Aldehyde Acrolein Research Articles

Mechanisms of acrolein-induced myocardial dysfunction: implications for environmental and endogenous aldehyde exposure

Aldehydes are ubiquitous pollutants generated during the combustion of organic materials and are present in air, water, and food. Several aldehydes are also endogenous products of lipid peroxidation and by-products of drug metabolism. Despite well-documented high reactivity of unsaturated aldehydes, little is known regarding their cardiovascular effects and their role in cardiac pathology. Accordingly, we examined the myocardial effects of the model unsaturated aldehyde acrolein. In closed-chest mice, intravenous acrolein (0.5 mg/kg) induced rapid but reversible left ventricular dilatation and dysfunction. In mouse myocytes, micromolar acrolein acutely depressed myofilament Ca(2+) responsiveness without altering catecholamine sensitivity, similar to the phenotype of stunned myocardium. Immunoblotting revealed increased acrolein-protein adducts and protein-carbonyls in both acrolein-exposed myocardium (1.8-fold increase, P < 0.002) and myocytes (6.4-fold increase, P < 0.02). Both the contractile dysfunction and adduct formation were markedly attenuated by pretreatment with the thiol donor N-acetylcysteine (5 mM). Two-dimensional gel electrophoresis and mass-assisted laser desorption/ionization time-of-flight mass spectrometry analysis revealed two groups of adducted proteins, sarcomeric/cytoskeletal proteins (cardiac alpha-actin, desmin, myosin light polypeptide 3) and energy metabolism proteins (mitochondrial creatine kinase-2, ATP synthase), indicating site-specific protein modification that was confirmed by immunohistochemical colocalization. We conclude that direct exposure to acrolein induces selective myofilament impairment, which may be, in part, related to the modification of proteins involved in myocardial contraction and energy metabolism. Myocardial dysfunction induced by acrolein and related aldehydes may be symptomatic of toxicological states associated with ambient or occupational exposures or drug toxicity. Moreover, aldehydes such as acrolein may mediate cardiac dysfunction in pathologies characterized by high-oxidative stress.

Kinetics and Mechanism of Protein Tyrosine Phosphatase 1B Inactivation by Acrolein

Human cells are exposed to the electrophilic alpha,beta-unsaturated aldehyde acrolein from a variety of sources. The reaction of acrolein with functionally critical protein thiol residues can yield important biological consequences. Protein tyrosine phosphatases (PTPs) are an important class of cysteine-dependent enzymes whose reactivity with acrolein previously has not been well-characterized. These enzymes catalyze the dephosphorylation of phosphotyrosine residues on proteins via a phosphocysteine intermediate. PTPs work in tandem with protein tyrosine kinases to regulate a number of critically important mammalian signal transduction pathways. We find that acrolein is a potent time-dependent inactivator of the enzyme PTP1B ( k inact = 0.02 +/- 0.005 s (-1) and K I = 2.3 +/- 0.6 x 10 (-4) M). The enzyme activity does not return upon gel filtration of the inactivated enzyme, and addition of the competitive phosphatase inhibitor vanadate slows inactivation of PTP1B by acrolein. Together, these observations suggest that acrolein covalently modifies the active site of PTP1B. Mass spectrometric analysis reveals that acrolein modifies the catalytic cysteine residue at the active site of the enzyme. Aliphatic aldehydes such as glyoxal, acetaldehyde, and propanal are relatively weak inactivators of PTP1B under the conditions employed here. Similarly, unsaturated aldehydes such as crotonaldehyde and 3-methyl-2-butenal bearing substitution at the alkene terminus are poor inactivators of the enzyme. Overall, the data suggest that enzyme inactivation occurs via conjugate addition of the catalytic cysteine residue to the carbon-carbon double bond of acrolein. The results indicate that inactivation of PTPs should be considered as a possible contributor to the diverse biological activities of acrolein and structurally related alpha,beta-unsaturated aldehydes.

Prostacyclin Prevents Pulmonary Endothelial Cell Apoptosis Induced by Cigarette Smoke

Impaired endothelial cell-dependent vasodilation, inflammation, apoptosis, and proliferation are manifestations of endothelial dysfunction in chronic obstructive pulmonary disease (COPD). Prostacyclin (PGI(2)) is a major product of the cyclooxygenase pathway with potent vasodilatory and antimitogenic properties and may be relevant to endothelial dysfunction in COPD. To determine if PGI(2) expression is altered in smoking-related lung disease and if it may be protective in COPD-associated endothelial dysfunction. We evaluated, by immunohistochemistry, Western blotting, and polymerase chain reaction, human emphysema tissue compared with normal tissue for expression of prostacyclin synthase (PGI(2)S). We examined the effects of cigarette smoke extract (CSE) and aldehyde components on eicosanoid expression in primary human pulmonary microvascular endothelial cells. Finally, we used a murine model of lung-specific PGI(2)S overexpression and in vitro studies to determine if PGI(2) expression has protective effects on cigarette smoke-induced endothelial apoptosis. Human emphysema lung tissue exhibited lower PGI(2)S expression within the pulmonary endothelium than in normal lung. In vitro studies demonstrated that CSE, and in particular the alpha,beta unsaturated aldehyde acrolein, suppressed PGI(2)S gene expression, whereas CSE significantly induced the upstream mediators COX-2 and cytosolic phospholipase A2 in human pulmonary microvascular endothelial cells. Mice with lung-specific PGI(2)S overexpression exhibited less endothelial apoptosis after chronic smoke exposure. In vitro, iloprost exhibited protective effects on CSE-induced apoptosis. PGI(2) has protective effects in the pulmonary vasculature after acute and chronic cigarette smoke exposure. An imbalance in eicosanoid expression may be important to COPD-associated endothelial dysfunction.

GSH-dependent regulation of Fas-mediated caspase-8 activation by acrolein

Activation of the cysteine protease caspase-8 by the death receptor Fas (CD95/APO-1) in B lymphoblastoid SKW6.4 cells or Jurkat T cells is associated with GSH depletion. Conversely, GSH depletion by the aldehyde acrolein (3–30 μM) was associated with inhibition of Fas-induced caspase-8 activation, although GSH depletion by buthionine sulfoximine (BSO) did not affect caspase-8 activation. In contrast to BSO, acrolein caused a loss of caspase-8 cysteine content in association with direct alkylation of caspase-8. Our findings indicate that inhibition of caspase-8 by thiol-reactive agents such as acrolein is not due to GSH depletion but caused by direct protein thiol modifications.

Activation of the death receptor pathway of apoptosis by the aldehyde acrolein

Reactive α,β-unsaturated aldehydes such as acrolein are major components of common environmental pollutants. As a toxic by-product of lipid peroxidation, acrolein has been implicated as a possible mediator of oxidative damage to cells and tissues in a wide variety of disease states, including atherosclerosis and neurodegenerative and pulmonary diseases. Although acrolein can induce apoptotic cell death in various cell types, the biochemical mechanisms are not understood. This study investigates the implication of the death receptor pathway in acrolein-induced apoptosis. Exposure of Chinese hamster ovary cells to acrolein caused translocation of adaptor protein Fas associated with death domain to the cytoplasmic membrane and caspase-8 activation. Kp7-6, an antagonist of Fas receptor activation, blocked apoptotic events downstream of caspase-8, such as caspase-7 activation and nuclear chromatin condensation. Acrolein activated the cross-talk pathway between the death receptor and mitochondrial pathways. Bid was cleaved to truncated-Bid, which was translocated to mitochondria. Activation of the mitochondrial pathway by acrolein was confirmed by caspase-9 activation. Inhibition of activation of either the Fas receptor or caspase-8 partially decreased acrolein-induced caspase-9 activation. These findings indicate that acrolein activates the Fas receptor pathway, which occurs upstream of the mitochondrial pathway. Caspase-9 activation still occurred despite inhibition of the Fas receptor pathway, suggesting that acrolein could also trigger the mitochondrial pathway independent of the receptor pathway. These findings improve our understanding of mechanisms of toxicity of the reactive aldehyde acrolein, which has widespread implications in multiple disease states which seem to be mediated by oxidative stress and lipid peroxidation.

Regulation of constitutive neutrophil apoptosis by the α,β-unsaturated aldehydes acrolein and 4-hydroxynonenal

Reactive alpha,beta-unsaturated aldehydes are major components of common environmental pollutants and are products of lipid oxidation. Although these aldehydes have been demonstrated to induce apoptotic cell death in various cell types, we recently observed that the alpha,beta-unsaturated aldehyde acrolein (ACR) can inhibit constitutive apoptosis of polymorphonuclear neutrophils and thus potentially contribute to chronic inflammation. The present study was designed to investigate the biochemical mechanisms by which two representative alpha,beta-unsaturated aldehydes, ACR and 4-hydroxynonenal (HNE), regulate neutrophil apoptosis. Whereas low concentrations of either aldehyde (<10 microM) mildly promoted apoptosis in neutrophils (reflected by increased phosphatidylserine exposure, caspase-3 activation, and mitochondrial cytochrome c release), higher concentrations prevented critical features of apoptosis (caspase-3 activation, phosphatidylserine exposure) and caused delayed neutrophil cell death with characteristics of necrosis/oncosis. Inhibition of caspase-3 activation by either aldehyde occurred despite increases in mitochondrial cytochrome c release and occurred in close association with depletion of cellular GSH and with cysteine modifications within caspase-3. However, procaspase-3 processing was also prevented, because of inhibited activation of caspases-9 and -8 under similar conditions, suggesting that ACR (and to a lesser extent HNE) can inhibit both intrinsic (mitochondria dependent) and extrinsic mechanisms of neutrophil apoptosis at initial stages. Collectively, our results indicate that alpha,beta-unsaturated aldehydes can inhibit constitutive neutrophil apoptosis by common mechanisms, involving changes in cellular GSH status resulting in reduced activation of initiator caspases as well as inactivation of caspase-3 by modification of its critical cysteine residue.

Research advances in the catalysts for the selective oxidation of ethane to aldehydes

Selective oxidation of ethane to aldehydes is one of the most difficult processes in the catalysis researches of low alkanes. The development of selective oxidation of ethane to aldehydes (formaldehyde, acetaldehyde and acrolein) is discussed. The latest progress of the catalysts, including bulk or supported metal oxide catalysts, highly dispersed and isolated active sites catalysts, and the photo-catalytic ethane oxidation catalysts, partial oxidation of ethane in the gas phase, and the proposed reaction pathways from ethane to aldehydes are involved.

Kinetics and products of the OH radical-initiated reaction of 3-methyl-2-butenal

Kinetics and products of the gas-phase reaction of OH radicals with 3-methyl-2-butenal [(CH3)2C=CHCHO] have been investigated at room temperature and atmospheric pressure of air. Using a relative rate method with methacrolein as the reference compound, a rate constant for the reaction of OH radicals with 3-methyl-2-butenal of (6.21 +/- 0.18) x 10(-11) cm3 molecule(-1) s(-1) at 296 +/- 2 K was measured, where the indicated error does not include the uncertainty in the rate constant for the methacrolein reference compound. Products of this reaction were investigated using in situ Fourier transform infrared (FT-IR) spectroscopy and solid phase microextraction (SPME) fibers coated with O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine for on-fiber derivatization of carbonyl compounds, with subsequent thermal desorption and analysis by gas chromatography. The products observed and the molar formation yields were: glyoxal, 40 +/- 3%; acetone, 74 +/- 6%; 2-hydroxy-2-methylpropanal, 4.6 +/- 0.7%; CO2, 39% initially, decreasing to 30% at greater extents of reaction; peroxyacyl nitrate(s) [RC(O)OONO2], 5-8%, increasing with the extent of reaction and with the sum of the CO2 and RC(O)OONO2 yields being 38 +/- 6%; and organic nitrates [RONO2], 8.5 +/- 2.3%. The formation of these products is readily explained by a reaction mechanism based on those previously formulated for the corresponding reactions of the alpha,beta-unsaturated aldehydes acrolein, crotonaldehyde and methacrolein. Based on the mechanism proposed, at room temperature H-atom abstraction from the CHO group accounts for 40 +/- 6% of the overall reaction, and OH radical addition to the carbon atoms of the C=C bond accounts for 53 +/- 4% of the overall reaction. Hence 93 +/- 8% of the reaction products and pathways are accounted for.

Reactions of CH-acids with , -unsaturated aldehydes in ionic liquids

Metal carbonate-catalyzed reactions of CH-acids (diethyl malonate, ethyl acetoacetate, ethyl cyanoacetate, and ethyl 2-acetyl- and 2-ethoxycarbonyl-5,9-dimethyldeca-4,8-dienoates) with α,β-unsaturated aldehydes (acrolein, crotonaldehyde, citral) were studied in an ionic liquid, 1-butyl-3-methylimidazolium hexafluorophosphate [bmim][PF6], and in a 1-butyl-3-methylimidazolium bromide ([bmim][Br]) — benzene system. The reactions with acrolein and crotonaldehyde afforded Michael addition products, those with citral resulted in Knoevenagel addition products. Sonication increased the yields of the Michael adducts. The ionic liquid [bmim][PF6] can be recovered and repeatedly used in the reactions.

Reactivity with Tris(hydroxymethyl)aminomethane confounds immunodetection of acrolein-adducted proteins.

The toxic alpha,beta-unsaturated aldehyde acrolein readily attacks proteins, generating adducts at cysteine, histidine, and lysine residues. In this study, rabbit antiserum was raised against acrolein-modified keyhole limpet hemocyanin in the expectation that it would allow immunodetection of adducted proteins in biological samples. Using slot-blot and enzyme-linked immunosorbent assays, the antiserum detected acrolein-modified protein with high sensitivity and specificity. Adduct immunodetection was strongly inhibited by acrolein-modified polylysine but not polyhistidine. Efforts to develop a Western blotting method for detecting adducted proteins in cell lysates were hampered by irreproducible outcomes, evidently due to adduct instability during SDS-PAGE. Indeed, adducts generated via brief exposure of a model protein to acrolein displayed pH- and concentration-dependent instability to tris(hydroxymethyl)aminomethane (Tris), a nucleophilic buffer used in protein electrophoresis. The effect was most striking when Tris solutions were buffered to pH 8.0 and higher. In contrast, adducts formed during extended exposure to acrolein (> or =60 min) were completely stable to Tris. The time dependence of susceptibility raised the possibility that Tris interfered with specific steps in lysine modification, which involves stepwise Michael addition of two molecules of acrolein to the same residue, followed by condensation and dehydration to form a heterocyclic adduct, N(epsilon)-(3-formyl-3,4-dehydropiperidino)lysine. We hypothesize that carbonyl-retaining Michael adducts may react with Tris by forming imines with the primary amine of the buffer. Consistent with this idea, triethanolamine, a tertiary amine buffer unable to form imines, had no effect on acrolein-adducted protein. These effects of Tris may explain difficulties in the detection of acrolein-adducted proteins during conventional Western blotting procedures.

Acrolein-sequestering ability of endogenous dipeptides: characterization of carnosine and homocarnosine/acrolein adducts by electrospray ionization tandem mass spectrometry.

Acrolein (ACR), the carbonyl toxin produced by lipid peroxidation, is significantly increased in Alzheimer's disease brain. Since ACR is one of the most reactive and neurotoxic aldehydes, and human brain contains both carnosine (beta-alanine-L-histidine) and homocarnosine (gamma-aminobutyryl-L-histidine), the aim of this work was first to evaluate the quenching ability of the two peptides towards ACR and then to characterize their reaction products by electrospray ionization tandem mass spectrometry (ESI-MS/MS; infusion experiments; positive-ion mode). The reaction progress of ACR with carnosine or homocarnosine was studied in phosphate buffer, by monitoring ACR consumption (by reverse-phase LC) and formation of the reaction products by ESI-MS/MS at different incubation times. N-Acetylcarnosine was used as reference compound to identify the sites of reaction. Both the dipeptides were able to quench ACR by almost 60% at 1 h and by more than 85% after 3 h incubation. Different reaction products between ACR and carnosine/homocarnosine were detected after 3 and 24 h, to indicate a complex reaction pathway involving sequential addition of 1, 2 and 3 moles of ACR/mole of the dipeptide to both the beta-alanine and histidine residues. The ESI mass spectra of ACR/carnosine reaction mixtures indicate formation of several molecular species, among which the predominant are: (a) the 14-membered macrocyclic derivatives, deriving from the formation of the iminic bond between the terminal amino group followed by intramolecular Michael addition of the C(3) of the ACR moiety to histidine; (b) the N(beta)-(3-formyl-3,4-dehydropiperidino) derivatives arising from the Michael addition of two acrolein molecules to the amino group of beta-alanine, followed by an aldol condensation and dehydration.The reaction of homocarnosine with ACR follows the same pathway, giving rise to the formation of homologous adducts. The results of this study shed light on the mechanism, until now never demonstrated, through which carnosine and homocarnosine detoxify the highly reactive aldehyde acrolein in a buffer system, and represent the starting point for further studies aimed at elucidating the biological role of these dipeptides in brain.

Inhibition of succinic semialdehyde dehydrogenase activity by alkenal products of lipid peroxidation

Lipid peroxidation causes the generation of the neurotoxic aldehydes acrolein and 4-hydroxy- trans-2-nonenal (HNE). These products are elevated in neurodegenerative diseases and acute CNS trauma. Previous studies demonstrate that mitochondrial class 2 aldehyde dehydrogenase (ALDH2) is susceptible to inactivation by these alkenals. In the liver and brain another mitochondrial aldehyde dehydrogenase, succinic semialdehyde dehydrogenase (SSADH/ALDH5A1), is present. In this study, we tested the hypothesis that aldehyde products of lipid peroxidation inhibit SSADH activity using the endogenous substrate, succinic semialdehyde (SSA, 50 μM). Acrolein potently inhibited SSADH activity (IC 50=15 μM) in rat brain mitochondrial preparations. This inhibition was of an irreversible and noncompetitive nature. HNE inhibited activity with an IC 50 of 110 μM. Trans-2-hexenal (HEX) and crotonaldehyde (100 μM each) did not inhibit activity. These data suggest that acrolein and HNE disrupt SSA metabolism and may have subsequent effects on CNS neurochemistry.

The influence of the solvents DMSO and ethanol on the genotoxicity of α,β-unsaturated aldehydes in the SOS chromotest

α,β-Unsaturated aldehydes are ubiquitous environmental pollutants, important industrial chemicals, have mani-fold biological functions in plants and insects and are natural products in food. They are endogenously formed in animals and humans during lipid peroxidation and arachidonic acid oxidation and are genotoxic, mutagenic and carcinogenic. Crotonaldehyde and 2-hexenal in food may contribute to general carcinogenicity in humans. The high bacterial toxicity of these compounds leads to problems in genotoxicity testing in bacterial systems. Recently, we have shown that using ethanol as solvent instead of dimethylsulfoxide (DMSO) results in an increase in the induction factors and the SOS-inducing potency of α,β-unsaturated ketones in the SOS chromotest. Here, we demonstrate that utilization of ethanol as solvent also improves the testing of α,β-unsaturated aldehydes. Five aldehydes out of nine tested were clearly positive in the SOS chromotest according to the criteria of Quillardet, i.e. acrolein, crotonaldehyde, 2,4-hexadienal, 2-methylacrolein and 2-ethylacrolein, three further, 2-hexenal, 2-heptenal and 2-propylacrolein showed a dose dependent increase of the induction factors which was however lower than 1.5 times that of the background. Only 2-butylacrolein did not lead to an increase in the induction factors. With DMSO as solvent only the three aldehydes acrolein, crotonaldehyde and 2,4-hexadienal showed an increase in the induction factor, which was however lower than 1.5 that of the background. Utilization of ethanol allows to establish structure genotoxicity relationships for α,β-unsaturated aldehydes in the SOS chromotest. Genotoxicity decreases with increasing degree of substitution. The decreasing genotoxicities can be explained (a) by increasing bacterial toxicity due to increasing lipophilicities of the higher substituted aldehydes and (b) by decreasing reactivity due to steric hindrance by the alkyl substituents.

In vitro effect of air pollutants on human bronchi.

A useful approach for constructing dose-response relationships and for studying the underlying mechanisms by which a xenobiotic agent enhances airway reactivity is to measure the response of an isolated airway following ex vivo exposure to a pollutant. We have in this way determined the dose-response relationship between ex vivo exposure to pollutants such as nitrogen dioxide (NO2), the aldehyde acrolein, and ozone (O3) and the reactivity to agonists in human isolated bronchial smooth muscle. We have also investigated the underlying alteration in the cellular mechanisms of airway smooth-muscle contraction induced by such exposure and found that it is related to alteration in calcium signaling at the site of the airway smooth-muscle cell. Finally, although there is epidemiological evidence that an increase in allergic diseases such as asthma may be linked to air pollution, there are few experimental data to address this issue. The final aim of this study was therefore to investigate the interaction between passive sensitization and exposure to pollutants in human isolated airways. We have examined (i) the effect of a pre-exposure to pollutants on the contraction of sensitized bronchi in response to a specific antigen and (ii) the effect of passive sensitization on the contraction in response to nonspecific agonists in bronchi pre-exposed to pollutants. The results indicate a combined effect of immunological sensitization and exposure to pollutants; that is, passive sensitization and exposure to pollutants act in a synergistic manner on human bronchial smooth-muscle reactivity in response to both specific antigens and nonspecific agonists.

Selected contribution: effect of the aldehyde acrolein on acetylcholine-induced membrane current in airway smooth muscle cells.

Acrolein administered to isolated airways has been shown to alter airway responsiveness as a consequence of its effect on Ca(2+) signaling. To examine the mechanisms involved, we studied the effect of acrolein on ACh- and caffeine-induced membrane currents (patch-clamp) in myocytes freshly isolated from rat trachea. In cells clamped at -60 mV, ACh (0.1-10 microM) induced a concentration-dependent inward current, which, in approximately 50% of the cells, was followed by current oscillations in response to high concentration of ACh (10 microM). Exposure to acrolein (0.2 microM) for 10 min significantly enhanced the amplitude of the low-ACh (0.1 microM) concentration-induced initial peak of current (318.8 +/- 28.3 vs. 251.2 +/- 40.3 pA; n = 25, P < 0.05). At a high-ACh concentration (10 microM), the frequency at which subsequent peaks occurred was significantly increased (13.2 +/- 1.1 vs. 8.7 +/- 2 min(-1); n = 20, P < 0.05). ACh-induced current was identified as a Ca(2+)-activated Cl(-) current. In contrast, similar exposure to acrolein, which does not alter caffeine-induced Ca(2+) release, did not alter caffeine-induced transient membrane currents (595 +/- 45 and 640 +/- 45 pA in control cells and in cells exposed to acrolein, respectively; n = 15). It is concluded that acrolein alters ACh-induced current as a consequence of its effect on the cytosolic Ca(2+) concentration response and that the protective role of inhibitors of Cl(-) channels in air pollutant-induced airway hyperresponsiveness should be examined.

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Interactions of α, β-unsaturated aldehydes and ketones with human glutathione S-transferase P1-1

In the present study the irreversible inhibition of human glutathione S-transferase P1-1 (GSTP1-1) by α, β-unsaturated aldehydes and ketones was studied. When GSTP1-1 was incubated with a 50-fold molar excess of the aldehydes acrolein (ACR) and 4-hydroxy-2-nonenal (HNE) and the ketones curcumin (CUR) and ethacrynic acid (EA) at 22°C, all of them inactivated GSTP1-1. The remaining activity after 4 h of incubation in all cases was lower than 10%. The aldehydes crotonaldehyde (CRA), cinnamaldehyde (CA) and trans-2-hexenal were found to inhibit GSTP1-1 only at a 5000-fold molar excess and even then, for example, for CA a higher remaining activity of 17% was observed. The same inhibition experiments were conducted with 3 mutants of GSTP1-1: the C47S and C101S mutants and the double mutant C47S/C101S. Remaining activity for C47S varied between ±40% for CRA, CA, CUR and HEX and ±80% for ACR, EA and HNE. For C101S it varied between 0 and 9% and for the double mutant C47S/C101S, activity after 4 h of incubation was variable. Again it varied between ±40% for CRA, CA, CUR and HEX and ±80% for ACR, EA and HNE. EA is known to react almost exclusively with cysteine 47. When [ 14C]EA was incubated with the GSTP1-1, modified by the α, β-unsaturated carbonyl compounds, no [ 14C]EA was incorporated in the enzyme, indicating that in all cases this cysteine residue was one of the major targets. Since Michael addition with these reagents is known to be reversible, the results of incubation of the inactivated enzymes with an excess of glutathione (GSH) were determined. For all compounds, a restoration of the catalytic activity was observed. The results indicate that α, β-unsaturated carbonyl derivatives inhibit GSTP1-1 irreversibly mainly by binding to cysteine residues of GSTP1-1, especially Cys-47. This means that some of these compounds (e.g. CUR) might modify GST activity in vivo when GSH concentrations are low by covalent binding to the enzyme.

Human bronchial smooth muscle responsiveness after in vitro exposure to oxidizing pollutants.

The aims of this work were (1) to determine the dose-response relationship between ex vivo exposure to oxidizing pollutants such as nitrogen dioxide (NO2), the aldehyde acrolein, and ozone (O3), and the reactivity to agonists in isolated human bronchial smooth muscle; and (2) to investigate the alterations in the cellular mechanisms of human airway smooth muscle contraction induced by such exposures. Experiments were performed in isolated human bronchi obtained at thoracotomy. Isometric contraction in response to a variety of agonists was compared between pollutant-exposed preparations and paired controls. Short exposures to NO2, acrolein, or O3 altered the subsequent airway smooth muscle responsiveness in a dose-dependent manner. The cellular mechanisms producing the airway hyperresponsiveness observed in vitro are shared by the three pollutants and include alterations in airway smooth muscle excitation-contraction coupling as well as indirect effects on neutral endopeptidase activity.

Glucose, glutathione, and cellular response to spermine oxidation products

Bovine serum amineoxidase (BSAO) oxidatively deaminates polyamines, which contain primary amine groups with formation of several toxic products, H 2O 2, and aldehyde (s). We evaluated the role of glucose metabolism via the pentose phosphate cycle and the level of intracellular glutathione on cytotoxicity induced by each of the toxic products in Chinese hamster ovary (CHO) cells. Glucose protected cells against cytotoxicity in the presence of BSAO at low spermine concentrations (< 50 pM), where H 20 2 was the only toxic species present. When catalase was present, cytotoxicity is attributed to spermine-derived aldehyde(s). Glucose did not protect cells against cytotoxicity induced by spermine-derived aldehyde (s), nor by the aldehyde acrolein. Hydrogen peroxide produced by spermine and BSAO stimulated pentose cycle activity, whereas the aldehyde(s) did not. Depletion of intracellular glutathione with L-buthionine sulfoximine (1 mM, 24 h) sensitized cells to the cytotoxic effects of both H 20 2 and the aldehyde(s) produced by spermine and BSAO. The pentose cycle and the glutathione redox cycle have an important role in protection against H 20 2 generated from spermine oxidation. Glutathione appears to have a role in protecting cells against cytotoxicity attributed to spermine-derived aldehyde(s), most likely by conjugation in a reaction catalyzed by glutathione S-transferase, whereas metabolism of glucose via the pentose cycle did not. The metabolism of both glucose and glutathione, affect the cellular response to H 20 2 and aldehyde (s) derived from spermine, although different pathways are involved.

Inhibition of Lecithin-Cholesterol Acyltransferase and Modification of HDL Apolipoproteins by Aldehydes

Experimental evidence suggests that aldehydes generated as a consequence of lipid peroxidation may be involved in the pathogenesis of atherosclerosis. It is well documented that aldehydes modify LDL: however, less is known concerning the effects of aldehydes on other plasma and interstitial fluid components. In the present study, we investigated the effects of five physiologically relevant aldehydes (acetaldehyde, acrolein, hexanal, 4-hydroxynonenal [HNE], and malondialdehyde [MDA]) on two key constituents of the antiatherogenic reverse cholesterol transport pathway, lecithin-cholesterol acyltransferase (LCAT) and HDL. Human plasma was incubated for 3 hours at 37 degrees C with each one of the five aldehydes at concentrations ranging from 0.16 to 84 mmol/L. Dose-dependent decreases in LCAT activity were observed. The short-chain (acrolein) and long-chain (HNE) alpha,beta-unsaturated aldehydes were the most effective LCAT inhibitors. Micromolar concentrations of these unsaturated aldehydes resulted in significant reductions in plasma LCAT activity. The short- and longer-chain saturated aldehydes acetaldehyde and hexanal and the dialdehyde MDA were considerably less effective at inhibiting LCAT than were acrolein and HNE. In addition to inhibiting LCAT, aldehydes increased HDL electrophoretic mobility and cross-linked HDL apolipoproteins. Cross-linking of apolipoproteins A-I and A-II required higher aldehyde concentrations than inhibition of LCAT. The alpha,beta-unsaturated aldehydes acrolein and HNE were fourfold to eightfold more effective cross-linkers of apolipoproteins A-I and A-II than the other aldehydes studied. These data suggest that products of lipid peroxidation, especially unsaturated aldehydes, may interfere with normal HDL cholesterol transport by inhibiting LCAT and modifying HDL apolipoproteins.