**7. New discoveries based on genome-wide screening (GWAS)**

Genome-wide association studies (GWAS) have been widely used in recent years to identify new genetic loci underlying chronic diseases. GWAS for endothelial function have been relatively limited due to the different phenotypes associated with it. The first such study was conducted by Vasan and colleagues for several cardiovascular traits, including FMD (%) and hyperemic flow velocity in 1345 individuals from the Framingham Heart Study, using a set of 100kSNPs [76]. They identified several SNPs associated with each trait in this study, including chloride channel (CFTR) and phosphodiesterase 5A (PDE5A) SNPs. Although these results have not been replicated, this was the first GWAS to directly examine endothelial function on a large population.

Yoshino and colleagues conducted an association study on the coronary vascular response to acetylcholine (ACh), a common index of coronary endothelial function,

#### *Genetic Markers of Endothelial Dysfunction DOI: http://dx.doi.org/10.5772/intechopen.109272*

in 643 female and male subjects [77]. They used a set of 1536 SNPs located in genes related to cardiovascular physiology and pathology. The results showed that variants in adenosine A1 receptor (ADORA1) were associated with endothelial dysfunction in the entire cohort, while variants in adenosine A3 receptor (ADORA3) and lipoprotein(a) (LPA) had the strongest associations with increased risk of endothelial dysfunction in women only.

In recent genome-wide association study (GWAS) studies in European population, three novel sites related to endothelial dysfunction were found [78, 79]: Vascular endothelial growth factor A (VEGFA) rs9472135, Faciogenital dysplasia 5 (FGD5) rs11128722, Zinc Finger C3HC-type Containing 1 (ZC3HC1) rs11556924.

Because many GWAS identify SNPs outside protein coding regions or in non-coding intervals, the contribution of small non-coding RNA (e.g., lncRNA, microRNA) in modulating endothelial function should be addressed. In 2011, genome-wide association studies (GWAS) identified ANRIL as a biomarker closely associated with coronary heart disease (CHD) [80]. These studies identified, locus 9p21 which contains many single nucleotide polymorphisms (SNPs) that are located in a "gene desert" without any protein-coding genes. A key portion of the SNPs at the 9p21 locus overlap with six exons in the ANRIL gene also known as CDKN2B-AS or CDKN2B-AS1, which is transcribed in the antisense direction in the INK4b-ARF-INK4a gene cluster. ANRIL is expressed in vascular endothelial cells, vascular smooth muscle cells, mononuclear phagocytes and atherosclerotic plaques and its variation is associated with vascular endothelial malfunction, vascular smooth muscle cell (VSMC) including proliferation, migration, senescence, apoptosis, mononuclear cell adhesion and proliferation, glycolipid metabolism disorders and DNA damage [81].

Heritable changes in gene activity and expression also can be the result of epigenetic changes. Recent evidence suggests epigenetic changes such as those induced by histonemethyltransferase Set7 are associated with endothelial dysfunction, including impaired FMD in diabetics [82].

The problem with the paucity of GWAS studies is that most disease-relevant single nucleotide polymorphisms (SNPs) cannot be assigned to a specific gene, and even demonstrating that a single SNP affects gene expression is not possible for most SNPs. This is a consequence of the complex architecture of the genome, in which enhancers are often located far from their target gene in a two-dimensional sequence-based projection. The second aspect is a consequence of the heterocellularity of the atherosclerotic lesion, such that a specific SNP is relevant in only one of the many different cell types expressed in the lesion.

In the future, thanks to the already initiated GWAS studies in single cells of the atherosclerotic lesion, this second problem may be solved.

*Endothelial Dysfunction - A Novel Paradigm*
