Single-nucleotide polymorphisms of VEGF gene are associated with risk of congenital valvuloseptal heart defects

doi:10.1016/j.ahj.2005.10.012

American Heart Journal

Volume 151, Issue 4, April 2006, Pages 878-881

https://doi.org/10.1016/j.ahj.2005.10.012 Get rights and content

Background

Disturbed vascular endothelial growth factor (VEGF) production during early heart morphogenesis causes endocardial cushion malformation, which results in congenital heart disease (CHD). We tested whether functional VEGF −460T/C and +405G/C polymorphisms that have an impact on VEGF levels were associated with CHD.

Methods

Dried blood samples were collected from 102 CHD children and 112 healthy control neonates. Genotyping was done with polymerase chain reaction–restriction fragment length polymorphism (VEGF +405G/C) and real-time polymerase chain reaction methods (VEGF −460T/C).

Results

VEGF −460C allele frequency was similar in control and CHD subjects. VEGF +405C allele was less prevalent in controls than in CHD subjects (0.21 vs 0.42, P < .001). Having VEGF +405C presented increased risk for CHD (odds ratio [OR] 1.72, 95% CI 1.32-2.26). VEGF −460CT/+405CC allele associations did not occur in controls but in CHD patients (0% vs 13%, OR 2.26, 95% CI 1.93-2.64), whereas −460CT/+405GG allele association was more prevalent in controls (32% vs 16%, OR 0.58, 95% CI 0.37-0.89).

Conclusions

VEGF gene and allele associations may be associated with increased risk of CHD.

Section snippets

Patients

We took one drop of blood apiece on filter paper from 102 unrelated children (52 boys and 50 girls between the ages of 2 and 10 years) with CHD, who were regularly checked at the Cardiology Outpatient Clinic of County Children's Hospital in the city of Pécs, Hungary.

Each patient had some form of valvular and/or septal defect that is associated with the maldevelopment of EC during early heart morphogenesis. Patients were, however, heterogeneous in term of CHD complexity and localization. Single

Results

No deviation from Hardy-Weinberg equilibrium was observed in the cases or the controls for the VEGF −460T/C (P = .16 and .13, respectively) and VEGF +405G/C SNPs (P = .24 and .43, respectively). Significant linkage disequilibrium coefficient was found between VEGF −460T/C and +405G/C SNPs in control subjects, but not in CHD patients (controls, D′ value 0.91, P < .0001; CHD patients, D′ value 0.18, P = 0.56); however, r values were not significant (controls r = 0.018, not significant; CHD

Discussion

In this study, we found that carrier state of VEGF +405C allele, especially that of +405CC genotype, and the −460TC/+405CC genotypic association are more prevalent in those children who have CHDs caused by EC malformation. This result is in line with previous findings that suggest a significance of these VEGF SNPs in other clinical conditions. According to clinical studies, VEGF −460C/CT SNP may be a marker for prostate cancer, susceptibility to endometriosis, and diabetic retinopathy.9, 10, 11

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It has been known for many years that cardiovascular developmental disorders have an important heritable component. In recent years the development of new technologies such as molecular labeling, the use of animal models, such as the mouse and zebrafish, and next-generation sequencing have revolutionized our understanding of normal heart development and the molecular processes which, when disturbed, lead to significant congenital heart disease (CHD). Although the precise cause of CHD remains unclear in most patients, this improved understanding of molecular processes is gradually clarifying our understanding of the interaction between genetic, epigenetic, and environmental phenomena. The aim of this chapter is to discuss the recent advances in our understanding of the molecular mechanisms that are associated with, and sometimes cause, CHD and to evaluate the link between genotype and phenotype.
Cellular decisions in cardiac outflow tract and coronary development: An act by VEGF and NOTCH
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Data from human pathologies, animal models and in vitro analyses have shed more light on the interaction between these two fascinating pathways, continuously confirming the role of VEGF and subsequent NOTCH-signaling in both vascular as well as cardiac development (Caolo et al., 2010; Chang et al., 2011; Iso et al., 2003a; Lawson et al., 2002; Olsson et al., 2006; Smedts et al., 2010). Within the human VEGF-gene, several polymorphisms in both coding and promoter region are currently known to be associated with diminished or elevated levels of VEGF-transcription (Lambrechts et al., 2005; Vannay et al., 2006). As defined by several mouse models (see below), relatively small changes in Vegf-expression can be lethal or result in severe abnormalities.
Congenital cardiac abnormalities are, due to their relatively high frequency and severe impact on quality of life, an important focus in cardiovascular research. Recently, various human studies have revealed a high coincidence of VEGF and NOTCH polymorphisms with cardiovascular outflow tract anomalies, such as bicuspid aortic valves and Tetralogy of Fallot, next to predisposition for cardiovascular pathologies, including atherosclerosis and aortic valve calcification. This genetic association between VEGF/NOTCH mutations and congenital cardiovascular defects in humans has been supported by substantial proof from animal models, revealing interaction of both pathways in cellular processes that are crucial for cardiac development.
This review focuses on the role of VEGF and NOTCH signaling and their interplay in cardiogenesis with special interest to coronary and outflow tract development. An overview of the association between congenital malformations and VEGF/NOTCH polymorphisms in humans will be discussed along with their potential mechanisms and processes as revealed by transgenic mouse models. The molecular and cellular interaction of VEGF and subsequent Notch-signaling in these processes will be highlighted.
Altered jugular vein and ductus venosus flow velocities in fetuses with increased nuchal translucency and distended jugular lymphatic sacs
2010, American Journal of Obstetrics and Gynecology
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Morphological studies of both aneuploid human and mouse embryos with nuchal edema showed a distended and abnormal endothelial differentiation of the jugular lymphatic system.4,7,25 Also, abnormal endothelial processes have been described to play a role in relation to development of cardiovascular defects.27,28 Previous research in trisomy 16 mouse embryos, an animal model for trisomy 21, showed a disturbed interaction between neuronal and endothelial pathways in the JLS, aortic arch, and ductus venosus region.29
We sought to assess blood flow in relation to jugular lymphatic distension in fetuses with increased and normal nuchal translucency (NT).
In all, 72 fetuses with normal NT and 71 fetuses with NT >95th percentile were evaluated. NT size, jugular lymphatic sacs (JLS), jugular vein and ductus venosus pulsatility index for veins (PIV), and intracardiac velocities were measured.
JLS were visualized in 22/72 fetuses with normal and in 55/71 fetuses with increased NT. Jugular vein and ductus venosus PIV was higher in fetuses with increased NT compared to normal NT (P < .01). Visibility of JLS was associated with a higher ductus venous PIV (P < .05), but not with a higher jugular vein PIV. Larger NT and larger JLS volumes were associated with higher jugular vein and ductus venosus PIV (P < .05).
This study shows a relation among increased NT, jugular lymphatic distension, and altered blood flow in jugular vein and ductus venosus.
Mutations in VEGFA are associated with congenital left ventricular outflow tract obstruction
2010, Biochemical and Biophysical Research Communications
Citation Excerpt :
In a recent work, Yang and colleagues [16] demonstrated that vascular endothelial growth factor A (VEGFA) and Notch1 play opposing roles in regulating endothelial-to-mesenchymal transformation, a critical event during embryonic valve development, in cloned endothelial cell populations isolated from ovine aortic valve leaflets. Furthermore, Vannay and colleagues [17] indicated that single nucleotide polymorphisms of the VEGF gene may be associated with an increased risk for congenital valvuloseptal heart defects. Most recently, a family-based case–control study suggested that VEGF single nucleotide polymorphisms contribute to a genetic predisposition to endocardial cushion defects [18].
Left ventricular outflow tract obstruction (LVOTO) comprises a spectrum of stenotic lesions. Previous studies have shown that the vascular endothelial growth factor (VEGF) signaling system plays a critical role in cardiac cushion formation, vasculogenesis, and angiogenesis. We hypothesize that VEGFA may be a potential candidate gene associated with the spectrum of LVOTO lesions. However, it remains unclear whether the VEGFA gene is responsible for the development of LVOTO malformations. In this study, we identified three exon mutations in the VEGFA gene in three of 192 nonsyndromic LVOTO patients, and the overall mutation frequency was 1.6% (3/192). The c.454C>T (p.Arg152X) nonsense mutation and c.19_22dupGACA (p.Thr8ArgfsX78) internal tandem duplication mutation each introduced a premature stop codon and are predicted to produce a truncated VEGFA protein. The c.998G>A missense mutation changes a highly conserved arginine to a glutamine at residue 333 (p.Arg333Gln). These mutations were carried by some family members, and average penetrance was 33.3%. The present study suggests, for the first time to our knowledge, that VEGFA mutations may be associated with congenital LVOTO malformations. We provide evidence that LVOTO is likely oligogenic.
Opposing actions of Notch1 and VEGF in post-natal cardiac valve endothelial cells
2008, Biochemical and Biophysical Research Communications
Citation Excerpt :
Hence, it appears that a finely-tuned balance of VEGF-A signaling is required to achieve appropriate levels of EMT and maintain a continuous endothelium as valve remodeling and leaflet elongation occur [7]. The importance of VEGF in valve development is supported by recent studies identifying single nucleotide polymorphisms in the VEGF gene associated with congenital valve defects [17]. Notch family members are transmembrane proteins that, upon ligand binding, undergo two proteolytic cleavage events to generate an intracellular Notch domain that can travel into the nucleus to regulate transcription and cell fate decisions.
The endothelium of the cardiac valves is unique compared the rest of the vasculature in its ability to undergo an endothelial-to-mesenchymal transformation (EMT) in vitro in response to transforming growth factor-β (TGF-β). EMT is a critical event during embryonic valve development, and both VEGF-A and Notch1 have been shown to function in this process. Here we investigate the effects of VEGF-A and Notch1 on EMT in clonal endothelial cell (EC) populations isolated from adult aortic valve leaflets. VEGF-A inhibited TGF-β-induced EMT. Endothelial growth, however, was not affected by VEGF-A or TGF-β. A positive role for Notch1 was revealed in three experiments: (1) TGF-β induced Notch1 mRNA in valve ECs, (2) a γ-secretase inhibitor of Notch1 signaling blocked EMT, and (3) overexpression of a ligand-independent form of Notch1 induced EMT. These results demonstrate, for the first time, that VEGF-A and Notch1 play opposing roles in regulating EMT in post-natal valve endothelium.
Association study between single nucleotide polymorphisms in the VEGF gene and polycystic ovary syndrome
2008, Fertility and Sterility
To investigate single nucleotide polymorphisms (SNPs) in vascular endothelial growth factor (VEGF) gene that have significant associations with the pathogenesis of polycystic ovary syndrome (PCOS) in a Korean population.
Case-control study.
University-based hospital.
One hundred thirty-four patients with PCOS and 100 healthy women as controls.
None.
Frequencies of genotypes for SNPs in VEGF gene, which were specifically expressed in a Korean population.
After genotypic analysis, we found that among 10 SNPs, one novel SNP at site +9812 and one known SNP at site +13553 have P values lower than .05 (+9812: odds ratio [95% confidence interval] 0.61 [0.39–0.95]; +13553: odds ratio [95% confidence interval] 0.59 [0.37–0.93]) and one haplotype (ht4) also has a P value in the significant range (odds ratio [95% confidence interval] 0.34 [0.16–0.74]).
We concluded that one novel SNP at +9812 site, one known SNP at +13553 site, and one selected haplotype in the VEGF gene have a high possibility of significant associations with the pathogenesis of PCOS in a Korean population.

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: This study was supported with funds from OTKA Grant T046086 and T 042956 and K+F 0134/2001.

^d: Ádám Vannay and Barna Vásárhelyi have equally contributed to this work.

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Clinical InvestigationGeneticsSingle-nucleotide polymorphisms of VEGF gene are associated with risk of congenital valvuloseptal heart defects

Background

Methods

Results

Conclusions

Section snippets

Patients

Results

Discussion

Cytokine

Urology

J Invest Dermatol

J Biol Chem

A novel role for VEGF in endocardial cushion formation and its potential contribution to congenital heart defects

Development

Elevated glucose inhibits VEGF-A–mediated endocardial cushion formation: modulation by PECAM-1 and MMP-2

J Cell Biol

Maternal diabetes: an independent risk factor for major cardiovascular malformations with increased mortality of affected infants

Teratology

Early events in valvulogenesis: a signaling perspective

Birth Defects Res Part C Embryol Today

Clinical Investigation
Genetics
Single-nucleotide polymorphisms of VEGF gene are associated with risk of congenital valvuloseptal heart defects