genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol...

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Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels Yasuyuki Nakamura a, *, Kenji Amamoto b , Shinji Tamaki a , Tomonori Okamura b , Yasuyuki Tsujita a , Yoshiki Ueno a , Yoshikuni Kita b , Masahiko Kinoshita a , Hirotsugu Ueshima b a First Department of Internal Medicine, Shiga University of Medical Science, Seta Tsukinowa-cho, Otsu City, Shiga 520-2192, Japan b Department of Health Science, Shiga University of Medical Science, Seta Tsukinowa-cho, Otsu City, Shiga 520-2192, Japan Received 5 December 2001; received in revised form 30 January 2002; accepted 8 February 2002 Abstract Moderate drinkers with a defective alcohol dehydrogenase type 3 (ADH3) genotype have higher high-density lipoprotein (HDL) levels and a decreased risk of coronary artery disease (CAD). We examined the interaction between the aldehyde dehydrogenase type 2 (ALDH2), alcohol intake, and HDL levels in 826 men and 1295 women in a rural town in Japan. The ALDH2 genotype of each subject was determined by polymerase chain reaction (PCR) analysis. HDL was adjusted for the alcohol intake, age, body mass index, smoking status, total cholesterol, triglycerides and HbA1c levels. None of the subjects had a history or ECG suggestive of CAD. The proportions of ALDH2, *1/*1, *1/*2, and *2/*2 (defective homozygote) were 45.8, 46.0, and 8.2%, respectively, for men. Drinking more than two drinks daily was associated with lower HDL levels in men with the defective genotypes compared with men with a normal genotype (55.69 /0.9 vs. 51.29 /0.9 mg/dl, mean9 /S.E., P B/0.0001). Also, drinking more than 0.5 drinks daily was not associated with beneficial effects on HDL levels in women with defective ALDH2 genotypes. Conclusions: Alcohol intake did not have beneficial effects on HDL levels in the defective ALDH2 genotype and may not protect against CAD in subjects with defective ALDH2 genotypes. # 2002 Elsevier Science Ireland Ltd. All rights reserved. Keywords: Alcohol; Polymorphism; Aldehyde dehydrogenase 2; High-density lipoprotein; Total cholesterol 1. Introduction Epidemiological studies have consistently shown that moderate alcohol consumption is associated with a reduced risk of myocardial infarction [1,2], despite the fact that alcohol consumption is closely related to the prevalence of hypertension [3], one of the major coronary risk factors. Although alcohol may be a surrogate for favourable socioeconomic or lifestyle factors associated with a reduction in coronary risk [4], and anti-inflammatory action of alcohol could contribute to the link between moderate consumption and lower cardiovascular mortality [5], most of the apparent benefit of alcohol consumption on the risk of myocardial infarction has been attributed to an increase in high-density lipoprotein (HDL) levels [2,6,7]. Two enzyme systems, alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), play key roles in alcohol metabolism [8]. Class I ADH isoenzymes, encoded by ADH1, ADH2 and ADH3, have low Michaelis /Menten constants (K m ) for ethanol. ADH2 and ADH3 show polymorphisms that produce isoen- zymes with distinct kinetic properties [8]. In Caucasian populations, while variant alleles at the ADH2 locus are relatively uncommon, but 40 /50% present at the ADH3 locus [9]. Recently, it has been reported in Caucasian subjects that moderate drinkers with a defective alcohol dehydrogenase ADH3 genotype have higher HDL levels, and a decreased risk of myocardial infarction [6]. Acetaldehyde, a metabolite of ethanol by ADH, is quickly metabolised into acetate by ALDH. Although there are multiple forms of ALDH in the liver [9,10], the mitochondrial enzyme encoded by the ALDH2 locus on * Corresponding author. Tel.: /81-77-548-2213; fax: /81-77-543- 5839. E-mail address: [email protected] (Y. Nakamura). Atherosclerosis 164 (2002) 171 /177 www.elsevier.com/locate/atherosclerosis 0021-9150/02/$ - see front matter # 2002 Elsevier Science Ireland Ltd. All rights reserved. PII:S0021-9150(02)00059-X

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Page 1: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

Genetic variation in aldehyde dehydrogenase 2 and the effect ofalcohol consumption on cholesterol levels

Yasuyuki Nakamura a,*, Kenji Amamoto b, Shinji Tamaki a, Tomonori Okamura b,Yasuyuki Tsujita a, Yoshiki Ueno a, Yoshikuni Kita b, Masahiko Kinoshita a,

Hirotsugu Ueshima b

a First Department of Internal Medicine, Shiga University of Medical Science, Seta Tsukinowa-cho, Otsu City, Shiga 520-2192, Japanb Department of Health Science, Shiga University of Medical Science, Seta Tsukinowa-cho, Otsu City, Shiga 520-2192, Japan

Received 5 December 2001; received in revised form 30 January 2002; accepted 8 February 2002

Abstract

Moderate drinkers with a defective alcohol dehydrogenase type 3 (ADH3) genotype have higher high-density lipoprotein (HDL)

levels and a decreased risk of coronary artery disease (CAD). We examined the interaction between the aldehyde dehydrogenase

type 2 (ALDH2), alcohol intake, and HDL levels in 826 men and 1295 women in a rural town in Japan. The ALDH2 genotype of

each subject was determined by polymerase chain reaction (PCR) analysis. HDL was adjusted for the alcohol intake, age, body mass

index, smoking status, total cholesterol, triglycerides and HbA1c levels. None of the subjects had a history or ECG suggestive of

CAD. The proportions of ALDH2, *1/*1, *1/*2, and *2/*2 (defective homozygote) were 45.8, 46.0, and 8.2%, respectively, for men.

Drinking more than two drinks daily was associated with lower HDL levels in men with the defective genotypes compared with men

with a normal genotype (55.69/0.9 vs. 51.29/0.9 mg/dl, mean9/S.E., P B/0.0001). Also, drinking more than 0.5 drinks daily was not

associated with beneficial effects on HDL levels in women with defective ALDH2 genotypes. Conclusions: Alcohol intake did not

have beneficial effects on HDL levels in the defective ALDH2 genotype and may not protect against CAD in subjects with defective

ALDH2 genotypes. # 2002 Elsevier Science Ireland Ltd. All rights reserved.

Keywords: Alcohol; Polymorphism; Aldehyde dehydrogenase 2; High-density lipoprotein; Total cholesterol

1. Introduction

Epidemiological studies have consistently shown that

moderate alcohol consumption is associated with a

reduced risk of myocardial infarction [1,2], despite the

fact that alcohol consumption is closely related to the

prevalence of hypertension [3], one of the major

coronary risk factors. Although alcohol may be a

surrogate for favourable socioeconomic or lifestyle

factors associated with a reduction in coronary risk

[4], and anti-inflammatory action of alcohol could

contribute to the link between moderate consumption

and lower cardiovascular mortality [5], most of the

apparent benefit of alcohol consumption on the risk of

myocardial infarction has been attributed to an increase

in high-density lipoprotein (HDL) levels [2,6,7]. Two

enzyme systems, alcohol dehydrogenase (ADH) and

aldehyde dehydrogenase (ALDH), play key roles in

alcohol metabolism [8]. Class I ADH isoenzymes,

encoded by ADH1, ADH2 and ADH3, have low

Michaelis�/Menten constants (Km) for ethanol. ADH2

and ADH3 show polymorphisms that produce isoen-

zymes with distinct kinetic properties [8]. In Caucasian

populations, while variant alleles at the ADH2 locus are

relatively uncommon, but 40�/50% present at the ADH3

locus [9]. Recently, it has been reported in Caucasian

subjects that moderate drinkers with a defective alcohol

dehydrogenase ADH3 genotype have higher HDL

levels, and a decreased risk of myocardial infarction [6].

Acetaldehyde, a metabolite of ethanol by ADH, is

quickly metabolised into acetate by ALDH. Although

there are multiple forms of ALDH in the liver [9,10], the

mitochondrial enzyme encoded by the ALDH2 locus on

* Corresponding author. Tel.: �/81-77-548-2213; fax: �/81-77-543-

5839.

E-mail address: [email protected] (Y. Nakamura).

Atherosclerosis 164 (2002) 171�/177

www.elsevier.com/locate/atherosclerosis

0021-9150/02/$ - see front matter # 2002 Elsevier Science Ireland Ltd. All rights reserved.

PII: S 0 0 2 1 - 9 1 5 0 ( 0 2 ) 0 0 0 5 9 - X

Page 2: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

chromosome 12 has a very low Km for acetaldehyde. The

gene for ALDH2 is located on the long arm of

chromosome 12, and the low metabolic function of

this isozyme is due to a point mutation at position 1510with a G to A transition in exon 12, which changes the

codon GAA encoding glutamic acid (Glu) at position

487 into a lysine codon (AAA) [11]. Harada and co-

workers found a polymorphism at �/357 with a G to A

substitution in the promoter region of the ALDH2 gene,

and reported that the ALDH2*2 allele was linked to the

G promoter allele and the ALDH2*1 allele was linked to

the A promoter allele [12]. Nearly half of the Asianpopulation lacks this enzyme, resulting in high acetal-

dehyde concentrations after drinking, as well as flushing

and other symptoms [11,13]. ALDH2 deficiency in-

cludes the heterozygote ALDH2*1/*2 and the atypical

homozygote ALDH2*2/*2. Although ALDH2 defi-

ciency can protect against alcoholism and alcohol liver

disease [11,14], the effects of moderate drinking on HDL

levels among ALDH2-deficient subjects are not known.Thus, we examined the interaction between the ALDH2

genotype, the level of alcohol intake, and HDL and total

cholesterol (TC) levels in 2121 Japanese men and

women.

2. Subjects and methods

2.1. Subjects

Of 2892 local residents who underwent a mass

medical examination in 1999 at S town (population of

about 15 000), a mountain farming community in

Western Japan, a total of 2395 subjects consisting of

917 men (mean age 589/15.7) and 1478 women (mean

age 56.39/15.6) from whom informed consent was

obtained after a full explanation of this genetic studywere enrolled. Of these, 274 were excluded because the

genotype was not determined (46 men, 44 women) or

they were taking lipid-lowering medications (45 men,

139 women). None of the subjects had a history or ECG

suggestive of coronary artery disease. This study was

approved by the Institutional Review Board of Shiga

University of Medical Science (No. 11�/15, 1999).

2.2. Genetic analysis

DNA was extracted from the buffy coat of whole

blood. ALDH2 genotypes were determined by the

polymerase chain reaction-the restriction fragment

length polymorphism (PCR-RFLP) method developed

by Takeshita and co-workers. [15]. Briefly, CAAATTA-

CAGGGTCAACTGCT and CCACACTCA-CAGTTTTCTCTT were used as 5? and 3? primers,

respectively. The corresponding DNA segment of

ALDH2 was amplified by the PCR on a Gene Amp

PCR System 9700 (Perkin�/Elmer, USA). After initial

denaturation at 94 8C for 60 s, annealing at 52 8C for

30 s, and extension at 72 8C for 30 s, samples were

incubated at 72 8C for 60 s. The amplified DNAfragment was cleaved by the restriction endonuclease

KSP6321, electrophoresed on a 3.5% Nusieve Agarose

gel (Biowhittaker Molecular Applications, USA), and

observed for a 112-bp fragment (ALDH2*2/*2 geno-

type), a 23-bp fragment (ALD2*1/*1 genotype), and

both 112-bp and 23-bp fragments (ALDH2*1/*2 geno-

type). Genotypes, determined by the PCR-RFLP

method on a total of 75 random samples consisting of25 PCR products of each genotype, were confirmed by

direct sequencing. Briefly, after fractionation of PCR-

RFLP products on 1% agarose gels, the desired DNA

bands were cut out, and the DNA was purified using a

QIAquick Gel Extraction Kit (QIAGEN, USA) and was

amplified with the above 5? primer, followed by analysis

on a ABI PRISM 310 Genetic Analyser (Perkin�/Elmer).

2.3. Measurements of serum cholesterol

Non-fasting blood was drawn and serum TC and

HDL levels were determined in one laboratory (Medic,

Shiga). The measurement precision and accuracy of

these serum lipids were certificated through a lipid

standardisation program by Osaka Medical Center for

Cancer and Cardiovascular Diseases, which is a member

of Cholesterol Reference Method Laboratory Network(CRMLN) controlled by the Centers for Disease Con-

trol and Prevention (Atlanta) [16].

2.4. Information on alcohol intake

Using a questionnaire, a well-trained public health

nurse obtained information on alcohol intake from each

subject. The frequency of drinking during an averageweek and the alcohol intake on each occasion were

determined, and the alcohol intake per week was

calculated, which was then divided by 7 to give the

alcohol intake per day. Two drink units were defined as

one GO (180 ml) of sake (23 g of ethanol), two bottles

(350�/2 ml) of beer, two single whiskeys, or two glasses

(180 ml) of wine.

2.5. Statistics

SAS version 6.12 on UNIX was used throughout the

study. The t -test was used to compare the means of

continuous variables in two groups and the chi-square

test was used to compare dichotomous variables. To

compare means among the three genotype groups, one-

way analysis of variance (ANOVA) was used. To assesswhether the ALDH2 genotype affected the relation

between the level of alcohol intake and HDL and TC

levels, we performed an analysis of covariance by

Y. Nakamura et al. / Atherosclerosis 164 (2002) 171�/177172

Page 3: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

adjusting for confounding factors: age, amount of

alcohol intake, BMI, and triglyceride levels (plus TC

levels in case of analysis for HDL), HbA1c value and

current smoking status. Since the average levels ofalcohol intake were lower among women than among

men, the categories of alcohol intake were dichotomised

at two drinks per day for men and at 0.5 drinks per day

for women.

Due to higher incidence of cholestryl ester transfer

protein (CETP) deficiency among subjects with high

HDL levels [17,18], separate analyses on the interaction

between the ALDH2 genotype, amount of alcoholintake, and HDL levels were performed after excluding

the subjects with HDL levels more than 80 mg/dl. There

were 136 subjects (29 men and 107 women) whose HDL

levels more than 80 mg/dl.

All P values were two-tailed, and P B/0.05 was

considered significant.

3. Results

3.1. Direct sequencing

Direct sequencing of 25 randomly selected samples of

each ALDH2 genotype identified by the PCR-RFLP

method did not show any discrepancy between the PCR-RFLP-determined genotypes in exon 12 of ALDH2 and

the 1510G/A sequence variations. In addition, no

sequence variations were found other than the 1510G/

A variations in exon 12 of ALDH2 [12]. Other reported

variations such as 1464G/A and 1486G/A were not

observed in the present study [19].

3.2. Characteristics of the subjects

Table 1 shows the characteristics of the subjects. Age,

height, weight, triglycerides, variables related to alcohol

consumption and the incidence of smoking were higher

in men, whereas TC and HDL levels were higher in

women. There were no difference in BMI and HbA1c.

Table 2 presents the variables in Table 1 grouped by

gender and the ALDH2 genotype. The frequencies ofthe ALDH2*1/*1, *1/*2 and *2/*2 genotypes were 45.8,

46.0 and 8.2% in men, and 50.6, 42.0 and 7.4% in

women, respectively, which indicated there were no

significant gender differences in the distribution. In

men, a comparison of ALDH2 genotypes showed

significant differences in weight, BMI, triglycerides,

and drinking-related variables. In comparison with

other genotypes, the subjects with the ALDH2*1/*1genotype consumed more alcohol on average and more

subjects had a habit of drinking. In addition, body

weight, BMI, and triglycerides were higher in this

subgroup. Age, height, TC and HDL levels, HbA1c

and the incidence of smoking were not different among

the three genotypes in men.

In women, subjects with the ALDH2*2/*2 genotype

were slightly older than those in the other groups. TClevels were lower in the ALDH2*1/*1 genotype.

Although the frequency and the average amount of

alcohol consumed were lower in women than in men,

women with the ALDH2*1/*1 genotype consumed more

alcohol on average and more had a habit of drinking. In

contrast to the results in men, significantly more women

with the ALDH2*2/*2 genotype were smokers com-

pared with women of the other genotypes. Althoughweight, BMI, and triglycerides were higher in men with

the ALDH2*1/*1 genotype, there were no similar

differences among the three genotypes in women.

3.3. HDL and TC levels adjusted for confounding factors

Among men who consumed less than two drinks per

day, the mean adjusted HDL levels were not signifi-

cantly different among the three ALDH2 genotypes.

However, among men who consumed at least two drinks

per day, the mean adjusted HDL levels were higher

Table 1

Characteristics of the subjects, grouped by gender

Variable gender Men Women P

N (Total�/2121) 826 1295

Age (year) 57.99/15.7 55.49/15.8 B/0.001

Height (cm) 165.79/6.7 152.89/7.0 B/0.001

Weight (kg) 62.29/9.7 52.49/8.4 B/0.001

BMI (m2) 22.69/3.0 22.49/3.2 0.19

TC 184.09/33.3 196.49/34.7 B/0.001

HDL 52.89/14.1 60.79/14.0 B/0.001

TG 147.49/102.5 114.59/66.4 B/0.001

HbA1c 5.19/0.8 5.19/0.7 0.23

EtOH intake (drinks per day) 1.99/1.9 0.39/0.6 B/0.001

Number of drinkers at �/0.5 drinks per week 593 (71.8%) 289 (22.3%) B/0.001

Number of drinkers at ]/two drinks per day 426 (51.6%) 63 (4.9%) B/0.001

Smokers 421 (51.0%) 97 (7.5%) B/0.001

Y. Nakamura et al. / Atherosclerosis 164 (2002) 171�/177 173

Page 4: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

among those with the ALDH2*1/*1 genotype than those

with the ALDH2*1/*2 genotype (55.69/0.9 vs. 51.29/0.9

mg/dl, mean9/S.E., P B/0.0001). In the ALDH2*1/*1

genotype A, men who consumed at least two drinks per

day showed higher mean adjusted HDL levels than men

who consumed less than two drinks per day (B/two

drinks per day: 52.69/1.1 vs. ]/two drinks per day:

55.69/0.9 mg/dl, P B/0.05) (Fig. 1A). These differences

were not seen in the defective ALDH2 genotypes.Among men who consumed less than two drinks per

day, the mean adjusted TC levels in the ALDH2*1/*1

genotype were lower than those in the ALDH2*2/*2

genotype (180.79/2.5 vs. 195.39/3.5 mg/dl, P�/0.001).

Among men who consumed at least two drinks per day,

the mean adjusted TC levels in the ALDH2*1/*1

genotype were lower than those in the ALDH2*1/*2

genotype (180.89/1.8 vs. 187.29/3.3 mg/dl, P�/0.031). A

comparison of mean adjusted TC levels in the ALDH2

genotype did not show any significant differences in all

the three subgroups (Fig. 1B).

Among women who consumed less than 0.5 drinks

per day, the mean adjusted HDL levels in the

ALDH2*1/*1 genotype were higher than those in the

ALDH2*1/*2 genotype (59.99/0.9 vs. 58.39/0.8 mg/dl,

P�/0.04) (Fig. 2A). Among women who consumed less

than 0.5 drinks per day, the mean adjusted TC levels in

the ALDH2*1/*1 genotype were lower than those in the

other genotypes (193.69/2.2 vs. 199.49/2.1 vs. 201.89/

3.3 mg/dl, P�/0.003 and 0.01, respectively) (Fig. 2B). Acomparison within the ALDH2 genotype showed sig-

nificant differences in the mean adjusted HDL and TC

levels in the ALDH2*1/*2, (HDL in ALDH2*1/*2: B/

0.5 drinks per day: 58.39/0.8 vs. ]/0.5 drinks per day:

63.39/1.6 mg/dl, P B/0.05; TC in ALDH2*1/*1: B/0.5

drinks per day: 199.49/2.1 vs. ]/0.5 drinks per day:

185.29/4.0 mg/dl, P B/0.05) (Fig. 2A and B).

Separate analyses on the interaction between theALDH2 genotypes, amount of alcohol consumption,

and HDL levels after excluding the subjects with HDL

levels more than 80 mg/dl did not cause any significant

differences in the results (data not shown).

4. Discussion

The present study showed that the beneficial effects of

moderate alcohol consumption on HDL levels, namely

an increase in HDL levels, were most prominent among

men with the ALDH2*1/*1 genotype. This finding is instriking contrast to the results of a study on the

interaction between moderate alcohol consumption

and the ADH3 genotype with regard to HDL choles-

Table 2

ALDH2 Genotype and characteristics of the subjects

Variable genotype *1/*1 *1/*2 *2/*2 P

Men

N (Subtotal�/826) 378 (45.8%) 380 (46%) 68 (8.2%)

Age (year) 57.89/15.5 58.69/15.4 53.89/17.7 0.07

Height (cm) 165.89/6.7 165.79/6.7 165.99/6.8 0.98

Weight (kg) 63.19/10.0 61.79/9.5 60.49/8.8 0.03

BMI (m2) 22.99/3.0 22.49/2.9 21.99/2.6 0.008

TC 183.79/35.3 183.69/31.6 188.39/31.7 0.54

HDL 53.99/14.1 52.19/14.1 50.49/13.4 0.07

TG 159.39/118.2 139.29/88.0 127.49/72.9 0.006

HbA1c 5.29/1.0 5.19/0.7 5.19/0.5 0.09

EtOH intake (drinks per day) 2.49/1.7 1.79/2.0 0.19/0.4 B/0.001

Number of drinkers at �/0.5 drinks per week 317 (83.9%) 270 (71.1%) 6 (8.8%) B/0.001

Number of drinkers at ]/two drinks per day 257 (68.0%) 167 (44.0%) 2 (3.0%) B/0.001

Smokers 192 (50.8%) 192 (50.7%) 36 (53.7%) 0.89

Women

N (Subtotal�/1295) 655 (50.6%) 544 (42.0%) 96 (0.7%)

Age (year) 55.29/16.1 55.09/15.6 59.29/13.9 0.04

Height (cm) 152.99/72.0 152.99/68.6 151.99/66.6 0.38

Weight (kg) 52.59/8.2 52.49/8.6 51.99/7.9 0.79

BMI (m2) 22.49/3.1 22.49/3.3 22.59/3.2 0.96

TC 193.89/34.8 197.99/34.7 205.89/32.9 0.003

HDL 61.29/13.7 60.09/14.0 61.09/16.3 0.35

TG 114.39/69.2 114.39/63.0 116.59/66.9 0.95

HbA1c 5.19/0.7 5.09/0.7 5.19/0.6 0.37

EtOH intake (drinks per day) 0.49/0.7 0.29/0.4 0.09/0.2 B/0.001

Number of drinkers at �/0.5 drinks per week 218 (33.3%) 69 (12.7%) 2 (2.1%) B/0.001

Number of drinkers at ]/two drinks per day 49 (7.5%) 13 (2.4%) 1 (1.0%) B/0.001

Smokers 38 (5.8%) 46 (8.5%) 12 (12.5%) 0.028

Y. Nakamura et al. / Atherosclerosis 164 (2002) 171�/177174

Page 5: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

terol levels and the risk of myocardial infarction

conducted by Hines and co-workers [9]. They found

that moderate drinkers who were homozygous for the

slow-oxidising ADH3 allele had higher HDL levels and

a substantially decreased risk of myocardial infarction.

Although we were not able to examine the effect of

alcohol consumption on the risk of myocardial infarc-

tion in the present study, alcohol intake may not protect

against myocardial infarction in subjects with defective

ALDH2 genotypes, as was the case in subjects with

defective ADH3 genotypes.

Association between alcohol consumption and an

increase in HDL levels has been well documented [20].

However, the mechanism by which alcohol increases

HDL is controversial. Studies in alcohol abusers whoconsume on average 80�/177 g/day of alcohol have

suggested that reductions in the plasma cholesteryl ester

transfer protein (CETP) concentration and activity were

responsible for the increase in the HDL levels by alcohol

Fig. 1. Adjusted High-Density Lipoprotein (HDL) and Total Choles-

terol (TC) levels according to the level of alcohol intake and the

ALDH2 genotype in the male subjects. Among men who consumed

less than two drinks per day, the mean adjusted HDL levels were not

significantly different among the three ALDH2 genotypes. However,

among men who consumed at least two drinks per day, the mean

adjusted HDL levels were higher among those with the ALDH2*1/*1

genotype than those with the ALDH2*1/*2 genotype (55.69/0.9 vs.

51.29/0.9 mg/dl, mean9/S.E., P B/0.0001). A comparison of mean

adjusted HDL levels in the ALDH2 genotype showed a significant

difference in the ALDH2*1/*1 genotype (B/two drinks per day: 52.69/

1.1 vs. ]/two drinks per day: 55.69/0.9 mg/dl, P B/0.05) (A). Among

men who consumed less than two drinks per day, the mean adjusted

TC levels in the ALDH2*1/*1 genotype were lower than those in the

ALDH2*2/*2 genotype (180.79/2.5 vs. 195.39/3.5 mg/dl, P�/0.001).

Among men who consumed at least two drinks per day, the mean

adjusted TC levels in the ALDH2*1/*1 genotype were lower than those

in the ALDH2*1/*2 genotype (180.89/1.8 vs. 187.29/3.3 mg/dl, P�/

0.031). A comparison of mean adjusted TC levels in the ALDH2

genotype did not show any significant differences in all the three

subgroups (B). Numeric P values are for the comparison with the

values in men who were in the ALDH2*1/*1 genotype in each category

of alcohol consumption. * denotes P B/0.05 and # denotes not

significant for the comparison between the values in men who

consumed less than two drinks daily versus the values in men who

consumed at least two drinks daily in each ALDH2 genotype.

Fig. 2. Adjusted High-Density Lipoprotein (HDL) and Total Choles-

terol (TC) levels according to the level of alcohol intake and the

ALDH2 genotype in the female subjects. Among women who

consumed less than 0.5 drinks per day, the mean adjusted HDL levels

in the ALDH2*1/*1 genotype were higher than those in the

ALDH2*1/*2 genotype (59.99/0.9 vs. 58.39/0.8 mg/dl, P�/0.04) (A).

Among women who consumed less than 0.5 drinks per day, the mean

adjusted TC levels in the ALDH2*1/*1 genotype were lower than those

in the other genotypes (193.69/2.2 vs. 199.49/2.1 vs. 201.89/3.3 mg/dl,

P�/0.003 and 0.01, respectively) (B). A comparison within the

ALDH2 genotype showed significant differences in the mean adjusted

HDL and TC levels in the ALDH2*1/*2, (HDL in ALDH2*1/*2: B/

0.5 drinks per day: 58.39/0.8 vs. ]/0.5 drinks per day: 63.39/1.6 mg/dl,

P B/0.05; TC in ALDH2*1/*1: B/0.5 drinks per day: 199.49/2.1 vs. ]/

0.5 drinks per day: 185.29/4.0 mg/dl, P B/0.05) (A and B). Numeric P

values are for the comparison with the values in women who were in

the ALDH2*1/*1 genotype in each category of alcohol consumption. *

denotes P B/0.05 and # denotes not significant for the comparison

between the values in women who consumed less than 0.5 drinks daily

versus the values in women who consumed at least 0.5 drinks daily in

each ALDH2 genotype.

Y. Nakamura et al. / Atherosclerosis 164 (2002) 171�/177 175

Page 6: Genetic variation in aldehyde dehydrogenase 2 and the effect of alcohol consumption on cholesterol levels

[21]. However, in another study that examined the effect

of moderate alcohol loading (0.5 g/kg body weight per

day) for 4 weeks after 3 weeks of abstinence, CETP

activity did not change, while HDL levels increased

significantly [22]. This study suggested that an increase

in lipoprotein lipase activity contributed the most to the

alcohol-induced rise in HDL levels. Thus, CETP may

not play an important role in alcohol-induced HDL

elevation in subjects who drink moderately. Whatever it

may be, the mechanism appears to work differently

when the acetaldehyde concentration is elevated in

subjects with the ALDH2-deficient genotype. More

recent studies indicated that alcohol increases HDL by

increasing the transport rate of apolipoprotein A-I

[23,24].

Plasma CETP deficiency was reported as a prevalent

genetic cause of elevated HDL levels in Japanese

subjects. Heterozygote frequencies of two common

mutation, intron 14G(1�/)-to-A (intron 14A) transition

and Asp442-to-Gly (D442G) missense mutation in exon

15, were 2 and 7%, respectively, in the Kanazawa area in

Japan [18]. Furthermore, the missense mutation was

found in Chinese population at a frequency of 4�/6%

[25]. Controversies exist as to the association between

the CETP deficiency and the coronary risk. Some

studies reported an increased risk of MI in the subjects

with CETP gene defects [26], the other studies reported

contrary [17,18,27]. Possible role of CETP deficiencies

was ruled out in the present study by the separate

analyses after excluding subjects with HDL levels more

than 80 mg/dl. However, studying an interaction be-

tween ALDH2 and CETP gene polymorphism is inter-

esting, and should be carried out.

A high percentage of the Asian population has a

defect in the ALDH2 genotype [11,13]. Subjects with a

defective ALDH2 genotype have a higher acetaldehyde

concentration after drinking alcohol, and show flushing

or other symptoms. It has also been suggested ALDH2

deficiency protects against alcoholism and alcohol-

induced liver damage because subjects with ALDH2

deficiency cannot tolerate large amount of alcohol

[17,20]. In our study, however, 44% of men with the

ALDH2*1/*2 genotype reported drinking more than

two drinks daily.

As seen in other studies, the women in the present

study consumed significantly less alcohol and signifi-

cantly fewer had a habit of drinking in comparison with

the men [9]. Therefore, a cut-off level of 0.5 drinks daily

was used for women, instead of two drinks daily, for the

analyses. The findings in the women were slightly

different from those in the men. Nevertheless, the

beneficial effects of alcohol intake were not at all

prominent in the ALDH2-deficient genotypes as in

men and women with ADH3 genotypes in the previous

study [9].

Problems in the self-reporting of drinking may

potentially limit the value of the present study, since

such a system may underestimate or overestimate the

true intake, especially in men who actually consumelarge amounts of alcohol [28,29]. However, the self-

reporting system has been shown to be useful [29], and

studies using such a system have provided important

information [2,9].

None of the subjects in our study had a history or

ECG suggestive of coronary artery disease. Therefore,

there is no direct evidence that alcohol intake protects

against coronary artery disease. These subjects need tobe followed-up for a longer period.

The polymorphism of other alcohol-metabolising

enzymes needs to be investigated. ADH2 polymorph-

isms are known to exist in Japanese. However, it has

been shown that 85% of Japanese possess an ADH

isozyme encoded by the ADH2*2 allele [30]. Therefore,

it is expected that most of the subjects in the present

study possessed the same ADH2 allele.Alcohol intake had quite different effects on HDL

levels in Japanese with a defective ALDH2 genotype in

comparison with a defective ADH3 genotype in Cauca-

sians. Unlike with the ADH3 genotype, alcohol intake

may not protect against MI in subjects with a defective

ALDH2 genotype.

Acknowledgements

This study was supported in part by a contract from

the Japanese Ministry of Education (Grant-in-aid for

Scientific Research on Priority Areas (C): 12204059,

Grant-in-aid for Scientific Research (B): 02454211,

Grant-in-aid for Scientific Research (C): 06670414).

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