ABC | Volume 110, Nº1, January 2018

Original Article Fischer et al Gene polymorphisms and coronary disease extension Arq Bras Cardiol. 2018; 110(1):16-23 number of genetic polymorphism is evaluated. Therefore, the use of genetic scores that evaluate the additional effect of each of these genetic variants may be relevant in the early identification of individuals at higher cardiovascular risk, who may benefit from differential attention. 23-25 We chose polymorphisms related to lipid metabolism, lipoprotein oxidation, blood pressure changes and vasoreactivity to determine their possible association with coronary atherosclerosis severity in patients with MS and recent ACS. Studies on genetic polymorphisms in MS patients who developed ACS are poorly described in the literature, which reinforces the importance of the present study. We examined the contribution of combined and isolated genetic polymorphisms of potential importance in cardiovascular disease, by a genetic score, to the extension and severity of coronary obstructive disease in patients at very high cardiovascular risk. 26 Methods Subjects A total of 116 consecutive patients of both sexes, mean age 56 ± 9 years, with 3 or more criteria for MS according to the NCEPIII 4 were prospectively assessed during hospitalization for ACS (acute myocardial or unstable angina). Inclusion criteria: 1. Men and women aged from 30 to 75 years; 2. Understanding and agreement in giving written consent. 3. Two or more ACS criteria including chest pain, increased enzyme levels and electrocardiographic changes; 4. Three or more MS criteria, 4 5. LDL-c < 130 mg/dL, HDL-c < 40 mg/dL in the first 24 hours of hospitalization; Exclusion criteria: 1. Use of hypolipidemic agents in the last 30 days; 2. Uncontrolled hypothyroidism (TSH > 8.0 μU/mL); 3. Infections, inflammatory diseases, active liver or kidney diseases; 4. Pregnant women, patients selected for surgeries, including myocardial revascularization in the next six weeks or patients who underwent surgeries during hospitalization. Ethical aspects The study protocol was approved by the Research Ethics Committee and the study was conducted according to the Helsinki Declaration. All patients signed the informed consent form before participating in any study procedure. Study design This was a prospective study that included hospitalized patients after ACS. Genetic polymorphisms related to lipid and lipoprotein metabolism, oxidative stress, endothelial function and blood pressure were evaluated. Clinical evaluation Clinical evaluation and blood collection for laboratory measurements were performed in the first three days of hospital discharge. Demographic data, risk factors, medical history, clinical examination, anthropometric data (weight, height, waist circumference), systolic and diastolic blood pressure were obtained. 27,28 Blood collection for laboratory tests Blood was collected after 12 hours of fasting and laboratory tests were performed at the Associação Fundo de Apoio à Psicobiologia (AFIP - Psychobiology Support Fund Association). Genetic tests were carried out at the Molecular Biology Laboratory of the Division of Lipids, Atherosclerosis and Vascular Biology. Biochemical tests Lipid profile (total cholesterol, LDL-c, HDL-c, triglycerides) was analyzed by automated method (Ópera, Bayer, Germany); glucose levels were determined by colorimetric reaction (ADVIA 1650 Chemistry System, USA), glycated hemoglobin (HbA1-c) was determined by high performance liquid chromatography (HPLC, Tosoh A1c 2,2 plus, USA). Mean insulin at -15min, -5min and 0min was assessed by direct chemiluminescence method (ADVIA Centaur, USA) with values expressed as μU/mL. Apolipoproteins A1, B and Lp(a) were analyzed by nephelometry (R100 analyzer, Behringer). Adiponectin was determined by ELISA (enzyme‑linked immunosorbent assay), using the Humam Adiponectin/ Acrp30 Immunoassay kit – Quantiquine, R&D Systems, and the BioTek ELx800 Absorbance Microplate Reader, with values expressed as ng/mL. Albuminuria was assessed in 12-hour overnight urine samples, and measured by immunoturbidimetric method (ADVIA 1650, Chemistry System, USA), with results expressed as mg/L. Plasma oxidative stress was analyzed by TBARS (Thiobarbituric acid reactive substances) test, as described by Ohkawa et al., 29 and measured using a spectrophotometer (Genesys 2, Spectronic); the results were expressed as nanomoles of malondialdehyde (MDA) per milliliter of plasma ( η moles/mL plasma). Vasoreactivity Endothelium-dependent (FMD, flow-mediateddilation) and endothelium-independent (EIR, endothelium‑independent relaxation) vasodilation tests were performed in the morning, after a 12-hour fast by an experient ultrasound technician according to the International Brachial Artery Reactivity Task Force guidelines. 30 An ultrasound system (Sonos 5500; Hewlett-Packard-Phillips, Palo Alto, CA) was used, equipped with a vascular software for two-dimensional imaging, color and spectral Doppler, internal electrocardiogram (ECG) monitor and a linear array transducer (with a frequency of 7.5-12.0 MHz). Image acquisition, FMD and EIR measures with isosorbide dinitrate (5mg; sublingual) were performed. Percentage variations in vessel diameter were calculated for 17