مجال
التميز
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تميز دراسي و بحثي
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البحوث المنشورة
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البحث (1):
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عنوان البحث:
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Identification of the Functional
Variant(s) that Explain the Low-Density Lipoprotein Receptor (LDLR) GWAS SNP
rs6511720 Association with Lower LDL-C and Risk of CHD
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رابط إلى البحث:
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Click Here
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تاريخ النشر:
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2016
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موجز عن البحث:
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Background
The Low-Density Lipoprotein
Receptor (LDLR) SNP rs6511720 (G>T), located in intron-1 of the
gene, has been identified in genome-wide association studies (GWAS) as being
associated with lower plasma levels of LDL-C and a lower risk of coronary
heart disease (CHD). Whether or not rs6511720 is itself functional or a
marker for a functional variant elsewhere in the gene is not known.
Methods
The association of LDLR SNP
rs6511720 with incidence of CHD and levels of LDL-C was determined by
reference to CARDIoGRAM, C4D and Global lipids genetics consortium (GLGC)
data. SNP annotation databases were used to identify possible SNP function
and prioritization. Luciferase reporter assays in the liver cell line Huh7
were used to measure the effect of variant genotype on gene expression.
Electrophoretic Mobility Shift Assays (EMSAs) were used to identify the
Transcription Factors (TFs) involved in gene expression regulation.
Results
The phenotype-genotype analysis
showed that the rs6511720 minor allele is associated with lower level of
LDL-C [beta = -0.2209, p = 3.85 x10-262], and lower risk of CHD
[log (OR) = 0.1155, p = 1.04 x10-7]. Rs6511720 is in complete
linkage. Rs6511720 is in complete linkage disequilibrium (LD) with three
intron-1 SNPs (rs141787760, rs60173709, rs57217136). Luciferase reporter
assays in Huh7 cells showed that the rare alleles of both rs6511720 and rs57217136
caused a significant increase in LDLR expression compared to
the common alleles (+29% and +24%, respectively). Multiplex Competitor-EMSAs
(MC-EMSA) identified that the transcription factor serum response element
(SRE) binds to rs6511720, while retinoic acid receptor (RAR) and signal
transducer and activator of transcription 1 (STAT1) bind to rs57217136.
Conclusion
Both LDLR rs6511720
and rs57217136 are functional variants. Both these minor alleles create
enhancer-binding protein sites for TFs and may contribute to increased LDLR expression,
which is consequently associated with reduced LDL-C levels and 12% lower CHD
risk.
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البحث (2):
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عنوان البحث:
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Identifying low density
lipoprotein cholesterol associated variants in the Annexin
A2 (ANXA2) gene
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رابط إلى البحث:
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Click Here
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تاريخ النشر:
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12 April 2017
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موجز عن البحث:
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Background and aims
Annexin-A2 (AnxA2) is an endogenous
inhibitor of proprotein convertase subtilisin/kexin type-9 (PCSK9). The
repeat-one (R1) domain of AnxA2 binds to PCSK9, blocking its ability to
promote degradation of low-density lipoprotein cholesterol-receptors (LDL-R)
and thereby regulate low-density lipoprotein cholesterol (LDL-C) levels. Here
we identify variants in ANXA2 influencing LDL-C levels and we determine the
molecular mechanisms of their effects.
Results
The ANXA2 single nucleotide polymorphism
(SNP) genotype-phenotype association was examined using the
Second-Northwick-Park Heart Study (NPHSII) (n∼2700) and the UCL-LSHTM-Edinburgh-Bristol
(UCLEB) consortium (n∼14,600). The
ANXA2-R1 domain coding-SNP rs17845226 (V98L) associated with LDL-C,
homozygotes for the minor allele having ≈18.8% higher levels of LDL-C (p =
0.004), and higher risk of coronary heart disease (CHD) (p = 0.04). The SNP
is in modest linkage disequilibrium (r2 > 0.5) with two intergenic SNPs,
rs17191344 and rs11633032. Both SNPs showed allele-specific protein binding,
and the minor alleles caused significant reduction in reporter gene
expression (≈18%, p < 0.001). In the expression quantitative trait loci
(eQTL) study, minor allele homozygotes have significantly lower levels of
ANXA2-mRNA expression (p = 1.36 × 10−05).
Conclusions
Both rs11633032 and rs17191344
SNPs are functional variants, where the minor alleles create
repressor-binding protein sites for transcription factors that contribute to
reduced ANXA2 gene expression. Lower AnxA2 levels could increase plasma
levels of PCSK9 and thus increase LDL-C levels and risk of CHD. This
supports, for the first time in humans, previous observations in mouse models
that changes in the levels of AnxA2 directly influence plasma LDL-C levels,
and thus implicate this protein as a potential therapeutic target for LDL-C lowering.
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البحث (3):
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عنوان البحث:
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The Genetic Spectrum of Familial
Hypercholesterolemia (FH) in the Iranian Population
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رابط إلى البحث:
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Click Here
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تاريخ النشر:
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2017
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موجز عن البحث:
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Familial hypercholesterolemia
(FH) is an autosomal dominant disorder associated with premature
cardiovascular disease (CVD). Mutations in the LDLR, APOB,
and PCSK9 genes are known to cause FH. In this study, we
analysed the genetic spectrum of the disease in subjects from the Iranian
population with a clinical diagnosis of FH. Samples were collected from 16
children and family members from five different cities of Iran. Probands
were screened for mutations in the LDLR, APOB,
and PCSK9 genes using next generation sequencing, with
results confirmed by Sanger sequencing. The likely pathology of identified
variants was examined using in silico tools. Of the
probands, 14 had a clinical diagnosis of homozygous FH and two of
heterozygous FH. No mutations were found in either APOB or PCSK9,
but nine probands were homozygous for seven different LDLR mutations,
with p.(Trp577Arg) occurring in three and p.Val806Glyfs*11 occurring in two
patients. Two mutations were novel: p.(Leu479Gln) and p.(Glu668*). Seven
probands with a clinical diagnosis of FH were mutation negative. This pilot
study, integrating clinical and molecular-based techniques, begins to
elucidate the FH heterogeneity and the mutation spectrum in the Iranian
population. Such information is important for future disease management and
cost savings.
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المؤتمرات العلمية:
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المؤتمر (1):
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عنوان المؤتمر:
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83rd European Atherosclerosis Society Congress
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تاريخ الإنعقاد:
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22-25/03/2015
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مكان
الإنعقاد:
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Glasgow,
UK
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طبيعة المشاركة:
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Poster presentation
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عنوان المشاركة:
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Identification of the functional
variants behind GWAS SNP rs6511720 for lower LDL-C and risk of CHD
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ملخص المشاركة:
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Background:
The LDLR (Low-Density Lipoprotein Receptor) SNP rs6511720 (G>T) (minor
allele frequency 0.10 to 0.13) has been identified in GWA studies as being
associated with lower plasma levels of LDL-C (size effect; -0.15 to
-0.26mmol/l) and a lower risk of CHD, myocardial infarction and Abdominal
Aortic Aneurysm. Rs6511720 is located in intron-1 of the LDLR gene, where
cis-acting gene regulating sites are commonly found.
Aim: To
identify the functional variant and the transcription factors (TFs) that bind
to the variant and are involved in gene expression regulation.
Methods:
SNP annotation databases were used to identify possible SNP functions,
luciferase reporter assays in the liver cell line Huh7, using the LDLR
promoter and the SNP in the downstream enhancer position of pGL3-basic, were
used to measure the effect of variant genotype on gene expression, and
Electrophoretic Mobility Shift Assays (EMSAs) using Huh7 nuclear proteins
were used to investigate variants affecting TF binding.
Result:
Rs6511720 is in strong linkage disequilibrium with three intron-1 SNPs, all
located in open chromatin structure, where TFs could bind. EMSAs on rs6511720
and these additional SNPs detected allele-specific protein binding to all
SNPs. Luciferase reporter assays showed rs6511720 and rs57217136 caused a
significant increase in LDLR gene expression (22.7% and 26.9% respectively).
Multiplex competitor EMSAs implicated Snai3 as binding rs6511720, while RAR
was binding rs57217136.
Conclusion:
Both LDLR rs6511720 and rs57217136 are functional variants. Both minor
alleles create enhancer-binding protein sites for TFs that contribute to
increased LDLR expression, which consequently reduces LDL-C levels and CHD
risk.
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المؤتمر (2):
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عنوان المؤتمر:
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85th European Atherosclerosis Society Congress
2017
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تاريخ الإنعقاد:
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23-26/04/2017
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مكان
الإنعقاد:
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Prague, Czech
Republic
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طبيعة المشاركة:
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Oral presentation
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عنوان المشاركة:
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Identifying low density lipoprotein cholesterol associated variants in
the Annexin A2 (ANXA2) gene
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ملخص المشاركة:
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Background: The
repeat-one (R1) domain of AnnexinxA2 binds to PCSK9, affecting the
degradation of LDLR and LDL-C levels.
Aims: To
identify functional variants in ANXA2 influencing lipid levels.
Methods: The
ANXA2 SNPs genotype-phenotype association was tested using the NPHS-II study.
SNP annotation databases identified possible SNP functions. ANXA2 selected
SNPs were inserted into the downstream enhancer position of the
pGL3-promoter. Luciferase assays in Huh7 cells were used to measure the
effect of variant genotype on gene expression. Electrophoretic mobility shift
assays (EMSAs) were used to investigate variants affecting transcription
factor binding.
Results: The
ANXA2-R1-domain coding-SNP rs17845226 (V98L) [MAF=≈12%] has a recessive
effect, with homozygotes for the minor allele having ≈16% higher levels of
LDL-C(p=0.004), and higher risk of CHD [HR (95% CI)=2.17 (1.03-4.60), p=0.04].
The SNP is in modest linkage disequilibrium (r2=0.5) with four SNPs
downstream of the ANXA2 gene-coding region. The eQTL study showed that
homozygotes for the minor allele of the rs11633032 SNP have significant lower
level of ANXA2-mRNA expression (p=1.36×10-05) in liver cells. All SNPs were
located in open chromatin structure, where TFs could bind. EMSAs on these
SNPs detected allele-specific protein binding to both rs11633032 and
rs17191344. Luciferase assays showed both SNPs caused significant reduction
in gene expression (≈18%). Multiplex-competitor-EMSAs implicated CTCF bind to
rs17191344, while GATA and Egr1 bind to rs11633032.
Conclusion: Both
rs11633032 and rs17191344 SNPs are functional variants, where minor alleles
create repressor-binding protein sites for TFs that contribute to reduced
gene expression of ANXA2, which consequently reduces LDL-C levels and CHD
risk.
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المؤتمر (3):
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عنوان المؤتمر:
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British
Atherosclerosis Society 2016
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تاريخ الإنعقاد:
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مكان
الإنعقاد:
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Cambridge, UK.
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طبيعة المشاركة:
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Poster presentation
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عنوان المشاركة:
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Identifying Low Density Lipoprotein Cholesterol Associated Variants in
the Annexin A2 (ANXA2) Gene
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ملخص المشاركة:
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Background: The
repeat-one (R1) domain of AnnexinxA2 binds to PCSK9, affecting the
degradation of LDLR and LDL-C levels.
Aims: To
identify functional variants in ANXA2 influencing lipid levels.
Methods: The
ANXA2 SNPs genotype-phenotype association was tested using the NPHS-II study.
SNP annotation databases identified possible SNP functions. ANXA2 selected
SNPs were inserted into the downstream enhancer position of the
pGL3-promoter. Luciferase assays in Huh7 cells were used to measure the
effect of variant genotype on gene expression. Electrophoretic mobility shift
assays (EMSAs) were used to investigate variants affecting transcription
factor binding.
Results: The
ANXA2-R1-domain coding-SNP rs17845226 (V98L) [MAF=≈12%] has a recessive
effect, with homozygotes for the minor allele having ≈16% higher levels of
LDL-C(p=0.004), and higher risk of CHD [HR (95% CI)=2.17 (1.03-4.60),
p=0.04]. The SNP is in modest linkage disequilibrium (r2=0.5) with four SNPs
downstream of the ANXA2 gene-coding region. The eQTL study showed that
homozygotes for the minor allele of the rs11633032 SNP have significant lower
level of ANXA2-mRNA expression (p=1.36×10-05) in liver cells. All SNPs were
located in open chromatin structure, where TFs could bind. EMSAs on these
SNPs detected allele-specific protein binding to both rs11633032 and
rs17191344. Luciferase assays showed both SNPs caused significant reduction
in gene expression (≈18%). Multiplex-competitor-EMSAs implicated CTCF bind to
rs17191344, while GATA and Egr1 bind to rs11633032.
Conclusion: Both
rs11633032 and rs17191344 SNPs are functional variants, where minor alleles
create repressor-binding protein sites for TFs that contribute to reduced
gene expression of ANXA2, which consequently reduces LDL-C levels and CHD
risk.
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