Alcohol Dehydrogenase Phenotypes Research Articles

GENETIC POLYMORPHISMS IN ETHANOL METABOLISM: ISSUES AND GOALS FOR PHYSIOLOGICALLY BASED PHARMACOKINETIC MODELING*

Chronic exposure to excessive ethanol consumption has adverse effects on virtually all organs and tissues in the body, including but not limited to the liver, pancreas, reproductive organs, central nervous system, and the fetus. Exposure to ethanol can also enhance the toxicity of other chemicals. Not all persons exposed to the same amount of ethanol experience the same degree of adverse effects. For example, only 12% to 13% of alcohol abusers develop cirrhosis. Possible factors which may alter susceptibility include age, sex, nutritional status, health status (i.e., smokers) and race. Some of these factors affect susceptibility because they alter ethanol metabolism, which occurs primarily in the liver by alcohol dehydrogenase (ADH). Genetic polymorphisms for ADH partially account for the observed differences in ethanol elimination rates among various populations but the relative contribution to susceptibility is not completely understood. Incorporation of the kinetic parameters associated with ADH polymorphisms into a physiologically based pharmacokinetic (PBPK) model for ethanol will aid in assessing the relative contribution to susceptibility. The specific information required to develop this model includes Km and Kcat values for each ADH isoform and the amount of each isoform present in the liver. Blood ethanol concentrations (BEC) from various populations with known ADH phenotypes are also necessary to validate the model. The impact of inclusion of these data on PBPK model predictions was examined using available information from adult white and African American males.

First pass metabolism of ethanol is strikingly influenced by the speed of gastric emptying

Background—Ethanol undergoes a first pass metabolism (FPM) in the stomach and liver. Gastric FPM of ethanol primarily depends on the activity of gastric alcohol dehydrogenase (ADH). In addition, the speed...

Genetic polymorphism and activities of human lung alcohol and aldehyde dehydrogenases: implications for ethanol metabolism and cytotoxicity.

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) exhibit genetic polymorphism and tissue specificity. ADH and ALDH isozyme phenotypes from 39 surgical Chinese lung specimens were identified by agarose isoelectric focusing. The identity of the lung beta-ADHs was further demonstrated by their characteristic pH-activity profiles for ethanol oxidation, Km values for NAD and ethanol, and inhibition by 4-methylpyrazole or 1,10-phenanthroline. The beta 2 allele, coding for beta 2 polypeptide, was found to be predominant in the lung specimens studied. The ADH activities in the lungs with the homozygous phenotype ADH2 2-2 (exhibiting beta 2 beta 2) and ADH2 1-1 (exhibiting beta 1 beta 1) and the heterozygous phenotype ADH2 2-1 (exhibiting beta 2 beta 2, beta 2 beta 1, and beta 1 beta 1) were determined to be 999 +/- 77, 48 +/- 17, and 494 +/- 61 nmol/min/g tissue, respectively. Fifty-one percent of the specimens studied lacked the ALDH2 activity band on the isoelectric focusing gels. The activities in the lung tissues with the ALDH2-active phenotype and the inactive phenotype were determined to be 30 +/- 3 and 17 +/- 1 nmol/min/g tissue, respectively. These findings indicate that human pulmonary ethanol-metabolizing activities differ significantly with respect to genetic polymorphism at both the ADH2 and the ALDH2 loci. The results suggest that individuals with high Vmax beta 2-ADH and deficient in low-Km mitochondrial ALDH2, accounting for approximately 45% of the Chinese population, may end up with acetaldehyde accumulation during alcohol consumption, rendering them vulnerable to tissue injury caused by this highly reactive and toxic metabolite.

Identification of tomato cultivars (Lycopersicon esculentum) by polyacrylamide isoelectric focusing

Isozyme analyses have been used for the definitive identification of many plant cultivars, but not for cultivated tomatoes. Six isozyme systems, namely alcohol dehydrogenase, acid phosphatase, phosphoglucomutase, esterase, phosphoglucoisomerase and 6-phosphogluconate dehydrogenase of tomato seed extracts were resolved by isoelectric focusing on polyacrylamide gels with a narrow pH gradient. Nine alcohol dehydrogenase phenotypes were distinguished which, with three acid phosphatase phenotypes, identified twelve of the seventeen cultivars. Fewer differences were found for the other isozymes. Since this method could differentiate between breeding parents and their progeny it is concluded that further investigations are warranted.

Liver alcohol and aldehyde dehydrogenase isozymes in a Chinese population in Hong Kong.

The distribution of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) isozymes in the livers of a Chinese population in Hong Kong was examined. Among the 90 livers examined, 7 were typical ADH phenotype consisting the normal beta 1-type isozymes and 83 were atypical phenotype consisting the beta 2-type isozymes. Livers of 48 subjects were of deficient type in ALDH containing ALDH-II alone and 42 were of normal type with both ALDH-I and ALDH-II. When the combination of ADH and ALDH isozymes is considered, the Chinese population in Hong Kong falls into 4 subgroups. For each group, the rates of ethanol and acetaldehyde clearance have a distinct and characteristic potential which is directly related to its particular combination of isozymes.

Genotyping of human alcohol dehydrogenases at the ADH2 and ADH3 loci following DNA sequence amplification

Humans are polymorphic at two of the alcohol dehydrogenase (ADH) loci important in ethanol metabolism, ADH2 and ADH3. Although the coding regions of these genes are 94% identical, they produce subunits that differ greatly in kinetic properties in vitro. These differences are likely to be reflected in the pharmacokinetics of alcohol metabolism, but studies have been hampered by the need to use liver biopsy specimens to determine the ADH phenotype. This problem has now been overcome by determining the genotype at these loci using DNA that has been amplifled in vitro by the polymerase chain reaction. We report here the identification of all three of the ADH2 alleles and both of the ADH3 alleles. Any pair of ADH2 or ADH3 alleles can be distinguished using allele-specific oligonucleotide probes directed at their single base pair difference. In addition, ADH2 2 can be distinguished from ADH2 1 and ADH2 3 by detecting a new MaeIII site created in the third exon by the single base pair alteration in ADH2 2 .

Chlorpropamide-alcohol flush reaction and isoenzyme profiles of alcohol dehydrogenase and aldehyde dehydrogenase.

To investigate the enzymatic basis of the chlorpropamide-alcohol flush reaction (CPAF) we compared the isoenzyme profiles of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) from liver biopsies, and of ALDH alone from erythrocytes and leucocytes, in CPAF-positive and CPAF-negative subjects. No differences were seen in ADH or ALDH phenotypes, or in the relative activities of the isoenzymes, between the two groups before chlorpropamide was given; in particular, no subjects showed the 'null' ALDH phenotype that is associated with the alcohol flush reaction in oriental subjects. There was a significant decrease in erythrocyte ALDH activity after 7 days' treatment with chlorpropamide in CPAF-positive individuals but no such difference was seen in CPAF-negative subjects. These results indicate that CPAF has a different enzymatic basis from the alcohol flush reaction of oriental subjects and suggest that in CPAF-positive subjects erythrocyte ALDH may be particularly susceptible to inhibition by chlorpropamide.

Alcohol dehydrogenase isozymes in baboons: tissue distribution, catalytic properties, and variant phenotypes in liver, kidney, stomach, and testis.

Isoelectric focusing and cellulose acetate electrophoresis were used to examine the multiplicity, tissue distribution, and variability of alcohol dehydrogenase (ADH) among baboons, a primate species used as a model for research on alcohol metabolism and alcohol-induced liver pathology. Five major ADH isozymes were resolved and distinguished on the basis of their isoelectric points, tissue distributions, relative activities with alcohol substrates, and sensitivities to inhibition with 4-methyl pyrazole. ADH-1 and ADH-2 exhibited class I kinetic properties and were observed in high activity in kidney and liver extracts, respectively. ADH-3 showed class II kinetic properties, exhibiting high activity in stomach extracts, and was widely distributed in extracts of other baboon tissues, including kidney, esophagus, heart, testis, brain, and male sex accessory tissues. ADH-4 also showed class II ADH properties but was found only in liver (similar to human "pi-ADH"). ADH-5 exhibited class III ADH kinetic properties, being inactive with ethanol up to 0.5 M (similar to human "chi-ADH") and was distributed widely in baboon tissue extracts. Major activity variation was observed for liver ADH-4 between different animals. An electrophoretic variant for ADH-3 was observed for the enzyme in stomach, kidney, and testis extracts, and activity variation existed for this isozyme in kidney extracts. It is apparent that baboon ADH shares a number of features with the human ADH phenotype; however, several species-specific differences were observed, particularly for the liver and kidney class I isozymes and for stomach ADH.

Alcohol and aldehyde dehydrogenase isoenzymes in North American Indians.

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) isoenzyme phenotypes were determined in autopsy liver samples from 50 North American Indians from New Mexico. Forty-six of the 50 livers had sufficient ADH activity to allow phenotyping at the ADH2 and ADH3 loci. All 46 livers possessed the "typical" ADH2 1-1 phenotype. The frequency of the ADH3(2) allele was 0.59 and is the highest thus far reported in any racial population. All 50 livers possessed the ALDH I isoenzyme which exhibits the greatest anodic mobility on starch gel electrophoresis at pH 7.6. The results show that ADH and ALDH phenotypes among American Indians living in New Mexico are very similar to those of Caucasian populations and quite different from those of Orientals.

MORPHOLOGICAL AND GENETIC VARIATION IN ECOLOGICALLY CENTRAL AND MARGINAL POPULATIONS OF RUMEX ACETOSELLA L. (POLYGONACEAE)

Six ecologically central (old field) and marginal (strip mine) populations of the hexaploid Rumex acetosella were collected, grown under uniform conditions, and examined for genetic and morphological variation. Extensive electrophoretic variation was found in alcohol dehydrogenase and phosphoglucose isomerase, while other enzymes surveyed showed little or no variation. Hedrick's genotypic measure of identity revealed mean values of 0.506 for central populations, 0.836 for marginal populations, and 0.633 for comparisons of central with marginal populations. Alcohol dehydrogenase phenotypes had significantly fewer electromorphs per individual in marginal populations. Clones of individuals from both environments were subjected to different watering regimes. No significant differences in root/shoot ratio, leaf number, total leaf area, or relative growth rate were found between strip mine and old‐field individuals within each watering treatment, although significant differences were found between watering treatments. There are small, but significant amounts of isozyme differentiation between central and marginal populations, while there was no such differentation for morphological characters.

Hepatic ADH and ALDH isoenzymes in different racial groups and in chronic alcoholism

A method has been developed for simultaneous analysis of alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) isoenzymes in small (2.5 mg) liver biopsy cores by starch gel electrophoresis. All the currently recognized hepatic isoenzymes coded by ADH 1, ADH 2, ADH 3 and ADH 4 can be detected as can the five ALDH isoenzymes. Using this technique we have investigated the isoenzyme composition of liver samples from English and Chinese subjects and a group of chronic alcoholics. Pronounced racial differences in frequency of ADH 2 and ALDH phenotypes were found—only 2 (4%) of English controls had the “atypical” ADH 2 variant whereas this was present in 42 (84%) of Chinese subjects, and whereas all the English subjects had the rapidly migrating mitochondrial isoenzyme of ALDH, this was absent in 27 (54%) of Chinese. No differences in ADH or ALDH phenotype were seen in the chronic alcoholics, all of whom were of English origin, compared with the English controls, but there was a reduction in overall ALDH activity and particularly in the mitochondrial isoenzymesin those with cirrhosis. The reduction in ALDH activity is probably acquired; by limiting acetaldehyde oxidation it could be responsible for the rapid deterioration in liver function in patients who continue drinking excessively.

Relationship between kinetics of liver alcohol dehydrogenase and alcohol metabolism

Since alcohol dehydrogenase (ADH) catalyzes the rate-limiting step for ethanol metabolism, knowledge of the steady-state kinetics of ADH in liver is fundamental to the understanding of the pharmacokinetics of ethanol elimination. Accordingly, we have determined the kinetic properties of purified ADH isoenzymes in rat and human liver. At low ethanol concentrations, rat liver ADH obeys the Theorell-Chance mechanism and the equation predicts that activity in vivo is limited below V max mainly by NADH inhibition. At ethanol concentrations above 10mM, substrate inhibition, consistent with the formation a dead-end ADH-NADH-ethanol complex, also becomes a rate-limiting factor. ADH activity, calculated from this equation and the concentrations of substrates and products present in liver during ethanol oxidation, agrees well with ethanol elimination rates measured in vivo. With human liver ADH, large differences are observed in the kinetic properties of 5 homodimeric isoenzymes: γ 1 γ 1 and γ 2 γ 2 exhibit negative cooperativity for ethanol saturation, while αα, β 1 β 1 and β Ind β Ind obey Michaelis-Menten kinetics. At pH 7.5, K m values for ethanol and V max values range from 0.048 mM and 9 min −1 for β 1 β 1 to 64 mM and 560 min −1 for β Ind β Ind , respectively. Therefore, individuals with different ADH phenotypes shoudl display different ethanol elimination profiles.

Isozyme variability in species of the genus Drosophila. VIII. The alcohol dehydrogenase polymorphism in North Carolina populations of D. melanogaster.

Adult Drosophila melanogaster flies collected from populations broadly dispersed over ecological and geographic strata of North Carolina, and over a period of 4 years, were analyzed for alcohol dehydrogenase phenotypes by gel electrophoresis. Gene frequencies in spring-summer-fall field collections were remarkably stable over all strata. Two winter collections exhibited contrasting gene frequency changes. In one case the results are interpreted in terms of long-distance migration from Florida, while the other is explicable by assignment of a causal role to environmental factors which accompany the winter season.

Developmental genetics of the alcohol dehydrogenase locus of the Japanese quail.

In Japanese quail ♀ × chicken ♂ hybrids, the preferential transcription and translation of a maternally derived quail allele appeared to occur at the autosomally inherited liver alcohol dehydrogenase (ADH) locus. In these hybrids, the product of a paternally derived chicken allele was not detectable until 3 days after the hatching (Castro-Sierra and Ohno, 1968). The present study was designed to answer the following two questions: (1) Exactly when and where in development does ADH first appear? (2) Does the appearance of the maternal gene product precede that of the paternal ADH in development of intraspecific heterozygotes as well? B and C electrophoretic variants of ADH of the Japanese quail were utilized. Developmental sequence was established. Whole embryo extracts of earlier developmental stages showed no ADH activity. At the 9th day of incubation, ADH activity was detected for the first time in newly organized liver. Until the time of hatching, no other tissue showed ADH activity. A few days after the hatching, ADH activity appeared in the kidney. Thus, the transcriptional activity by the ADH locus of Japanese quail appears highly tissue-specific. Four hundred and seven progeny resulting from 9 breeding pairs of the 5 heterozygote-producing mating types were analyzed at the 9th day (−8 days) of incubation and at the day of hatching (day 0) for their ADH phenotypes. It is concluded that so far as B/C heterozygotes of Japanese quail are concerned, the synchronous appearance of both parental gene products is a rule at this locus.