INTRODUCTION

Cardiovascular disease (CVD) is the primary cause of death worldwide. One of the leading disorders that are linked to premature development of CV disease is metabolic syndrome (MetS), a condition characterized by multiple alterations including abdominal obesity, hypertriglyceridemia, elevated fasting glucose, hypertension, and low HDL-C levels (¹). A key event that precedes CVD is the formation of the atheroma plaque, caused by the unregulated uptake of oxidized low-density lipoproteins (oxLDLs) by macrophages that progressively turn into foam cells, the main trigger of the atherosclerotic lesion (²) saturable pathway related to the pathway for macrophage uptake of acetylated LDL. Conditioning LDL with cultured aortic smooth muscle cells had a qualitatively similar but smaller effect; conditioning with fibroblasts had no effect. Conditioning very low density lipoproteins or high density lipoproteins with endothelial cells did not affect subsequent metabolism of these lipoproteins by macrophages. Endothelial cell-modified LDL, while degraded more rapidly than control LDL by macrophages, was degraded more slowly by cultured smooth muscle cells and by human skin fibroblasts. Degradation of endothelial cell-modified LDL by macrophages was accompanied by stimulation of cholesterol esterification, inhibition of cholesterol synthesis, and a net increment in total cellular cholesterol content. Thus, a biologically generated modification of LDL is described that markedly alters cholesterol metabolism of macrophages and, consequently, may play a role in foam cell formation during atherogenesis, “container-title”: Arteriosclerosis (Dallas, Tex.. These oxLDLs elicit the formation of immunogenic epitopes in the LDL molecule, and the subsequent production of anti-oxidized low-density lipoprotein antibodies (anti-oxLDL Abs) (³). However, the physiological role of anti-oxLDL Abs remains controversial since these are present in healthy subjects as well as in different diseases. Therefore, it has been suggested that they may have a pro-atherogenic role, an anti-atherogenic function, or no correlation at all (^{4 5}-⁶).

The atheroprotective role of HDL particles results from their antioxidant and anti-inflammatory properties, mainly conferred by the paraoxonase-1 (PON1) enzyme, which circulates chemically bound to the apolipoprotein A-1 (Apo A1), the largest protein constituent of HDL particles (⁷). PON1 is a 345 amino-acid glycoprotein, and its serum activity is inversely associated with oxidative stress, as it can hydrolyze lipid peroxides, cholesteryl esters, and oxidized phospholipids, thus preventing the formation of atherosclerotic lesions (⁸). The circulating levels of HDLs are mainly mediated by reverse cholesterol transport (RCT), of which several pathways have been described—one of them is HDL particles traveling towards peripheral tissues, where their Apo A1 protein region stimulates cholesterol efflux from ATP-binding cassette transporters, ABCA1 and ABCG1, promoting its clearance and/or HDL biogenesis (⁹). Furthermore, it has been suggested that the PON1 enzyme enhances RCT by facilitating the union of HDL particles to macrophages and the ABCA1 protein (¹⁰).

The ABCA1 transporter protein is coded for by the ABCA1 gene, located in the long arm of the human chromosome 9 (9q22-31). This protein plays a key role in HDL particle formation and cholesterol efflux from macrophages, thereby precluding the formation of macrophage foam cells (¹¹). Several single-nucleotide polymorphisms (SNPs) in the ABCA1 gene have been reported—R219K (rs2230806), R1587K (rs2230808), and I883M (rs4149313) are the most extensively studied, and have been associated with CV risk and plasma lipid profile (¹²). Particularly, the R1587K SNP, which is located in exon 35 and induces a change from arginine to lysine in position 1,587 of the ABCA1 protein, has been associated with increased levels of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and TGs, and reduced HDL-C levels (¹²,¹³).

Nevertheless, the relationships between R1587K genotypes and lipids levels are inconsistent among all populations, and have not been studied in Mexicans, especially in the northern region population, where dyslipidemias are common, mainly hyperalphalipoproteinemia, hypertriglyceridemia, and hypercholesterolemia (¹⁴)middle (India. Therefore, we aimed to evaluate the potential association between the R1587K ABCA1 polymorphism with CV risk and MetS presence, as well as with plasma lipid profile, PON1 activity and concentration, and anti-oxLDL titers in a northern Mexican population.

MATERIAL AND METHODS

RESULTS

PARTICIPANT CHARACTERISTICS AND DIFFERENCES BY R1587K GENOTYPE

The study population was integrated by 57 participants (29 men and 28 women) with an average age of 38 ± 10 years. Genotypes were distributed as follows: 63.2% had the RR genotype, 26.3% the RK genotype, and 10.5% the KK genotype. Therefore, for statistical purposes we decided to group the RK and KK genotypes (34.1% had RK + KK) for comparison with RR carriers, assuming that the presence of one or two K alleles would modify the risk for the assessed variables.

BMI showed that both study groups were overweight, had an excess of body fat percentage, elevated LDL-C levels, and were also sedentary according to their physical activity level (²¹). The RR and the RK + KK genotypes carriers had elevated LDL-C and decreased HDL-C levels as compared to the NCEP ATP III guidelines (²²). RR carriers showed a moderate CV risk, whereas the RK + KK group had a high CV risk according to the AIP.

As seen in table I, triglyceride levels were significantly higher among the RK/KK genotype carriers than among the RR genotype carriers. Also, the RK/KK carriers showed a tendency to greater waist circumference, TC levels, anti-oxLDL Abs, and atherogenic index of plasma when compared to RR carriers. However, no significant relationship was detected between ABCA1 R1587K genotypes (RR and RK + KK) with BMI, fat-free mass, fat mass, blood pressure, physical activity, smoking, alcohol consumption, fasting glucose, fasting insulin, HOMA-IR index, LDL-C levels, HDL-C levels, and PON1 concentration and activity (Table I).

Table I.  Demographic, body composition, clinical and lifestyle characteristics of the study population, stratified by genotype 

Data are given as mean ± standard deviation or median [25-75 interquartile range]. BMI: body mass index; WC: waist circumference; FFM: fat-free mass; SBP: systolic blood pressure; DBP: diastolic blood pressure; BMR: basal metabolic rate; HOMA-IR: homeostatic model assessment-insulin resistance; TC: total cholesterol; LDL-C: low-density lipoprotein cholesterol; HDL-C: high-density lipoprotein cholesterol; TG: triglycerides; anti-OxLDL Abs: anti-oxidized low-density lipoprotein autoantibodies; PON1: paraoxonase-1; AIP: atherogenic index of plasma.

ASSOCIATION OF R1587K GENOTYPES WITH CV RISK AND METS

The RK + KK genotypes were associated with MetS presence (OR = 4.566, 95% CI = 1.386-14.92, p = 0.010). Also, the RK + KK genotypes showed a tendency to association with higher CV risk based on the AIP score (OR = 2.800, 95% CI = 0.910-8.611, p = 0.069).

Table II.  Association of ABCA1 R1587K genotypes with CV risk and metabolic syndrome presence 

Data are represented in counts (percentages within parentheses). AIP: atherogenic index of plasma; MetS: metabolic syndrome; p-values and ORs with 95% CIs were calculated using Pearson's chi² test; p < 0.05 was considered significant.

DISCUSSION

The major pathway in HDL particle biogenesis is the RCT, in which the ABCA1 protein plays a pivotal role by stimulating the cholesterol efflux, and precluding the foam cell formation, that induce the atherosclerotic lesion. Several SNPs in the ABCA1 gene have been described. In particular, the R1587K polymorphism has been associated with dyslipidemias and CV risk, but no information exists regarding its relationship with a highly related predecessor of these diseases such as the MetS, and other non-common metabolic and cardiovascular parameters such as lipid-related enzymes and antibodies involved in these pathways. Therefore, we decided to analyze the R1587K SNP in the ABCA1 gene and to evaluate its association with CV risk and MetS presence, as well as its possible role in the lipid profile, PON1 activity and concentration, and anti-oxLDL titers of a northern Mexican population. We found an association between the RK + KK genotype and the presence of MetS, as well as higher TG levels and a tendency towards high CV risk among the RK + KK genotype carriers. However, this polymorphism showed no association with HDL-C levels, PON1, and anti-oxLDL titers.

The frequency of R187K genotypes in ABCA1 observed in our study was 63.2% for RR and 36.8% for the RK + KK genotypes; to our knowledge, this is the first time that R1587K genotypes have been related to MetS in a Mexican population, having previously been mainly described in Caucasians. These frequencies are similar to that previously reported in children and adolescents from Poland in remission of nephrotic syndrome, reporting a frequency of 58% of the RR genotype and 42% of the RK + KK genotype (²³). A similar distribution of these genotypes was reported in a Dutch population of men with proven coronary artery disease, as well as in the Danish general population (²⁴,²⁵). However, these frequencies differ from those in a group of young Greek nurses, showing a reduced frequency of the RR genotype (47.08 %) in comparison to the RR + KK genotypes (52.92%) (²⁶). These differences are likely explained by the genetic heterogeneity extant among populations, which translates into different phenotypes, highlighting the importance of replicating these genetic studies in different populations.

The R1587K SNP, specifically the K allele, in the ABCA1 gene has been previously related with dyslipidemias, specifically with high TC, LDL-C, and TG levels, as well as with lower HDL-C levels in both men and women of the general population or with proven CVD of different age groups (¹²,¹³). Although we observed low HDL-C levels among the participants, the concentrations of this lipid fraction showed no relationship with the R1587K SNP; however, TG level turned out to be related with this SNP, being higher among RK + KK carriers. Also, we observed that carriers of the RK + KK genotypes tended to have higher TC and anti-oxLDL Ab levels, abdominal obesity, and high CV risk (as evaluated by the AIP), although not reaching statistical significance. This difference in plasmatic TG levels between the RK + KK genotype carriers as compared to RR genotype carriers is consistent with that reported by Kolovou et al. in Greek women without a history of coronary artery disease (²⁶), and with the tendency described by Ksiazek et al. in children and adolescents undergoing nephrotic syndrome remission (²³). Conversely, Jensen et al. reported a lower concentration of TGs in women without CV disease who were carriers of the K allele of this SNP (²⁷).

Even though the direct connection between the ABCA1 protein and TG levels is not fully understood, there is a direct relationship between HDL-C levels and TGs, seen through HDL modeling, where several processes are involved. One of these is mediated by the cholesteryl ester transfer protein (CETP), where an exchange of cholesteryl esters (CE) from HDL to TGs in apoB-containing lipoproteins (LDL/VLDL) occurs (²⁸). This exchange ends in apoB-containing lipoproteins enriched with CEs and drained of TGs, and in HDL particles depleted of CEs and high in TGs. The clearance of these HDL particles enriched with TGs is faster than that of those enriched with CEs, thus an increase in TG levels triggers a decrease in HDL-C levels (²⁹). This reduction in HDL-C levels may also result in changes in the contents and activities of related proteins. Nevertheless, whether the low HDL-C levels observed in our study are due to an increase in TG levels or to the presence of a R1587K SNP remains unknown. Therefore, further and larger-scale studies including different genetic variants and proteins involved in lipid metabolism are required.

Another interesting finding was the association between the RK + KK genotypes and the presence of MetS, in addition to the marginal association detected between the RK + KK genotypes and high CV risk, according to the AIP. This is consistent with the fact that hypertriglyceridemia, alone or in combination with low HDL-C concentrations, has been described as a major factor related to MetS (³⁰). Even though LDL-C has been widely known as one of the major atherogenic lipids, the association between hypertriglyceridemia and risk of CVD has been extensively described (³¹). Nevertheless, this high risk cannot be completely attributed to the presence of the R1587K SNP, since this population presents with other CV risks including elevated LDL-C and low HDL-C levels, low physical activity, and altered body composition (elevated body fat percentage, abdominal obesity, and high BMI), all of which contribute to increased CV risk.

A limitation of this study was its relatively small number of participants, so it is necessary to further perform wider studies where these associations with the R1587K genotypes may be corroborated. However, this study adds new information regarding the role of this SNP in the northern Mexican population. Also, we included a novel and common cardiovascular risk biomarker to help elucidate the findings.

In summary, the R1587K SNP of the ABCA1 gene was related to TG level; specifically, RK + KK genotype carriers had higher TG concentrations than RR genotype carriers, while HDL-C levels, PON1 activity and concentration, and anti-oxLDL levels did not differ between genotypes. Further, an association was found between the RK + KK genotypes and the presence of MetS, as well as a tendency towards high CV risk, indicating that RK + KK carriers are more likely to develope CVD. These findings provide novel information about the presence and implications of the R1587K SNP of the ABCA1 gene in lipid metabolism and CV risk, and could contribute to understand and devise further interventions to prevent the clinical alterations commonly observed in this population.