ABC | Volume 113, Nº1, July 2019

Original Article Chaves-Markman et al A novel mutation in Fabry disease Arq Bras Cardiol. 2019; 113(1):77-84 seem to be present in the early stages of life; however, it is usually not clinically detectable until the third or fourth decade of life. Deposits of glycosphingolipids in valves and coronary vessels are frequent, whichmay cause complete atrioventricular block, mitral insufficiency, left ventricular hypertrophy (LVH), or myocardial ischemia. 10 Currently, specific mutations are associated with the cardiac variant showing myocardial hypertrophy that is clinically similar to HCM. 11,12 Patients with FD are at risk for developing cerebrovascular disease (CVD), cardiac sudden death, and renal failure, and these patients can benefit from specific treatments. 13–15 In this study, a screening for the GLA gene was performed in a group of patients with echocardiographic diagnosis of hypertrophic cardiomyopathy (HCM). Methods Subjects A cross-sectional study was conducted in a convenience sample of patients with HCM treated at the cardiology outpatient clinic at a university hospital in Recife, Pernambuco, Brazil. All patients with an echocardiographic diagnosis of HCM based on the European Society of Cardiology 16 criteria were included. Patients with coronary artery disease, valvulopathies and hypertensive cardiomyopathy were excluded. Definitions of HCM A Transthoracic Echocardiography (TTE) was used to establish the diagnosis of HCM based on the following criteria: unexplained maximum ventricular thickness (MVT) ≥ 15 mm in any cardiac segment or a MVT ≥ 13 mm in a patient with family history of HCM. The obstructive presentation was defined by a left ventricular outflow gradient ≥ 30 mmHg at rest or after Valsalva maneuver or in orthostatism. 13,16 Echocardiographic assessment The TTE was previously performed in all subjects to establish the diagnosis of HCM (Vivid 7 or Vivid E9 GE) with offline dataset analysis (EchoPac®, GE Healthcare, Little Chalfont, United Kingdom). The diameters of the interventricular septum and the left ventricular (LV) posterior wall, LV end diastolic volume, and LV end systolic volume were determined through M-mode or 2D imaging. The MVT of all segments were measured in the parasternal short axis. The ejection fraction (EF) and diastolic function were calculated using Simpson’s method and pulsatile Doppler, respectively. 17,18 Magnetic resonance imaging Magnetic resonance imaging (MRI) was performed in all patients with GLA mutation to assess the presence of myocardial fibrosis using the delayed myocardial enhancement technique (DMET). A 1.5-T MRI scanner (MAGNETOM Essenza, Siemens Healthcare, Erlangen, Germany) with an eight-channel phased array coil was used. The images were acquired in the cardiac short axis, from the base to the apex, with 8-mm slices and 2-mm intervals using the T1-weighted echo-gradient sequence. An inversion recovery prepulse and adjusted inversion time were used to neutralize the myocardial signal. Image acquisition was started at approximately 8–10 min after the infusion of gadolinium contrast at a dosage of 0.2 mmol/ kg. DMET increases the amount of contrast between the normal tissue (dark due to signal neutralization) and the fibrotic tissue (white due to enhancement of gadolinium in the T1-weighted sequence). 19 Electrocardiography and Dynamic electrocardiography (24-h Holter) A resting 12-lead electrocardiogram was performed in all patients. A continuum digital recorder ( Cardio Light ®, Cardio Sistemas Comercial Industrial, São Paulo, Brazil) was used to record and analyze the Holter tests for 24 h. The electrodes were positioned in the electrocardiographic derivationsMV1,MV4, and MV6. The results were analyzed using the Cardiomanager 540® (Cardio Sistemas Comercial Industrial, São Paulo, Brazil) software by an independent observer searching for cardiac arrhythmias. Molecular and enzymatic essays All included patients underwent digit blood capillary puncture and blood samples were collected on filter paper. The samples were dried for 3–4h at room temperature, stored in a plastic envelop at 4°C, and sent to the Centogene Laboratory (Rostock, Germany). Molecular analysis to determine GLA gene mutations was performed in the samples from the female subjects, whereas mutation analysis was performed after evidence of low α -Gal A activity in male subjects. The expression level and enzymatic activity of the biomarker lyso-Gb3 were identified through high-performance liquid chromatography and tandem mass spectrometry. The GLA gene was analyzed using polymerase chain reaction (PCR) and sequencing of all coding regions and highly conserved exon–intron boundaries through next-generation sequencing with Illumina HiSeq (Illumina, California, USA). GLA gene analysis was performed in all patients with HCM. Statistical analyses Data were analyzed descriptively using the Statistical Package for the Social Sciences version 20.0 (IBM Company, Armonk, NY, USA). Before analyzing the continuous variables, the data sets were tested for normality by performing the Shapiro-Wilk test. Normally-distributed continuous variables were presented with measures of central tendency and dispersion (mean and standard deviation), and categorical variables were described as absolute (n) and relative (%) frequencies. Comparative analysis was performed using Pearson’s chi-square test for categorical variables. Numerical variables were analyzed using the paired Student’s t -test. The significance level was defined as 5% throughout the entire statistical analysis (p < 0.05). 78