Balkan Journal of Medical Genetics

DIFFICULTIES IN DIAGNOSING FABRY DISEASE IN PATIENTS WITH UNEXPLAINED LEFT VENTRICULAR HYPERTROPHY (LVH): IS THE NOVEL GLA GENE MUTATION A PATHOGENIC MUTATION OR POLYMORPHISM?
Aladağ N, Ali Barman H, Şipal A, Akbulut T, Özdemir M, Ceylaner S
*Corresponding Author: Nesim Aladağ, MD, nesimaladag@hotmail.com, Van Yüzüncü yıl University, Faculty of Medicine, Department of Cardiology, Van, Turkey. Phone: +90 544 961 31 69 / Fax: +90 432 486 54 07
page: 43

MATERIALS AND METHODS

Over 120 patients older than 30 years who were diagnosed with unexplained left ventricular (LV) wall thickness of ≥ 13 mm through echocardiography were screened for FD between March 2020 and March 2021. Patients were excluded who were suffering from major heart valve disease, significant hypertension, coarctation, strain conditions like aortic stenosis, earlier diagnosis of FD, previous history of any disease that was linked to LVH, or a family history of autosomal dominant hypertrophic cardiomyopathy (HCM) or FD. All participants provided their written informed consent. The study was conducted in conformance to the Helsinki Protocol and obtained the approval of the local ethics committee. The collection of peripheral venous blood samples from all the participants of the study was carried out in the EDTA tubes (2 ml) for performing mutation analysis. These samples were sent to a laboratory specialized in diagnosing genetic disorders. In all patients, GLA gene sequence analysis was carried out. If a GLA gene mutation was identified, measurements for α-Gal A enzyme activity and lyso-Gb3 levels were obtained. In patients with gene mutations, a family screening was also carried out. Trans-Thoracic Echocardiography Trans-thoracic echocardiography (TTE) on the left lateral decubitus position was performed in all patients after they had rested for at least 15 minutes (Philips iE33 Healthcare, Andover, Massachusetts, USA). Echocardiography images were obtained from four standard views (two-chamber apical, long-axis parasternal, short-axis parasternal and four-chamber apical). M-mode recording was used in 2D echocardiography to achieve the standard value of LV diameter and function. According to the recommendations of the American Society of Echocardiography, an average of at least three cardiac cycles was obtained to evaluate the M-mode echocardiogram, (13). The M-mode echocardiography was used to compute the left ventricular end-diastolic diameter (LVEDD), left ventricular endsystolic diameter (LVESD), left ventricular posterior wall thickness (LVPW) and interventricular septum (IVS) from the long-axis parasternal view in millimeters. The biplanar disc technique (modified Simpson’s rule) was used to compute the LV ejection fraction (EF). α-Gal A enzyme activity and lyso-Gb3 measurements α-Gal A DBS card study is performed by fluorimetric method. 4-Methylumbelliferyl-α-D-galactopyranoside (TRC, M334475) was used as substrate and N-Acetyl-Dgalactosamine (Sigma, A2795) was used as inhibitor. The reaction is stopped after a 3 mm dried blood spot (DBS) punch, inhibitor, and substrate incubation for 17 hours at 37°C. Fluorescence is recorded at Ex: 366 nm, Em: 442 nm in the fluorimeter. The calibration curve is created with 4-Methylumbelliferone (Sigma M1381) and the results are evaluated. The LC-MS/MS system is used to measure the lyso-Gb3 level. A 5 mm DBS punch is taken from standards, controls and samples, internal standard N-Gly- Lyso-Gb3 is added. After extraction, it is taken into vials and analyzed in 10 μL LC-MS/MS system. Mutation analysis - Polymerase Chain Reaction - Sequencing Peripheral blood samples of 200 μl were obtained from the study participants. These samples were stored at a temperature of -200 °C till the polymerase chain reaction (PCR) step was carried out. The design of the in-house PCR primers was done for the coding region and the exoneintron borders of the GLA gene. The Sanger technique was used to perform sequencing on a genetic analyser (Applied Biosystems Inc.). SeqScape 2.5.0 was used to evaluate the data (Applied Biosystems Inc.). Statistical Analysis Analysis was performed on the demographic properties and echocardiographic parameters of all the screened patients. The Statistical Package for the Social Sciences (SPSS) program, version 19.0 (SPPS Statistics IBM®) was used to perform the descriptive analysis of the data, and the outcomes were presented as percentages, numbers, or mean values ± a standard deviation.

Number 27 VOL. 27 (2), 2024	Number 27 VOL. 27 (1), 2024
Number 26 Number 26 VOL. 26(2), 2023 All in one	Number 26 VOL. 26(2), 2023
Number 26 VOL. 26, 2023 Supplement	Number 26 VOL. 26(1), 2023
Number 25 VOL. 25(2), 2022	Number 25 VOL. 25 (1), 2022
Number 24 VOL. 24(2), 2021	Number 24 VOL. 24(1), 2021
Number 23 VOL. 23(2), 2020	Number 22 VOL. 22(2), 2019
Number 22 VOL. 22(1), 2019	Number 22 VOL. 22, 2019 Supplement
Number 21 VOL. 21(2), 2018	Number 21 VOL. 21 (1), 2018
Number 21 VOL. 21, 2018 Supplement	Number 20 VOL. 20 (2), 2017
Number 20 VOL. 20 (1), 2017	Number 19 VOL. 19 (2), 2016
Number 19 VOL. 19 (1), 2016	Number 18 VOL. 18 (2), 2015
Number 18 VOL. 18 (1), 2015	Number 17 VOL. 17 (2), 2014
Number 17 VOL. 17 (1), 2014	Number 16 VOL. 16 (2), 2013
Number 16 VOL. 16 (1), 2013	Number 15 VOL. 15 (2), 2012
Number 15 VOL. 15, 2012 Supplement	Number 15 Vol. 15 (1), 2012
Number 14 14 - Vol. 14 (2), 2011	Number 14 The 9th Balkan Congress of Medical Genetics
Number 14 14 - Vol. 14 (1), 2011	Number 13 Vol. 13 (2), 2010
Number 13 Vol.13 (1), 2010	Number 12 Vol.12 (2), 2009
Number 12 Vol.12 (1), 2009	Number 11 Vol.11 (2),2008
Number 11 Vol.11 (1),2008	Number 10 Vol.10 (2), 2007
Number 10 10 (1),2007	Number 9 1&2, 2006
Number 9 3&4, 2006	Number 8 1&2, 2005
Number 8 3&4, 2004	Number 7 1&2, 2004
Number 6 3&4, 2003	Number 6 1&2, 2003
Number 5 3&4, 2002	Number 5 1&2, 2002
Number 4 Vol.3 (4), 2000	Number 4 Vol.2 (4), 1999
Number 4 Vol.1 (4), 1998	Number 4 3&4, 2001
Number 4 1&2, 2001	Number 3 Vol.3 (3), 2000
Number 3 Vol.2 (3), 1999	Number 3 Vol.1 (3), 1998
Number 2 Vol.3(2), 2000	Number 2 Vol.1 (2), 1998
Number 2 Vol.2 (2), 1999	Number 1 Vol.3 (1), 2000
Number 1 Vol.2 (1), 1999	Number 1 Vol.1 (1), 1998

About the journal ::: Editorial ::: Subscription ::: Information for authors ::: Contact