Nutrigenetic association of the 5-lipoxygenase gene with myocardial infarction

Abstract

Background: 5-Lipoxygenase (5-LO) catalyzes the rate-limiting step of the biosynthesis of proinflammatory leukotrienes from arachidonic acid (AA) and has been associated with atherosclerosis in animal models and humans. We previously reported that variants of a 5-LO promoter repeat polymorphism were associated with carotid atherosclerosis in humans, an effect that was exacerbated by high dietary AA but mitigated by high dietary n-3 fatty acids.

Objective: We sought to confirm these initial observations with a more clinically relevant phenotype such as myocardial infarction (MI).

Design: The 5-LO polymorphism was genotyped in 1885 Costa Rican case-control pairs and tested for association with MI. Functional experiments were carried out to determine whether the associated alleles had differences in mRNA expression.

Results: The frequency of variant genotype groups did not differ significantly between cases and controls. However, a significant gene x diet interaction was observed, in which, relative to the common 5 repeat allele, the 3 and 4 alleles were associated with a higher MI risk in the high (> or = 0.25 g/d) dietary AA group (odds ratio: 1.31; 95% CI: 1.07, 1.61) and with a lower risk in the low (<0. 25 g/d) AA group (0.77; 0.63, 0.94) (P for interaction = 0.015). Using allele-specific quantitation, the short alleles had expression approximately twice that of the 5 allele (P < 0.0001).

Conclusions: The 3 and 4 variants lead to higher 5-LO expression and provide additional evidence that these alleles are associated with greater risks of atherosclerosis and MI in the context of a high-AA diet.

FIGURE 1

Mean (±SEM) [in relative units (ru)] real-time quantitation of 5-lipoxygenase (5-LO) mRNA concentrations in control subjects with different 5-LO promoter genotypes. 5-LO expression did not differ significantly between persons who were homozygous or heterozygous for the shorter 3 and 4 promoter alleles and those who were 55 homozygotes. Total RNA was isolated from 123 buffy coat samples from controls and reverse-transcribed into cDNA. Real-time polymerase chain reaction was carried out in triplicate, and expression levels were normalized to β-actin as an endogenous control.

FIGURE 2

Mean (±SEM) allele-specific quantitation of 5-lipoxygenase mRNA concentrations in heterozygous persons. RNA from the 3A (A) and 4T (B) 5-lipoxygenase promoter alleles are ≈2.5-fold and 1.7-fold as abundant as is RNA from the 5G and 5C alleles, respectively (P < 0.0001 for both comparisons). Coding single-nucleotide polymorphisms in exon 1 and exon 2 were used to compared the expression of 4T and 3A haplotypes to that of the 5C and 5G haplotypes. Total RNA from buffy coat samples of persons with haplotypes 3A/5G (n = 45) and 4T/5C (n = 41) was reverse-transcribed into cDNA and subjected to allele-specific quantitation using pyrosequencing technology, as described in the Methods section. Genomic DNA from the same persons for each haplotype was used as control reactions, in which the ratio from the 2 alleles is 1, as expected. Data are expressed as the ratio of the 3A or 4T allele to the 5G or 5C allele, respectively.