Clinica Chimica Acta 411 (2010) 26–30
Contents lists available at ScienceDirect
Clinica Chimica Acta
j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / c l i n c h i m
Genetic variant in visfatin gene promoter is associated with decreased risk of
coronary artery disease in a Chinese population
Jian-Jun Yan a, Na-Ping Tang a,b, Jian-Jin Tang a, En-zhi Jia a, Ming-Wei Wang a, Qi-Ming Wang a, Jun Zhu a,
Zhi-Jian Yang a, Lian-sheng Wang a,⁎, Jun Huang a,⁎
a
Department of Cardiology, First Affiliated Hospital of Nanjing Medical University, 300 Guangzhou Road, Nanjing, 210029, Jiangsu Province, China
National Shanghai Center for New Drug Safety Evaluation and Research, Shanghai Institute of Pharmaceutical industry, 199 Guoshoujing Road, Zhangjiang Hi-Tech Park,
Pudong, Shanghai 201203, China
b
a r t i c l e
i n f o
Article history:
Received 28 March 2009
Received in revised form 27 September 2009
Accepted 29 September 2009
Available online 3 October 2009
Keywords:
Coronary artery disease
Visfatin
Polymorphism
a b s t r a c t
Background: Visfatin is a newly identified pro-inflammatory adipokine expressed predominantly in visceral
fat. Previous studies have suggested a role for visfatin in low-grade inflammation and regulation of lipid
metabolism. Most recently, a genetic polymorphism − 1535C > T located in the visfatin gene promoter has
been identified, and suggested to be associated with the regulation of visfatin expression, lipid levels.
However, it is unclear whether this polymorphism has a linkage with CAD.
Methods: We conducted a hospital-based case-control study with 257 CAD patients and 292 controls to
examine the potential association of the Visfatin − 1535C > T polymorphism with CAD.
Results: The frequencies of the CC, CT, and TT genotypes in cases were significantly different from those of
controls (χ2 = 6.223, P = 0.045). Subjects with the variant genotypes (CT + TT) had a 40% decreased risk of
CAD relative to CC carriers (adjusted OR = 0.60, 95%CI = 0.40–0.89). Furthermore, the adjusted OR of a TT
genotype for CAD was 0.52 (95%CI = 0.31–0.87). There was a significant association between Visfatin
− 1535C > T polymorphism and triglyceride levels in both CAD patients and controls (P = 0.003, 0.018,
respectively). In stratified analyses, the T allele was significantly associated with reduced risk of CAD in
males, subjects with age < 59 years, and non-smokers. Moreover, a borderline statistical significance
(P = 0.058 for trend) was observed between the variant genotypes and severity of CAD.
Conclusion: Our results suggested that Visfatin − 1535C > T polymorphism might be associated with reduced
risk of CAD in a Chinese population.
© 2009 Elsevier B.V. All rights reserved.
1. Introduction
Coronary artery disease (CAD) is one of the major causes of death in
most countries, including China [1]. Atherosclerosis, the main cause of
CAD, is an inflammatory disease in which immune mechanisms interact
with environmental and genetic risk factors to initiate, propagate, and
activate lesions in the arterial tree, eliciting various acute and sever
events such as acute coronary syndromes [2]. Most recently, genetic
studies have revealed a series of candidate gene susceptibility variants
that may also contribute to the pathogenesis of CAD [3,4].
Visfatin (also known as pre-B cell colony-enhancing factor or PBEF)
is a novel pro-inflammatory adipokine expressed markedly in visceral
fat [5]. Visfatin is a 52- to 55-kDa protein, which was originally cloned
as a growth factor enhancing the effect of stem cell factor and IL-7 on
the colony formation activity of early stage B cells [6]. Visfatin has a
potential insulin-mimetic action that may cause insulin sensitivity [5],
⁎ Corresponding authors. Tel./fax: +86 25 83724440.
E-mail addresses: [email protected] (L. Wang), [email protected]
(J. Huang).
0009-8981/$ – see front matter © 2009 Elsevier B.V. All rights reserved.
doi:10.1016/j.cca.2009.09.033
however, which remains controversial [5,7]. Recent studies have
indicated that visfatin is involved in the processes of inflammation, an
important component of atherosclerosis. Serum visfatin levels were
positively correlated with the serum levels of IL-6 and CRP, indicating
that circulating visfatin may reflect inflammation status [8,9]. Visfatin
has also been shown to be localized to foam cell macrophages within
unstable atherosclerotic lesions, which potentially play a role in
plaque destabilization [10]. Moreover, it has been suggested that
visfatin plays a key part in nicotinamide adenine dinucleotide (NAD)
biosynthesis pathway [7,11,12]. Furthermore, visfatin might be an
independent CAD risk factor [13].
The visfatin gene is located on chromosome 7q22.2, which is
composed of 11 exons and 10 introns, spanning 34.7 kb of genomic
DNA. The proximal and distal regions of visfatin gene promoter
contain several transcription initiation sites, and binding sites for Sp1
and ubiquitous transcription factors such as nuclear factor 1 (NF-1),
AP-1, AP-2 [14]. To date, several single nucleotide polymorphisms in
the visfatin gene have been described. The visfatin promoter
polymorphisms have been reported as a linkage with low-grade and
acute inflammation [15,16]. One of the most frequently studied
J.-J. Yan et al. / Clinica Chimica Acta 411 (2010) 26–30
27
polymorphisms, −1535C > T, was reported to be associated with the
regulation of visfatin gene expression and serum lipid level [16–18].
However, there were some conflicting results regarding whether this
variant is functional or not [16,17,24]. To the best of our knowledge,
no data have been published on the association of the − 1535C > T
polymorphism with the risk of CAD. We investigate the potential
association between the − 1535C > T polymorphism and the risk of
CAD in a hospital-based case-control study in a Chinese population.
Biotech Co., Ltd.). Subsequently, LDR products were analyzed by DNA
sequencing (Model 377, Applied Biosystems). All assays were conducted blindly without the knowledge of case or control status.
Additionally, about 10% of the samples were randomly selected and
retested by direct DNA sequencing on a 3730xl DNA analyzer (Applied
Biosystems) and the results were 100% concordant.
2. Materials and methods
Statistical analyses were conducted with Stata ver. 8.0 (STATA Corp.,
College Station, TX) and SPSS 13.0 (SPSS Inc., Chicago, IL). Normality was
tested using the Kolmogorov–Smirnov test. Differences of continuous
variables without skewness (presented as mean± SD) between 2
groups were calculated by the Student's t-test. Differences of continuous
variables departing from the normal distribution even after transformation between 2 groups were analyzed by Mann–Whitney U-test.
Pearson χ2-test was used to compare allele distribution and qualitative
variables represented as frequencies. Hardy–Weinberg equilibrium was
assessed for both subjects and controls by a χ2 goodness-of-fit test.
Odds ratio (OR) and 95% confidence interval (CI) were calculated to
estimate the correlation between the −1535C> T polymorphism and
the risk of CAD. A binary logistic regression analysis was used for the
evaluation of the independent effect of visfatin genotypes on the
development of CAD, adjusted for the presence of risk factors including
age, sex, body mass index (BMI), smoking status, hypertension, diabetes
and dyslipidemia. Odds ratio (OR) and 95% confidence interval (CI) were
calculated. The linear trend in the association of −1535C> T polymorphism with CAD severity was evaluated by χ2-test for trend. Two-tailed
P < 0.05 were considered as statistical significance.
2.1. Study population
The association study enrolled 257 unrelated patients of CAD, who
were all admitted to the First Affiliated Hospital of Nanjing Medical
University. The diagnosis of CAD was certified by coronary angiography
performed with the Judkins technique using a quantitative coronary
angiographic system [19]. CAD was defined as an angiography with
≥50% luminal narrowing in one main coronary artery or more. Patients
with CAD were divided into 1-, 2-, and 3-vessel disease subgroups
according to the number of significantly stenosed vessels with reference
to the Coronary Artery Surgery Study classification. Two cardiologists
who were unaware of the patients included in this study assessed the
angiograms. This study included 292 unrelated control subjects
randomly selected from outpatients (staff of local companies and
administration agencies) who underwent regular physical examinations during the same time in the same hospital. After considering it
unethical to perform coronary angiography to rule out the presence of
asymptomatic CAD, those control subjects with a history of angina,
symptoms or signs of other atherosclerotic vascular diseases and an
abnormal electrocardiogram were excluded. All subjects enrolled in this
study were of Han Chinese origin and residing in or near Jiangsu
Province. They had no history of significant concomitant diseases
including cardiomyopathy, bleeding disorders, renal failure, previous
thoracic irradiation therapy and malignant diseases. Hypertension was
defined as resting systolic blood pressure >140 mm Hg and/or diastolic
blood pressure >90 mm Hg or in the presence of active treatment with
antihypertensive agents. Diabetes was defined as fasting blood glucose
>7.0 mmol/l or a diagnosis of diabetes needing diet or antidiabetic drug
therapy. Dyslipidemia was defined as total cholesterol concentration of
>5.72 mmol/l or on drugs. Individuals who formerly or currently
smoked ≥10 cigarettes per day for at least 2 years were defined as
smokers. This study was approved by the First Affiliated Hospital Ethics
Committee of Nanjing Medical University and informed consent was
obtained from each participant.
2.3. Statistical analysis
3. Results
3.1. Demographic information
The characteristics of our study subjects are given in Table 1.
Patients with CAD were much older, smoked more cigarettes, had
significantly higher weight, TG, TC , LDL-C, body mass index (BMI),
and were more likely to be diabetic, hypertensive and dyslipidemic
than the control subjects. In terms of coronary angiographic findings,
100 (38.9%) CAD cases had single-vessel disease, 87 (33.9%) had
double-vessel disease and 70 (27.2%) had triple-vessel disease.
Table 1
Baseline characteristics of cases and controls.
Characteristics
2.2. DNA extraction and genotyping
Peripheral venous blood was drawn from each participant. Genomic
DNA was extracted using the AxyPrep DNA Blood kit (Axygen Scientific
Inc., Union City, CA, USA). The −1535C > T (rs61330082) polymorphism was genotyped by the PCR-LDR Sequencing method, as reported
previously [20,21]. A 216 bp DNA fragment (nucleotide positions
−1637 to −1422 of the promoter) containing the polymorphic site
was amplified by PCR in the ABI 9600 (Applied Biosystems, Foster City,
CA) using the forward primer 5′-ACACAGGGAAGATCAACCAA-3′ and
the reverse primer 5′-AGACAACCTCAGTCAACACTA-3′. The PCR was
carried out in a total volume of 15 μl containing 1.5 μl 10 × PCR buffer,
0.2 μl 10 pmol each primer, 0.3 μl dNTP, 0.25 μl Taq polymerase (MBI
fermentas), 2 μl of genomic DNA and 10.75 μl H2O. The PCR cycling
parameters were 35 cycles of 30 s at 94 °C, 55 °C for 30 s and 72 °C for
45 s. Ligase Detection Reaction (LDR) was performed in a total volume of
10 μl containing 2 μl PCR product, 1 μl 10 × Taq DNA ligase buffer,
0.125 μl 40 U/μl Taq DNA ligase (NEB), 1 μl 10 pmol probes (0.33 μl each
of probe), and 5.875 μl H2O. LDR probes were composed of 1 common
probe and 2 discriminating probes (designed by the Shanghai Generay
Age (years)
Sex (male), n (%)
BMI (kg/m2)
Hypertension, n (%)
Diabetes, n (%)
Dyslipidemia, n (%)
Smoking, n (%)
TC (mmol/l)
TG (mmol/l)
HDL-C (mmol/l)
LDL-C (mmol/l)
Glucose (mmol/l)
Number of diseased vessels
Single vessel, n (%)
Double vessels, n (%)
Triple vessels, n (%)
Cases
Controls
(n = 257)
(n = 292)
P
63.9 ± 9.6
210 (81.7)
24.9 ± 3.3
128 (49.8)
56 (21.8)
64 (24.9)
96 (37.4)
4.14 ± 1.16
1.73 ± 0.96
1.12 ± 0.43
2.58 ± 0.89
5.86 ± 2.69
59.3 ± 12.4
211 (72.3)
24.1 ± 2.7
98 (33.6)
23 (7.9)
33 (11.3)
42 (14.4)
3.96 ± 1.17
1.42 ± 1.01
1.33 ± 0.35
2.28 ± 0.76
5.00 ± 0.92
< 0.001
0.009
0.002
< 0.001
< 0.001
< 0.001
< 0.001
0.009
< 0.001
< 0.001
< 0.001
< 0.001
100 (38.9)
87 (33.9)
70 (27.2)
–
–
–
–
–
–
BMI, body mass index; HDL-C, high density lipoprotein cholesterol; LDL-C, low density
lipoprotein cholesterol; TC, total cholesterol; TG, triglyceride.
Age, TC, TG, HDL-C, LDL-C and glucose (expressed as mean ± SD) were abnormal
distributed and analyzed by Mann–Whitney U-test. BMI (expressed as mean ± SD) was
normally distributed and analyzed by Student's t-test. Other data were expressed as
frequencies and percentages and evaluated by χ2-test.
28
J.-J. Yan et al. / Clinica Chimica Acta 411 (2010) 26–30
Table 2
Distribution of the visfatin genotype and the risk estimates for the variant visfatin genotypes.
Adjusteda
Cases
Controls
Crude
(n = 257)
(n = 292)
OR (95%CI)
P
OR (95%CI)
P
92 (35.8)
118 (45.9)
47 (18.3)
165 (64.2)
78 (26.7)
143 (49.0)
71 (24.3)
214 (73.3)
1.00
0.70 (0.48–1.03)
0.56 (0.35–0.90)
0.65 (0.45–0.94)
0.071
0.017
0.022
1.00
0.64 (0.42–0.98)
0.52 (0.31–0.87)
0.60 (0.40–0.89)
0.042
0.013
0.011
302 (58.8)
212 (41.2)
299 (51.2)
285 (48.8)
–
–
–
–
–
–
–
–
b
Genotype , n (%)
CC
CT
TT
CT + TT
Allelec, n (%)
C allele
T allele
Distributions of the visfatin genotype in cases and controls were in Hardy–Weinberg equilibrium (P = 0.399 and P = 0.733, respectively).
CI, confidence interval; OR, odds ratio.
a
Adjusted for age, sex, body mass index, smoking status, hypertension, diabetes, dyslipidemia.
b
Pearson χ2 = 6.223, P = 0.045 for genotype.
c
Pearson χ2 = 6.300, P = 0.012 for allele.
3.2. Genotypes and allele frequencies of cases and controls and their
associations with CAD
The allele frequencies of SNP −1535C > T in CAD group and control
group were 58.8%/41.2% and 51.2%/48.8%, respectively. The genotype
distribution in our study subjects showed no deviation from Hardy–
Weinberg equilibrium (P = NS). The distribution of C > T genotypes was
also markedly different between cases and controls (P = 0.045) as
shown in Table 2. Table 2 also shows the risk estimates for the variant
−1535C > T genotypes among CAD patients compared with controls.
Overall, after being adjusted for the risk factors including age, sex, BMI,
smoking status, hypertension, diabetes and dyslipidemia, the OR for
subjects with the variant genotypes (CT and TT) was 0.60 (95%
CI= 0.40–0.89, P = 0.011). We also found that TT genotype carriers
had a 48% reduction in risk of CAD compared with the CC carriers
Table 3
BMI, lipid profiles and glucose of cases and controls in the visfatin genotypes.
Characteristics
BMI (kg/m2)
TC (mmol/l)
TG (mmol/l)
HDL-C (mmol/l)
LDL-C (mmol/l)
Glucose (mmol/l)
Cases
P
CC
CT + TT
24.9 ± 3.4
4.26 ± 1.20
1.91 ± 0.93
1.11 ± 0.41
2.67 ± 0.90
5.84 ± 2.59
24.9 ± 3.3
4.06 ± 1.14
1.63 ± 0.97
1.12 ± 0.44
2.54 ± 0.88
5.87 ± 2.74
NS
NS
0.003
NS
NS
NS
Controls
CT + TT
24.2 ± 2.39
4.04 ± 1.25
1.67 ± 1.35
1.33 ± 0.34
2.45 ± 0.73
5.00 ± 0.81
24.1 ± 2.86
3.94 ± 1.14
1.33 ± 0.83
1.34 ± 0.35
2.21 ± 0.77
5.00 ± 0.95
3.3. BMI, lipid profiles and glucose of cases and controls in the visfatin
genotypes
Table 3 shows BMI, lipid profiles and glucose levels according to the
−1535C> T genotypes in the CAD patients and controls. We found a
significant association between the genotypes and serum triglyceride
levels in these 2 groups (P = 0.003 and 0.018, respectively). In addition,
we also noted that there was a borderline significance between the
genotypes and serum LDL-C levels in controls (P = 0.062). However,
there were no significant differences between the genotypes and BMI or
glucose levels in both groups.
3.4. Stratified analyses of the polymorphism and CAD
P
CC
(adjusted OR = 0.52, 95% CI = 0.31–0.87, P = 0.013). We used a
dominant model for analysis in the present study. In addition, we
have attempted to conduct analysis with other models. However, no
other statistical significance was observed (data not shown).
NS
NS
0.018
NS
NS
NS
BMI, body mass index; CAD, coronary artery disease; HDL-C, high density lipoprotein
cholesterol; LDL-C, low density lipoprotein cholesterol; TC, total cholesterol; TG,
triglyceride.
TC, TG, HDL-C, LDL-C and glucose (expressed as mean ± SD) were abnormal distributed
and analyzed by Mann–Whitney U-test. BMI (expressed as mean ± SD) was normal
distributed and analyzed by Student's t-test.
We conducted stratification analyses according to median age of
controls (59 years), gender, smoking status (Table 4). We noted that
the T allele was significantly associated with a reduction in risk of CAD
in male (adjusted OR = 0.49, 95% CI = 0.31–0.77, P = 0.002); subjects
with age < 59 years (adjusted OR = 0.43, 95% CI = 0.21–0.88,
P = 0.021) and non-smokers (adjusted OR = 0.57, 95% CI = 0.37–
0.87, P = 0.009), while not in female (adjusted OR = 1.27, 95%
CI = 0.52–3.08, P = NS); subjects with age ≥ 59 years (adjusted
OR = 0.68, 95% CI = 0.41–1.12, P = NS) and smokers (adjusted
OR = 0.82, 95% CI = 0.36–1.85, P = NS). Patients with CAD were
subclassified into 3 subgroups (single-, double- and triple-vessel
disease) on the basis of the number of affected coronary arteries. The
frequency of patients with variant genotypes decreased gradually
Table 4
Stratified analyses by age and sex for the variant visfatin genotypes in cases and controls.
Variable
Age (median)
<59 years
≥59 years
Sex
Males
Females
Smoking
Yes
No
TT + CT/CC
Adjusteda
Crude
Case
Controls
OR (95%CI)
P
OR (95%CI)
P
44/25
121/67
108/38
106/40
0.62 (0.34–1.15)
0.68 (0.43–1.09)
NS
NS
0.43 (0.21–0.88)
0.68 (0.41–1.12)
0.021
NS
131/79
34/13
158/53
56/25
0.56 (0.37–0.85)
1.17 (0.53–2.58)
0.006
NS
0.49 (0.31–0.77)
1.27 (0.52–3.08)
0.002
NS
67/29
98/63
31/11
183/67
0.72 (0.31–1.66)
0.58 (0.38–0.89)
NS
0.012
0.82 (0.36–1.85)
0.57 (0.37–0.87)
NS
0.009
CI, confidence interval; OR, odds ratio.
a
Adjusted for age, sex, body mass index, smoking status, hypertension, diabetes, dyslipidemia.
J.-J. Yan et al. / Clinica Chimica Acta 411 (2010) 26–30
Table 5
Visfatin genotypes and CAD severity.
CAD severity
SVD (n = 100)
DVD (n = 87)
TVD (n = 70)
P for trenda
Genotype, n (%)
CC
CT + TT
31 (33.7)
29 (31.5)
32 (34.8)
69 (41.8)
58 (35.2)
38 (23.0)
0.058
CAD, coronary artery disease; DVD, double-vessel disease; SVD, single-vessel disease;
TVD, triple-vessel disease.
a
χ2 = 3.601 for trend.
from single- to triple-vessel disease with a borderline significance
(P = 0.058 for trend) (Table 5).
4. Discussion
The present case-control study analyzed the association between
CAD risk and genetic polymorphism at position −1535 in the promoter
of visfatin gene. According to our data, the −1535 C to T variant was
associated with risk reduction of CAD development in a Chinese
population.
Recent studies have also indicated that visfatin, whose plasma
concentrations are altered in obesity and obesity related disorders,
might be involved in the processes of low-grade inflammation [22], an
important component of atherosclerosis. Visfatin might induce
endothelial VEGF and MMP production and activity via multiple
signalling pathways including MAPK, PI3K/Akt, and VEGFR2, suggesting a vital role of visfatin in the pathogenesis of vascular inflammation
[23]. Visfatin has also been shown to be localized to foam cell
macrophages within unstable atherosclerotic lesions, which presumptively play a role in plaque destabilization [10]. In addition,
plasma visfatin levels are significantly correlated with CAD, particularly ACS, independent of well-known CAD risk factors [13].
The −1535C > T polymorphism is located on the distal promoter
region, which contains several CAAT boxes and TATA-like sequences
and binding sites for CCAAT/NF-1, NF- B, NF-IL-6, GR, and AP-1 [14].
More importantly, −1535C > T, was reported to be associated with the
regulation of visfatin gene expression [16,24] and serum lipid level
[17,18].
Then, we hypothesized that the −1535C > T polymorphism might
be correlated well with the susceptibility of CAD. In the present study,
we conducted a hospital-based case-control study with 257 CAD
patients and 292 controls to examine the potential association of Visfatin −1535C> T polymorphism with CAD. A 40% decreased risk of CAD
was observed in subjects with −1535CT/TT compared with the CC
carriers (adjusted OR= 0.60, 95% CI = 0.40–0.89, P = 0.011), indicating
the T allele might confer a protective factor. The study of Ye et al. showed
that the T variant in the T-1535C SNP (described as C-1543T in their
study) resulted in nearly a 2-fold decrease in the reporter gene
expression and transcription of visfatin was reduced in patients with
the T variant, suggesting that the −1535C > T polymorphism is
associated with a decreased risk of acute respiratory distress syndrome
(ARDS) [16]. Thereby, the T allele was expected to have lower levels of
visfatin gene transcription and enzymatic activity than C allele. There is a
strong likelihood that the C to T substitution at this position may reduce
activity of transcription factors (such as NF- B and AP-1), thereby
downregulating visfatin gene transcriptional activity. Recently, they also
confirmed that, compared to its common −1535C allele, the −1535T
variant significantly attenuated its binding to an IL-1β induced
unknown transcription factor in pulmonary vascular endothelial cells,
which may underlie reduced expression of PBEF and lower susceptibility to acute lung injury [24]. Whereas, another investigation among
Japanese suggested that this variation might be not functional [17].
Differences in experimental conditions, ethnicity and geographic
variation may account for this discrepancy. Overall, we could not
29
exclude the possibility that the association between −1535C > T
polymorphism and CAD was due to the effect of this variant on the
expression of visfatin gene, however, more studies are needed to
confirm this hypothesis.
We also estimated the impact of − 1535C > T polymorphism on
the BMI, lipid profiles and glucose levels in CAD patients and controls.
Our data showed that this promoter variant was associated with a
lower serum triglyceride concentration in CAD patients and in
controls. Likewise, Tokunaga et al. also found that the − 1535 T/T
genotype was associated with lower serum triglyceride levels and
higher HDL-C levels in the nondiabetic subjects [17]. Additionally, Jian
et al. also reported that the haplotype containing − 1535C > T
polymorphism was correlated significantly with lipid metabolism
[18]. Bailey et al. observed a remarkable association between the
rs7899066 and rs11977021 variants and dyslipidemia associated with
insulin resistance [25]. Though the precise mechanism by which this
variant influences lipid metabolism remains to be determined, there
are still several plausible explanations. First, it has been shown that
lipid phenotypes (HDL-C, TG) have a linkage with chromosome 7
[26,27], where the visfatin gene is located. It is likely that the T variant
might have an impact on gene transcriptional activity, thus inducing
different lipid phenotypes. Second, the −1535C > T may be in linkage
disequilibrium with other SNPs such as the rs7899066 and
rs11977021 in visfatin promoter, exerting influence on lipid metabolism. Third, visfatin has been reported as the rate limiting component
in the mammalian nicotinamide adenine dinucleotide (NAD) biosynthesis pathway [11,12]. As a result, NAD regulated by visfatin has an
effect on lipid metabolism. However, our study indicated that there
were no significant differences between the genotypes and BMI or
glucose levels in CAD patients and controls. This finding is in
concordance with the observations made by Tokunaga et al. [17]
and Zhang YY et al.[15], revealing that visfatin gene might not be
related to glucose metabolism directly. But Jian et al. revealed that
haplotype containing − 1535C > T polymorphism was significantly
associated with 2-h glucose concentrations [18]. These discrepancies
may be explained by several factors: first, differences in confounding
factors (such as age, drug therapy and lifestyle) due to different
criteria for recruitment; and second, the possibility of this variant's
linkage disequilibrium with other SNPs. Nonetheless, additional
studies are required to further define the results in detail.
According to stratified analyses, we also observed an interaction
between the −1535C > T polymorphism and sex. In male subjects,
those who carried the variant genotypes had a significant reduction in
risk of CAD than CC carriers. Epidemiological data show that the risk of
CAD differs between female and male patients. There are several
possible mechanisms for sex differences in gene expression, such as
imprinting [28] and hormonal effects [29]. Kowalska et al. reported that
visfatin remains an independent predictor of serum testosterone and
free androgen index in the lean polycystic ovary syndrome (PCOS)
subjects [30]. Moreover, testosterone has been found to reduce the
expression of proinflammatory cytokines, such as TNF-a, IL-1, and IL-6 in
human vascular endothelium and monocytes, and imbalance of sex
steroids contributes to adverse cardiac effects in men [31]. Thereby, that
testosterone might synergize the influence of the genetic variants,
would account for this observation, inducing CAD risk reduction in male
subjects.
We also noticed that decreased risk of CAD with the variant visfatin
genotypes was pronounced in younger subjects (age <59 years), but
not in older subjects. Overwhelming accumulated exposure to environmental risks in older individuals may weaken the influence of this
polymorphism in our study. Besides, de Luis et al. [32] and Jin et al. [33]
also found that age has an inverse correlation with serum visfatin
concentrations. It is possible that lower serum visfatin concentrations
might also temper the impact of this polymorphism in the older group.
We noted that among non-smokers, possession of the variant
genotypes had a 43% reduced risk of CAD than CC genotype carriers.
30
J.-J. Yan et al. / Clinica Chimica Acta 411 (2010) 26–30
Exposure to tobacco smoke has been clearly accepted as a major risk
factor for CAD. Excessive exposure to smoking may veil the protective
function of the variant genotypes in our study. Our result suggests that
the − 1535C > T polymorphism may not be a protective factor against
smoking-related CAD in the Chinese population. Nevertheless, this
result might be a coincidence due to the relatively small numbers in
the subgroups.
Our study has several limitations. Firstly, selection bias in the
present study might have affected our results, although, the genotype
distribution of patients and controls in our study was compatible with
the Hardy–Weinberg expectations. Secondly, the relatively small
sample size may underpower the results of our study. Thirdly, our
data were obtained at the time of diagnosis, thus prospectively
followed-up clinical outcome including severe cardiac events may be
required to analyze the association between the − 1535C > T polymorphism and the CAD prognosis. Lastly, our study was performed in
a Chinese population. Data should be extrapolated to other regions
and ethnic groups cautiously. However, this internally consistent pilot
study should provide valuable information to future studies in this
area. This study demonstrates that Visfatin –1535C > T polymorphism
is associated with reduced risk of CAD. Especially in younger, male
subjects and non-smokers, the –1535C > T polymorphism is correlated well with risk reduction of CAD.
Acknowledgments
Project was supported by grants from the Natural Science Foundation of Jiangsu Province (No. BK2007254), Ministry of Personnel of
China for returned student (No. DG216D5021) and the National Natural
Science Foundation of China (No. 30871078).
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