Elevated serum interleukin-6 is predictive of coronary artery disease in intermediate risk overweight patients referred for coronary angiography

We have conducted a prospective cross-sectional study with patients referred for coronary angiography due to non-acute chest pain in a reference cardiology center at Hospital de Clínicas of Porto Alegre, Brazil. Between 2013 and 2014 all patients referred to elective coronary angiography for non-acute chest pain were screened. Patients who fulfilled inclusion and exclusion criteria and accepted to participate were included in the study. All participants were in agreement to have their data published and signed a written informed consent, revised by the local ethics committee which than approved the study protocol.

Inclusion criteria were: age between 40 and 70 years and chronic chest pain. We have excluded patients with a known history of diabetes, previous acute coronary syndrome or stroke, previous coronary artery revascularization, estimated glomerular filtration rate below 45 mL/min/L. 73 m², presence of class IV-NYHA congestive heart failure, chronic obstructive pulmonary disease, body mass index above 44 kg/m², previous organ transplantation, current evaluation for any organ transplantation and those with presence of rheumatic, endocrine or infectious chronic diseases. We also excluded patients using any medication that could modify glucose-insulin metabolism such as insulin, metformin, sulfonylureas and patients using corticosteroids, HIV anti-retrovirals, carbamazepine, phenytoin, drugs for cancer, immunosuppressor drugs, nitrofurantoin, anti-malarics, lithium and anti-psicotic drugs.

Blood pressure was measured at the catheterization laboratory after 15 min of rest, in the sitting position, in the right arm, in 3 sequential measurements, using an automatic aneroid sphygmomanometer (OMRON Comfort III Visomat Incoterm, Germany). We considered the lowest blood pressure result as the final measure. Blood samples were collected between 12 and 24 h after coronary angiography. For serum IL-6 measurement, a custom Luminex^® assay was employed (Invitrogen^®, #LHB0001CM) following the manufacturer orientation. Briefly, 50 µL of undiluted sample was added in a well containing buffers and magnetic beads. After 2 h of incubation (550 rpm, room temperature), the wells were washed and the detection antibody was added for further 1 h. After washing, streptavidin-RPE was added for further 30 min, the wells were washed again and the beads were suspended in 125 µL of wash buffer. Beads were read in Luminex^® x-Map 200 and a minimum of 100 events were recorded for each bead. The limit of detection was defined by the lowest standard value (0.08, pg/mL). Values were expressed as pg/mL.

Serum high sensitive C-reactive protein (hs-CRP) measurements were determined through turbidimetric immunoassay method (Roche^®). Serum creatinine (Jaffé method), lipid profile, glycated hemoglobin (HPLC) were also measured.

The atherosclerotic cardiovascular disease (ASCVD) risk score was calculated based on AHA/ACC 2013 guidelines to estimate the 10-year risk score for men and women from 40 to 79 years of age for a first hard ASCVD event. The variables to estimate each patient risk included were age, gender, race, total cholesterol, high-density lipoprotein cholesterol, systolic blood pressure, actual treatment for high blood pressure, diabetes and current smoking status [3]. The ASCVD risk score calculator was obtained at: http://​tools.​acc.​org/​ASCVD-Risk-Estimator/​. We considered intermediate risk when the 10 years score was between 7.5 and 20%.

Coronary angiography was performed using the Axiom Artis Siemens^® equipment (Germany) in all patients. Two experienced interventional cardiologists who were blinded to all other clinical variables made all angiographic measurements. Angiographic analyses were made by visual (non-quantitative) estimates of luminal narrowing in at least two different orthogonal projections.

The presence of CAD was defined as any lesion causing more than 30% reduction in  artery diameter of any epicardial vessel. We considered not significant coronary disease (NCAD) when lesions were undetectable or below to 30% stenosis.

The severity of CAD was further assessed through the “anatomic burden score” obtained from the COURAGE trial [16]. This score consisted in a grading scale of 17 progressive degrees of severity starting from zero, corresponding to complete absence of coronary disease with stenosis above 50%, to 17, which corresponds to severe 3-vessels disease, including lesions at proximal left anterior descending artery, plus left circumflex artery and right coronary involvement. To meet criteria, each lesion must represent at least 50% diameter stenosis. We divided patients into three groups: NONE for patients with score zero; MILD–MODERATE, for patients with score 1–5, corresponding to isolate lesions in right coronary artery, left circumflex artery and left anterior descending artery, increasing score in this order; and SEVERE, for patients with scores between 6 and 17, corresponding to two-vessel and three-vessel artery disease.

Continuous variables with parametric distribution were expressed as mean ± standard deviation, whereas non-parametric variables levels were expressed as median [95% confidence interval (CI)] and analyzed using Mann–Whitney’s test. Categorical data were expressed as frequencies and their differences were analyzed using the Chi-squared test in the general characteristic Table 1. Receiver operating characteristic (ROC) curves were used to evaluate the discriminatory power of IL-6 to determine CAD. The severity of CAD (burden score) analysis was performed through ANOVA with Bonferroni post-tests. Comparison of ROC curves was performed by comparing area under the curves through trapezoid method and C statistics. We used the following cut-off values for analysis: IL-6: 1.0 pg/mL, hs-CRP and ASCVD score. Statistical analyses were performed using SPSS version 15.0 (SPSS Inc., Chicago, Illinois).