caudal response relative to the T-10 injury site. Both HNE/protein complex and IgG staining revealed that the caudal levels T-11 and T-12 stained significantly more intensely than the rostral levels T-9 and T-8, respectively. A higher percentage of neurons positive for HNE/protein immunostaining was observed in spinal cord levels caudal to the injury site compared with equidistant rostral regions. Protein dot-blot assays also revealed a similar asymmetrical rostral-caudal HNE/protein content. To analyze the timing of the BSCB breach, another group of animals received identical contusions, and horseradish peroxidase (HRP) was injected 10 minutes before or at various times after injury (1, 3, and 6 hours, and 1, 2, and 7 days). Maximum HRP permeability was seen immediately after injury, with a significant decrease occurring by 1 hour and a return to control levels by 2 days posttrauma. CONCLUSIONS: Data from this study indicate possible compromise of neuronal, axonal, glial, and synaptic function after trauma, which may be a factor in motor deficits seen in animals after spinal cord contusion. The colocalization of the IgG stain with the HNE/protein stain is consistent with the hypothesis of a mutual cause-effect relationship between BSCB and oxidative stress in central nervous system trauma.

Aldehydes↗

PubMed1997

4-Hydroxynonenal, a product of lipid peroxidation, inhibits dephosphorylation of the microtubule-associated protein tau.

In Alzheimer's disease (AD) the microtubule-associated protein tau is excessively phosphorylated in degenerating neurons, but the mechanisms underlying the increased phosphorylation are unknown. Recent findings suggest that oxidative stress, and membrane lipid peroxidation in particular, contributes to the neurodegenerative process in AD. We now report that following exposure of cultured rat hippocampal neurons to 4-hydroxynonenal (HNE), an aldehydic product of membrane lipid peroxidation, tau is resistant to dephosphorylation. Immunocytochemical and Western blot analyses using phosphorylation-sensitive tau antibodies showed that HNE treatment causes a moderate increase in basal levels of tau phosphorylation, and prevents tau dephosphorylation by alkaline phosphatase in neurons pretreated with the phosphatase inhibitor okadaic acid. Studies with anti-HNE antibodies showed that HNE binds directly to tau, and that HNE immunoreactivity localizes to cell bodies and axons, cell compartments that contain tau. These data suggest a role for HNE in altered tau phosphorylation and neurofibrillary degeneration in AD.

Aldehydes↗

PubMed1997

Evidence that 4-hydroxynonenal mediates oxidative stress-induced neuronal apoptosis.

Oxidative stress is believed to play important roles in neuronal cell death associated with many different neurodegenerative conditions (e.g., Alzheimer's disease, Parkinson's disease, and cerebral ischemia), and it is believed also that apoptosis is an important mode of cell death in these disorders. Membrane lipid peroxidation has been documented in the brain regions affected in these disorders as well as in cell culture and in vivo models. We now provide evidence that 4-hydroxynonenal (HNE), an aldehydic product of membrane lipid peroxidation, is a key mediator of neuronal apoptosis induced by oxidative stress. HNE induced apoptosis in PC12 cells and primary rat hippocampal neurons. Oxidative insults (FeSO4 and amyloid beta-peptide) induced lipid peroxidation, cellular accumulation of HNE, and apoptosis. Bcl-2 prevented apoptosis of PC12 cells induced by oxidative stress and HNE. Antioxidants that suppress lipid peroxidation protected against apoptosis induced by oxidative insults, but not that induced by HNE. Glutathione, which binds HNE, protected neurons against apoptosis induced by oxidative stress and HNE. PC12 cells expressing Bcl-2 exhibited higher levels of glutathione and lower levels of HNE after oxidative stress. Collectively, the data identify that HNE is a novel nonprotein mediator of oxidative stress-induced neuronal apoptosis and suggest that the antiapoptotic action of glutathione may involve detoxification of HNE.

Aldehydes↗

PubMed1997

4-Hydroxynonenal, an aldehydic product of membrane lipid peroxidation, impairs glutamate transport and mitochondrial function in synaptosomes.

Removal of extracellular glutamate at synapses, by specific high-affinity glutamate transporters, is critical to prevent excitotoxic injury to neurons. Oxidative stress has been implicated in the pathogenesis of an array of prominent neurodegenerative conditions that involve degeneration of synapses and neurons in glutamatergic pathways including stroke, and Alzheimer's, Parkinson's and Huntington's diseases. Although cell culture data indicate that oxidative insults can impair key membrane regulatory systems including ion-motive ATPases and amino acid transport systems, the effects of oxidative stress on synapses, and the mechanisms that mediate such effects, are largely unknown. This study provides evidence that 4-hydroxynonenal, an aldehydic product of lipid peroxidation, mediates oxidation-induced impairment of glutamate transport and mitochondrial function in synapses. Exposure of rat cortical synaptosomes to 4-hydroxynonenal resulted in concentration- and time-dependent decreases in [3H]glutamate uptake, and mitochondrial function [assessed with the dye 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)]. Other related aldehydes including malondialdehyde and hexanal had little or no effect on glutamate uptake or mitochondrial function. Exposure of synaptosomes to insults known to induce lipid peroxidation (FeSO4 and amyloid beta-peptide) also impaired glutamate uptake and mitochondrial function. The antioxidants propyl gallate and glutathione prevented impairment of glutamate uptake and MTT reduction induced by FeSO4 and amyloid beta-peptide, but not that induced by 4-hydroxynonenal. Western blot analyses using an antibody to 4-hydroxynonenal-conjugated proteins showed that 4-hydroxynonenal bound to multiple cell proteins including GLT-1, a glial glutamate transporter present at high levels in synaptosomes. 4-Hydroxynonenal itself induced lipid peroxidation suggesting that, in addition to binding directly to membrane regulatory proteins, 4-hydroxynonenal potentiates oxidative cascades. Collectively, these findings suggest that 4-hydroxynonenal plays important roles in oxidative impairment of synaptic functions that would be expected to promote excitotoxic cascades.

ATP-Binding Cassette Transporters↗

PubMed1997

4-hydroxynonenal, a lipid peroxidation product, rapidly accumulates following traumatic spinal cord injury and inhibits glutamate uptake.

Traumatic injury to the spinal cord initiates a host of pathophysiological events that are secondary to the initial insult. One such event is the accumulation of free radicals that damage lipids, proteins, and nucleic acids. A major reactive product formed following lipid peroxidation is the aldehyde, 4-hydroxynonenal (HNE), which cross-links to side chain amino acids and inhibits the function of several key metabolic enzymes. In the present study, we used immunocytochemical and immunoblotting techniques to examine the accumulation of protein-bound HNE, and synaptosomal preparations to study the effects of spinal cord injury and HNE formation on glutamate uptake. Protein-bound HNE increased in content in the damaged spinal cord at early times following injury (1-24 h) and was found to accumulate in myelinated fibers distant to the site of injury. Immunoblots revealed that protein-bound HNE levels increased dramatically over the same postinjury interval. Glutamate uptake in synaptosomal preparations from injured spinal cords was decreased by 65% at 24 h following injury. Treatment of control spinal cord synaptosomes with HNE was found to decrease significantly, in a dose-dependent fashion, glutamate uptake, an effect that was mimicked by inducers of lipid peroxidation. Taken together, these findings demonstrate that the lipid peroxidation product HNE rapidly accumulates in the spinal cord following injury and that a major consequence of HNE accumulation is a decrease in glutamate uptake, which may potentiate neuronal cell dysfunction and death through excitotoxic mechanisms.

ATP-Binding Cassette Transporters↗

PubMed1997

Impairment of glucose and glutamate transport and induction of mitochondrial oxidative stress and dysfunction in synaptosomes by amyloid beta-peptide: role of the lipid peroxidation product 4-hydroxynonenal.

Deposits of amyloid beta-peptide (A beta), reduced glucose uptake into brain cells, oxidative damage to cellular proteins and lipids, and excitotoxic mechanisms have all been suggested to play roles in the neurodegenerative process in Alzheimer's disease. Synapse loss is closely correlated with cognitive impairments in Alzheimer's disease, suggesting that the synapse may be the site at which degenerative mechanisms are initiated and propagated. We report that A beta causes oxyradical-mediated impairment of glucose transport, glutamate transport, and mitochondrial function in rat neocortical synaptosomes. A beta induced membrane lipid peroxidation in synaptosomes that occurred within 1 h of exposure; significant decreases in glucose transport occurred within 1 h of exposure to A beta and decreased further with time. The lipid peroxidation product 4-hydroxynonenal conjugated to synaptosomal proteins and impaired glucose transport; several antioxidants prevented A beta-induced impairment of glucose transport, indicating that lipid peroxidation was causally linked to this adverse action of A beta. FeSO4 (an initiator of lipid peroxidation), A beta, and 4-hydroxynonenal each induced accumulation of mitochondrial reactive oxygen species, caused concentration-dependent decreases in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction, and reduced cellular ATP levels significantly. A beta also impaired glutamate transport, an effect blocked by antioxidants. These data suggest that A beta induces membrane lipid peroxidation, which results in impairment of the function of membrane glucose and glutamate transporters, altered mitochondrial function, and a deficit in ATP levels; 4-hydroxynonenal appears to be a mediator of these actions of A beta. These data suggest that oxidative stress occurring at synapses may contribute to the reduced glucose uptake and synaptic degeneration that occurs in Alzheimer's disease patients. They further suggest a sequence of events whereby oxidative stress promotes excitotoxic synaptic degeneration and neuronal cell death in a variety of different neurodegenerative disorders.

Aldehydes↗

PubMed1997

4-Hydroxynonenal, an aldehydic product of lipid peroxidation, impairs signal transduction associated with muscarinic acetylcholine and metabotropic glutamate receptors: possible action on G alpha(q/11).

Considerable data indicate that oxidative stress and membrane lipid peroxidation contribute to neuronal degeneration in an array of age-related neurodegenerative disorders. In contrast, the impact of subtoxic levels of membrane lipid peroxidation on neuronal function is largely unknown. We now report that 4-hydroxynonenal (HNE), an aldehydic product of lipid peroxidation, disrupts coupling of muscarinic cholinergic receptors and metabotropic glutamate receptors to phospholipase C-linked GTP-binding proteins in cultured rat cerebrocortical neurons. At subtoxic concentrations, HNE markedly inhibited GTPase activity, inositol phosphate release, and elevation of intracellular calcium levels induced by carbachol (muscarinic agonist) and (RS)-3,5-dihydroxyphenyl glycine (metabotropic glutamate receptor agonist). Maximal impairment of agonist-induced responses occurred within 30 min of exposure to HNE. Other aldehydes, including malondialdehyde, had little effect on agonist-induced responses. Antioxidants that suppress lipid peroxidation did not prevent impairment of agonist-induced responses by HNE, whereas glutathione, which is known to bind and detoxify HNE, did prevent impairment of agonist-induced responses. HNE itself did not induce oxidative stress. Immunoprecipitation-western blot analysis using an antibody to HNE-protein conjugates showed that HNE can bind to G alpha(q/11). HNE also significantly suppressed inositol phosphate release induced by aluminum fluoride. Collectively, our data suggest that HNE plays a role in altering receptor-G protein coupling in neurons under conditions of oxidative stress that may occur both normally, and before cell degeneration and death in pathological settings.

Aldehydes↗

PubMed1997

The lipid peroxidation product, 4-hydroxy-2-trans-nonenal, alters the conformation of cortical synaptosomal membrane proteins.

Alzheimer's disease (AD) is widely held to be a disorder associated with oxidative stress due, in part, to the membrane action of amyloid beta-peptide (A beta). A beta-associated free radicals cause lipid peroxidation, a major product of which is 4-hydroxy-2-trans-nonenal (HNE). We determined whether HNE would alter the conformation of synaptosomal membrane proteins, which might be related to the known neurotoxicity of A beta and HNE. Electron paramagnetic resonance spectroscopy, using a protein-specific spin label, MAL-6 (2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl), was used to probe conformational changes in gerbil cortical synaptosomal membrane proteins, and a lipid-specific stearic acid label, 5-nitroxide stearate, was used to probe for HNE-induced alterations in the fluidity of the bilayer domain of these membranes. Synaptosomal membranes, incubated with low concentrations of HNE, exhibited changes in protein conformation and bilayer order and motion (fluidity). The changes in protein conformation were found to be concentration- and time-dependent. Significant protein conformational changes were observed at physiologically relevant concentrations of 1-10 microM HNE, reminiscent of similar changes in synaptosomal membrane proteins from senile plaque- and A beta-rich AD hippocampal and inferior parietal brain regions. HNE-induced modifications in the physical state of gerbil synaptosomal membrane proteins were prevented completely by using excess glutathione ethyl ester, known to protect neurons from HNE-caused neurotoxicity. Membrane fluidity was found to increase at higher concentrations of HNE (50 microM). The results obtained are discussed with relevance to the hypothesis of A beta-induced free radical-mediated lipid peroxidation, leading to subsequent HNE-induced alterations in the structure and function of key membrane proteins with consequent neurotoxicity in AD brain.

Aldehydes↗

PubMed1997

Immunological evidence for hypochlorite-modified proteins in human kidney.

Oxygen radicals and oxidatively modified proteins seem to participate in degenerative vascular and inflammatory diseases. Factors that contribute to the development of atherosclerosis, eg, oxidation of low-density lipoproteins (LDLs), may also contribute to glomerulosclerosis. Although the nature of the in vivo oxidants remains unknown, recent findings indicated that the myeloperoxidase (MPO)-H2O2-halide system could play an important role in modification of (lipo)proteins in human tissues. MPO, the enzyme responsible for hypochlorite (HOCl/OCl-) formation, is present in human atherosclerotic lesions and in inflammatory conditions. In the present study, MPO was identified by Western blot analysis and immunohistochemical technique in diseased human kidney either with primarily sclerotic or inflammatory lesions. Furthermore, the presence of HOCl-modified proteins was demonstrated in diseased renal tissues using a specific monoclonal antibody (clone 2D10G9), raised against HOCl-modified LDL, that does not cross-react with native LDL or Cu(2+)-, 4-hydroxynonenal-, or malondialdehyde-modified LDL. The antibody recognized HOCl-modified proteins in glomerular and tubulointerstitial inflammatory and fibrotic lesions and pronounced immunostaining was demonstrated in mononuclear cells. LDL or human serum albumin oxidized by HOCl in vitro, but not native LDL or human serum albumin, effectively competed with epitopes in diseased kidney for antibody binding. Western blot analysis in diseased kidney protein samples revealed at least two major proteins recognized by the anti-HOCl-modified protein monoclonal antibody. Densitometric evaluation of immunoreactive bands obtained under these conditions demonstrated that expression of HOCl-modified proteins is tightly coupled to expression of immunoreactive MPO in the same tissue samples. From our studies it is proposed that oxidation of proteins by HOCl might be a leading event in glomerular and tubulointerstitial injury. By this mechanism, mononuclear cells, a permanent source for MPO, may play a key role in the development of nephrosclerosis, glomerulo-clerosis, and tubulointerstitial fibrosis, respectively.

Adult↗

PubMed1996

Cytotoxic and chemotactic potencies of several aldehydic components of oxidised low density lipoprotein for human monocyte-macrophages.

We have investigated the cytotoxic and chemotactic potencies of malondialdehyde (MDA), hexanal, 4-hydroxyhexenal (HHE), 4-hydroxynonenal (HNE) and 4-hydroxyoctenal (HOE), which are aldehydes found in oxidised low density lipoprotein (LDL), for human monocyte-macrophages. They were toxic in the following order: hexanal<HHE= HOE< HNE. HNE was toxic at 20 microM and chemotactic at 2.5 microM. The other aldehydes tested had no chemoattractant activity. Our results suggest that HNE arising from LDL oxidation could attract monocytes into the human atherosclerotic lesion. A direct cytotoxic role of aldehydes in foam cell death in the lesion is less likely.

Aldehydes↗