of a stroke on twin studies (comparison of the incidence of stroke in monozygotic
versus dizygotic twins) and, even more, in family history studies.^16
2.3 Recombinant DNA Biotechnology Methods
in the Detection of “Stroke Genes”
To identify actually the genes for stroke, three main methods have been used:
linkage analysis, candidate-gene approach, and genome-wide association studies
(GWAS). The linkage analysis, in which the association between different chro-
mosomal markers (and thus proximally located genes) and disease phenotype
within families is searched for, was very useful in detecting genes causing single-
gene disorders related to monogenic stroke variants, such as notch3 gene, causing
CADASIL, or the variants in phosphodiesterase 4D, associated with ischemic
stroke in Islandic population.^17 The linkage analysis is, in general, less successful
in detecting genes associated with polygenic diseases, such as stroke, in which
many genes contribute, usually in a small amount, to overall risk. Even less
satisfactory were the results obtained using candidate-gene approach, in which
genetic variants were searched in candidate genes that have been previously
associated with stroke risk.^18
The real breakthrough in searching the genes for stroke came with the use of
affordable and reliable high-throughput microarray technology and the develop-
ment of GWAS approach, in which a huge number (one million or more) of single
nucleotide polymorphisms (SNPs), spanning the whole human genome, are tested
in the individual subjects with the disease, and the results are compared with control
cases.^19 The GWAS genotyping revolutionized personalized medicine approach
not only in the field of stroke and other CNS diseases and disorders, but also in
general: many GWAS have been successfully conducted to identify DNA variants
associated with complex human diseases and traits, such as cancer, autoimmune
diseases, height, blood pressure, body mass index, etc.^20 GWAS have identified
many novel genetic associations in different cerebrovascular diseases, including
stroke, but most of those associations, however, account for only a small increase in
disease risk, probably due to heterogeneity of the disease.^21
The results of GWAS for ischemic stroke are very recent, dating from 2007
onward. The variant of genePITX2(paired-like homeodomain transcription factor
2 or pituitary homeobox 2), encoding for transcription factor involved in the
(^16) Markus ( 2012 ) and Jerrard-Dunne et al. ( 2003 ).
(^17) Markus ( 2012 ), Joutel et al. ( 1996 ), and Helgadottir et al. ( 2004 ).
(^18) Markus ( 2012 ) and Dichgans and Markus ( 2005 ).
(^19) Hardy and Singleton ( 2009 ) and Schierding et al. ( 2014 ).
(^20) Hou and Zhao ( 2013 ).
(^21) Campione et al. ( 1999 ).
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