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In this cross-sectional study a
total of 453 genetically unrelated adult subjects of both genders, ranging from
18 to 60 years of age, were enrolled from our previous
study in which the relationship of two
LXR? SNPs with obesity was investigated on individuals who were randomly
selected from participants of a cohort study
of metabolic syndrome and cardiovascular disease risk
factors performed in Mashhad as a second largest city in Iran (15). The world
health organization-body mass index (WHO-BMI) was calculated as a weight to height ratio (kg/m2), and
a BMI of 18.5-24.99, 25–29.99 and ?30 kg/m2
were taken as cut-off values defining normal weight, overweight and obesity,
respectively. With this in mind, all subjects were classified into three
groups: 209 subjects as overweight, 168 subjects as obese, and 76 healthy
individuals as normal weight. Before participating
voluntarily in research work, informed written
consent was obtained from all participants. After
that, a complete
physical assessment and anthropometric measurements were performed by trained
health workers. Blood samples were drowned after a fasting period, and data on age, sex, geographical birthplace, ethnicity, past medical and
family history, and dietary habits were
collected. All
subjects with a history of Stroke, MI (Myocardial infarction),
endocrinological abnormalities, diabetes mellitus, alcohol consumption, heart,
liver and/or renal disease, or those who were under
high blood pressure medication and lipid or
glucose lowering treatment were excluded from the study. This research project was approved by the Ethical Committee of the Shahid Beheshti
University of Medical Science (SBUMS: Research Project No. 648).


2.2. Anthropometric and
Biochemical measurements

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Anthropometric and clinical measurements were measured according to
international standard procedures as described in
details in our previous studies (11, 15).
In brief, all parameters including height, weight, waist circumference
(WC), hip circumference (HC), waist/hip ratio, systolic blood pressure (SBP)
and diastolic blood pressure (DBP), total cholesterol (TC), high- density
lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C),
triglycerides (TG), fasting blood glucose (FBG) and serum C-reactive protein
(CRP) were measured.


2.3. Genotyping

Genomic DNA was isolated from
the peripheral blood sample with FlexiGene DNA Kit (Qiagen) in accordance with the manufacturer’s
instructions. Genotyping of single
nucleotide polymorphisms (SNPs) was performed by allelic discrimination
assays using TaqMan probes (C_16059177_10; Applied
Biosystems, USA). Allelic discrimination was performed
on Applied Biosystems 7500 real-time PCR System. The following conditions
were used for the polymerase chain reaction: initial denaturation step at 95°C for 10 minutes, and 40 cycles of 92°C for 15 seconds and 60°C
for 1 minute. Approximately 10% of the samples were regenotyped to ensure reproducibility
and accuracy.


2.4. Statistical analysis

All statistical analysis was performed using SPSS v.16.0 (SPSS Inc., Ill., USA). The descriptive statistics
were determined for all variables and are presented
as mean (SD) for the normally distributed variables (or as the median (IQR) for
non-normal variables). The chi-square
or Fisher’s exact tests were applied to evaluate differences in allele and genotype
distribution among the examined groups. It was also used to assess any deviation from the
Hardy-Weinberg equilibrium (HWE) for the study SNP. Independent-samples
t-test was used to compare the means of demographic
and clinical variables between independent groups. Analyses
of variance (ANOVA) followed by the
Bonferroni posthoc test, was performed to
compare anthropometric traits for differences
between genotype groups. Using data from our previous report
regarding the other common variant (rs17373080) of
LXR? gene (16), haplotype
frequencies for two sites and any potential relationship with excess weight were
estimated using the SNPStats program (17). Logistic regression analyses were used to calculate the
odds ratios (ORs) with 95% con?dence interval (CI) of
each related parameter, both in crude or adjusted for age and gender.
All the analyses were two-sided and statistical significance was set at p-value of < 0.05.

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