Showing papers on "Aldehyde dehydrogenase published in 2005"

Analysis and update of the human aldehyde dehydrogenase (ALDH) gene family

Abstract: The aldehyde dehydrogenase (ALDH) gene superfamily encodes enzymes that are critical for certain life processes and detoxification via the NAD(P)+-dependent oxidation of numerous endogenous and exogenous aldehyde substrates, including pharmaceuticals and environmental pollutants. Analysis of the ALDH gene superfamily in the latest databases showed that the human genome contains 19 putatively functional genes and three pseudogenes. A number of ALDH genes are upregulated as a part of the oxidative stress response and inexplicably overexpressed in various tumours, leading to problems during cancer chemotherapy. Mutations in ALDH genes cause inborn errors of metabolism -- such as the Sjogren - Larsson syndrome, type II hyperprolinaemia and γ-hydroxybutyric aciduria -- and are likely to contribute to several complex diseases, including cancer and Alzheimer's disease. The ALDH gene products appear to be multifunctional proteins, possessing both catalytic and non-catalytic properties.

Characterization of cells with a high aldehyde dehydrogenase activity from cord blood and acute myeloid leukemia samples.

Abstract: Aldehyde dehydrogenase (ALDH) is a cytosolic enzyme that is responsible for the oxidation of intracellular aldehydes. Elevated levels of ALDH have been demonstrated in murine and human progenitor cells compared with other hematopoietic cells, and this is thought to be important in chemoresistance. A method for the assessment of ALDH activity in viable cells recently has been developed and made commercially available in a kit format. In this study, we confirmed the use of the ALDH substrate kit to identify cord blood stem/progenitor cells. Via multicolor flow cytometry of cord blood ALDH+ cells, we have expanded on their phenotypic analysis. We then assessed the incidence, morphology, phenotype, and nonobese diabetic/ severe combined immunodeficiency engraftment ability of ALDH+ cells from acute myeloid leukemia (AML) samples. AML samples had no ALDH+ cells at all, an extremely rare nonmalignant stem/progenitor cell population, or a less rare, leukemic stem cell population. Hence, in addition to identifying nonmalignant stem cells within some AML samples, a high ALDH activity also identifies some patients' CD34+/ CD38- leukemic stem cells. The incidence of normal or leukemic stem cells with an extremely high ALDH activity may have important implications for resistance to chemotherapy. Identification and isolation of leukemic cells on the basis of ALDH activity provides a tool for their isolation and further analysis.

An essential role for mitochondrial aldehyde dehydrogenase in nitroglycerin bioactivation.

Abstract: The identity of the cellular mechanisms through which nitroglycerin (glyceryl trinitrate, GTN) elicits nitric oxide (NO)-based signaling to dilate blood vessels remains one of the longest standing foci of investigation and sources of controversy in cardiovascular biology. Recent evidence suggests an unexpected role for mitochondria. We show here that bioconversion by mitochondria of clinically relevant concentrations of GTN results in activation of guanylate cyclase, production of cGMP, vasodilation in vitro, and lowered blood pressure in vivo, which are eliminated by genetic deletion of the mitochondrial aldehyde dehydrogenase (mtALDH). In contrast, generation of vasoactivity from alternative nitro(so)-vasodilators is unaffected. In mtALDH-/- mice and their isolated vascular tissue, GTN bioactivity can still be generated, but only at substantially higher concentrations of GTN and by a mechanism that does not exhibit tolerance. Thus, mtALDH is necessary and sufficient for vasoactivity derived from therapeutic levels of GTN, and, more generally, mitochondria can serve as a source of NO-based cellular signals that may originate independently of NO synthase activity.

Detailed expression analysis of selected genes of the aldehyde dehydrogenase (ALDH) gene superfamily in Arabidopsis thaliana.

Abstract: Aldehyde dehydrogenase (ALDH) genes have been identified in almost all organisms from prokaryotes to eukaryotes, but particularly in plants knowledge is very limited with respect to their function. The data presented here are a contribution towards a functional analysis of selected Arabidopsis ALDH genes by using expression profiles in wild types and mutants. The Arabidopsis thaliana genome contains 14 genes which represent 9 families. To gain insight into the possible roles of aldehyde dehydrogenases from Arabidopsis, the expression patterns of five selected ALDH genes were analyzed under defined physiological conditions. Three genes (ALDH3I1, 3H1 and ALDH7B4) that belong to two different families are differentially activated by dehydration, high salinity and ABA in a tissue-specific manner. The other two genes (ALDH3F1 and ALDH22A1) are constitutively expressed at a low level. Transcript analysis of ALDH3I1 and ALDH7B4 in Arabidopsis mutants suggests that stress responses are differentially controlled by the phytohormone ABA as well as by pathways that affect sugar metabolism and fatty acid composition of membrane lipids. Our results indicate that the stress-associated ALDH genes participate in several pathways and that their regulation involves diverged signal transduction pathways.

Aldehyde dehydrogenase 2 gene targeting mouse lacking enzyme activity shows high acetaldehyde level in blood, brain, and liver after ethanol gavages.

Abstract: Background: Previously, we created an aldehyde dehydrogenase 2 gene transgenic (Aldh2−/−) mouse as an aldehyde dehydrogenase (ALDH) 2 inactive human model and demonstrated low alcohol preference. In addition, after a free-choice drinking test, no difference in the acetaldehyde level was observed between the Aldh2−/− and wild type (Aldh2+/+) mice. The actual amounts of free-choice drinking were so low that it is uncertain whether these levels are pharmacologically and/or behaviorally relevant in either strain. To elucidate this uncertainty, we compared the ethanol and acetaldehyde concentration in the blood, brain, and liver between the Aldh2−/− and Aldh2+/+ mice after ethanol gavages at the same dose and time. Method: We measured differences in the ethanol and acetaldehyde levels between the Aldh2−/− and Aldh2+/+ mice by headspace gas chromatography-mass spectrometry (GC-MS) after ethanol gavages at the same dose and time. Results: Significantly higher blood acetaldehyde concentrations were found in the Aldh2−/− mice than in the Aldh2+/+ mice 1 hr after the administration of ethanol gavages at doses of 0.5, 1.0, 2.0, and 5.0 g/kg. The blood acetaldehyde concentrations in the two strains were 2.4 vs. 0.5, 17.8 vs. 1.9, 108.3 vs. 4.3, and 247.2 vs. 14.0 (μM), respectively. In contrast, no significant difference was observed in the blood ethanol concentrations between the Aldh2+/+ and Aldh2−/− mice. The aldehyde dehydrogenase 2 enzyme metabolized 94% of the acetaldehyde produced from the ethanol as calculated from the area under the curve (AUC) of acetaldehyde when ethanol was administered at a dose of 5.0 g/kg. Conclusions: These data indicate that mouse ALDH2 is a major enzyme for acetaldehyde metabolism, and the Aldh2−/− mice have significantly high acetaldehyde levels after ethanol gavages.

Retinoic Acid Down-Regulates Aldehyde Dehydrogenase and Increases Cytotoxicity of 4-Hydroperoxycyclophosphamide and Acetaldehyde

Abstract: Multiple prior studies have identified aldehyde dehydrogenases (ALDH) that are capable of oxidizing retinal to retinoic acid. In this study, we test the hypothesis that the accumulation of intracellular retinoic acid may lead to the suppression of ALDH expression and thus increase cytotoxicity to 4-hydroperoxycyclophosphamide (4-HC) in vitro. Mainly A549, but also other lung cancer cell lines, were used in our experiments, with the former having high levels of two ALDH isozymes expressed. Dose-response and time-course experiments were performed by incubating the cells with all-trans retinoic acid (ATRA) as well as other commercially available retinoids. The results show that incubation of A549 cells with any of the retinoids at pharmacologic doses for ≥48 h results in a significant decrease in ALDH-1A1 and ALDH-3A1 enzyme activity and protein levels but not the corresponding mRNAs. Such a decrease in ALDH activity was seen in all cell lines tested and results in a significant increase in toxicity of 4-HC and acetaldehyde, both of which are substrates for the enzymes. Prior incubation with ATRA also results in increased cytotoxicity, although to a lesser degree, of phenylketophosphamide and melphalan, neither of which is a substrate for ALDHs. These results suggest a post-translational mechanism through which retinoids decrease both ALDH expression, which results in increased cytotoxicity of 4-HC and acetaldehyde, although other previously described effects of these retinoids may contribute to the slight increase in cytotoxicity seen with other chemotherapy agents. These results may have clinical implications in regard to the use of retinoids in lung cancer prevention and treatment.

Isolation and Characterization of an Aldehyde Dehydrogenase Encoded by the aldB Gene of Escherichia coli

Abstract: An aldehyde dehydrogenase was detected in crude cell extracts of Escherichia coli DH5α. Growth studies indicated that the aldehyde dehydrogenase activity was growth phase dependent and increased in cells grown with ethanol. The N-terminal amino acid sequence of the purified enzyme identified the latter as an aldehyde dehydrogenase encoded by aldB, which was thought to play a role in the removal of aldehydes and alcohols in cells that were under stress. The purified enzyme showed an estimated molecular mass of 220 ± 8 kDa, consisting of four identical subunits, and preferred to use NADP and acetaldehyde. MgCl2 increased the activity of the NADP-dependent enzyme with various substrates. A comparison of the effect of Mg2+ ions on the bacterial enzyme with the effect of Mg2+ ions on human liver mitochondrial aldehyde dehydrogenase revealed that the bacterial enzyme shared kinetic properties with the mammalian enzyme. An R197E mutant of the bacterial enzyme appeared to retain very little NADP-dependent activity on acetaldehyde.

Role of aldehyde dehydrogenase isozymes in the defense of rat lens and human lens epithelial cells against oxidative stress.

Abstract: PURPOSE. 4-Hydroxynonenal (HNE), a metastable lipid peroxidation product, is highly toxic to various cell types if not detoxified. Because of its constant exposure to light, the ocular lens continuously generates reactive oxygen species which, under conditions of oxidative stress, may lead to excessive lipid peroxidation and consequent formation of lipid-derived aldehydes (LDAs) such as HNE. The contribution of various isozymes of aldehyde dehydrogenase (ALDH) to the oxidation of LDAs has never been systematically investigated in the lens. The present study was undertaken to ascertain the role of ALDH1A1 and -3A1 in HNE metabolism and HNE-induced toxicity in cultured human lens epithelial cells (HLECs) and in rat and mouse lenses. METHODS. The metabolism of 3 H-HNE was studied in ALDH3A1-knockout mouse lens and in HLECs transfected with ALDH1A1-or -3A1-specific antisense RNA and short interfering (Si)RNA. Appropriate controls were used, including wild-type mouse lens, scrambled oligonucleotides, and a transfection reagent. Transfected HLECs were exposed to oxidative stress (Fenton reaction) or HNE (30 μM) for 3 hours. Toxicity parameters, such as cell viability, apoptosis, and protein-HNE adducts and oxidation of exogenously added 3 H-HNE were measured. Rat lenses were transfected with the SiRNA specific to ALDH1A1, and oxidation of 3 H-HNE and the susceptibility of the transfected lenses to oxidation-induced opacification were measured. RESULTS. Rat lenses transfected with ALDH1A1-specific SiRNA, or cultured in the presence of the ALDH inhibitor cyanamide/ disulfiram and subjected to oxidative stress displayed accelerated loss of transparency and a diminished capacity to oxidize HNE. Similarly, inhibition of ALDH1A1 in HLECs by ALDH1A1-specific antisense RNA or SiRNA was associated with decreased oxidation of 3 H-HNE and increased susceptibility of the cells to oxidative damage, including apoptosis. Furthermore, 3 H-HNE metabolism and HNE-induced toxicity were not affected in ALDH3A1-specific SiRNA- or antisense RNA-treated rat lenses, HLECs, or ALDH3Al-null mouse lenses. CONCLUSIONS. The results suggest that, under oxidative stress, HNE produced in the lens epithelium can cause toxicity and thus contribute to oxidation-induced cataractogenesis. Furthermore, the studies indicate that ALDH1A1 is a critical isozyme for maintaining clarity in human, rat, and mouse lenses.

Tissue-distribution of aldehyde dehydrogenase 2 and effects of the ALDH2 gene-disruption on the expression of enzymes involved in alcohol metabolism.

Abstract: In alcohol metabolism, acetaldehyde, a highly reactive intermediate that may cause cellular and DNA damages, is converted to acetate by mitochondrial aldehyde dehydrogenase ALDH2. Although the majority of ingested alcohol is eliminated in the liver, the first-pass metabolism of ethanol in the upper digestive tract is also important for prevention and management of ethanol-related gastrointestinal diseases. However, the tissue-distribution of Aldh2 in mice has been poorly investigated. In this study, therefore, we investigated the tissue-distribution of Aldh2 as well as Aldh1, Cyp1a1, Cyp2e1, and Cyp4b1 in wild type and Aldh2-null mice by immuno-histochemical analysis. The human liver and esophageal tissues were also examined. In mice, the Aldh2 protein was detected in the liver, lung, heart, kidney, testis, esophagus, stomach, colon, and pancreas, suggesting that the tissue-distribution of Aldh2 in mice is similar to that in humans. Therefore, Aldh2-null mice may be useful model animals for the investigation of alcohol metabolism and related diseases. Compared with the wild type, the expression level of Cyp2e1 was increased in the liver from Aldh2-null mice based on Western blot analysis, whereas the levels of Aldh1, Cyp1a1, and Cyp4b1 were indistinguishable. This observation suggests that a metabolite(s) of Aldh2 might down-regulate the expression of Cyp2e1 gene.

Genetic associations of alcohol and aldehyde dehydrogenase with alcohol dependence and their mechanisms of action.

Abstract: Two alcohol dehydrogenase genes (ADHIB and ADH1C on chromosome 4) and one aldehyde dehydrogenase gene (ALDH2 on chromosome 12) exhibit functional polymorphisms that are associated with lower rates of alcohol dependence. The ALDH2*2 allele,found almost exclusively in Asian populations, has the strongest relationship. The ADH1B*2, ADH1B*3, and ADHlC*i alleles, found in varying prevalence in different ethnic groups, have also been associated with lower rates of alcohol dependence. Studies of the ADHIBand ADH1C haplotypes, however, have shown that ADH1C*I is in linkage disequilibrium with ADHiB*2, and the ADH1C*i allele does not appear to have significant unique associations with alcohol dependence. The hypothesized mechanism underlying the associations of the ADH1B and ALDH2 polymorphisms with alcohol dependence is that the isoenzymes encoded by these alleles lead to an accumulation of acetaldehyde during alcohol metabolism. Based on their kinetic properties, ALDH2 *2 theoretically should lead to a slower removal of acetaldehyde than ALDH2*1, whereas ADH1B*2 and ADH1B*3 should lead to a more rapid production of acetaldehyde than ADHIB*I. It is further hypothesized that elevations in acetaldehyde cause more intense reactions to alcohol and lead to lower levels of alcohol intake. Data are consistent with the hypothesis that elevations in acetaldehyde, increased sensitivity to alcohol, and lower levels of drinking reflect the mechanism by which the ALDH2*2 allele reduces risk for alcohol dependence. There is also some evidence supporting this mechanism for the ADH1B*2 and ADHIB*3 alleles, but the results are less consistent. These findings highlight the value of trying to elucidate the mechanism by which genes ultimately give rise to differences in alcohol dependence through the examination of mediating behaviors.

Aldehyde dehydrogenase activity as a marker for the quality of hematopoietic stem cell transplants.

Abstract: Aldehyde dehydrogenase activity as a marker for the quality of hematopoietic stem cell transplants

Molecular cloning, baculovirus expression, and tissue distribution of the zebrafish aldehyde dehydrogenase 2.

Abstract: Ethanol is metabolized to acetaldehyde mainly by the alcohol dehydrogenase pathway and, to a lesser extent, through microsomal oxidation (CYP2E1) and the catalase-H2O2 system. Acetaldehyde, which is responsible for some of the deleterious effects of ethanol, is further oxidized to acetic acid by aldehyde dehydrogenases (ALDHs), of which mitochondrial ALDH2 is the most efficient. The aim of this study was to evaluate zebrafish (Danio rerio) as a model for ethanol metabolism by cloning, expressing, and characterizing the zebrafish ALDH2. The zebrafish ALDH2 cDNA was cloned and found to be 1892 bp in length and encoding a protein of 516 amino acids (Mr = 56,562), approximately 75% identical to mammalian ALDH2 proteins. Recombinant zebrafish ALDH2 protein was expressed using the baculovirus expression system and purified to homogeneity by affinity chromatography. We found that zebrafish ALDH2 is catalytically active and efficiently oxidizes acetaldehyde (Km = 11.5 μM) and propionaldehyde (Km = 6.1 μM). Similar kinetic properties were observed with the recombinant human ALDH2 protein, which was expressed and purified using comparable experimental conditions. Western blot analysis revealed that ALDH2 is highly expressed in the heart, skeletal muscle, and brain with moderate expression in liver, eye, and swim bladder of the zebrafish. These results are the first reported on the cloning, expression, and characterization of a zebrafish ALDH, and indicate that zebrafish is a suitable model for studying ethanol metabolism and, therefore, toxicity.

Enzymatic oxidation of vanillin, isovanillin and protocatechuic aldehyde with freshly prepared Guinea pig liver slices.

Abstract: Background/Aims: The oxidation of xenobiotic-derived aromatic aldehydes with freshly prepared liver slices has not been previously reported. The present investigation compares the relative contributio

Cutaneous metabolism of glycol ethers.

Abstract: The toxicity of glycol ethers is associated with their oxidation to the corresponding aldehyde and alkoxyacetic acid by cytosolic alcohol dehydrogenase (ADH; EC 1.1.1.1.) and aldehyde dehydrogenase (ALDH; 1.2.1.3). Dermal exposure to these compounds can result in localised or systemic toxicity including skin sensitisation and irritancy, reproductive, developmental and haemotological effects. It has previously been shown that skin has the capacity for local metabolism of applied chemicals. Therefore, there is a requirement to consider metabolism during dermal absorption of these compounds in risk assessment for humans. Cytosolic fractions were prepared from rat liver, and whole and dermatomed skin by differential centrifugation. Rat skin cytosolic fractions were also prepared following multiple dermal exposure to dexamethasone, ethanol or 2-butoxyethanol (2-BE). The rate of ethanol, 2-ethoxyethanol (2-EE), ethylene glycol, 2-phenoxyethanol (2-PE) and 2-BE conversion to alkoxyacetic acid by ADH/ALDH in these fractions was continuously monitored by UV spectrophotometry via the conversion of NAD+ to NADH at 340 nm. Rates of ADH oxidation by rat liver cytosol were greatest for ethanol followed by 2-EE >ethylene glycol >2-PE >2-BE. However, the order of metabolism changed to 2-BE >2-PE >ethylene glycol >2-EE >ethanol using whole and dermatomed rat skin cytosolic fractions, with approximately twice the specific activity in dermatomed skin cytosol relative to whole rat skin. This suggests that ADH and ALDH are localised in the epidermis that constitutes more of the protein in dermatomed skin than whole skin cytosol. Inhibition of ADH oxidation in rat liver cytosol by pyrazole was greatest for ethanol followed by 2-EE >ethylene glycol >2-PE >2-BE, but it only inhibited ethanol metabolism by 40% in skin cytosol. Disulfiram completely inhibited alcohol and glycol ether metabolism in the liver and skin cytosolic fractions. Although ADH1, ADH2 and ADH3 are expressed at the protein level in rat liver, only ADH1 and ADH2 are selectively inhibited by pyrazole and they constitute the predominant isoforms that metabolise short-chain alcohols in preference to intermediate chain-length alcohols. However, ADH1, ADH3 and ADH4 predominate in rat skin, demonstrate different sensitivities to pyrazole, and are responsible for metabolising glycol ethers. ALDH1 is the predominant isoform in rat liver and skin cytosolic fractions that is selectively inhibited by disulfiram and responds to the amount of aldehyde formed by the ADH isoforms expressed in these tissues. Thus, the different affinity of ADH and ALDH for alcohols and glycol ethers of different carbon-chain length may reflect the relative isoform expression in rat liver and skin. Following multiple topical exposure, ethanol metabolism increased the most following ethanol treatment, and 2-BE metabolism increased the most following 2-BE treatment. Ethanol and 2-BE may induce specific ADH and ALDH isoforms that preferentially metabolise short-chain alcohols (i.e. ADH1, ALDH1) and longer chain alcohols (i.e. ADH3, ADH4, ALDH1), respectively. Treatment with a general inducing agent such as dexamethasone enhanced ethanol and 2-BE metabolism suggesting induction of multiple ADH isoforms.

Tissue- and species-specific expression patterns of class I, III, and IV Adh and Aldh1 mRNAs in rodent embryos

Abstract: Alcohol and aldehyde dehydrogenases (ADHs and ALDHs) may be of interest in the pathology of Parkinson's disease (PD) because of their role in protection against toxins and in retinoid metabolism, which is required for growth and development of the mesencephalic dopamine system. In the present study, the spatial and temporal expression patterns of Adh1, Adh3, Adh4, and Aldh1 mRNAs in embryonic C57BL/6 mice (E9.5-E19.5) and Sprague-Dawley rats (E12.5-P0) have been investigated by using radioactive oligonucleotide in situ hybridization. High expression of Aldh1 mRNA was found in the developing mesencephalic dopamine neurons of both mice and rats. Expression of Adh1 and Adh4 mRNAs was observed in adrenal cortex and olfactory epithelium in mice. Additionally, Adh1 was expressed in epidermis, liver, conjunctival, and intestinal epithelium. In rat embryos, expression was less extensive, with Adh1 mRNA being found in liver and intestines. Adh3 expression was ubiquitous in both mouse and rat embryos, suggesting a housekeeping function of the gene. Consistent with previous studies in adult rats and mice, our data suggest that Adh3 is the only ADH class present in rodent brain. Adh and Aldh gene activity in mouse and rat embryos indicate the possible involvement of the respective enzymes in retinoid metabolism and participation in defense against toxic insults, including those that may be involved in the pathogenesis of PD.

Characterization of Sphingomonas aldehyde dehydrogenase catalyzing the conversion of various aromatic aldehydes to their carboxylic acids

Abstract: An aldehyde dehydrogenase gene, designated phnN, was isolated from a genome library of the 1,4-dimethylnaphthalene-utilizing soil bacterium, Sphingomonas sp. 14DN61. Escherichia coli expressing the phnN gene converted 1,4-dihydroxymethylnaphthalene to 1-hydroxymethyl-4-naphthoic acid. The putative amino acid sequence of the phnN gene product had 31-42% identity with those of NAD(+)-dependent short-chain aliphatic aldehyde dehydrogenases and a secondary alcohol dehydrogenase. The NAD(P)(+)-binding site and two consensus sequences involved in the active site for aldehyde dehydrogenase are conserved among these proteins. The PhnN enzyme purified from recombinant E. coli showed broad substrate specificity towards various aromatic aldehydes, i.e., 1- and 2-naphaldehydes, cinnamaldehyde, vanillin, syringaldehyde, benzaldehyde and benzaldehydes substituted with a hydroxyl, methyl, methoxy, chloro, fluoro, or nitro group were converted to their corresponding carboxylic acids. Interestingly, E. coli expressing phnN was able to biotransform a variety of not only aromatic aldehydes, but also aromatic alcohols to carboxylic acids.

Sesamin ingestion regulates the transcription levels of hepatic metabolizing enzymes for alcohol and lipids in rats.

Abstract: Background: Sesamin, a major lignan in sesame seeds, has multiple functions such as stimulation effect of ethanol metabolism in mice and human, and prevention of ethanol-induced fatty liver in rats. However, the mechanism has not been clarified yet. Methods: The changes of gene expression were investigated in rats given 250 mg/kg of sesamin (sesamin rats) or vehicle (control rats) for three days by using a DNA microarray analysis. At 4 hr after the final ingestion, the profiles of gene expression in rat livers were compared. Results: The analysis showed that 38 transcripts were up-regulated with a significant change of more than two-fold and eight transcripts were down-regulated with a significant change to less than half in the livers of sesamin rats versus control rats. The gene expression levels of the early stage enzymes of (3-oxidation including long-chain acyl-CoA synthetase, very long-chain acyl-CoA synthetase and carnitine palmitoyltransferase were not changed, however, those of the late stage enzymes of β-oxidation including trifunctional enzyme in mitochondria, and acyl-CoA oxidase, bifunctional enzyme and 3-ketoacyl-CoA thiolase in peroxisomes, were significantly increased by sesamin ingestion. Also, in sesamin rats, the gene expression of aldehyde dehydrogenase was increased about three-fold, whereas alcohol dehydrogenase, liver catalase and CYP2E1 were not changed. Changes in the gene expression of alcohol- and aldehyde-metabolizing enzymes observed in a DNA microarray were also confirmed by a real-time PCR method. Conclusions: These results suggested that sesamin ingestion regulated the transcription levels of hepatic metabolizing enzymes for alcohol and lipids.

Expression of aldehyde dehydrogenase 2 in the normal esophageal epithelium and alcohol consumption in patients with esophageal cancer.

Abstract: Alcohol consumption is a risk factor for esophageal cancer. Acetaldehyde, a highly toxic intermediate produced from ethanol, is converted to acetic acid mainly by aldehyde dehydrogenase 2 (ALDH2) in the metabolic pathway of ethanol. Fifty percent of Japanese have inactive ALDH2 due to genetic polymorphism, which is considered to be a risk factor associated with esophageal cancer. In our previous study, we have demonstrated that ALDH2 is expressed in the esophagus with a considerable variation among individuals. In this study, we further investigated the expression of ALDH2 in esophagus and its relationship with risk factors of esophageal cancer. Tissue specimens resected from 51 patients with esophageal cancer were analyzed by immunohistochemistry using ALDH2-antibody. The immuno-staining of ALDH2 in the esophageal epithelium was compared with both the drinking habit and the occurrence of flushing that is closely associated with the ALDH2 deficiency. ALDH2 was not detectable in 8 (16%) among 51 specimens. All of the 8 patients were non- or light-drinkers but not heavy-drinkers. Among 18 patients showing the high level ALDH2 expression in the esophagus, 15 patients (83%) were heavy-drinkers. Although the relationship between the ALDH2 deficiency and drinking habit is not clear, the patients with ALDH2 deficiency tend to be non- or light drinkers while heavy-drinkers tend to have the active form of ALDH2. These results suggest that both inactive and active forms of ALDH2 are induced in the esophagus by heavy drinking and also support a hypothesis that ALDH2 deficiency might be a high-risk factor of esophageal cancer for the individuals having a heavy-drinking habit. To our knowledge, this is the first study demonstrating the induction of ALDH2 in the esophagus by ethanol consumption.

Characterization of a structure close to the coenzyme-binding site of liver aldehyde dehydrogenase.

Abstract: NAD analogues with the nicotinamide moiety exchanged for acetylpyridino-pentyl or acetylpyridino-butyl groups function as coenzymes in the enzymatic reaction with liver aldehyde dehydrogenase. The corresponding bromoacetyl derivatives bind to the coenzyme-binding site of the enzyme and inactive the protein by covalent modification of single residues close to the active site. Protection by coenzymes and substrate against the inactivation differs slightly for the two coenzyme analogues, suggesting the presence of more than one reactive residue. This is consistent with the results of differential carboxymethylation of cysteine residues of the basic isozyme in the presence and absence of the inhibitor disulfiram. The amino acid sequence around one reactive cysteine residue close to the active site of the acidic isozyme was determined after labeling with the butyl coenzyme analogue. This structure bears no extensive homology to corresponding known structures of dehydrogenases working on other types of aldehyde substrates.

Erythrocyte aldehyde dehydrogenase activity: lack of association with alcohol use and dependence or alcohol reactions in australian twins

Abstract: Aim: Aldehyde dehydrogenase 1 (ALDH1) has been advocated as a marker of alcohol intake. The absence or low levels of ALDH1 may be associated with alcohol-induced flushing or other reactions to alcohol in Europeans and therefore, with reduced alcohol use. This study tested whether variation in erythrocyte ALDH1 activity was associated with alcohol use, alcohol dependence or reactions to alcohol in unselected subjects of European descent, and whether variation in ALDH1 activity was subject to genetic influences. Methods: ALDH activity was measured in erythrocytes from 677 men and women who had participated in a twin study of alcohol use and dependence. Results: There were no significant effects of sex, alcohol consumption or alcohol dependence on ALDH activity. Subjects who reported reactions to alcohol did not have low activity. Women aged below 45 years had lower ALDH activity than men or older women. The heritability of ALDH activity was 56% (95% confidence interval = 42-67%). Conclusions: Previous reports that erythrocyte ALDH activity is low in alcoholics were not substantiated in this community-based sample. Associations with alcohol reactions were not found. ALDH activity varies widely between subjects, largely because of genetic factors.

Phenylacetaldehyde Oxidation by Freshly Prepared and Cryopreserved Guinea Pig Liver Slices: The Role of Aldehyde Oxidase

Abstract: Phenylacetaldehyde is formed when the xenobiotic and biogenic amine 2-phenylethylamine is inactivated by a monoamine oxidase-catalyzed oxidative deamination. Exogenous phenylacetaldehyde is found in certain foodstuffs such as honey, cheese, tomatoes, and wines. 2-Phenylethylamine can trigger migraine attacks in susceptible individuals and can become fairly toxic at high intakes from foods. It may also function as a potentiator that enhances the toxicity of histamine and tyramine. The present investigation examines the metabolism of phenylacetaldehyde to phenylacetic acid in freshly prepared and in cryopreserved guinea pig liver slices. In addition, it compares the relative contribution of aldehyde oxidase, xanthine oxidase, and aldehyde dehydrogenase in the oxidation of phenylacetaldehyde using specific inhibitors for each oxidizing enzyme. The inhibitors used were isovanillin for aldehyde oxidase, allopurinol for xanthine oxidase, and disulfiram for aldehyde dehydrogenase. In freshly prepared liver slices, phenylacetaldehyde was converted mainly to phenylacetic acid, with traces of 2-phenylethanol being present. Disulfiram inhibited phenylacetic acid formation by 80% to 85%, whereas isovanillin inhibited acid formation to a lesser extent (50% to 55%) and allopurinol had little or no effect. In cryopreserved liver slices, phenylacetic acid was also the main metabolite, whereas the 2-phenylethanol production was more pronounced than that in freshly prepared liver slices. Isovanillin inhibited phenylacetic acid formation by 85%, whereas disulfiram inhibited acid formation to a lesser extent (55% to 60%) and allopurinol had no effect. The results in this study have shown that, in freshly prepared and cryopreserved liver slices, phenylacetaldehyde is converted to phenylacetic acid by both aldehyde dehydrogenase and aldehyde oxidase, with no contribution from xanthine oxidase. Therefore, aldehyde dehydrogenase is not the only enzyme responsible in the metabolism of phenylacetaldehyde, but aldehyde oxidase may also be important and thus its role should not be ignored.

Metabolism of isovanillin by aldehyde oxidase, xanthine oxidase, aldehyde dehydrogenase and liver slices.

Abstract: Aromatic aldehydes are good substrates of aldehyde dehydrogenase activity but are relatively poor substrates of aldehyde oxidase and xanthine oxidase. However, the oxidation of xenobiotic-derived aromatic aldehydes by the latter enzymes has not been studied to any great extent. The present investigation compares the relative contribution of aldehyde dehydrogenase, aldehyde oxidase and xanthine oxidase activities in the oxidation of isovanillin in separate preparations and also in freshly prepared and cryopreserved liver slices. The oxidation of isovanillin was also examined in the presence of specific inhibitors of each oxidizing enzyme. Minimal transformation of isovanillin to isovanillic acid was observed in partially purified aldehyde oxidase, which is thought to be due to residual xanthine oxidase activity. Isovanillin was rapidly metabolized to isovanillic acid by high amounts of purified xanthine oxidase, but only low amounts are present in guinea pig liver fraction. Thus the contribution of xanthine oxidase to isovanillin oxidation in guinea pig is very low. In contrast, isovanillin was rapidly catalyzed to isovanillic acid by guinea pig liver aldehyde dehydrogenase activity. The inhibitor studies revealed that isovanillin was predominantly metabolized by aldehyde dehydrogenase activity. The oxidation of xenobiotic-derived aromatic aldehydes with freshly prepared or cryopreserved liver slices has not been previously reported. In freshly prepared liver slices, isovanillin was rapidly converted to isovanillic acid, whereas the conversion was very slow in cryopreserved liver slices due to low aldehyde dehydrogenase activity. The formation of isovanillic acid was not altered by allopurinol, but considerably inhibited by disulfiram. It is therefore concluded that isovanillin is predominantly metabolized by aldehyde dehydrogenase activity, with minimal contribution from either aldehyde oxidase or xanthine oxidase.

Aldehyde dehydrogenase (ALDH2) activity in hepatoma cells is reduced by an adenoviral vector coding for an ALDH2 antisense mRNA.

Abstract: Background: Individuals carrying the Glu487Lys coding mutation in the gene for mitochondrial aldehyde dehydrogenase (ALDH2) have a diminished capacity to metabolize acetaldehyde This deficiency leads to increases in blood acetaldehyde levels when they consume ethanol, which results in an aversion to alcohol and in marked protection against alcoholism In the present studies, we aimed to mimic the high-acetaldehyde low-ALDH2 activity phenotype in a rat hepatoma cell line by inhibiting Aldh2 gene expression by an Aldh2 antisense-coding gene carried by an adenoviral vector Methods: We designed and produced elevated titers of adenoviral vectors (1013 virions/ml) carrying Aldh2 cDNA cloned in the reverse orientation preceded by a CMV promoter and followed by a poly-A termination signal Rat hepatoma cells were infected with these vectors Results: Studies showed that 1) the antisense gene is actively transcribed in the cells and high levels of antisense mRNA are attained, 2) the antisense gene reduced ALDH2 activity by 65%, and 3) when incubated with 10 mM ethanol, acetaldehyde accumulation by cells increased 8-fold to levels (80–90 μM) known to be aversive to animals and humans Conclusions: Data presented show that antialcohol drugs that inhibit Aldh2 gene expression can be generated endogenously in liver cells infected by an adenoviral vector carrying an antisense-coding gene, thus mimicking the high-acetaldehyde phenotype that exists in humans carrying the Glu487Lys mutation who are protected against alcoholism

Nitrate-based vasodilators inhibit multiple vascular aldehyde dehydrogenases

Abstract: Nitrate-based vasodilators (NBVs) are commonly used to treat multiple sequelae of atherosclerosis. A commonly used NBV, glyceryl trinitrate (GTN) is bioactivated by mitochondrial, class 2 aldehyde dehydrogenase (ALDH2). ALDH2 and other ALDHs are NAD(P)+-dependent enzymes critical to the detoxification of cytotoxic lipid-aldehydes elevated in atherosclerotic lesions, such as trans-4-hydroxy-2-nonenal (HNE). The GTN bioactivation step, however, inac-tivates ALDH2 and may alter the metabolism of these aldehydes. In this study, we tested the hypothesis that multiple ALDH enzymes are inhibited by different NBVs. ALDH2, ALDH3A, and ALDH5A were present in aorta with ALDH2 and ALDH3A localized to the smooth muscle layers. GTN (1 microM) inhibited ALDH2 activity (55 +/- 6% of control) and ablated ALDH3 activity. In contrast, isosorbide-2,5-dinitrate (ISDN, 1 microM) inhibited ALDH3 activity (1.1 +/- 0.4% of control) but did not inhibit ALDH2 activity even up to 50 microM ISDN. In homogenates of rat aorta, GTN (1 microM) inhibited the NAD+-dependent (41 +/- 5% of control) and NADP+-dependent (25 +/- 6% of control) detoxification of HNE. The inhibition of ALDH3A, but not ALDH2, could be prevented by the addition of dithiothreitol. These studies demonstrate that GTN and ISDN possess selectivity for ALDH inactivation with different mechanisms of inactivation.

Mechanisms in alcohol-associated carcinogenesis.

Abstract: This review represents an overview on some aspects of pathogenetic mechanisms in alcohol-associated carcinogenesis and is based on presentations held on the symposium Mechanisms in alcohol-associated carcinogenesis at the 2004 ISBRA Meeting in Heidelberg/Mannheim, Germany. The chairs were Nils Homann and Hiromasa Ishii. The presentations were (1) Genetic polymorphisms of alcohol and aldehyde dehydrogenases, mean corpuscular volume and cancer risk of the upper aerodigestive tract in Japanese by Akira Yokoyama; (2) Retinoids, alcohol and carcinogenesis by Xiang-Dong Wang; (3) Bacterial ethanol metabolism and cancer by Nils Homann; (4) The role of ethanol metabolism in alcohol-associated carcinogenesis by Helmut K. Seitz; (5) Alcohol and breast cancer: potential mechanisms by Keith W. Singletary.

A novel test for identifying genes involved in aldehyde detoxification in the yeast. Increased sensitivity of superoxide-deficient yeast to aldehydes and their metabolic precursors.

Abstract: A novel test for the identification of genes involved in aldehyde metabolism is proposed, based on detection of altered sensitivity of the yeast to corresponding alcohols, metabolic precursors of the aldehydes. This attitude enabled to an unexpected detection increased sensitivity of mutants devoid of CuZn-superoxide dismutase (CuZnSOD) to allyl alcohol (precursor of acrolein) and nonenol. We interpret this finding as due to inactivation of some important element of aldehyde detoxification by increased flux of superoxide in DeltaCuZnSOD mutants.