Research Article Open AccessKhalil et al., Int J Genomic Med 2014, 2:1 [601834]

Research Article Open AccessKhalil et al., Int J Genomic Med 2014, 2:1
http://dx.doi.org/10.4172/2332-0672.100011 7
Research Article Open AccessInternational Journal of Genomic Medicine
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
Keywords: Resistin gene polymorphism; Insulin resistance; Type 2
diabetes mellitus
Introduction
In 2010, 285 million people had been diagnosed with diabetes
mellitus worldwide, a prevalence of 6.4%. This is predicted to increase
to 439 million, a prevalence of 7.7% by 2030 [1]. The number of deaths
indirectly linked to diabetes mellitus is estimated to be 3.96 million per
year for all age groups, a prevalence of 6.8% [2].
Insulin resistance is an early and strong determinant of T2DM.
According to the American Diabetes Association 2013 guidelines,
insulin resistance and persistent hyperinsulinemia are found in a variety
of medical conditions, including dyslipidemia and hypertension [3,4].
Resistin is a peptide hormone highly expressed in mouse adipose
tissue [5]. Expression of resistin appears to increase along with
adipocyte differentiation [6]. In humans, the expression pattern of
resistin is not totally consistent with the expression in mice and resistin
is known to be expressed at higher levels in pre-adipocytes than
in mature adipocytes [7]. In addition to fat tissue [8] resistin is also
expressed in human peripheral blood monocytes [9].
Resistin has been shown to cause “high levels of Low-Density
Lipoprotein (LDL); increasing the risk of heart disease [10]. Resistin
increases the production of LDL in human liver cells and also degrades
LDL receptors in the liver. As a result, the liver is less able to clear LDL-
cholesterol from the body. Resistin accelerates the accumulation of
LDL in arteries, increasing the risk of heart disease. Resistin adversely
impacts the effects of statins, the main cholesterol-reducing drug used
in the treatment and prevention of cardiovascular disease [11].
Resistin plays a role in the regulation of energy, glucose, and
lipid homeostasis [12] and the maintenance of fasting blood glucose
levels [13] by modulating hepatic insulin action [14]. Resistin has low
circulating levels [15], but in some studies it has been reported to be upregulated in insulin resistance, T2DM, and Cardiovascular Diseases
(CVD) [16].
Some studies have reported increased resistin expression levels
in type 2 diabetes mellitus, insulin resistance or obesity, Metabolic
Syndrome (MS), and cardiovascular disease [17], while some failed to
detect any change in resistin levels in these conditions [18].
Resistin may increase the susceptibility of Metabolic Syndrome
(MS) by regulating adiponectin secretion from adipocytes and
enhancing hepatic gluconeogenesis by inhibiting the enzymes
involved in gluconeogenesis through AMP-activated protein kinase
activation [19]. Another study reported that subjects with premature
atherosclerosis have higher levels of plasma resistin compared with
subjects with established atherosclerosis [20].
Resistin gene (RETN) located on chromosome 19p13.3 and up to
70% of the variation in serum resistin levels explained by genetic factors.
RETN, a gene coding for resistin, is located at chromosome location
19p13.3 [21]. Several single-nucleotide polymorphisms (SNPs) have
been described in the RETN promoter, intron and 3’UTR (un-translated
region) regions. Only one non-synonymous substitution (C63R) in the
coding region of exon 3 has been reported. Due to its frequency (<2.0%),
this polymorphism has not been studied further [22].
*Corresponding author: Alsayed alnahal, Internal Medicine Department, Zagazig
University Hospital, Egypt, Tel: 00201113174798; E-mail: alnahal2002@yahoo.com
Received April 18, 2014 ; Accepted June 25 , 2014 ; Published July 07 , 2014
Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014)
Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin Resistance
in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi:
10.4172/2332-0672.100011 7
Copyright: © 2014 Khalil O, et al. This is an open-access article distributed under
the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
source are credited.Abstract
Background: Insulin Resistance (IR) is a condition which precedes the development of type 2 diabetes mellitus
(T2DM). Resistin is a hormone secreted by adipocytes. Resistin gene (RETN) polymorphisms has been found to be
associated with obesity and insulin resistance. We choose 60 patients have known type 2 diabetes mellitus against
45 healthy subjects to investigate the relationship between RETN +299 gene polymorphisms and insulin resistance,
in non-obese patients with T2DM.
Results: The present study revealed statistically significant increase in AA and combined GA+AA genotypes
(with ODD Ratio 4.04 and 4.75 respectively), and statistically significant decrease in GG genotype in non-obese
T2DM as compared to the control subjects. Also we found statistically significant increase in A allele and serum
resistin in T2DM group as compared to the control group. In addition there were statistically significant increase
in mean value ± SD of fasting blood glucose, insulin, HOMA-IR, HbA1C%, Resistin, total cholesterol, triglyceride,
LDL-C and statistically significant decrease in mean value ± SD in serum HDL-C in AA combined AA+GA subgroups
as compared to GG subgroup of T2DM group.
Conclusions: Our study has shown that resistin +299G/A is an important genetic regulator that resulted
hyperresistinemia and subsequently may be predisposing factor the development of T2DM in non-obese Egyptian
population.Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption Non-obese Type 2 Diabetes?
Osama Khalil1, Alsayed Alnahal1*, Mohamed Ghonium1, Samy Fawzy2, Magdy Ibrahem2,Nermin Raafat2 and Walaa Samy2
1Department of Internal Medicine, Zagazig University Hospital, Egypt
2Department of Medical biochemistry, Zagazig university Hospital, Egypt

Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
Page 2 of 7
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
Results of different studies investigating genetic variations in the
RETN were controversial. [23].
The allele and genotype frequencies at several RETN SNPs between
subjects with Type 2 diabetes and healthy controls have been compared
[24] and no difference has been observed, except in one study. Tan
et al. reported that subjects with Type 2 diabetes have more often allele
A than allele G compared with healthy controls in 3’UTR +62G>A
SNP[25]. RETN polymorphisms have also been associated with other
obesity-related phenotypes. An association has been found with blood
pressure [26], Body Mass Index (BMI), body weight and body fat
percentage, serum triglycerides, fasting blood glucose concentration
and lower insulin resistance index [24].
Therefore, we aimed to investigate the relationship between RETN
+299 gene polymorphisms and insulin resistance, in non-obese patients
with T2DM.
Subjects and Methods
Subjects
This study is observational, case contro l study included (105)
unrelated subjects recruited from diabetes and endocrinology
outpatient clinic of Internal Medicine Department of Zagazig University
Hospitals . Subjects were classified into two groups; Group I: include 60
non-obese patients known Type 2 diabetes mellitus (T2DM) and non-
complicated, (31 males and 29 females) with age ranged from 40-60
with a mean value ± SD of 50.8 ± 6.33 years).24 of them were on anti
hyperlipidemic drugs, they were on diet control in addition to oral
hypoglycemic drugs. 22 of them received lipid lowering drugs (e.g.
statins).Group II : include 45 apparently healthy patients (non-diabetic
group) as control group, who were (23 males and 22 females) matched
to cases by gender and ethnic origin. We exclude the obese participants
with BMI>25 kg/m2, uncontrolled hypertension (BP>160/100), liver,
kidney, thyroid, cardiovascular orany active inflammatory diseases, as
well as receiving medications that affect glucose metabolism(except in
T2DM group). The ethical committee of Faculty o f Medicine, Zagazig
University approved our study protocol, and all participants assigned
written informed consent.
Methods
All patients were subjected to thorough history taking including
family history of diabetes, complete physical and full clinical assessment
including vital signs and anthropometric variables (BMI was calculated
as weight in kg/ height in (meters).
Biochemical analysis: Routine investigations which include
(Complete blood picture, Liver and Kidney function tests, admission
blood glucose (RBG). fasting blood glucose (FBG), HbA1c [7], serum
electrolytes, urine analysis, fasting Lipid profile
According to Friedewald formula, LDL cholesterol was calculated
from the following [27]: LDL cholesterol = total cholesterol − (HDL
cholesterol + 0.47 triglycerides).
Fasting serum insulin by ELISA
Determination of insulin resistance (IR) by Homeostasis model
assessments of insulin resistance (HOMA-IR) were performed and
calculated by the following formula: HOMA index = [Fasting insulin
(μU/ml) ×fasting plasma glucose (mmol/l)]/22.5. Subjects were
considered insulin resistance if HOMA was greater than 1.64 [28].Specific investigations
a. Serum resistin was measured by ELISA kit provided by (Biosource
Europe S.A., Nivelles, Belgium).
b. Genomic DNA extraction
DNA was isolated and purified from whole blood (EDTA) using
QIAamp-spin-columns according to the protocol provided by the
manufacturer (QIAamp Blood Kit; Qiagen GmbH, Hilden, Germany).
DNA was stored at −20°C till the time of use.
c. Polymorphism analysis of the RETN +299G>A (rs3745367)
Determination of +299 resi stin g ene polymorphism by PCR
amplification according to Kunnari et al, followed by restriction
site enzyme digestion and gel electrophoresis [30] (Figures 1 and
2). An intron variant, at position +299 (G>A) (rs3745367) from a
translation start site, in RETN gene was amplified using forward
primer 5’-GAGAGGATCCAGGAGGTCG-3’ and the reverse primer
5’-GTGAGACCAAACGGTCCCT-3’ . The amplification was carried
out in a total 50 μl volume reaction contained 50 ng of genomic DNA,
0.2 μM of each primer (Promega, Madison, WI) and 1× PCR mix (Taq
PCR Master Mix Kit, QIAGEN, GmbH, Hilde n, Germany). PCR was
Figure 1: Picture of gel electrophoresis showing amplified PCR product of
resistin gene sequence of 373 bp length. NC: Negative control, M: (100 bp –
1.5 kb) DNA ladder.
Figure 2: Picture of gel electrophoresis showing G Allele and A Allele variant of
+299 G/A resistin gene polymorphism. NC: Negative control, M: (100 bp – 1.5
kb) DNA ladder.

Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
Page 3 of 7
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
significant at P<0.05. All data were evaluated usi ng statistical package
for social sciences (SPSS) for windows version 17.
Results
Our study included 2 groups: T2DM group which included 60
patients , against 45 healthy subjects (control group), both of them were
age and sex matched ,no statically significant difference as regards to
Body mass index , systolic and diastolic blood pressure as shown in
Table 1.
Using student t- test (unpaired), there were statistically significant
increase in the mean value ± SD of fasting blood glucose, fasting serum
insulin , HOMA-IR Total cholesterol ,triglyceride, LDL, HbA1C % and
serum Resistin also statistically significant decrease in the mean value ±
SD of serum HDL between T2DM group and healthy control as shown
in Table 1.
In Table 2,we studied distribution of Resistin SNP genotype and
alleles in both groups, we found the frequencies of RETN+299 AA
genotypes were significantly increased inT2DM patients compared to
control group (23(38.4%) versus 6(13.4%). Carriers of the AA genotypes
were significantly more likely to develop T2DM (O.R=4.041, 95%
CI=1.47- 11.04, P=0.004). The frequency of the A allele of RETN +299
was increased in T2DM patients as compared to control (62.5% versus
35.6%), and this difference was statistically significant (p=0.0001).
A GA genotype frequency was increased in T2DM as compared to performed as follows: denaturation at 95°C for 5 min, preannealing at
59°C for 1 min, and then elongation at 72°C for 2 min followed by 35
cycles of 30 s at 95°C, 30 s at 59°C and1 min 15 s at 72°C, and elongation
at 72°C for 10 min. The 173-bpPCR product w as digested with 5 U AluI
(MBI-Fermentas, United Kingdom) at 37°C for 16 h. The digestion
products were separated on a 2% agarose gel stained with of ethidium
bromide to visualize the bands. Quality control measures include
blinded analyses and replicates of 10% of samples, and negative controls
for contamination (no DNA) were run routinely with patient samples.
Blood sampling: Five ml of fasting peripheral venous blood sample
were withdrawn from each subject under complete aseptic conditions
and divided into 3 portions: 1 ml of whole blood was collected into
evacuated tubes containing EDTA, for genomic DNA extraction
andHbA1c; 1 ml of whole blood was collected into evacuated tubes
containing fluoride for fasting blood glucose. Serum were separated
immediately from remaining part of the sample and stored at -20°C
until analysis.
Statistical analysis: The results were expressed as the mean ±
SD and were analyzed using Student’s t-test, ANOV A. The statistical
significances of differences in the frequencies of variants between the
groups were tested using the χ2 test. In addition, the Odds Ratios (ORs)
and 95% confidence interval (95% CI) were calculated as a measure
of the association of the RETN +299G>A genotypes with insulin
resistance and development of T2DM.A difference was considered
Parameter Control
(n = 45)T2DM Patients
(n = 60)t p
Age (years) 51.26 ± 5.54 50.8 ± 6.22 0.387 0.6996
Gender
Male
Female 20 (44%)
25 (56%)22 (40%)
28 (60%)Chi=0.32
P=0.32
SBP (mmHg) 124.2 ± 4.6 125.1 ± 4.8 -0.94 0.534
DBP(mmHg) 76.5 ± 4.1 75.8 ± 3.8 0.861 0.39
B.M.I(kg/m2) 23.8 ± 1.02 23.2 ± 1.8 -1.06 0.29
Creatinine (mg/dl) 0.78 ± 0.152 0.75 ± 0.13 1.023 0.30
Fasting blood glucose (mg/dL) 89.7 ± 10.2 195.7 ± 55.9 -14.33 <0.001
Fasting serum insulin (μU/dl) 7.42 ± 1.92 12.43 ± 5.6 -6.38 <0.001
Total cholesterol (mg/dl) 182.32 ± 16.24 204.8 ± 33.2 4.89 <.001
Triglycerides (mg/dl) 180.36 ± 14.53 205.6 ± 32.3 -5.30 <0.001
LDL (mg/dl) 100.70 ± 20.16 121.8 ± 33.6 -3.73 <0.001
HDL (mg/dL) 45.36 ± 6.2 40.6 ± 6.95 3.581 <0.001
HOMA-IR 1.43 ± 0.48 3.46 ± 2.8 -5.496 <0.001
HbA1C % 5.16 ± 0.67 9.24 ± 0.1 -10.41 < 0.001
Resistin (ng/ml) 1.34 ± 0.3 4.5 ± 1.2 -19.59 < 0.001
SBP: Systolic Blood Pressure; DBP: Diastolic Blood Pressure
Table 1: Comparison of some socio-demographic data, clinical and biochemical characteristics of the studied subjects.
Control
(n = 45)T2DM Patient
(n = 60)χ2POdd ratio
O.R(95%CI)
GG 19(42.2%) 8(13.3%) 11.13 0.001 0.210 0.08136-0.5448
GA 20(44.4%) 29(48.3%) 0.154 0.69 1.169 0.54-2.54
AA 6(13.4%) 23(38.4%) 7.96 0.004 4.041 1.47-11.04
GA+AA 26(57.8%) 52(86.7%) 11.3 0.0004 4.75 1.84-12.29
G allele 58(64.4%) 45(37.5%)
14.87 0.0001 3.021 1.711-5.332 A allele 32(35.6%) 75(62.5%)
Table 2: Frequency and distribution of resistin SNP genotype and alleles frequencies in diabetics’ patients and controls.

Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
Page 4 of 7
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
the control (48.3% and 44.4% respectively) but this difference was
statistically non-significant. In Table 3, we divided each of the control
and T2DM into 3 subgroups according to genotype frequencies and we
compared the mean value ± SD of the biochemical parameters, there
were statistically significant difference As regards to mean value ±
SD of fasting blood glucose, fasting serum insulin, HOMA-IR- Total
cholesterol, triglyceride, LDL-C, HDL-C, HbA1C % and serum Resistin
in AA subgroups (of T2DM group) as compared to both GG and GA
subgroups. As well as there were statistically significant difference when
comparing GG and GA subgroups as fasting blood glucose, fasting
serum insulin, HOMA-IR, triglyceride, LDL-C, HbA1C % and serum
Resistin. No statistically significant differences were detected as regards
to the biochemical parameters when comparing AA, GA, GG subgroups
in control group. Using student t test we found statistically significant
increase in mean value ± SD of serum total chole sterol, triglyceride,
LDL, fasting blood glucose (mg/dL) Fasting blood insu lin (μU/ml),
HOMA-IR, HbA1C % and serum Resistin, as well as decrease in mean value ± SD of HDL in combine d GA+ AA subgroup as compared to
GG subgroup of T2DM Patients in Table 4. Table 5 showed multiple
regression analysis of when resistin gene polymorphism +299 (G>A)
was used as dependent variable and, HbA1c, HOMA-IR, cholesterol,
triglyceride, HDL-c, as well as LDL-c were taken as independent
variables. Our results revealed that RETN serum resistin (P=0.001),
was independently associated with resistin gene polymorphism +299
(G>A).
Discussion
Egypt has a high prevalence rate of diabetes 15.56%, with high
incidence in urban than the rural areas, and obesity 20% in urban areas
[31].
Up till now, there are contradictions among the results of studies
investigating genetic variations in resistin, including Single Nucleotide
Polymorphisms (SNPs)
GG GA AA F value significance
Total cholesterol(mg/dl) control 182 ± 16.2 180 ± 17.8 183 ± 14.56 0.081 p=.92
T2DM Patient 185 ± 23200 ± 22
-229 ± 37
ab 10.27 P<0.001
Triglycerides(mg/dl)control 176 ± 14 177 ± 15 181 ± 13 0.277 p=.75
T2DM Patient 186 ± 18203 ± 23
a230 ± 34
ab10.55 p<0.001
LDL (mg/dl)control 101.18 ± 8.21 99.7 ± 8.3 101.6 ± 6.39 0.22 p=.80
T2DM Patient 103.8 ±
13.2121.6 ± 11.1 a145.9 ± 23
ab 23.11 p<0.001
HDL (mg/dL) control 45.62 ± 5.44 44.9 ± 6.5 45.2 ± 5.66 0.068 p=.93
T2DM Patient 44 ± 5.4 38.2 ± 6.337.1 ± 7.2
a 3.56 p<0.05
Fasting blood glucose (mg/dL)control 89.7 ± 9.2 90.7 ± 8.2 89.8 ± 10.5 0.005 p=.99
T2DM Patient 172 ± 45 190.7 ± 50.9 a220 ± 51.3
ab 3.77P<0.05
Fasting blood insulin (μU/ml) Control 4.4 ± 1.85 4.5 ± 1.76 6.0 ± 1.79 1.93 p=0.157
T2DM Patient 6.8 ± 2.22 11.7 ± 2.34
a20.75±2.44
ab143.04p<0.001
HOMA-IRControl 1.07 ± 0.55 1.21 ± 0.46 1.29 ± 0.84 0.76 p=0.47
T2DM Patient 2.61 ± 0.695.9 ± 1.08
a11.27 ± 2.18
ab 307.23p<0.001
HbA1C %Control 5.06 ± 0.47 5.16 ± 0.37 5.10 ± 0.57 0.25 P=0.77
T2DM Patient 6.60 ± 1.237.68 ± 1.58
a9.68 ± 2.38
ab 10.41 P<0.001
Resistin(ng/ml)Control 1.30 ± 0.29 1.29 ± 0.4 1.32 ± 0.35 .0195 P 0.98
T2DM Patient 2.50 ± 1.023.44 ± 1.22
a4.46 ± 1.6
ab
ab 7.24P <0.001
Table 3: Statistical analysis of biochemical parameters in association with genotype distribution in studied groups.
GG AA +GA T value p-value
Total cholesterol(mg/dl) 185 ± 23 215 ± 30 3.29 <0.001
Triglycerides(mg/dl) 186±18 216.5 ± 29 4.05 <0.001
LDL(mg/dl) 103.8 ± 13.2 134 ± 17 5.77 <0.001
HDL(mg/dL) 44 ± 5.4 37.65±6.6 -2.99 <0.05
Fasting blood glucose(mg/dL) 172 ± 45 206 ± 51 2.139 <0.05
Fasting blood insulin (μU/ml) 6.8 ± 2.22 16.2 ± 2.39 11.3 <0.0001
HOMA-IR 2.61 ± 0.69 8.85 ± 1.63 18.76 <0.001
HbA1C % 6.60 ± 1.23 8.68 ± 1.98 4.044 <0.001
Resistin (ng/ml) 2.50±1.02 3.95 ± 1.41 3.53 <0.001
Table 4: Statistical analysis of biochemical parameters in GG subgroup as compared to combined GA+AA subgroup (in T2DM group).

Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
Page 5 of 7
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
Some authors reported the genetic variant at nucleotide+299
(G>A) and obesity as a risk factors for T2DM in Caucasians [32]. As
well as, some studies have found associations between resistin gene
polymorphism, and T2DM. Therefore, the genetic variations of the
resistin gene in humans require clarification. Previous studies tried to
clarify the functions of resistin and stated that resistin might impair
insulin action, glucose tolerance, and decreased glucose uptake in
skeletal muscle cells [33].
Previous study found that resistin was highly expressed in human
omental and abdominal subcutaneous adipocytes than in adipocytes
from the thigh, suggesting a possible role of resistin in obesity related
insulin resistance [34]. Most of pa tients with T2DM were related to
obesity and insulin resistance. Some authors stated that serum resistin
levels were increased in T2DM subjects [35]. Other studies have
reported no association between resistin levels and markers of insulin
resistance in T2DM patients [36]. Such findings have been re-tested by
this study.
The purposes of the current study were to investigate the possible
associations of RETN +299 gene polymorphism with insulin resistance
in non-obese subjects with T2DM, and to detect whether this
polymorphisms is associated with glucose intolerance. The present
study revealed statistically significant increase in AA genotype and
combined GA+AA (with ODD Ratio 4.04and 4.75 respectively), and
statistically significant decrease in GG genotype in non-obese T2DM as
compared to the control subjects, The distribution of +299 resistin allele
frequencies G and A was 37.5% and 62.5% respectively in the diabetic
group whereas in the control group was 64.4% and 35.6% respectively.
There was a statistically significant association of A allele in the diabetic
group when compared with the control group (P=0.0001).
Our results are suggesting an association between the presence
of the polymorphism and the presence of T2DM. In agreement with
Miyamoto et al. and Suriyaprom et al. who reported an association
of RETN +299 with T2DM among Japanese and Thais subjects,
respectively [20-23]. Also Tan et al. reported that subjects with Type
2 diabetes have more often allele A than allele G compared with
healthy controls [25]. Serum resistin was statistically increased in
both AA and GA subgroups as compared to the GG subgroup of
type 2 diabetics, also in combined AA+GA subgroup as compared
to GG subgroup of diabetic group. Our results agreed with Asano et
al. [37] they have shown that the plasma resistin concentration was
strongly influenced by the rs34861192 and rs374536 8 polymorphisms
of RETN in an aged Japanese population. Also this goes in agreement
with the results obtained by El-Shal et al. who reported that the
frequency of the RETN +299 AA genotype was significantly increased
in obese diabetic patients compared to control group, the odds ratio (OR=3.53, P=0.005) suggested an association between the presence
of the polymorphism and the prevalence of disease. They revealed
that the genotypes and alleles of resistin polymorphisms +299(G>A)
were significantly associated with increased risk of impaired glucose
tolerance and T2DM compared to normal glucose tolerant patients
[38].
This polymorphism is in an intron, +299 (G>A), which generally has
not been considered to have regulatory functions. However, it has been
shown that SNPs in the non-coding region, such as the 3’-untranslated
gene region, canaffect gene expression [39]. Tsukada et al. said that intron
polymorphism of TFAP2B, a susceptibility gene to T2DM, influence
adipocytokinegene expression transcriptional activity [40]. Suriyaprom
et al. concluded that resistin gene polymorphism at +299 (G>A) maybe
a marker in linkage disequilibrium with other polymorphism affecting
gene expression and may contribute to increased resistin levels in Thai
diabetic subjects, which may be involved in the pathogenesis of type 2
diabetes by impaired insulin action [23].
Ma et al. demonstrated that, a resistin genotype at position +299
(G>A) and obesity was a significant determinant of type 2 diabetes risk
in Caucasians [32]. Also, the G/A or A/A genotype of SNP +299 G>A
was found to be associated with increased risk of metabolic syndrome
in a Japanese cohort study [20], but h ad a protective effect against
hypertension in a Finnish population based cohort study [41].
Whereas resistin gene +299 (G>A) was not associated with type
2 diabetes in Japanese subjects [42]. This polymorphism is in an
intron, +299 (G>A), which generally has not been considered to have
regulatory functions. However, it has been shown that SNPs in the non-
coding region, such as the 3’-untranslated gene region, can affect gene
expression [43].
The contradictions of resistin gene polymorphism among different
studies might be explained by the different genetic backgrounds or
environmental conditions of the population studied [37].
The present study reveals that there was a statistically significant
association between hyperglycemia and resistin gene polymorphism at
positions +299 (G>A). These findings cope with Suriyaprom, et al. [23].
Conflicting findings between these studies could be due to true
differences in allelic association with the disease phenotype in different
populations. In agreement with this notion were the differences in allele
frequencies of these SNPs in various populations.
Lau CH, et al. demonstrated in their study that SN P+299 G>A at
the RETN locus appeared to affect the serum resistin concentration,
but their association with the risk of T2DM as well as metabolic risk
factors might not be due to alterations in resistin level only, but also Model Unstandardized CoefficientsStandardized
Coefficients t Sig. 95% Confidence Interval for B
B Std. Error Beta Lower Bound Upper Bound
(Constant) -1.665 1.455 -1.144 0.258 -4.585 1.256
cholesterol 0.007 0.004 0.189 1.869 0.067 0 0.014
Triglyceride -0.003 0.003 -0.083 -0.848 0.4 -0.01 0.004
HDL-c 0.013 0.021 0.059 0.609 0.545 -0.03 0.056
LDL-c -0.003 0.003 -0.099 -1.009 0.317 -0.009 0.003
HOMA -0.006 0.044 -0.014 -0.143 0.887 -0.095 0.082
HbA1c% -0.017 0.059 -0.029 -0.281 0.78 -0.135 0.102
Resistin 0.696 0.102 0.721 6.848 0 0.492 0.9
Table 5: Multiple regression analysis of when resistin gene polymorphism +299 (G>A) was used as dependent variable and , HbA1c, HOMA-IR, cholesterol,triglyceride,HDL-c,
as well as LDL-c.

Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
Page 6 of 7
Volume 2 • Issue 1 • 1000117Int J Genomic Med
ISSN: 2332-0672 IJGM, an open access journal
other mechanisms than an effect on serum resistin levels by which
SNP+299 G>A at the RETN locus could be involved in the association
with susceptibility to T2DM [44].
Resistin circulates in two distinct assembly states, trimers and
hexamers, Hexamersis formed by intertrimer disulfide bonds. Ethnicity
and metabolic changes possibly modulate resistin action by affecting its
assembly states [45].
In this study, a significant increase in indices of insulin resistance
(including fasting blood glucose and HOMA index) was increased in
T2DM group as compared to control group (p<0.001). According to
genotype in patient group, subjects with AA and combined GA+AA
genotypes have a statistically signific ant increase of fasting blood
glucose, serum insulin and HOMA index compared to subjects with
GG genotype.
This finding confirmed the results of El-Shalet al. who found a
significant increase in HOMA-IR, fasting blood glucose level, serum
insulin, among T2DM and obese subjects with GA, AA genotypes of
RETN +299 when compared to GG and CC genotypes, respectively
[38]. But all subjects of this study with average body mass index,
because they were on diet control regimen for treatment of T2DM.
Our results revealed statistically significant increase in HbA1c
(%) in both AA and GA subgroups as compared to GG subgroup of
diabetic group coinciding with uncontrolled diabetes. In summary, we
found statistically significant increase in AA and combined GA+AA
genotypes, and decrease in GG genotype in non-obese T2DM as
compared to the control subjects. As well as statistically significant
increase in A allele and serum resistin in T2DM group as compared
to the control group. Resistin +299G/A polymorphism is in an intron,
which generally has not been considered to have regulatory functions.
However, it has been shown that SNPs in the noncoding region, such as
the 3’-untranslated gene region, can affect gene expression [46].
In conclusion, the results of our study supported the suggestion
thatRETN+299 G>A SNP contributed to increase the susceptibility and
the development of type 2 diabetes mellitus. This conclusion is based on
the observation that our results among diabetic patients demonstrated
elevated fasting blood glucose level, serum insulin level and HOMA-
IR in both AA /GA carriers of RETN +299.Thismay be considered
as genetic biomarker for type 2 diabetes in non-obes e with potential
impact on the family counseling and management.
Further studies are needed to fully clarify the role of resistin gene in
type 2 diabetes, by investigation of other populations including different
ethnic groups, how resistin polymorphism at +299 (G>A) affects gene
expression and to compare both resistin gene polymorphism and serum
resistin level in obese and non-obese patients with T2DM.
Limitation
This study was carried out on a small sample of type 2diabetes of
Egyptiation citizen, and studied RETN +299 only. Some diabetics were
not obese due diet regimen after diagnosis of diabetes. A larger study in
needed to compare other genetic polymorphism e.g RETN-420, as well
as obese and non-obese type 2 diabetes, considering also environmental
factor in urban and rural areas in Egypt.
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Citation: Khalil O, Alnahal A, Ghonium M, Fawzy S, Ibrahem M, et al. (2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2: 117. doi: 10.4172/2332-0672.100011 7
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(2014) Does Resistin Gene Polymorphisms +299 (G>A) Participate in Insulin
Resistance in Egyption non-Obese Type 2 Diabetes? Int J Genomic Med 2:
117. doi: 10.4172/2332-0672.100011 7