Atherosclerosis 203 (2009) 192200

Relationship of the ApoE polymorphism to plasma lipid traits among

South Asians, Chinese, and Europeans living in Canada
Debika Burman a,1 , Andrew Mente a,1 , Robert A. Hegele c,d,1 , Shofiqul Islam a,1 ,
Salim Yusuf a,b,1 , Sonia S. Anand a,b,,1
a Population Health Research Institute, Hamilton Health Sciences and McMaster University, Hamilton, ON, Canada
b Department of Medicine, McMaster University, Hamilton, ON, Canada
c Blackburn Cardiovascular Genetics Laboratory, Robarts Research Institute, London, ON, Canada
d Department of Medicine, University of Western Ontario, London, ON, Canada

Received 16 January 2008; received in revised form 11 June 2008; accepted 11 June 2008
Available online 20 June 2008

Abstract
Background: The prevalence of cardiovascular risk factors and atherosclerosis vary between ethnic groups. We examined the apolipoprotein
E (ApoE) polymorphism, its association with lipid traits and atherosclerosis, and its influence on ethnic variations on lipid traits.
Methods: In a randomly sampled cross-sectional study of 985 South Asian, Chinese, and European Canadians, three common isoforms of
ApoE (E2, E3 and E4), plasma lipid concentrations and atherosclerosis of the carotid artery were measured.
Results: The E2, E3 and E4 allele frequencies were 5.7%, 84.0%, and 10.2%, respectively, and differed significantly between ethnic groups.
There was a strong, stepwise association between ApoE and each plasma lipid trait, except triglycerides. The E4 genotype was associated with
higher low-density lipoprotein cholesterol (p for trend < 0.001), ApoB (p < 0.001), ApoB/ApoA ratio (p < 0.001) and lipoprotein (a) (p < 0.001),
and lower ApoA (p < 0.001) and high-density lipoprotein cholesterol (HDL-C) (p = 0.005). A similar pattern of effects was observed across
all ethnic groups. Ethnicity accounted for modest variation in ApoA, ApoB/ApoA ratio and HDL-C (4.25.0%), whereas ApoE isoforms
explained only a small proportion of variability (0.31.4%). Dietary and lifestyle factors accounted for modest variation in several traits
including HDL-C (5.6% and 5.0%, respectively). Carotid atherosclerosis was lower among individuals with E2 isoform in keeping with the
effect of E2 on lipid levels.
Conclusions: The ApoE isoform is associated with plasma lipoproteins in all ethnic groups, yet explains only a small proportion of the
inter-ethnic variation for most plasma lipoproteins. Additional genetic variants and/or health behaviors likely contribute to ethnic variations
in plasma lipid traits.
2008 Elsevier Ireland Ltd. All rights reserved.

Keywords: Atherosclerosis; Apolipoprotein E; Ethnicity; Genetics; Lipids

1. Introduction

Cardiovascular disease (CVD) prevalence and mortal-

Abbreviations: CVD, cardiovascular disease; ApoE, apolipoprotein E;
LDL-C, low-density lipoprotein; HDL-C, high-density lipoprotein choles-
terol; Apo B, apolipoprotein B; Apo A, apolipoprotein A; Lp(a), lipoprotein
(a); TG, triglycerides; IMT, intimal medial thickness.
Corresponding author at: Population Health Research Institute, Hamilton
General Hospital, 4-EAST Room 441, 237 Barton St. E, Hamilton, ON,
Canada L8L 2X2. Tel.: +1 905 527 4322x44557; fax: +1 905 577 1490.
E-mail address: anands@mcmaster.ca (S.S. Anand).
1 For the SHARE Investigators.
ity rates vary substantially between ethnic groups living in
Canada. The lowest prevalence has been observed among
the Chinese, whereas the highest prevalence occurs among
people who originate from the Indian subcontinent (South
Asians) [1]. The determinants of CVD are likely the same
across ethnic populations although the prevalence, and hence
the population attributable risk of each factor varies between
ethnic groups [2]. It is likely that modifiable CVD risk factors
such as plasma lipids have genetic determinants (Fig. 1).

0021-9150/$ see front matter 2008 Elsevier Ireland Ltd. All rights reserved.
doi:10.1016/j.atherosclerosis.2008.06.007
 D. Burman et al. / Atherosclerosis 203 (2009) 192200
Fig. 1. Carotid atherosclerosis [mean intimal medial thickness (IMT)] by
ApoE allele status overall and among South Asians, Chinese, and Europeans
living in Canada. Bars are S.E.
Apolipoprotein E (ApoE) is a structural component of
most classes of lipoprotein particles, and its functions include
serving as a ligand for cell-surface receptor mediated uptake
of lipoproteins. A common polymorphism of the ApoE gene
(rs429358, rs7412) encoding human ApoE results in three
major isoforms called Apo E2, Apo E3, and Apo E4, which
are coded for by three codominant alleles (E2, E3, and
E4, respectively) [3,4]. Population studies have consistently
demonstrated an association between ApoE and plasma con-
centrations of total cholesterol (TC), low-density lipoprotein
cholesterol (LDL-C) and ApoB [3,511]. In general, the
ApoE alleles (when ordered E2, E3, and E4) are positively
associated with higher concentrations of these plasma lipids
in both European and non-European populations [511].
However previous studies of ApoE and plasma lipids in ethnic
minorities were carried out in the United States, focusing pri-
marily on African-Americans and Hispanic-Americans [12],
or were conducted in ethnic populations living in Asia [6].
Past studies did not assess these associations in South Asian
and Chinese populations exposed to the influences of a West-
ern society.
The objectives of this study are to: (1) compare ApoE
genotype and allele frequency distributions in South Asians,
Chinese, and Europeans; (2) investigate associations between
ApoE and plasma lipid concentrations, overall and among
ethnic groups; (3) determine how much of the between
group differences in lipid concentrations is attributable to the
ApoE polymorphism and dietary or lifestyle factors believed
to influence lipid levels; and (4) examine the association
between ApoE variants and atherosclerosis across ethnic
groups.
2. Materials and methods
2.1. Study population
The study population was comprised of Canadians of
South Asian, Chinese or European origin who participated
193
in the Study of Health Assessment and Risk in Ethnic groups
(SHARE), a cross-sectional prevalence study of CVD risk
factors conducted between 1996 and 1998 [1]. Individuals
were randomly selected based on unique last names from
Toronto, Hamilton, and Edmonton using a previously vali-
dated method [13]. Subjects were aged 3575 years, and had
lived in Canada for 5 years. Ethics approval was obtained
from the McMaster University and University of Western
Ontario Institutional Review Boards, and all participating
institutions. Informed consent was obtained from each sub-
ject. The investigation conforms with the principles outlined
in the Declaration of Helsinki.
2.2. Nutrient analysis
Dietary assessment was conducted using validated
culture-specific, self-administered, quantitative food fre-
quency questionnaires (FFQs) [14]. The FFQ measures the
usual dietary intake over the previous 12 months to estimate
nutrient and energy intake. We determined nutrient content
by analyzing diet records using the Food Processor nutri-
ent analysis software (version 6.11; ESHA, Salem, OR),
which incorporated the 1991 Canada Nutrient File and US
Department of Agriculture databases [14]. We examined the
influence of dietary protein, carbohydrates, saturated fat and
trans fat, since these factors vary by ethnic origin [14] and are
associated with lipid traits [15]. Nutrient values were adjusted
for total energy intake using the residual method [16].
2.3. Laboratory measurements and ultrasonography
Fasting blood samples were obtained from all participants
in the same period. The samples were centrifuged, frozen at
70 C, and transferred on dry ice to a central core laboratory
in Hamilton, Canada for analysis using standard methodol-
ogy. All blood measurements were analyzed blinded to the
participants ethnicity and clinical history. The laboratory
procedures for assessing triglycerides (TG), HDL-C, LDL-
C and lipoprotein (a) (Lp(a)) have been described previously
[1]. ApoB was measured using the rate nephelometic method
on the Beckman Array Protein System, using reagents sup-
plied by Beckman Instruments (Brea, CA, USA). ApoA
was measured using Roche Immunoturbidimetric assay per-
formed on the Hitachi 917 (Indianapolis, IN, USA). The
laboratory maintains traceability with the National Refer-
ence System for Cholesterol by direct comparison with the
cholesterol reference method using fresh serum, and has
demonstrated the ability to meet National Cholesterol Educa-
tion Program performance criteria. We measured extracranial
carotid artery atherosclerosis using B-mode ultrasound (Acu-
son, Mountain View, CA, USA), as described previously [1].
2.4. ApoE genotype determination
Genomic DNA was extracted from whole blood using
the Puregene DNA purification kit (Gentra Systems, Min-
194 D. Burman et al. / Atherosclerosis 203 (2009) 192200

neapolis MN, USA). ApoE genotypes were determined using
the restriction isotyping method of Hixson and Vernier [17].
Genotyping was performed with blinding to subject identity
and phenotype. Sequence-proven controls were run with each
reaction. A random 5% of samples were genotyped again on
another day; no discrepancies were observed.
2.5. Statistical analysis
All statistical analyses were performed using SAS soft-
ware version 9.1. Allele frequencies were determined using
the gene counting method. The chi-square goodness-of-fit
test was used to examine whether observed genotype fre-
quencies differed from those expected by HardyWeinberg
equilibrium. The chi-square contingency test was used to
compare allele and genotype frequency distributions across
ethnic groups and between sexes.
Plasma Lp(a) and TG values were logarithmically trans-
formed before all statistical analyses to improve the normality
of the distributions. The ApoE genotype groups were as fol-
lows: E2 subjects had either the E2/2 or E2/3 genotype.
E3 subjects were E3/3 homozygotes. E4 subjects had
either the E4/3 or E4/4 genotype. As in many other studies
of this nature, subjects with the E4/2 genotype (n = 6) were
excluded from the subsequent analyses since they could not
be simply classified into any single allele group. Associations
of lipid traits and atherosclerosis with ApoE genotype within
and between ethnic groups were determined with analysis
of covariance (ANCOVA) models with p-values for trend,
adjusting for age, sex, center, ethnicity, smoking, alcohol
intake, physical activity, and dietary factors. The ANCOVA
models also provided estimates of (1) the effect size of the
E3 and E4 alleles compared with E2, (2) the effect of South
Asian and Chinese ethnicity coded as categorical dummy
variables compared with European ethnicity, and (3) the inter-
action between ApoE genotype and South Asian or Chinese
ethnicity with Europeans as the reference group [18].
Linear regression was used to estimate the proportion of
variance in plasma traits attributable to ethnicity, ApoE allele
status, dietary factors, and lifestyle. Five models were con-
structed: Model 1 included only age, sex and center; Model
2 included the variables in Model 1 plus ethnicity; Model 3
included Model 2 plus ApoE allele status; Model 4 included
Model 3 plus dietary factors; Model 5 included Model 4
plus lifestyle factors (smoking, alcohol intake, and physi-
cal activity). We examined the change in R2 progressively
from Model 1 to Model 5 using the coefficient of partial
determination (partial R2 ) [18]. Thus, comparing Model 1 to
Model 2, the partial R2 estimates the proportion of variance
in plasma traits attributable to ethnicity, adjusting for age,
sex and center. Similarly, a comparison of Model 2 to Model
3 reflects the proportion of between-ethnic group variation
attributable to ApoE isoforms, while controlling for covari-
ates. Comparisons of Model 3 vs. Model 4, and Model 4 vs.
Model 5 (the full model) were conducted in the same fashion.
The corresponding p-values for the partial R2 estimates were
determined using the multiple partial F-test [18].
3. Results
Of the 985 subjects who were randomly sampled, 17 could
not be genotyped, leaving a final sample with complete data
for 968 subjects, including 336 South Asians, 311 Chinese,
and 321 Europeans.
The characteristics of participants are displayed in Table 1.
The mean age of the overall population was 49.5 years,
and men and women are equally represented. Chinese and
European subjects had similar plasma lipid concentrations
(all p = N.S.). South Asian subjects had significantly higher

Table 1
Distribution of risk factors among South Asians, Chinese and Europeans living in Canada [mean (S.E.)a ]
Variableb Overall (n = 985) South Asians Chinese (CH) Europeans Overall p SA vs. CH SA vs. EU CH vs. EU
(SA) (n = 342) (n = 317) (EU) (n = 326)
Age (years) 49.5 (0.3) 49.4 (0.5) 47.8 (0.5) 51.3 (0.6) <0.001 0.112 0.041 <0.001
Men [n (%)] 506 (51.4) 187 (54.7) 162 (51.1) 157 (48.2) 0.240
LDL-C (mmol/L) 3.21 (0.03) 3.30 (0.04) 3.14 (0.05) 3.18 (0.04) 0.036 0.014 0.061 0.555
ApoB (g/L) 1.02 (0.01) 1.07 (0.01) 0.99 (0.01) 1.00 (0.01) <0.001 <0.001 <0.001 0.574
ApoA (g/L) 1.37 (0.01) 1.30 (0.01) 1.41 (0.01) 1.41 (0.01) <0.001 <0.001 <0.001 0.984
ApoB/ApoA 0.78 (0.01) 0.87 (0.02) 0.73 (0.02) 0.74 (0.02) <0.001 <0.001 <0.001 0.658
HDL-C (mmol/L) 1.15 (0.01) 1.05 (0.02) 1.20 (0.02) 1.20 (0.02) <0.001 <0.001 <0.001 0.959
Lp(a) (g/L)c 5.00 (0.04) 5.24 (0.06) 4.89 (0.07) 4.88 (0.07) <0.001 <0.001 <0.001 0.912
TG (mmol/L)c 0.30 (0.01) 0.40 (0.02) 0.24 (0.02) 0.25 (0.02) <0.001 <0.001 <0.001 0.804
BMI (kg/m2 ) 25.9 (0.1) 26.2 (0.2) 23.9 (0.2) 27.5 (0.2) <0.001 <0.001 <0.001 <0.001
WHR 0.87 (0.002) 0.88 (0.004) 0.86 (0.004) 0.86 (0.004) <0.001 <0.001 <0.001 0.185
Alcohol intake (times/week) 1.23 (0.06) 0.80 (0.10) 0.69 (0.10) 2.20 (0.10) <0.001 0.44 <0.001 <0.001
Physical activity (h/week)d 7.51 (0.05) 7.30 (0.08) 7.15 (0.09) 8.09 (0.08) <0.001 0.19 <0.001 <0.001
a Concentrations adjusted for age, sex, and center.
b LDL-C indicates low-density lipoprotein cholesterol; ApoB, apolipoprotein B; HDL-C, high-density lipoprotein cholesterol; Lp(a), lipoprotein (a); TG,
triglycerides; BMI, body mass index; WHR waisthip ratio.
c Natural log.
d Physical activity was defined as the number of h/week spent exercising or playing sports during the past year.
 D. Burman et al. / Atherosclerosis 203 (2009) 192200 195

Table 2a
ApoE allele frequency distribution among South Asians, Chinese, and Europeans living in Canada
Allele (%) Total sample South Asians Chinese Europeans Overall pa SA vs. CHb SA vs. EUb CH vs. EUb
N 968 336 311 321
E2 5.7 1.9 8.4 7.2 <0.001 <0.001 <0.001 0.54
E3 84.0 87.6 86.0 78.3 0.03 0.11 0.009 0.32
E4 10.2 10.4 5.6 14.5 <0.001 0.001 0.07 <0.001
ApoE, apolipoprotein E; SA, South Asian; CH, Chinese; EU, European.
a Overall p, 2 2 d.f.
b Pairwise p, 2 1 d.f.

plasma concentrations of LDL-C compared to the Chinese, trend <0.05). For HDL-C and TG, the associations with ApoE
and significantly higher plasma ApoB, ApoB/ApoA ratio, alleles were not consistent among ethnic groups. The asso-
Lp(a) and TG concentrations and significantly lower ApoA ciations between ApoE and each adjusted plasma lipid trait
and HDL-C compared to both Chinese and Europeans. South are shown in Appendix A.
Asian and Chinese subjects consumed significantly less alco- In a multivariate regression analysis that included ApoE
hol and were less physically active than Europeans (all genotype, ethnicity and covariates, the E4 isoform was iden-
p < 0.05). tified as a significant determinant for each plasma trait, except
ApoE allele frequency distributions differed significantly TG (p = 0.44) (Table 3). Compared to E2, E4 was asso-
between ethnic groups (Table 2a). Observed genotype fre- ciated with higher LDL-C (p < 0.001), ApoB (p < 0.001),
quencies (Table 2b) did not deviate from HardyWeinberg ApoB/ApoA ratio (p < 0.001) and Lp(a) (p = 0.006), and with
equilibrium in South Asians (2 = 2.74, p = 0.80), Chinese lower ApoA (p = 0.006) and HDL-C (p = 0.044). E3 was a
(2 = 5.92, p = 0.46), or Europeans (2 = 4.14, p = 0.62). significant determinant of plasma LDL-C (p < 0.001), ApoB
Allele and genotype frequency distributions were similar (p < 0.001), ApoB/ApoA ratio (p = 0.001), Lp(a) (p < 0.001)
between sexes (Pearson chi-square p = 0.31 for genotype, and and TG (p = 0.002).
p = 0.26 for allele frequency; data not shown). As shown in Table 3, while adjusting for ApoE geno-
The E3 allele was the most common in each ethnic group type and covariates, South Asians had a higher ApoB
(87.6% in South Asians, 86.0% in Chinese, and 78.3% in (p = 0.002), ApoB/ApoA ratio (p < 0.001), Lp(a) (p = 0.002),
Europeans). South Asians had a lower frequency of the E2 and TG (p < 0.001), and a lower ApoA (p < 0.001) and
allele (1.9%) compared to Chinese and Europeans (8.4% and HDL-C (p < 0.001) compared to Europeans. The interaction
7.2%, respectively, both p < 0.001). Chinese subjects had a between ApoE and South Asian or Chinese ethnicity was
lower frequency of the E4 allele (5.6%) compared to South non-significant for each plasma trait except HDL-C (p = 0.03
Asians and Europeans (10.4% and 14.5%, respectively, both for South Asians, p = 0.004 for Chinese) and TG (p = 0.01 and
p < 0.001). p < 0.001, respectively) after adjustment for lifestyle factors.
There was a strong, stepwise association between the As indicated by the partial R2 for Model 1 vs. Model
ApoE polymorphism and each of the adjusted plasma lipid 2 (shown in Table 4), ethnicity accounted for 0.4% of the
traits, except for TG. The E4 genotype was associated with variance in LDL, 1.9% in ApoB, 5.0% in ApoA, 4.2% in
a significantly higher LDL-C, ApoB, ApoB/ApoA ratio and ApoB/ApoA ratio, 4.5% in HDL, 2.1% in TG, and 1.7%
Lp(a), and lower ApoA and HDL-C. Plasma TG concentra- in Lp(a). The addition of ApoE polymorphism (Model 2 vs.
tion was highest among E2 and E4 subjects (p < 0.01). Model 3) accounted for an additional 3.2% of the variation in
Stepwise trends were also observed for LDL-C, ApoB and LDL-C, 2.5% in ApoB, 0.6% in ApoA, 1.4% in ApoB/ApoA
ApoB/ApoA ratio separately in all three ethnic subgroups, ratio, 0.3% in HDL-C, 1.3% in TG, and 1.7% in Lp(a),
and for ApoA in South Asians and Chinese (all p-values for over and above the contribution of ethnicity. Dietary factors
accounted for 4.0% of the variation in ApoB and 5.6% in
Table 2b HDL-C, but 0.12.6% of the variation in other traits. Lifestyle
ApoE genotype frequency distribution among South Asian, Chinese, and factors explained 5.0% of the variation in HDL-C and 6.0%
Europeans living in Canada
in ApoA, but only 0.11.4% of the variation in other mea-
Genotype Total population South Asians Chinese Europeans sures (Table 4). Alcohol intake accounted for 3.6% of the
[n (%)]
variation in HDL-C and 4.4% in ApoA, while physical activ-
N 968 336 311 321 ity explained 1.5% of the variation in HDL-C and 1.7% in
E2/2 0 0 0 0
E3/2 104 (10.7) 11 (3.3) 51 (16.4) 42 (13.1)
ApoA. Smoking explained negligible variation in these traits
E4/2 7 (0.7) 2 (0.6) 1 (0.3) 4 (1.2) (<0.1%) (data not shown).
E3/3 673 (69.5) 256 (76.2) 225 (72.3) 192 (59.8) The ApoE gene polymorphism was not significantly
E4/3 177 (18.3) 66 (19.6) 34 (10.9) 77 (24.0) associated with atherosclerosis, neither overall (p = 0.31 for
E4/4 7 (0.7) 1 (0.3) 0 6 (1.9) trend) nor within ethnic subgroups. In each group, however,
ApoE indicates apolipoprotein E. there was a non-significant trend for less atherosclerosis in
 196
Table 3
Effect of ethnicity and ApoE allele status on plasma traits
Multivariate predictors LDL-C ApoB ApoA ApoB/ApoA HDL-C

(S.E.) p (S.E.) p (S.E.) p (S.E.) p (S.E.) p

Ethnicity
European Reference Reference Reference Reference Reference
South Asian 0.07 (0.06) 0.26 0.06 (0.02) 0.002 0.11 (0.02) <0.001 0.12 (0.02) <0.001 0.15 (0.03) <0.001
Chinese 0.002 (0.06) 0.97 0.001 (0.02) 0.95 0.004 (0.02) 0.85 0.002 (0.02) 0.926 0.002 (0.03) 0.93
ApoE statusb <0.001 <0.001 0.002 <0.001 0.009
E2 Reference Reference Reference Reference Reference
E3/3 0.43 (0.08) <0.001 0.10 (0.02) 0.100 0.04 (0.02) 0.07 0.10 (0.03) 0.001 0.02 (0.03) 0.46

D. Burman et al. / Atherosclerosis 203 (2009) 192200

E4 0.52 (0.10) <0.001 0.15 (0.03) <0.001 0.08 (0.03) 0.01 0.14 (0.04) <0.001 0.08 (0.04) 0.04
South Asian ApoE interaction 0.50 0.37 0.11 0.36 0.03
Chinese ApoE interaction 0.77 0.12 0.09 0.16 0.004
Age 0.01 (0.003) <0.001 0.01 (0.000) <0.001 0.004 (0.000) <0.001 0.001 (0.001) 0.314 0.003 (0.001) 0.01
Male gender 0.36 (0.05) <0.001 0.14 (0.02) <0.001 0.19 (0.02) <0.001 0.21 (0.001) <0.001 0.28 (0.02) <0.001

Multivariate predictors Lp(a)a TGa

(S.E.) p (S.E.) p
Ethnicity
European Reference Reference
South Asian 0.27 (0.09) 0.002 0.22 (0.04) <0.001
Chinese 0.02 (0.09) 0.87 0.05 (0.04) 0.30
ApoE statusb <0.001 0.001
E2 Reference Reference
E3/3 0.47 (0.11) <0.001 0.17 (0.06) 0.002
E4 0.36 (0.13) 0.006 0.05 (0.07) 0.44
South Asian ApoE interaction 0.77 0.01
Chinese ApoE interaction 0.66 <0.001
Age 0.002 (0.003) 0.64 0.01 (0.002) <0.001
Male gender 0.21 (0.07) 0.003 0.27 (0.03) <0.001
LDL-C, low-density lipoprotein cholesterol; ApoB, apolipoprotein B; HDL-C, high-density lipoprotein cholesterol; Lp(a), lipoprotein (a); TG, triglycerides; CHO, carbohydrates; ApoA, apolipoprotein A;
indicates unstandardized regression coefficient. Covariates include age, sex, and center; Adjustment for additional covariates (dietary predictors, smoking, alcohol intake, and physical activity) produced similar
results. c Nutrient intake levels have been log-transformed, adjusted for total caloric-intake, and exponentiated for ease of interpretation.
a Natural log.
b E2 indicates subjects with E3/2 or E2/2 genotypes and E4 indicates subjects with E4/3 or E4/4 genotypes. E2 homozygotes were the referent for each group.
 D. Burman et al. / Atherosclerosis 203 (2009) 192200 197

Table 4
The coefficients of partial determination (partial R2 ) indicating the proportion of variance in plasma lipid concentrations attributable to ethnicity, ApoE
genotype, and dietary factors
Predictor variable LDL-C ApoB ApoA ApoB/ApoA HDL-C

Partial R2 p Partial R2 p Partial R2 p Partial R2 p Partial R2 p

1b
Ethnic origin (Model vs. 2c ) 0.004 0.02 0.019 <0.001 0.050 <0.001 0.042 <0.001 0.045 <0.001
ApoE genotype (Model 2c vs. 3d ) 0.032 <0.001 0.025 <0.001 0.006 0.006 0.014 <0.001 0.003 0.02
Dietary factors (Model 3d vs. 4e ) 0.001 0.02 0.040 <0.001 0.025 0.002 0.026 <0.001 0.056 <0.001
Lifestyle factors (Model 4e vs. 5f ) 0.005 0.051 0.007 0.03 0.060 <0.001 0.014 <0.001 0.050 <0.001

Predictor variable Lp(a)a TGa

Partial R2 p Partial R2 p

Ethnic origin (Model 1b vs. 2c ) 0.017 <0.001 0.021 <0.001

ApoE genotype (Model 2c vs. 3d ) 0.017 <0.001 0.013 <0.001
Dietary factors (Model 3d vs. 4e ) 0.005 0.070 0.088 <0.001
Lifestyle factors (Model 4e vs. 5f ) 0.001 0.32 0.011 0.004
LDL-C indicates low-density lipoprotein cholesterol; ApoB, apolipoprotein B; HDL-C, high-density lipoprotein cholesterol; Lp(a), lipoprotein (a); TG,
triglycerides; CHO, carbohydrates. R2 indicates adjusted coefficient of determination.
a Natural log.
b Model 1 included the variables age, sex, and center.
c Model 2 included Model 1 plus ethnicity.
d Model 3 included Model 2 plus ApoE.
e Model 4 included Model 3 plus energy adjusted nutrient intake levels of saturated fat, trans fat, protein, and carbohydrates.
f Model 5 included Model 4 plus smoking, alcohol intake, and physical activity.
The coefficient of partial determination (partial R2 ) shows the marginal reduction in the variation of Y associated with a new variable X (e.g., ethnicity)

which is added to a simpler model that includes other predictors (e.g., age, sex, and center). For example, a partial R2 value of 0.05 for ApoA comparing Model
1 vs. Model 2 means that adding ethnicity to the model which already includes age, sex, and center reduces the variation in ApoA by 5%. Corresponding p
values for the partial R2 estimates were determined using the multiple partial F-test.

subjects with the E2 allele. An assessment of adjusted plasma
ApoB/ApoA ratio as a predictor of atherosclerosis revealed
a positive and significant association ( = 0.053, p = 0.001).
4. Discussion
To our knowledge this is the first study to examine the asso-
ciation between the ApoE polymorphism and plasma lipid
traits in South Asian and Chinese populations exposed to the
influences of a Western society. Previous studies of ApoE
and plasma lipids in ethnic minorities were conducted in the
United States, focusing primarily on African-Americans and
Hispanic-Americans [12], or were carried out in ethnic popu-
lations living in Asia [6]. In this study, participants from three
well-defined ethnic groups were randomly selected from
three Canadian cities, and detailed information was obtained
on lifestyle factors including dietary intake, smoking his-
tory, physical activity, and alcohol consumption. Sampling
in SHARE was random and participants in each ethnic group
were proportionately recruited [1], making the associations
in this study unlikely to be the result of sampling bias.
The frequency of ApoE alleles in our subjects differed
significantly across ethnic groups, a finding compatible with
that of previous studies [7,19,20]. We observed in each eth-
nic group and overall, the E3 allele was the most frequent,
with E4 less frequent, and E2 least frequent. This finding
is ubiquitous for human populations [3,4,7,19,20]. We also
noted that the E2 allele frequency was highest in Chinese and
lowest in South Asians, whereas the E4 allele frequency was
lowest in Chinese, intermediate in South Asians, and highest
in Europeans. Similar contrasts have been observed between
Chinese and South Asians in Malaysia [6,21].
Our study shows that while frequencies of ApoE alleles
differ significantly between ethnic groups, the associations
between ApoE and LDL-C, ApoB, ApoA, ApoB/ApoA ratio,
and Lp(a) are consistent across ethnic groups. The ascending
E2 < E3 < E4 gradient for these plasma traits across different
ethnic subgroups, despite varying ApoE allele frequencies,
is consistent with the premise highlighted in a recent meta-
analysis by Ioannidis et al. [22], which showed that while the
frequencies of genetic variants across racial groups are often
heterogeneous, the effects of the variants are more likely to be
homogenous [22]. These associations previously have been
reported predominantly in Caucasians [3,5,8,9,11,23,24] as
well as in Asian populations residing overseas [6]. Our find-
ings suggest that the association between ApoE and plasma
lipid traits is universal across different ethnic subgroups,
irrespective of lifestyle influences that might be related to
increased Westernization.
This study also demonstrates that the ApoE polymor-
phism accounts for only a small proportion of the inter-ethnic
group differences for most plasma lipid traits. While ethnic-
ity explains a modest amount of inter-individual variability
in ApoA, ApoB/ApoA ratio, and HDL-C, the contribution of
ApoE genotype above and beyond the influence of ethnicity
is relatively small. Thus, genetic variation at this locus alone
likely cannot account for the differences in mean plasma
198 D. Burman et al. / Atherosclerosis 203 (2009) 192200
lipid concentrations observed between ethnic groups. Dietary
and lifestyle factors have also been incriminated previously
[25,26], but prior studies did not assess their effect on plasma
lipid traits beyond the influence of ethnic origin. In our study,
dietary factors contributed modestly to the prediction of sev-
eral plasma traits including HDL-C and ApoB, while lifestyle
factors (predominantly alcohol intake and physical activ-
ity) modestly predicted HDL-C and ApoA. We previously
reported the effect of carbohydrate intake on HDL cholesterol
[27] and individuals who consumed the greatest proportion of
energy from carbohydrates had the lowest HDL cholesterol
independent of other HDL correlates. The beneficial effect
of alcohol intake on lipid profile has been shown [28] and
E2 and E3 carriers improve plasma lipoprotein/lipid profiles
more with exercise training than Apo E4 individuals [29].
Thus ethnic variations in health behavior appear to influence
plasma lipids and should be considered along with genetic
variants in the explanation of inter-ethnic variations in plasma
lipids [30,31].
The associations between the ApoE polymorphism and
plasma concentrations of Lp(a), ApoA, HDL-C, and TG
have been inconsistent in the literature, with results that
range from no association [3,9,23,32,33] to associations that
vary by sex [10]. Unlike our study, de Knijff et al. [34]
reported that the E4 allele, rather than the E3 allele, was
associated with higher mean Lp(a). The biology underlying
these associations is complex, however, since the majority of
genetically determined variance in Lp(a) results from varia-
tion at the ApoA locus on chromosome 6 [35]. Conversely,
our observation that ApoA is highest in E2 subjects sup-
ports previous findings by Kahraman et al. [36], although
others reported similar or lower ApoA concentration in E2
subjects [32,33]. Similarly, in Chinese and Asian Indian pop-
ulations residing in Asia, the E2 isoform was associated with
higher HDL-C [6], as in our study, although others did not
find an association in Japanese [37,38]. The present study
extends these observations to Asians living in North Amer-
ica who are likely exposed to different dietary and lifestyle
influences.
Our study is supportive of an association between ApoE
isoform and mean IMT in all ethnic groups. While our data
shows a non-significant trend for lower mean IMT in the sub-
jects with the E2 isoform, the small number of subjects with
this genotype may partly explain the null findings. Similarly,
in their recent meta-analysis of 22 studies, Paternoster et al.
reported a significant protective association between the E2
allele and carotid IMT [39]. However, these studies mostly
focused either on Caucasians or evaluated a small number of
subjects in Eastern Asia, but did not assess South Asians. Our
study also shows that the ApoE isoform is significantly asso-
ciated with ApoB/ApoA ratio, which itself is significantly
associated with carotid atherosclerosis. Similarly Bennet et
al. showed that the E2 genotype is associated with a 20%
reduction in coronary heart disease compared to the E3 geno-
type, an effect that is likely mediated by the influence of E2
on plasma lipids [11].
The main strengths of this study are the recruitment of
a random population, inclusion of three ethnic groups using
similar criteria, and use of standard methods of phenotyping
and genotyping. The inclusion criteria that both parents and
grandparents must have originated from South Asian, Chi-
nese or European nations [1] likely ensured that admixture
in the sample was negligible [40].
The study has some limitations. The inclusion of partici-
pants receiving medication for CVD, diabetes, hypertension
or hypercholesterolemia can alter physiologic lipid levels
and confound the lipid associations. However, in a sensi-
tivity analysis that excluded these patients (n = 352), the
associations between ApoE isoforms and plasma lipid traits
among ethnic groups were unaltered (data not shown).
Our analyses involved testing for associations with multi-
ple outcome measures (plasma traits) across several ethnic
subgroups, which increases the likelihood of detecting
significant associations where none exist (type 1 error).
However, since strong associations were observed for mul-
tiple plasma traits across ethnic subgroups despite the
small number of E2 subjects (n = 55), and given that the
effects of the ApoE isoforms were consistent with previ-
ous observations, it is unlikely that our findings were due
to chance.
In conclusion, ApoE isoform is significantly associated
with plasma lipoproteins in all ethnic groups, but explains
only a small proportion of the inter-ethnic variation for
most plasma lipoproteins. Additional genetic variants and/or
health behaviors likely contribute to ethnic variations in
plasma lipid traits.
Conict of interest
D. Burman, A. Mente, S. Islam, S. Yusuf, and S. Anand
declare no conflict of interest. R. Hegele has spoken on the
genetic determinants of dyslipidemia and received payment
from those funding the events (Merck Schering, Pfizer, and
Oryx) in amounts less than $10,000.
Acknowledgements
D. Burman contributed to the analysis and interpretation
of the data, and the drafting of the manuscript. A. Mente con-
tributed to the analysis, interpretation of the data, and drafting
of the manuscript. R. A. Hegele performed DNA extraction
and genotyping, and contributed to interpretation and drafting
of the manuscript. S. Islam was responsible for database man-
agement and provided comments on the statistical analyses.
S. Yusuf contributed to the design of the study, interpreta-
tion, and drafting of the manuscript. S. S. Anand contributed
to the design of the study, interpretation, and drafting of the
manuscript.
We acknowledge Dr. Koon K. Teo and Dr. Vlad Vuksan for
their efforts in recruiting participants in SHARE for Edmon-
 D. Burman et al. / Atherosclerosis 203 (2009) 192200 199

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