Acute coronary syndrome (ACS) is a cardiovascular disease (CD) characterized by occlusion of the coronary arteries and most often begins with the rupture of an atherosclerotic plaque in this artery, inducing the formation of a thrombus that occludes partially or totally the vessel.1 Among the risk factors that influence its development, the presence of diabetes, dyslipidemia, smoking, obesity, sedentarism, hypertension and stress are highlighted. Age, gender and family history are also considered risk factors.2 In addition, genetic factors may also contribute to a greater susceptibility to disease.3

Some genetic polymorphisms have already been linked to the risk of developing CDs in specific populations. Interleukin (IL) -8 is produced by a variety of cell types involved in atherosclerosis, in addition to being expressed in areas rich in macrophages of atherosclerotic lesions.3^-5 In addition to a potential role in the initiation and progression of atherosclerosis, IL-8 can also participate in the complications of this process by acting on destabilization of atherosclerotic plaque and specifically on thrombosis.3^,4 Single nucleotide polymorphism (SNP) in IL8 gene (rs 4073) has been shown to be associated with SCA in different populations. Its frequency is related to a high plasma level of its respective cytokine, which may contribute to an inflammation associated with the infiltration of macrophages and neutrophils, thus providing a substrate for the development of ACS.5^,6 This SNP has the ability to alter the transcriptional activity of the gene, contributing to the increase of IL-8 cytokine levels.7^-9

IL-16 is a pleiotropic and immunoregulatory proinflammatory cytokine.10 Elevated levels of this cytokine were detected in patients with CAD.11 In previous studies, this cytokine is related to the regulation of recruitment and activation of CD4 + T cells in the inflammatory process of CVDs.12

Some studies report that the G mutant allele in its gene (rs11556218) is associated with a significantly increased risk of CDs.13^-15 The exact role of IL-16 in the pathogenesis of ACS remains unclear, but it is known that the rs11556218 T/G SNP may influence the expression of IL-16 cytokine and alteration of its serum levels may promote the production of other cytokines IL-1, IL-2 and IL-6 that are involved in the development of ACS.15

These findings suggest that SNPs in the IL8 (rs4073) and IL16 (rs11556218) genes may be useful markers of genetic susceptibility to ACS, allowing the identification of high-risk individuals by selecting for more invasive therapies16 and closer monitoring.

The lack of uniformity in positive results in several populations is the major problem for the study of genetic association.17 Thus, it is necessary to study SNPs in specific populations to identify their association with diseases in a region.

The aim of this study was to evaluate the association between SNPs in the IL8 (rs4073) and IL16 genes (rs11556218) and the risk of developing ACS. In addition it was our intention to compare our blood donor population with other healthy populations from different countries, to contribute to the identification of molecular markers in disease susceptibility.

The distributions of the genotypic frequencies are in accordance with the Hardy-Weinberg equilibrium.

The mean age found in the groups with ACS, without ACS and blood donors was 62 (± 13.0), 58 (± 18.9) and 48 (± 6.3) years old, respectively. The male gender was the most frequent in the three groups evaluated (76.5%, 58% and 85.4%, respectively).

The majority of patients with ACS and non-ACS were non-smokers (69.5% and 82%, respectively) and non-diabetics (55.5% and 68%, respectively). Hypertensive patients accounted for 80.5% of patients with ACS and dyslipidemia was present in 64% of ACS patients and only 14% of those without ACS.

Results for the rs4073 (IL8) SNP demonstrated a higher frequency of the AT genotype in all groups analyzed.

For SNP rs11556218 (IL16), the most frequent genotype in all 3 groups was TT. In addition, TG genotype showed higher frequency in blood donor individuals (35.5%) compared to ACS patients (21.0%; p = 0.002). G allele carriers (TG + GG) were more present in donor subjects (36%) than in patients with ACS (23%) (p = 0.0052). When comparing the groups with ACS and without ACS, no difference was found in the genotypic distribution (Table 2).

When genetic polymorphisms were evaluated in relation to the main risk factors for ACS (smoking, diabetes, hypertension and dyslipidemias), the IL8 gene showed no association. The rs11556218 SNP in the IL16 gene showed statistical association when analyzed against risk factors hypertension (p = 0.002) and dyslipidemias (p = 0.01) (Table 3).

From the results of the genotyping for the polymorphisms and considering the significant association between the SNP rs11556218 in the IL16 gene and SCA, the cytokine IL-16 was dosed. For this, 20 samples from patients with ACS (TT = 9), (TG = 10), (GG = 1) and 20 samples from donor individuals (TT = 9), (TG = 10) and (GG = 1) were used.

The dosages of IL-16 were similar among the three different genotypes in patients with ACS (Figure 1B).

Figure 1

Box-plot (median and quartiles) of IL-16 dosages in patients with ACS and blood donors in different IL16 genotypes. A) Median IL-16 in blood donors (TT = 447.7 pg/mL; TG + GG = 709.2 pg/mL, p = 0.001); B) Median of IL-16 in patients with ACS (TT = 381.4 pg/ml; TG + GG = 314.9 pg/ml, p = 0.288); C) Median IL-16 in blood donors (563.0 pg/mL) and patients with ACS (355.4 pg/ mL) (p = 0.04); p = Mann-Whitney.

The age presents itself as one of the most important risk factors for ACS.28 According to Dutra et al.,29 cardiovascular diseases affect mostly elderly individuals. The mean age found in the present study was 62 years old, corroborating the data obtained by Marino et al.,30 in a study performed with Brazilian patients with ACS who found a mean age of 63 years old and by Shah et al.,31 studying Scottish patients with ACS with a mean age of 62 years old.

Besides age, gender is also important in the epidemiology of ACS. According to Maas and Appelman,32 cardiovascular diseases, where ACS is included, usually manifest later in women than in men, partly because of differences in exposure to risk and hormonal factors.33^,34

In our study, males accounted for 76.5% of ACS patients. Similar data were obtained by Huang et al.,15 and Bray et al.,35 who observed the frequency of 80.4% and 68.0% of male in ACS patients of their studies, respectively.

Smoking, diabetes, hypertension and dyslipidemias are considered important in increasing the risk for ACS, however, most of the patients in the present study were neither smokers nor diabetics in both groups (with or without ACS). These data corroborate the study by Guedes et al.,36 in which the majority of Brazilian patients with CD were not smokers (93.3%). In a study by Vogiatzi et al.,5 with Greek patients diagnosed with ACS, the majority was not diabetic (64.3%).

Despite this, diabetes and smoking may not have their importance excluded as risk factors for ACS.

Generally, the majority of patients (with or without ACS) had systemic arterial hypertension. These results corroborate the data obtained from patients with ACS in different countries (Australia, Pakistan, Mexico and Japan) who indicated that the majority of the patients were hypertensive, varying between 56.5% in Japan and 68.0% in the Mexico.14^,35^,37^,38 For the Ministry of Health, this disease is responsible for at least 25% of the deaths due to ACS.39

In the present study it was verified that the majority of patients with ACS are dyslipemic (64.0%), similar data were verified by Feijó et al.,40 Rodrigues et al.,41 and Andrade et al.,42 who studied Brazilian patients with CDs and found that 50.5%, 73.9% and 60.1%, respectively, were dyslipemic.

The data about risks factors reinforce that this disease has a multifactorial character. Thereby the genetic component directly participates in the predisposition to ACS.

Regarding the polymorphism in the IL8 gene (rs4073), we observed a higher frequency of the AT heterozygous genotype in the three groups analyzed. Similarly, in a study by Zhang et al.,6 a prevalence of AT genotype was 47.4% in patients with ACS and 45.9% in the control group.

For the SNP in the IL16 gene (rs11556218), a difference in heterozygous TG genotype frequencies was observed between the group of patients with ACS and the group of blood donors (p = 0.0026). In addition, carriers of the G mutant allele were more present in blood donors than in patients with ACS, suggesting that this allele may have a protective action in ACS.

Discordant data were found in a study by Chen et al..8 in which the G allele was more frequent in patients with ACS (84.9%) than in the control group (64.2%). However, the authors point that the lack of success in replicating the positive result in other populations is the main problem for genetic association studies.8

When comparing the serum levels of IL-16 cytokine, donor individuals presented higher values than patients with ACS (p = 0.04). In addition, among the blood donors, the G (TG + GG) allele presented higher IL-16 values (p = 0.01). These results corroborate the data obtained by Gronberg et al.,11^,43 where the presence of IL-16 on the carotid plaque is associated with a decreased risk of cardiovascular events in studies performed with a Swedish population. This is due to the stabilization of the atherosclerotic plaques, avoiding the ACS disease.11^,43

The cytokine IL-16 has been described as a pro-inflammatory cytokine. However, there are few studies supporting its possible anti-inflammatory function, identified by the present study.11^,43

In addition, one hypothesis for the lack of association found between ACS and non-ACS patients is that the SNP in the IL16 gene (rs11556218) is involved with CDs in general, not only with ACS, and further studies are needed, with different populations to answer this question.

An association was found regarding the polymorphism in IL16 gene (rs11556218) and the risk factors hypertension and dyslipidemia. These results corroborate the data found by Chen et al.,7 with 300 patients with ACS who showed a significant association between the homozygous variant (GG) genotype and the hypertension, diabetic and smoker individuals. In addition, the study by Wu et al.,15 with 326 Chinese patients with ACS also found an association between the GG genotype and the disease in hypertensive, diabetic, smoker, obese and dyslipemic individuals.

The analysis of the frequencies of polymorphisms in genes of medical interest for a disease in a population can become an important tool to understand and evaluate the risks that this disease can bring, as well as its morbidity and mortality. The results can be used in future studies of association between these polymorphisms and different pathologies.

Thus, the present study also evaluated a comparison between the frequencies of polymorphisms in the IL8 and IL16 genes of our healthy, blood donor individuals with several other healthy populations from other countries.

Regarding the IL8 polymorphism, a similarity was observed in the genotypic distribution with two Brazilian populations (Maranhão and São Paulo) and two from other countries (Iran and Greece). With respect to the Brazilian populations, it can be interpreted that despite the intense miscegenation in the country, the genetic characteristics in relation to the SNP evaluated, have been conserved in these populations.

Regarding the SNP in the IL16 gene, no similarities were found when comparing the frequencies of our study with other populations.

The genotypic variety found in the comparison between the results of the present study and other countries leaves us the important observation that further population studies should be carried out in order to understand these similarities or differences found.

The mutant G allele of rs11556218 polymorphism in the IL-16 gene presents a possible protective action in relation to the development of ACS in the studied population.

Regarding the SNP rs4073 (IL8), no association was found with SCA.

The comparison between the frequency of the SNPs rs4073 (IL8) and rs11556218 (IL16) of the present study and other world populations demonstrated a wide genetic variety.

The main limitation of the study refers to the composition of the HEMOPE donor group, since it was not possible to match the ages between this group and the patients. Through literature search, it was possible to observe that the use of blood donors to compose a group of comparisons in genetic studies is adequate, since this group of individuals represents a healthy population in general.

Although it is known that the age for donation ranges from 16 to 69 years, and therefore the age of patients with ACS would be within this age group, during the collection of samples it was observed that the age obtained from the group of was less than 62 years. This fact generated a selection bias.

Therefore, the HEMOPE donor group was used only for comparison of the genotypic and allelic frequencies using the Williams G test and not to establish the odds ratios for the development of ACS.