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At least 343 records · Page 19Linked to original sources

PubMed2000

Kinetics and mechanism of the reaction of aminoguanidine with the alpha-oxoaldehydes glyoxal, methylglyoxal, and 3-deoxyglucosone under physiological conditions.

Aminoguanidine (AG), a prototype agent for the preventive therapy of diabetic complications, reacts with the physiological alpha-oxoaldehydes glyoxal, methylglyoxal, and 3-deoxyglucosone (3-DG) to form 3-amino-1,2,4-triazine derivatives (T) and prevent glycation by these agents in vitro and in vivo. The reaction kinetics of these alpha-oxoaldehydes with AG under physiological conditions pH 7.4 and 37 degrees was investigated. The rate of reaction of AG with glyoxal was first order with respect to both reactants; the rate constant k(AG,G) was 0.892 +/- 0.037 M(-1) sec(-1). The kinetics of the reaction of AG with 3-DG were more complex: the rate equation was d[T](o)/dt (initial rate of T formation) = [3-DG](k(AG,3-DG)[AG] + k(3-DG)), where k(AG,3-DG) = (3. 23 +/- 0.25) x 10(-3) M(-1) sec(-1) and k(3-DG) = (1.73 +/- 0.08) x 10(-5) sec(-1). The kinetics of the reaction of AG with methylglyoxal were consistent with the reaction of both unhydrated (MG) and monohydrate (MG-H(2)O) forms. The rate equation was d[T](o)/dt = ¿k(1)k(AG,MG)/(k(-1) + k(AG,MG)[AG]) + k(AG, MG-H(2)O)¿[MG-H(2)O][AG], where the rate constant for the reaction of AG with MG, k(AG,MG), was 178 +/- 15 M(-1) sec(-1) and for the reaction of AG with MG-H(2)O, k(AG,MG-H(2)O), was 0.102 +/- 0.001 M(-1) sec(-1); k(1) and k(-1) are the forward and reverse rate constants for methylglyoxal dehydration MG-H(2)O right harpoon over left harpoon MG. The kinetics of these reactions were not influenced by ionic strength, but the reaction of AG with glyoxal and with methylglyoxal under MG-H(2)O dehydration rate-limited conditions increased with increasing phosphate buffer concentration. Kinetic modelling indicated that the rapid reaction of AG with the MG perturbed the MG/MG-H(2)O equilibrium, and the ratio of the isomeric triazine products varied with initial reactant concentration. AG is kinetically competent to scavenge the alpha-oxoaldehydes studied and decrease related advanced glycated endproduct (AGE) formation in vivo. This effect is limited, however, by the rapid renal elimination of AG. Decreased AGE formation is implicated in the prevention of microvascular complications of diabetes by AG.

Deoxyglucose↗

PubMed2001

Cytotoxic action of methylglyoxal on insulin-secreting cells.

Methylglyoxal is a spontaneous product of glucose metabolism which is known to have cytotoxic actions and to be present in raised concentrations in hyperglycaemia. It could therefore play an important role in glucose toxicity. We have investigated the cytotoxic effects of methylglyoxal on insulin-secreting cells, which are particularly sensitive to glucose toxicity. Methylglyoxal caused a concentration-dependent increase in the number of apoptotic RINm5F cells within 4-6 hours. A similar effect was observed with rat pancreatic beta-cells. tert-butylglyoxal, which is a poor substrate for the glyoxalase pathway, exerted a similar, though more potent apoptotic action. Dexamethasone and NaF were also found to induce apoptosis in RINm5F cells. Flow cytometric analysis suggested a degree of necrosis in addition to apoptosis resulting from treatment with methylglyoxal. The cytotoxic effect of methylglyoxal could contribute towards glucose toxicity in insulin-secreting cells.

Animals↗

PubMed2002

Identification of alpha-dicarbonyl scavengers for cellular protection against carbonyl stress.

Tissue deterioration and aging have long been associated with the accumulation of chemically induced protein and DNA damage. Reactive oxygen species (ROS) and reactive carbonyl species (RCS), especially alpha-dicarbonyl compounds, are key mediators of damage caused by oxidative stress, glycation, and UV-irradiation. The toxic effects of ROS are counteracted in vivo by antioxidants and antioxidant enzymes, and the deleterious effects of one RCS, methylglyoxal, are counteracted by a ubiquitous glyoxalase system. Carbonyl stress as a result of toxic effects of various mono-dicarbonyls (e.g. 4-hydroxynonenal) and alpha-dicarbonyls (e.g. glyoxal and deoxyosones) cannot be directly antagonized by antioxidants, and only a small number of biological carbonyl scavengers like glutathione (GSH) have been identified to date. We have developed a new screening method for the identification of carbonyl scavengers using a rapid glycation system that proceeds independent of oxygen and therefore, excludes identification of inhibitory compounds acting as antioxidants. Using this screening assay adapted to 96-well microtiter plates, we have identified the cysteine derivative 3,3-dimethyl-D-cysteine as a potent inhibitor of non-oxidative advanced glycation. Comparative kinetic analyses demonstrated the superior alpha-oxoaldehyde-scavenging activity of D-penicillamine over that of aminoguanidine. D-Penicillamine traps alpha-oxoaldehydes by forming a 2-acylthiazolidine derivative as shown by structure elucidation of reaction products between D-penicillamine and methylglyoxal or phenylglyoxal. We demonstrated that upon co-incubation, D-penicillamine protects human skin keratinocytes and fibroblasts (CF3 cells) against glyoxal- and methylglyoxal-induced carbonyl toxicity. Our research qualifies alpha-amino-beta-mercapto-beta,beta-dimethyl-ethane as a promising pharmacophore for the development of related alpha-dicarbonyl scavengers as therapeutic agents to protect cells against carbonyl stress.

Cells, Cultured↗

PubMed1997

Inactivation of glutathione reductase by 4-hydroxynonenal and other endogenous aldehydes.

4-Hydroxynonenal, a product of oxidative degradation of unsaturated lipids, is an endogenous reactive alpha,beta-unsaturated aldehyde with numerous biological activities. 4-Hydroxynonenal rapidly inactivated glutathione reductase in an NADPH-dependent reaction. Inactivation appears to involve the initial formation of an enzyme-inactivator complex, K(D) = 0.5 microM, followed by the inactivation reaction, k = 1.3 x 10(-2) min(-1). alpha,beta-Unsaturated aldehydes such as acrolein, crotonaldehyde, and cinnamaldehyde also inactivated glutathione reductase, although rates varied widely. Inactivation of glutathione reductase by alpha,beta-unsaturated aldehydes was followed by slower NADPH-independent reactions that led to formation of nonfluorescent cross-linked products, accompanied by loss of lysine and histidine residues. Other reactive endogenous aldehydes such as methylglyoxal, 3-deoxyglucosone, and xylosone inactivated glutathione reductase by an NADPH-independent mechanism, with methylglyoxal being the most reactive. However, 2-oxoaldehydes were much less effective than 4-hydroxynonenal. Inactivation of glutathione reductase by these 2-oxoaldehydes was followed by slower reactions that led to the formation of fluorescent cross-linked products over a period of several weeks. These changes were accompanied by loss of arginine residues. Thus, the sequence of events is different for inactivation and modification of glutathione reductase by alpha,beta-unsaturated aldehydes compared with 2-oxoaldehydes with respect to kinetics, NADPH requirements, fluorescence changes, and loss of amino acid residues. The ability of 4-hydroxynonenal at low concentrations to inactivate glutathione reductase, a central antioxidant enzyme, suggests that oxidative degradation of unsaturated lipids may initiate a positive feedback loop that enhances the potential for oxidative damage.

Acrolein↗

PubMed1998

Effects of methylglyoxal on rat pancreatic beta-cells.

The addition of the alpha-ketoaldehyde methylglyoxal (0.5 or 1 mmol/L) to single isolated rat pancreatic beta-cells caused a rapid, marked depolarization resulting in electrical activity. This effect of methylglyoxal on beta-cell was reversible upon removal of the alpha-ketoaldehyde, and could be inhibited by the anion channel blockers 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) and 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB). Methylglyoxal also resulted in elevated cytosolic [Ca2+] and an intracellular acidification in intact rat islets. In perifused islets, methylglyoxal provoked a modest, transient stimulation of secretion but inhibited glucose-induced insulin release. Incubation of islets with methylglyoxal resulted in the formation of large quantities of D-lactate, indicating metabolism of the alpha-ketoaldehyde via the glyoxalase pathway. The effects of methylglyoxal on beta-cell membrane potential, cytosolic [Ca2+] and intracellular pH were also observed in response to phenylglyoxal which is also effectively metabolized via the glyoxalase pathway. However, t-butylglyoxal which is poorly metabolized via the glyoxalase pathway, caused neither depolarization of the membrane potential nor intracellular acidification, but did inhibit glucose-induced insulin release. These findings suggests that the depolarization and acidification evoked by methyl- and phenylglyoxal are dependent upon their metabolism via the glyoxalase pathway. The possible mechanisms coupling alpha-ketoaldehyde metabolism via the glyoxalase pathway with membrane depolarization are discussed.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1999

Trioses and related substances: tools for the study of pancreatic beta-cell function.

Trioses, such as D-glyceraldehyde, have been used extensively for the study of stimulus-secretion coupling mechanisms in pancreatic beta-cells. It is generally assumed that trioses enter the glycolytic pathway at the triose phosphate level, and stimulate insulin release in a manner analogous to glucose. This review focuses on a number of triose effects that are not entirely consistent with this model. These effects are likely to result, at least in part, from the actions of alpha-ketoaldehydes. One such compound, methylglyoxal, appears to be a major contaminant of triose preparations, and exerts effects on the beta-cell identical to some of those evoked by glyceraldehyde. A related substance, hydroxypyruvaldehyde, is a product of triose autoxidation, which could exert similar effects. Study of the actions of trioses and alpha-ketoaldehydes could assist our understanding of cellular physiology, in general, and beta-cell function, in particular. These substances are also likely to be of pathophysiological importance, especially in the context of sugar toxicity and autoxidative cell damage.

Animals↗

PubMed1999

Accumulation of alpha-oxoaldehydes during oxidative stress: a role in cytotoxicity.

Glyoxal, methylglyoxal (MG), and 3-deoxyglucosone (3-DG) are physiological alpha-oxoaldehydes formed by lipid peroxidation, glycation, and degradation of glycolytic intermediates. They are enzymatically detoxified in cells by the cytosolic glutathione-dependent glyoxalase system (glyoxal and MG only) and by NADPH-dependent reductase and NAD(P)+-dependent dehydrogenase. In this study, the changes in the cellular and extracellular concentrations of these alpha-oxoaldehydes were investigated in murine P388D1 macrophages during necrotic cell death induced by median toxic concentrations of hydrogen peroxide and 1-chloro-2,4-dinitrobenzene (CDNB). Alpha-oxoaldehyde concentrations were determined by derivatization with 1,2-diamino-4,5-dimethoxybenzene. There were relatively small increases in cellular and extracellular glyoxal concentration, except that extracellular glyoxal was decreased with hydrogen peroxide. The cytosolic concentration of 3-DG and the cytosolic and extracellular concentrations of MG, however, increased markedly. Aminoguanidine inhibited alpha-oxoaldehyde accumulation and prevented cytotoxicity induced by hydrogen peroxide and CDNB. The accumulation of glyoxal and MG in toxicant-treated cells was a likely consequence of decreased in situ activity of glyoxalase 1. The effect was marked for MG but not for glyoxal, suggestive of a greater metabolic flux of MG formation than of glyoxal. The accumulation of 3-DG in toxicant-treated cells was probably due to the decreased availability of pyridine nucleotide cofactors for the detoxification of 3-DG. Impairment of alpha-oxoaldehyde detoxification is cytotoxic, and this may contribute to toxicity associated with GSH oxidation and S conjugation in oxidative stress and chemical toxicity, and to chronic pathogenesis associated with diabetes mellitus where there is oxidative stress and the formation of glyoxal, MG, and 3-DG is increased.

Animals↗

PubMed1999

Reaction of metformin with dicarbonyl compounds. Possible implication in the inhibition of advanced glycation end product formation.

Dicarbonyl compounds such as methylglyoxal and glyoxal are extremely reactive glycating agents involved in the formation of advanced glycation end products (AGEs), which in turn are associated with diabetic vascular complications. Guanidino compounds such as aminoguanidine appear to inhibit AGE formation by reacting with alpha-dicarbonyl compounds. The aim of this work was to study whether the antihyperglycemic agent metformin (a guanidine-like compound) might react with reactive alpha-dicarbonyls. Metformin was incubated at pH 7.4 and 37 degrees in the presence of either methylglyoxal or glyoxal and reaction products analysed by HPLC coupled to mass tandem spectrometry. AGE formation on albumin by methylglyoxal and glyoxal in the presence or absence of metformin was also studied by measuring the fluorescence at 370/440 nm after albumin-AGE isolation by ultrafiltration. As a standard for mass spectra analysis, a metformin-methylglyoxal adduct was chemically synthesised and characterised as a triazepinone (2-amino-4-(dimethyl-amino)-7-methyl-5,7-dihydro-6H-[1,3,5]triazepin+ ++-6-one). The results obtained showed that metformin strongly reacted with methylglyoxal and glyoxal, forming original guanidine-dicarbonyl adducts. Reaction kinetic studies as well as mass fragmentation spectra of the reaction products were compatible with the presence of triazepinone derivatives. In the presence of metformin, AGE-related fluorescence after albumin incubation with either glyoxal or methylglyoxal was decreased by 37% and 45%, respectively. These results suggest that besides its known antihyperglycemic effect, metformin could also decrease AGE formation by reacting with alpha-dicarbonyl compounds. This is relevant to a potential clinical use of metformin in the prevention of diabetic complications by inhibition of carbonyl stress.

Animals↗

PubMed2000

Selective vulnerability of spinal motor neurons to reactive dicarbonyl compounds, intermediate products of glycation, in vitro: implication of inefficient glutathione system in spinal motor neurons.

We investigated the effects of two reactive dicarbonyl compounds, methylglyoxal (MG) and 3-deoxyglucosone (3-DG), on cultured spinal cord neurons. Incubation of cortical and spinal neurons with MG and 3-DG for 24 h induced neuronal death in a dose-dependent manner. Spinal motor neurons were more vulnerable than spinal non-motor neurons and cortical neurons. Treatments with glutathione (GSH)-augmenting agents showed protective effects against MG and 3-DG neurotoxicity. Motor neurons were better protected than non-motor neurons. Cotreatment, but not pretreatment, of aminoguanidine (AG), a known inhibitor of advanced glycation end-products (AGEs) from crosslinking, showed a protective effect on spinal neurons with no difference in protective rates between motor and non-motor spinal neurons. Treatments with GSH depleting agents enhanced the neurotoxicity of MG and 3-DG on spinal neurons. Motor neurons were more vulnerable than non-motor neurons with GSH-depleting treatments prior to MG and 3-DG exposures. These data demonstrate that spinal motor neurons are more vulnerable to dicarbonyl compounds, and this selectivity might be related to the relatively inefficient GSH system in spinal motor neurons.

Amyotrophic Lateral Sclerosis↗

PubMed2003

Kinetic modelling of Amadori N-(1-deoxy-D-fructos-1-yl)-glycine degradation pathways. Part I--reaction mechanism.

The fate of the Amadori compound N-(1-deoxy-D-fructos-1-yl)-glycine (DFG) was studied in aqueous model systems as a function of pH and temperature. The samples were heated at 100 and 120 degrees C with initial reaction pH of 5.5 and 6.8. Special attention was paid to the formation of the free amino acid, glycine; parent sugars, glucose and mannose; organic acids, formic and acetic acid and alpha-dicarbonyls, 1- and 3-deoxyosone together with methylglyoxal. For the studied conditions decreasing the initial reaction pH with 1.3 units or increasing the temperature with 20 degrees C has the same effect on the DFG degradation as well as on glycine formation. An increase in pH seems to favour the formation of 1-deoxyosone. The lower amount found comparatively to 3-deoxyosone, in all studied systems, seems to be related with the higher reactivity of 1-deoxyosone. Independently of the taken pathway, enolization or retro-aldolization, DFG degradation is accompanied by amino acid release. Together with glycine, acetic acid was the main end product formed. Values of 83 and 55 mol% were obtained, respectively. The rate of parent sugars formation increased with pH, but the type of sugar formed also changed with pH. Mannose was preferably formed at pH 5.5 whereas at pH 6.8 the opposite was observed, that is, glucose was formed in higher amounts than mannose. Also, independently of the temperature, at higher pH fructose was also detected. pH, more than temperature, had an influence on the reaction products formed. The initial steps for a complete multiresponse kinetic analysis have been discussed. Based on the established reaction network a kinetic model will be proposed and evaluated by multiresponse kinetic modelling in a subsequent paper.

Acetic Acid↗

PubMed2001

Metabolism of the 2-oxoaldehyde methylglyoxal by aldose reductase and by glyoxalase-I: roles for glutathione in both enzymes and implications for diabetic complications.

Numerous physiological aldehydes besides glucose are substrates of aldose reductase, the first enzyme of the polyol pathway which has been implicated in the etiology of diabetic complications. The 2-oxoaldehyde methylglyoxal is a preferred substrate of aldose reductase but is also the main physiological substrate of the glutathione-dependent glyoxalase system. Aldose reductase catalyzes the reduction of methylglyoxal efficiently (k(cat)=142 min(-1) and k(cat)/K(m)=1.8x10(7) M(-1) min(-1)). In the presence of physiological concentrations of glutathione, methylglyoxal is significantly converted into the hemithioacetal, which is the actual substrate of glyoxalase-I. However, in the presence of glutathione, the efficiency of reduction of methylglyoxal, catalyzed by aldose reductase, also increases. In addition, the site of reduction switches from the aldehyde to the ketone carbonyl. Thus, glutathione converts aldose reductase from an aldehyde reductase to a ketone reductase with methylglyoxal as substrate. The relative importance of aldose reductase and glyoxalase-I in the metabolic disposal of methylglyoxal is highly dependent upon the concentration of glutathione, owing to the non-catalytic pre-enzymatic reaction between methylglyoxal and glutathione.

Aldehyde Reductase↗

PubMed2001

Inhibitory effects of S-nitrosoglutathione on cell proliferation and DNA synthesis: possible role of glyoxalase I inactivation.

We investigated the inhibitory effects of S-nitrosoglutathione (GSNO) on cell proliferation, DNA synthesis and several enzymatic activities using spontaneously immortalized human endothelial cells (ECV304). Proliferation of ECV304 was inhibited by GSNO in a dose-dependent manner (125-1000 microM). DNA synthesis was decreased 2 h after addition of GSNO to cells and was markedly repressed from 20 h after the addition. The activity of ribonucleotide reductase, a rate-limiting enzyme for DNA synthesis, was unchanged in GSNO-treated cells. GSNO inhibited less than 40% of mitochondrial respiration activity, and the membrane potential and cellular levels of ATP were not significantly decreased by GSNO. GSNO had no inhibitory effect on activities of glutathione peroxidase, glutathione S-transferase and glutathione reductase. However, glyoxalase I (Glo I) activity was decreased to 20% of the control level within 60 min, and was consistently repressed during exposure to GSNO for 20 h. A membrane-permeable Glo I inhibitor, S-bromobenzylglutathione diethylester, inhibited proliferation of ECV304 cells, while methylglyoxal (MG), a toxic metabolite generated during glycolysis and a substrate for Glo I, failed to inhibit the cell growth even at 100 microM. Glo I in several mammalian cell lines was inactivated by GSNO with a pI shift. Although we failed to detect accumulation of MG under conditions of Glo I inactivation, these results suggest that the inhibitory effects of GSNO on cell proliferation and DNA synthesis might be at least partly due to inactivation of Glo I.

Animals↗

PubMed2001

Role of iron in the interaction of red blood cells with methylglyoxal. Modification of L-arginine by methylglyoxal is catalyzed by iron redox cycling.

Diabetes mellitus is characterized by increased methylglyoxal (MG) production. The aim of the present study was to investigate the role of iron in the cellular and molecular effects of MG. A red blood cell (RBC) model and L-arginine were used to study the effects of MG in the absence and presence of iron. Intracellular free radical formation and calcium concentration were measured using dichlorofluorescein and Fura-2-AM, respectively. Effects of MG were compared to the effect of ferrous iron. Reaction of L-arginine with MG was investigated by electron spin resonance (ESR) spectroscopy and by a spectrophotometric method. MG caused an iron dependent oxidative stress in RBCs and an elevation of the intracellular calcium concentration due to formation of reactive oxygen species. Results of co-incubation of MG with ferrous iron in the RBC model suggested an interaction of MG and iron; one interaction was a reduction of ferric iron by MG. A role of iron in the MG-L-arginine reaction was also verified by ESR spectroscopy and by spectrophotometry. Ferric iron increased free radical formation as detected by ESR in the MG-L-arginine reaction; however, ferrous iron decreased it. The reaction of MG with L-arginine yielded a brown product as detected spectrophotometrically and this reaction was catalyzed at a lower rate with ferric iron but at a higher rate with ferrous iron. These data suggest that MG causes oxidative stress in cells, which is due at least in part to ferric iron reduction by MG and to the modification of amino acids e.g. L-arginine by MG, which is catalyzed by iron redox cycling.

Arginine↗

PubMed2002

Expression of rat liver glutathione-S-transferase GSTA5 in cell lines provides increased resistance to alkylating agents and toxic aldehydes.

The glutathione-S-transferases (GST) are a major contributor to the eukaryotic cell's defences against chemical and oxidative stress. However, the role of individual GST isoenzymes in conferring resistance to xenobiotics has not been fully determined. We have examined the effect of the rat GSTA5 isoenzyme in the detoxication of alkylating agents and aldehydes by constructing a cell line in which it is stably expressed. The hamster fibroblast cell line V79 was transfected with a construct expressing GSTA5 from the CMV promoter. A stable clone (V79-GSTA5) was isolated after selecting for the neomycin phosphotransferase gene present on the introduced DNA. The cell line showed significantly increased levels of resistance towards the alkylating agents chorambucil and melphalan. Levels of resistance were 4-6-fold greater in V79-GSTA5 cells than in control cells. Increased levels of resistance were also observed towards the lipid peroxidation product acrolein (IC(50)=80 microM compared with 17 microM in control cells). The V79-GSTA5 cells also showed a 4-fold increase in resistance to trans, trans muconaldehyde (IC(50)=4 micro compared with l microM for control cells). GSTA5 did not protect against 4-hydroxynonenal, but it did provide greater levels of protection to hydrogen peroxide, with an IC(50) of 380 microM in V79-GSTA5 compared with 180 microM in control cells. In contrast, V79-GSTA5 cells were more sensitive to methyl glyoxal, suggesting that a methyl glyoxal-glutathione conjugate is more toxic that the parental compound. These data contribute towards the evaluation of the role of GSTA5 in the detoxication of these compounds.

Aldehydes↗

PubMed2003

Different activity of glyoxalase system enzymes in specimens of Sparus auratus exposed to sublethal copper concentrations.

The present study regards possible changes in the activity of glyoxalase system enzymes (glyoxalase I, GI, and glyoxalase II, GII) in tissues (brain, liver and white muscle) of the mediterranean bony fish Sparus auratus after a 20 days exposure to sublethal concentrations (0.1 or 0.5 ppm) of Cu in the marine water and on control untreated animals. The experiments also included measurements of copper concentration in the tissues, as well as of lactate dehydrogenase (LDH) activity, to evaluate possible Cu accumulation and changes in glycolytic activity respectively. Cu accumulation only occurs in the liver. GI, GII and LDH activities kept unchanged in the brain after copper exposure. GI activity in liver and muscle of copper-exposed animals decreases probably for a slackening in the glycolytic rate, as suggested by the lowering of LDH activity. GII activity remains unchanged or increases (liver extract, 0.5 ppm of Cu), maybe to safeguard enough cellular levels of GSH.

Animals↗

PubMed2003

Up-regulation of aldose reductase by the substrate, methylglyoxal.

Methylglyoxal (MG), a reactive dicarbonyl produced during glucose metabolism, is known as a preferred substrate of aldose reductase (AR; AKR1B1) that concomitantly catalyzes the reduction of glucose in the polyol pathway. MG modifies cellular proteins to form cross-links of amino groups, generating so-called advanced glycation end products. Increased rates of MG formation under hyperglycemic conditions and ensuing high serum levels of MG are reported in diabetic patients. As involvement of AR in the pathogenesis of diabetic complications has been suggested, we investigated the effects of MG on AR activity using cultured vascular smooth muscle cells (SMC) isolated from rat aorta. MG-induced a dose- and time-dependent increase in AR mRNA levels to a maximum of 4.5-fold. This increase in mRNA was accompanied by elevated enzyme activity and protein levels. Pretreatment of SMC with N-acetyl-L-cysteine significantly suppressed the MG-induced AR expression, while DL-buthionine-(S,R)-sulfoximine further augmented the MG-induced increase in AR mRNA level. Intracellular levels of reactive oxygen species determined using 2',7'-dichlorofluorescein diacetate were significantly elevated in SMC treated with MG, suggesting the involvement of oxidative stress in this process. Under oxidative stress generated by hydrogen peroxide, the major signaling pathway mediating the up-regulation of AR expression was demonstrated to be the epidermal growth factor receptor-ERK pathway. In contrast, the p38 kinase pathway appears to mediate MG-induced AR expression. The cytotoxic effect of MG was significantly enhanced in the presence of the AR inhibitor ponalrestat, indicating a protective role of AR against MG-induced cell damage.

Aldehyde Reductase↗

PubMed2003

Methylglyoxal metabolism and diabetic complications: roles of aldose reductase, glyoxalase-I, betaine aldehyde dehydrogenase and 2-oxoaldehyde dehydrogenase.

The 2-oxoaldehyde methylglyoxal (MeG) is the precursor to a number of the known advanced glycation endproducts (AGE) implicated in the development of diabetic complications. Other 2-oxoaldehydes that are important in AGE formation, such as glyoxal, glucosone, deoxyglucosone, xylosone and deoxyxylosone, are produced by nonenzymatic reactions. By contrast, MeG is produced by both enzymatic and nonenzymatic processes, most of which appear to be enhanced in diabetes. MeG may be a major precursor to formation of AGE, and rates of production of MeG depend upon physiological conditions such as hyperglycemia and ketoacidosis. MeG is also unique compared to the other 2-oxoaldehydes in its complex metabolism. At least four pathways contribute to detoxification of MeG: (1) aldose reductase, a member of the aldo-keto reductase superfamily, catalyzes the NADPH-dependent reduction of a wide range of aldehydes. MeG is the best of the known physiological aldehyde substrates of aldose reductase. The distribution of aldose reductase in human tissue is restricted; there is little expression in liver; (2) the ubiquitous and highly active glyoxalase system converts MeG into D-lactate. However, this system depends upon the availability of glutathione; activity is severely limited by conditions of oxidative stress that impact levels of glutathione; (3) betaine aldehyde dehydrogenase, also known as ALDH9, is able to catalyze the oxidation of MeG to pyruvate, although less efficiently than with its substrate betaine aldehyde; (4) the long-known but not widely studied 2-oxoaldehyde dehydrogenases (2-ODHs) catalyze the oxidation of MeG to pyruvate, primarily in liver. There are two NADP-dependent 2-ODHs in human liver. Both of these require an activating amine. The physiological activator is unknown.

Aldehyde Oxidoreductases↗

PubMed1998

Formation of DNA adducts in human buccal epithelial cells exposed to acetaldehyde and methylglyoxal in vitro.

Acetaldehyde (AA) and methylglyoxal (MG) are reactive, ubiquitous aldehydes, present in the environment and endogenously formed in animals and humans. They have both been shown to readily form DNA adducts under simulated physiological conditions. We report here on the use of cultured normal and SV40T antigen-immortalized human buccal epithelial cells as model systems for aldehyde exposure of the oral epithelium, occurring through the ingestion of alcoholic beverages and brewed coffee, as well as by inhalation of tobacco smoke and automobile exhaust. By the application of recently developed 32P-postlabeling methods, the presence of both endogenous and induced AA and MG DNA adducts was demonstrated in cultured human epithelial cells. Furthermore, these DNA adducts were formed in a dose-dependent manner at aldehyde concentrations that were relatively nontoxic to the cells.

Acetaldehyde↗