| Research Abstract |
Coronary artery disease (CAD) and ischemic cerebrovascular disease (CVD) are typical human attributes that have a complex multifactorial etiology. This study aimed to establish systems to comprehensively evaluate the contribution of genetic factors to the pathogenesis of atherosclerosis and thrombosis in the Japanese populations, where allele frequencies are known to be quite different than Caucasians, using "candidate gene approach".
Polymorphisms in platelet activating factor (PAF) receptor, PAF acetylhydrolase, and platelet GPIba receptor were investigated in relation to their functional consequences and association with CVD. PAF is a lipid mediator that has potent biologic effects on a variety of cells. For platelets, binding of PAF to its specific receptor (PAF-R) results in activation of phospholipases and protein kinases, and increase in intracellular Ca^<++> concentration. A common single nucleotide polymorphism was recently identified that results in an amino-acid substitution
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in the cytoplasmic region of PAF-R, Ala/Asp at residue 224 (^<224>A/D, Fukunaga K et al, 2001). Our analysis of 280 general Japanese populations showed the genotype frequencies of 75.7%, 21.8%, and 2.5% for A/A, A/D, and D/D, respectively. We measured PAF-induced platelet aggregation in healthy volunteers with different genotypes. Two healthy volunteers with PAF-R D/D genotype showed impaired aggregating response to a wide range of PAF concentrations. Platelets from one of the two subjects did not undergo irreversible aggregation even at higher concentration of PAF (6 micro M). The frequencies in the patient group were 78.9%, 17.2%, and 3.9% for A/A, A/D, and D/D genotypes, respectively, which were not statistically different from the control group. Genetic deficiency of plasma PAF-AH is relatively common in the Japanese but is not found in white populations. PAF-AH deficiency is attributed to a single point mutation, Val279Phe. It has been reported that subjects with the homozygous mutation (Phe/Phe) have near-absent plasma PAF-AH activity, whereas heterozygotes (Val/Phe) have about half of the PAF-AH activity as compared to the wild-type (Val/Val). The clinical relevance of Val279Phe is still controversial. We examined the relationships of this mutation with in vitro platelet function, and the frequencies of Val279Phe in CVD patients and control subjects. Platelet-rich plasma from healthy volunteers with heterozygous mutation (Val/Phe) showed apparently higher platelet aggregating response to threshold concentrations (〜0.01 micro M) of PAF as compared to Val/Val, suggesting the role of PAF-AH in the regulation of platelet activation. Genotype frequencies for CVD patients were 60.3, 35.9, and 3.8% for Val/Val, Val/Phe, and Phe/Phe, respectively, which were significantly different from the control group (68.8, 27.9, and 3.4%, respectively, p<0.05 when compared between Val/Val vs Val/Phe+Phe/Phe). When analysis was confined to those with age<60, the difference was more significant (55.7, 39.3, and 4.9% for CVD and 70.6, 26.4, and 3.1% for controls, p<0.01, OR=1.9, Val/Val vs Val/Phe+Phe/Phe). The genotype-effect became stronger when subjects without smoking were compared (p<0.01, OR=2.4). The present study suggests that heterozygous deficiency of PAF-AH with ^<279>Val/Phe which is common in Japanese may induce impaired regulation of platelet activation, and is associated with increased risk of CVD at younger age. We also analyzed a Connexin 37 gene polymorphism C1016T in CVD patients, and found that the frequency of the T allele was higher in the patient group, and that hypertension interacts with this association.
Combinations of multiple genetic factors are believed to be crucial for the development of thrombotic disorders. The ultimate goal of the clinical appreciation of polymorphic markers is to identify subgroups of individuals who are best prevented from developing diseases, or who respond best to dietary, behavioral, or pharmacologic interventions. For this purpose, polymorphisms need to be investigated not only in relation to disease susceptibility, but also with regard to responsiveness to treatment, and gene-environment interactions. Less
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