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The influence of the adiponectin gene on adiponectin concentrations and
parameters of metabolic syndrome in non-diabetic Korean women
Yangsoo Jang
a,b,c,1
, Jey Sook Chae
c,d,1
, Soo Jeong Koh
c,d
, Yae Jung Hyun
c,d
, Ji Young Kim
c,d
,
Yeo Jin Jeong
d,e
, Sungha Park
b
, Chul-Min Ahn
b
, Jong Ho Lee
c,d,
⁎
a
Cardiovascular Genome Center, Cardiovascular hospital, Severance hospital, Seoul, Korea
b
Cardiology Division, Yonsei University College of Medicine, Yonsei Cardiovascular Center, Severance hospital, Seoul, Korea
c
Yonsei University Research Institute of Science for Aging, Yonsei Univ., Seoul, Korea
d
National Research Laboratory of Clinical Nutrigenetics and Nutrigenomics, Yonsei Univ., Seoul, Korea
e
Department of Food and Nutrition, Brain Korea 21 Project, Yonsei University College of Human Ecology, Yonsei Univ., Seoul, Korea
Received 21 December 2007; received in revised form 11 February 2008; accepted 11 February 2008
Available online 16 February 2008
Abstract
Background: Concentrations of adiponectin, the protein product of the adipoctye C1q and collagen-domain-containing (ADIPOQ) gene are
associated with type 2 diabetes and coronary artery disease. We investigate the association of single-nucleotide polymorphisms (SNPs) in the
ADIPOQ gene with adiponectin concentrations, and to parameters of metabolic syndrome.
Methods: 867 unrelated, non-diabetic Korean women, 20 to 69 y, were genotyped for 8 SNPs in the ADIPOQ gene (- 11391GNA, - 11377C NG,
H241P, Y111H, G90S, R221S, 45TNG, 276G NT). Adiponectin, a homeostasis model assessment of insulin resistance (HOMA-IR), and
metabolic parameters were measured.
Results: Carriers of genotype T/T at position 276 had significantly higher adiponectin concentrations than G/G carriers (P=0.005). Homozygous
carriers of the TG haplotype (i.e., individuals who were T/T at 45 and G/G at 276) and heterozygous carriers of the TG haplotype (TG/X) had
lower adiponectin concentrations than non-TG carriers (P b0.001). Significant associations between SNP at 276 and serum concentrations of
triglyceride (P = 0.013), insulin (P = 0.013) and HOMA-IR (P = 0.012) were found. The 45-276 haplotypes had associations identical to the
276G NT SNP. In subgroup analysis, subjects carrying the TG haplotype had significantly lower adiponectin concentrations than non-TG carriers
in both normal weight (P b0.001) and overweight-obese (P= 0.009) subgroups. The association of the TG haplotype with increasing insulin
concentrations was significant among overweight-obese subjects (P = 0.004), but was not significant among normal weight subjects. A similar
association was found between the 45-276 haplotype and HOMA-IR.
Conclusion: There is a strong association of the adiponectin SNP276 genotypes and the adiponectin 45-276 haplotypes with circulating
adiponectin concentrations in non-diabetic Korean women. In addition, this haplotype is associated with increased insulin concentrations and
insulin resistance index only in overweight-obese individuals.
© 2008 Elsevier B.V. All rights reserved.
Keywords: Adipocyte C1q and collagen domain-containing (ADIPOQ); Adiponectin; Single nucleotide polymorphism (SNP); Insulin; Homeostasis model
assessment of insulin resistance (HOMA-IR)
1. Introduction
Persons with low concentrations of adiponectin, the protein
product of the adipoctye C1q and collagen-domain-containing
(ADIPOQ) gene, are more likely to develop type 2 diabetes
and coronary artery disease [1–3]. Single-nucleotide polymor-
phisms (SNPs) as well as haplotypes at the ADIPOQ locus
are reported to be associated with circulating adiponectin
A
vailable online at www.sciencedirect.com
Clinica Chimica Acta 391 (2008) 85–90
www.elsevier.com/locate/clinchim
⁎Corresponding author. Department of Food & Nutrition, Yonsei Univ., 134
Shinchon-Dong, Sudaemun-Gu, Seoul, 120-749, Korea. Tel.: +82 2 2123 3122;
fax: +82 2 364 9605.
E-mail address: jhleeb@yonsei.ac.kr (J.H. Lee).
1
These two authors equally contributed to this work.
0009-8981/$ - see front matter © 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.cca.2008.02.011
concentrations, but the results are controversial. Menzaghi et al.
[4] previously described an association between 2 SNPs in
the ADIPOQ gene (i.e., SNPs 45T NG, 276G NT, considered
together as “TG”haplotype) and several features of insulin
resistance in non-diabetic Caucasians from Italy, including low
serum adiponectin concentrations. Nakatani et al. [5] also
reported that TG haplotype had association with obesity and
insulin sensitivity in non-diabetic Japanese subjects. Vasseur
et al. [6] have reported that a different ADIPOQ haplotype,
defined by SNPs -11391G NA and -11377C NG in the promoter
region (“GG”haplotype), was associated with low adiponectin
concentrations in French Caucasians, although no associa-
tion with insulin-resistance indexes and type 2 diabetes was
observed. The reasons for these partially discrepant results are
unknown, and may result from the different genetic background
of the study populations.
2. Methods
2.1. Study subjects
The study population included 867 unrelated, non-diabetic Korean women,
aged 20 to 69 y, who were selected from participants in a nutritional genomic
study conducted by the National Research Laboratory of Clinical Nutrigenetics
and Nutrigenomics (Program #2005-01572) at Yonsei University. Subjects were
screened by medical history questionnaire, physical examination, and fasting
blood profiles. All women were sedentary (b2 h/wk of structured exercise),
nonsmoker, and non to low alcohol consumers. Subjects were excluded from the
study if they displayed clinical evidence of the following: 1) cardiovascular
disease (CVD), peripheral vascular disease, or stroke; 2) diabetes; 3) orthopedic
limitations; 4) body weight fluctuation ≥±2 kg in the past year; 5) thyroid or
pituitary disease; 6) infection by medical questionnaire examination and com-
plete blood count; 7) acute or chronic inflammatory disease; and 8) medication
that could affect metabolism. Among postmenopausal women, none were
receiving hormone replacement therapy with estrogen.
Before participation, the purpose of the study was carefully explained to all
participants, and their informed consent was obtained. The protocol was
approved by the Institutional Review Board of Yonsei University and the study
was carried out in accordance with the Helsinki Declaration.
2.2. Anthropometric parameters, blood pressure measurements, and
blood collection
Body weight and height were measured unclothed and without shoes in the
morning. Waist circumference was measured with paper tape horizontally at the
umbilicus in the standing position after normal expiration. Blood pressure was
read from the left arm of seated subjects with an automatic blood pressure
monitor (TM-2654, A&D, Tokyo, Japan) after 20 min of rest. The average of
three measurements was recorded for each subject. Venous blood specimens
were collected in EDTA-treated and plain tubes after a 12-hr fast. The tubes were
placed on ice until they arrived at the laboratory (within 1-3 h) and were stored at
-70°C until analysis.
2.3. Genotyping
Genomic DNA was extracted from 5 ml of whole blood using a DNA isolation
kit (WIZARD
R
Genomic DNA purification kit, Promega Corp., Madison, WI)
according to the manufacturer's protocol. Eight SNPs (- 11391GNA and
-11377CNG at promoter region, 45TNG at exon 2, 276GNTatintron 2,and
H241P, Y111H, G90S and R221S at exon 3) were examined for analysis of the
association of these SNPs with plasma adiponectin and features of metabolic
syndrome. Each genotyping was performed by SNP-IT™assays using single
primer extension technology (SNPstream 25 K™System, Orchid Biosystems,
Princeton NJ). The DNA fragments were visualized by UV illumination using
Image Analyzer (AlphaImager™1220, Alpha Innotech Corp., San Leandro CA)
respectively. pUC19 DNA/Msp I (Hpa II) Marker (MBI Fermentas, Vilnius,
Lithuania) served as a control standard.
2.4. Serum lipid profile
Blood fasting serum concentrations of total cholesterol and triglycerides
were measured using commercially available kits on a Hitachi 7150 analyzer
(Hitachi Ltd. Tokyo, Japan). After using dextran sulfate magnesium to
precipitate serum chylomicron, low-density lipoprotein (LDL), and high-density
lipoprotein (HDL) cholesterol from the supernatant was measured by an
enzymatic method. LDL cholesterol was indirectly estimated in subjects with
serum triglyceride concentrations b4.52 mol/l (400 mg/ml) using the Friedewald
formula. In subjects with serum triglyceride concentrations ≥4.52 mol/l, LDL
cholesterol was measured by an enzymatic method on a Hitachi analyzer
directly. Serum apolipoprotein A-I and B were determined by turbidometry at
340 nm using a specific anti-serum (Roche, Switzerland).
2.5. Glucose, insulin, and HOMA-IR
Fasting glucose was measured by the glucose oxidase method using a Beckman
Glucose Analyzer (Beckman Instruments, Irvine, CA). Insulin was measured by
radio-immunoassays with kits from Immuno-Nucleo Corp. (Stillwater, MN). Insulin
resistance (IR) was calculated with the homeostasis model assessment (HOMA)
using the following equation: HOMA-IR={fasting insulin (μIU/ml) × fasting
glucose (mmol/l)}/22.5 [7].
2.6. Plasma adiponectin concentrations
Plasma adiponectin concentration was measured using an enzyme immunoas-
say (Human Adiponectin ELISA kit, B-Bridge International Inc., CA, USA) which
were read using a Victor
2
(Perkin Elmer life sciences, Turku, Finland) at 450 nm
and wavelength correction was set to 540 nm.
2.7. Statistical analyses
Hardy Weinberg Equilibrium (HWE), linkage Disequilibrium (LD), and
haplotype frequencies were determined by Haploview ver. 3.32 (http://www.
broad.mit.edu/mpg/haploview/) on the basis of the EM algorithm [8] and was
examined using the Executive SNP Analyzer 1.2A (http://snp.istech.info/istech/
board/login_form.jsp). χ
2
tests were used to determine whether individual
variants were in HWE at each locus in the samples.
Differences in clinical and metabolic variables between individuals with
different genotypes and haplotypes were tested by Student's ttests (to test
the difference between 2 genotype groups) and 1-way analysis of variance
(ANOVA) or analysis of covariance (ANCOVA) (with Tukey's test for post hoc
comparisons of each genotype or haplotype) using SPSS ver. 12.0 for Windows
(Statistical Package for the Social Science, SPSS Ins., Chicago, IL). It was first
determined whether each variable presented a normal distribution before
statistical testing, and then logarithmic transformation was performed on the
skewed variables. For descriptive purposes, mean values are presented using
untransformed values. Results are expressed as mean±S.E. A 2-tailed value of
Pb0.05 was considered statistically significant.
3. Results
3.1. Detection of SNPs in the ADIPOQ gene
The allele frequencies of the 8 SNPs in the ADIPOQ
were as follows: - 11391G NA (G:A = 1:0), - 11377C NG(C:G=
0.73:0.27), 45T NG (T:G = 0.71:0.29), 276G NT (G:T= 0.70:0.30),
86 Y. Jang et al. / Clinica Chimica Acta 391 (2008) 85–90
H241P (A:C = 1:0), Y111H (T:C = 1:0), G90S (G:A = 1:0), and
R221S (C:A = 0.98:0.02). We included SNPs - 11377C NG,
45T NG, and 276G NT in further analysis because the others
were rare mutation with minor allele frequency (MAF) b2% in this
study subjects.
3.2. Distribution of - 11377C NG, 45T NG and 276G NTgenotype
and 45-276 haplotype analyses in the entire population
All SNPs were in Hardy-Weinberg equilibrium (p N0.05).
53.3% of C/C, 39.8% of C/G, and 6.9% of G/G was at SNP
-11377, 51.7% of T/T, 39.1% of T/G, and 9.2% of G/G was at
SNP 45, and 49.1% of G/G, 41.3% of G/T, and 9.6% of T/T was
at SNP 276. These frequencies are similar to those reported
previously in non-diabetic Koreans (T frequency at position
45 = 0.71; G frequency at position 276 = 0.71) [9] and Japanese
(0.71; 0.70, respectively) [10].
There was low LD between the SNP + 45T NGandSNP
- 11377C NG(D′=0.692;Pb0.001) and between SNP +276GNT
and SNP -11377CNG(D′=0.003; P= 0.092). SNP +45TNG
and SNP + 276G NT were found to be highly linked by the linkage
disequilibrium test (D′=1; Pb0.001). In light of the results of
the LD analysis, haplotype analysis was performed for
SNP 45T NG combined with SNP 276G NT. There were esti-
mated 45-276 haplotype frequencies of 16.3% for TG/TG, 25.8%
for TG/TT, 23.6% for TG/GG, 9.6% for TT/TT, 15.5% for TT/GG,
and 9.2% for GG/GG. For subsequent statistical analyses, subjects
were divided into 3 haplotype groups: homozygous for the TG
haplotype (TG/TG; n = 141), heterozygous carriers of the TG
haplotype (TG/X; n= 429) and non-TG haplotype carriers (X/X;
n = 297).
3.3. Clinical characteristics according to ADIPOQ SNP genotypes
The general characteristics of the subjects were presented in
Tab l e 1. Participantshad a mean age of 41± 0.5 y old, witha mean
body mass index (BMI) of 24.6±0.1 kg/m
2
and other clinical
parameters were within a normal range. The clinical character-
istics of the study subjects were analyzed adjusted for age, BMI,
smoking, and alcohol consumption according to genotypes at
the - 11377C NG, 45TNG, and 276G NTSNPs(Table 2). There
were no significant differences in age, anthropometric parameters,
systolic and diastolic blood pressure and serum concentrations of
glucose, HDL- and LDL-cholesterol according to SNP geno-
types. Total- (P=0.004) and LDL-cholesterol (P=0.033) con-
centrations in G allele carriers of the 276GNT SNP is higher than
T/T genotype group after an adjustment. No other SNPs were
associated with total- or LDL-cholesterol. Similarly, no associa-
tion was found with 45-276 haplotypes (data were not shown). The
adiponectin concentrations were different between the major allele
carriers than a homozygotes for minor allele of SNP -11377C NG
(P = 0.036), 45T NG (P = 0.050), and 276G NT (P = 0.004), respec-
tively (Tab l e 2).
3.4. Association of the ADIPOQ SNPs with serum triglyceride
and insulin concentrations
Serum concentrations of triglyceride, insulin, HOMA-IR, and
adiponectin were analyzed according to genotypes at the
Table 1
General characteristics of the study subjects
n = 867
Age (y) 41.2 ± 0.45
BMI (kg/m
2
)
1
24.6 ± 0.12
Waist (cm) 83.6 ± 0.32
Blood Pressure (BP)
Systolic BP (mmHg) 117.9± 0.58
Diastolic BP (mmHg) 75.2 ± 0.40
Triglyceride (mg/dl)⁎118.5± 2.04
Total cholesterol (mg/dl) 197.4 ± 1.27
LDL-cholesterol (mg/dl) 121.5 ± 1.20
HDL-cholesterol (mg/dl) 51.7± 0.44
Glucose (mg/dl) 83.8 ± 0.34
Insulin (μU/ml) 9.08 ± 0.19
HOMA-IR
2⁎
1.90 ± 0.04
Adiponectin (ug/ml)⁎7.39 ± 0.13
⁎log-transformed.
1
body mass index.
2
HOMA-IR = {fasting insulin (μIU/ml) × fasting glucose (mmol/l)}/22.5.
Table 2
Clinical characteristics according to the genotype of ADIPOQ SNPs in healthy Korean women
SNP -11377 SNP 45 SNP 276
C/C+C/G G/G P T/T+T/G G/G P G/G+G/T T/T P
n 807 60 787 80 784 83
Age (years) 41 ± 0.5 40 ± 2.0 NS 41 ± 0.5 41 ± 1.5 NS 41 ± 0.5 43 ± 1.5 NS
BMI (kg/m
2
)
1
24.7 ± 0.1 24.4 ± 0.5 NS 24.7 ± 0.1 24.2± 0.4 NS 24.7 ± 0.1 24.6 ± 0.4 NS
Waist (cm) 83.6 ± 0.3 83.1 ± 1.4 NS 83.7 ± 0.3 82.7± 1.1 NS 83.6 ± 0.3 83.3 ± 1.0 NS
Blood Pressure (BP)
Systolic BP (mmHg) 118± 0.6 118± 2.1 NS 118 ± 0.6 116 ± 1.8 NS 118 ± 0.6 120 ± 1.8 NS
Diastolic BP (mmHg) 75 ± 0.4 75 ± 1.4 NS 75 ± 0.4 75 ± 1.2 NS 75 ± 0.4 76 ± 1.3 NS
Glucose (mg/dL) 84 ± 0.4 84 ± 1.2 NS 84 ± 0.4 83 ± 1.3 NS 84 ± 0.4 84 ± 1.2 NS
HDL-cholesterol (mg/dL) 52 ± 0.5 51 ± 1.6 NS 52 ± 0.5 53 ± 1.5 NS 52 ± 0.5 52 ± 1.0 NS
Total-cholesterol (mg/dL) 197± 1.3 199 ± 5.1 NS 197 ± 1.3 196 ±4.1 NS 198 ± 1.3 191± 3.9 0.004
LDL-cholesterol (mg/dL) 121 ± 1.2 122 ± 5.0 NS 122 ± 1.3 119± 3.7 NS 122 ± 1.3 117 ± 3.8 0.033
Adiponectin (ug/mL) 7.49 ± 0.14 6.16± 0.41 0.036 7.28± 0.13 8.40 ± 0.50 0.050 7.28± 014 8.35± 0.42 0.004
Data are mean ± S.E.
1
body mass index. P values were from Student's t-test comparing the differences between the genotype groups of each SNPs adjusted for age,
BMI, smoking and alcohol consumption.
87Y. Jang et al. / Clinica Chimica Acta 391 (2008) 85–90
-11377CNG, 45T NG, and 276G NTSNPs(Ta ble 3 ). There were
significant associations between SNP at 276 and the serum
concentrations of triglyceride (P= 0.013), insulin (P= 0.013) and
HOMA-IR (P= 0.012) as well as the adiponectin (P= 0.001). The
subjects with the T/T genotype at position 276 had lower serum
triglyceride concentration (T/T; 105± 5 mg/dl vs G/G; 123 ± 3,
P= 0.005) and HOMA-IR (T/T; 1.65± 0.09 vs G/G; 2.01±0.07,
P =0.049) than those with the G/G genotype. SNPs -11377 and 45
were not associated with these metabolic variables. The 45/276
haplotypes showed a similar association as the 276G NTSNP
(Table 3).
3.5. Association of ADIPOQ SNPs with adiponectin
concentrations
Differences in plasma adiponectin concentrations were ob-
served across genotypes of the three ADIPOQ SNPs (Table 3,
Fig. 1). There was a difference between C/C and G/G group
of -11377 (P= 0.025) but the differences among every genotype
group could not reach the statistical significance. A strong statistical
significance was reached at position 276 (P= 0.01), with carriers of
genotype T/T having 20% higher adiponectin concentrations than
G/G carriers (G/G; 7.0 ± 0.18 vs T/T; 8.4 ± 0.42 μg/ml, P = 0.005).
Table 3
Association of the ADIPOQ SNPs and with serum triglyceride and insulin concentrations
N Triglyceride (mg/dl) P Insulin (μU/ml) P HOMA-IR
1
P Adiponectin (ug/ml) P
- 11377 C/C 454 115± 3 NS 8.88 ± 0.22 NS 1.85 ± 0.05 NS 7.54 ± 0.19 NS
C/G 340 121 ± 4 9.38 ± 0.35 1.97 ± 0.08 7.42 ± 0.20
G/G 58 126 ± 8 9.18± 0.55 1.90 ± 0.12 6.16± 0.41
45 T/T 438 119 ± 3 NS 8.97 ± 0.28 NS 1.88 ± 0.06 0.342 7.25 ± 0.18 NS
T/G 336 119 ± 3 9.37± 0.27 1.97 ± 0.06 7.31± 0.21
G/G 78 113 ± 6 8.40± 0.49 1.75 ± 0.11 8.40± 0.50
276 G/G 416 123 ±3 0.013 9.54 ± 0.30 0.013 2.01± 0.07 0.012 7.01 ± 0.18 0.001
G/T 352 116 ± 3 8.77± 0.25 1.84 ± 0.06 7.59± 0.21⁎
T/T 84 105 ± 5⁎⁎ 8.01 ± 0.41⁎1.65 ± 0.09⁎8.35 ± 0.42⁎⁎
45-276 TG/TG 141 125 ± 5 0.004 9.54 ± 0.68 NS 2.01 ± 0.14 0.055 6.73 ± 0.30 b0.001
TG/X 429 123 ± 3 9.43± 0.24 1.98 ±0.06 6.89 ± 0.17
X/X 297 109 ± 3⁎8.34 ± 0.25 1.74± 0.06⁎8.36 ± 0.24⁎⁎⁎
To comply with normality assumption, the data were tested after log-transformation.
P values were from one-way analysis of covariance (ANCOVA) comparing the differences among the genotype groups adjusted for age, BMI, smoking, and alcohol
consumption.
⁎Pb0.05, ⁎⁎Pb0.01, ⁎⁎⁎Pb0.001 compared to G/G group of SNP 276 or TG/TG group of 45-276 haplotype from Tukey's test for post hoc comparisons.
1
HOMA-IR = {fasting insulin (μIU/ml) × fasting glucose (mmol/l)}/22.5.
Fig. 1. Association of the ADIPOQ SNPs and plasma adiponectin concentrations in 867 non-diabetic Korean women. Mean ±S.E. Serum adiponectin concentrations
were compared among genotype groups by ANOVA after log-transformation. ⁎Pb0.05, ⁎⁎Pb0.01, ⁎⁎⁎Pb0.001 compared to wild type group from Tukey's test for
post hoc comparisons after ANCOVA adjusted for age, BMI, smoking and alcohol consumption.
88 Y. Jang et al. / Clinica Chimica Acta 391 (2008) 85–90
Much stronger associations with adiponectin concentrations were
observed when SNP 45 and SNP 276 were considered together as
haplotypes (P b0.001). Homozygous carriers of the TG haplotype
(i.e., individuals who were T/T at 45 and G/G at 276) and
heterozygous carriers of the TG haplotype (TG/X) had lower
adiponectin concentrations than non-TG carriers (TG/TG; 6.7 ±
0.30 vs TG/X; 6.9 ± 0.17 vs X/X; 8.4 ± 0.24 μg/ml, P b0.001). A
tendency to be associated with lower adiponectin concentrations
was found for homozygotes for 45G/G genotype but this was not
statistically significant.
3.6. Effect of ADIPOQ 45-276 haplotype on adiponectin and
insulin concentrations according to BMI
We evaluated association of the 45-276 haplotype with
serum adiponectin concentrations in the 2 subgroups according
to individual BMI [i.e., below (normal weight) or above
(overweight-obese) 25 kg/m
2
](Table 4). As expected, the 2
subgroups differed significantly for mean plasma adiponectin
(8.1 ± 0.19 vs 6.4 ± 0.16 μg/ml, P b0.001), serum insulin (7.7 ±
0.21 vs 10.7 ± 0.29 μU/ml, P b0.001) and HOMA-IR (1.56 ±
0.04 vs 2.28 ± 0.07, P b0.001). In subgroup analysis, subjects
carrying the TG haplotype had significantly lower adiponectin
concentrations than non-TG carriers (X/X) in both normal
weight (TG/TG + TG/X; 7.4 ± 0.22 vs X/X; 9.3 ± 0.33 μg/ml,
Pb0.001) and overweight-obese (TG/TG + TG/X; 6.1 ± 0.19 vs
X/X; 6.9 ± 0.30 μg/ml, P = 0.009) subgroups. The association of
the TG haplotype with lower adiponectin concentrations was
more pronounced in the normal weight subjects. The associa-
tion of the TG haplotype with higher insulin concentrations than
non-TG haplotype was significant among overweight-obese
subjects (TG/TG + TG/X; 11.2 ± 0.37 vs X/ X; 9.5 ± 0.42 μU/ml,
P = 0.004), but not significant among normal weight subjects
(TG/TG + TG/X; 7.8 ± 0.31 vs X/X; 7.4 ± 0.27 μU/ml,
P = 0.352). The differences in serum insulin concentrations
between the subjects carrying the TG haplotype and the non-TG
carriers in two subgroups became large with overweight-obesity
(ranging from 0.3 μU/ml in the normal weight individuals to
1.7 μU/ml in the overweight-obese individuals). A similar
association was found between the 45-276 haplotype and
HOMA-IR (Table 4).
4. Discussion
In this study, SNP 276G NT had the strongest association
with adiponectin concentrations, with allele G being associated
with low and allele T with high adiponectin concentrations.
Allele G of SNP276 was more strongly associated with
adiponectin concentrations, especially when occurring in
conjunction with allele T at position 45. This present findings
are confirmative result for our previous reports that the
haplotype TG defined by the SNPs 45T NG and 276G NT was
associated with lower adiponectin concentrations in healthy
individuals [4,11].
Although the overweight-obese subgroup showed lower
mean values of plasma adiponectin than the normal weight
subgroup, subjects carrying the TG haplotype had significantly
lower adiponectin concentrations than non-TG carriers in both
subgroups. This result, which is unaffected by consideration of
age and BMI across haplotype, supports the recent finding of
Menzaghi et al. [4] that adiponectin heritability is not affected
by the inclusion of body weight in the genetic model. However,
neither SNP 45 nor SNP 276 affects known regulatory regions,
one being a synonymous substitution and the other being
located in an intron [12]. Thus, these variants are probably
markers of one or more haplotypes containing a causal
polymorphism affecting plasma adiponectin concentrations.
A tendency to be associated with low adiponectin was
observed for the G/G genoty pe of the -11377C NG SNP in this
study. Similarly, the G allele of the - 11377 SNP was found to be
associated with low adiponectin in healthy postmenopausal
Danish women [13]. In addition, a haplotype including SNPs
-11391 and -11377 was associated with adiponectin concentra-
tions and with type 2 diabetes in French Caucasians [14,15].
However, -11391 SNP at ADIPOQ promoter region was mono-
morphic in this study population (G:A = 1:0). Also -11391 SNP
was not found in Japanese and Chinese population [10,16].
Thus, differences among populations in the linkage disequili-
brium structure at this locus may result in association of the
adiponectin concentration haplotype with different SNPs in
different populations.
We found an association between the 276G allele as well as
the 45-276 TG haplotype with significantly higher serum
insulin and insulin resistance index. The present findings are
consistent with previous reports [4] that the ADIPOQ gene may
play an important role in insulin resistance syndrome. The
association in this study also extends to other features of the
insulin resistance syndrome, such as increased serum concen-
trations of triglyceride. This might be a manifestation of the
same genetic effect underlying the association with low
adiponectin concentrations due to the role of adiponectin as a
potent insulin enhancer [17–19]. It has been suggested that
ADIPOQ gene variants are associated with insulin resistance,
Table 4
Association of the SNP45-276 haplotype with circulating concentrations of
adiponectin and insulin and HOMA-IR in BMI categorized subgroups
Normal wt Overwt-obese
TG/TG+
TG/X
X/X P TG/TG+
TG/X
X/X P
n 313 175 257 122
Age (y) 40 ± 1 40± 1 0.709 42 ± 1 44 ± 1 NS
BMI
(kg/m
2
)
1
22.3 ±0.12 22.1 ± 0.16 0.310 27.9 ± 0.16 27.5± 0.20 NS
Adiponectin
(ug/ml)⁎
7.4 ± 0.22 9.3 ± 0.33 b0.001 6.1 ± 0.19 6.9± 0.30 0.009
Insulin
(μU/ml)⁎
7.8 ± 0.31 7.4 ± 0.27 0.352 11.2 ± 0.37 9.5± 0.42 0.004
HOMA-IR
2⁎
1.6 ± 0.06 1.5 ± 0.06 0.171 2.4 ± 0.09 2.0± 0.09 0.008
⁎Tested after log-transformation.
P values were from Student's t-test comparing the differences between the
haplotype groups adjusted for age, smoking and alcohol consumption.
1
body mass index.
2
HOMA-IR = {fasting insulin (μIU/ml) × fasting glucose (mmol/l)}/22.5.
89Y. Jang et al. / Clinica Chimica Acta 391 (2008) 85–90
which might be mediated through alterations in the expression
concentration and serum concentrations of adiponectin [4,10].
In subgroup analysis, the high serum insulin concentrations
and HOMA-IR observed among subjects with the 45-276 TG
haplotype was only observed in the overweight-obese subgroup.
This suggests that a significant proportion of the gene effect on
serum insulin concentrations depends on changes in body fat
mass. One possible explanation for our results might be that the
effect of the adiponectin gene on serum insulin or insulin
resistance index is exaggerated by environmental factors such as
overweight-obesity. However, a contrasting result has been
reported. In 253 Italian subjects, an association of T/T genotype
at SNP 276 with high insulin resistance was only found in the
group of participants with a BMI below the median of 26.2 kg/m
2
[20]. This discrepancy might be partly due to ethnic specificity,
the difference in the entry criteria of subjects (e.g., women only
vs women and men) or a limited number of subjects. Further
study would be needed to clarify which is the case; that certain
ADIPOQ variants predispose to obesity and low adiponectin
concentrations causing low insulin sensitivity or thatobesity leads
to low adiponectin concentration and low insulin sensitivity and
certain ADIPOQ variants merely exacerbate this effect.
Because gender is an important determinant for adiponectin
concentrations, in our study we specifically focused on a repre-
sentative group of non-diabetic, healthy Korean women of age
20-69 y. Therefore, our data cannot be easily generalized to
other men or other ethnic, age, or geographical groups. Despite
this limitation, our data replicated a strong association of
the SNP276 genotypes as well as the 45-276 haplotypes with
circulating adiponectin concentrations. In addition, this haplo-
type or an unknown nearby functional variant in linkage dis-
equilibrium would increase insulin concentrations and insulin
resistance index only in overweight-obese individuals.
Acknowledgements
This work as supported by the Korea Science and Engineer-
ing Foundation (KOSEF), the Korea government Ministry of
Science and Technology (M10642120002-06N4212-00210),
National Research Laboratory project # R0A-2005-000-
10144-0, Ministry of Science and Technology, Korea Health
21 R&D Projects, Ministry of Health & Welfare (A000385,
A020593, A050376), Korea Research Foundation Grant funded
by Korea Government (MOEHRD, Basic Research Promotion
Fund) (KRF-2006-311-C00640), and Brain Korea 21 Project,
Yonsei University College of Human Ecology, Yonsei University.
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