Prolonged Right Ventricular Ejection Delay in Brugada Syndrome Depends on the Type of SCN5A Variant　― Electromechanical Coupling Through Tissue Velocity Imaging as a Bridge Between Genotyping and Phenotyping ―

Abstract

Background: Patients with Brugada syndrome (BrS) and a history of syncope or sustained ventricular arrhythmia have longer right ventricular ejection delays (RVEDs) than asymptomatic BrS patients. Different types of SCN5A variants leading to different reductions in sodium current (I_Na) may have different effects on conduction delay, and consequently on electromechanical coupling (i.e., RVED). Thus, we investigated the genotype-phenotype relationship by measuring RVED to establish whether BrS patients carrying more severe SCN5A variants leading to premature protein truncation (T) and presumably 100% I_Na reduction have a longer RVED than patients carrying missense variants (M) with different degrees of I_Na reduction.

Methods and Results: There were 34 BrS patients (mean [±SD] age 43.3±12.9 years; 52.9% male) carrying an SCN5A variant and 66 non-carriers in this cross-sectional study. Patients carrying a SCN5A variant were divided into T-carriers (n=13) and M-carriers (n=21). Using tissue velocity imaging, RVED and left ventricular ejection delay (LVED) were measured as the time from QRS onset to the onset of the systolic ejection wave at the end of the isovolumetric contraction. T-carriers had longer RVEDs than M-carriers (139.3±15.1 vs. 124.8±11.9 ms, respectively; P=0.008) and non-carriers (127.7±17.3 ms, P=0.027). There were no differences in LVED among groups.

Conclusions: Using the simple, non-invasive echocardiographic parameter RVED revealed a more pronounced ‘electromechanical’ delay in BrS patients carrying T variants of SCN5A.

Brugada syndrome (BrS) is an autosomal dominant inherited disease associated with syncope, life-threatening ventricular arrhythmias (VA), and sudden cardiac death (SCD).¹ Diagnosis of BrS depends on the occurrence of a spontaneous or drug-induced coved-type electrocardiogram (ECG) in the right precordial leads (V1–V3).^2,3 To date, 18 genes have been linked to BrS.^4,5 SCN5A, the first gene discovered and still the major gene associated with BrS, encodes the pore-forming α-subunit of the cardiac Nav1.5 voltage-gated sodium channel and is located on chromosome 3p21.^6,7 Loss-of-function mutations in SCN5A are identified in 20–30% of patients with BrS.⁷ These mutations result in reduced sodium channel availability, through either decreased membrane surface channel expression due to trafficking impairment or altered channel gating properties.^7–9 Because the sodium current (I_Na) plays a central role in the initiation and propagation of cardiac impulses,¹⁰ loss-of-function SCN5A mutations result in slowing of atrial and ventricular conduction.^11,12

Slowing of right ventricular (RV) conduction has been suggested as a pathophysiological mechanism underlying right precordial ST segment elevation and its relation to arrhythmogenesis in BrS.¹³ Previously, we demonstrated that BrS patients with a history of syncope or spontaneously sustained VA had significantly longer RV ejection delays (RVEDs) than asymptomatic BrS patients and controls.¹⁴ Surprisingly, the mean RVED of BrS patients with any SCN5A variant in that cohort was not significantly prolonged compared with non-carriers.¹⁴ We hypothesized that this lack of difference between SCN5A variant carriers and non-carriers could be attributed to the fact that different subtypes of SCN5A variants resulting in different degrees of I_Na reduction^12,15,16 may have different effects on conduction delay and RVED measurements. To date, this detailed genotype-phenotype relationship has not been investigated. Therefore, the aim of the present study was to determine whether BrS patients carrying a more severe SCN5A variant leading to premature protein truncation (T) and consequent abolishment of I_Na also had a longer electromechanical coupling time, based on measures of RVED, than patients carrying missense variants (M) with different degrees of I_Na reduction.

Methods

The present cross-sectional study was performed in accordance with the Declaration of Helsinki and was approved by the Ethics Committee of UZ Brussel. Written informed consent was obtained from all patients.

Study Population

The study was performed on 34 BrS patients carrying an SCN5A variant and 66 non-SCN5A carriers. All subjects had been diagnosed with BrS based on either a spontaneous or drug-induced ST segment elevation with a Type 1 morphology of >2 mm in more than 1 lead among the right precordial leads (V1–3). All patients were over 18 years of age and none had a previous history of pericarditis, ischemic heart disease, cardiomyopathy of any origin, structural heart disease, or any other channelopathy; nor did any subjects exhibit atrial fibrillation (AF) or pacing during echocardiography. To avoid confounding effects on measurements of ejection delays, only patients who were not using β-blockers (BB) or antiarrhythmic drugs (AAD) were included in the study. All available clinical data (gender, age, ethnicity, previous malignant events, such as syncope, spontaneous sustained VA, or aborted cardiac arrest) up to time that transthoracic echocardiography (TTE) was performed were collected.

Transthoracic Echocardiography

TTE was performed in patients in a left lateral decubitus position at rest using a commercial cardiac ultrasound system (Vivid 9; GE Vingmed Ultrasound, Horten, Norway) equipped with a 2-dimensional (2D) broad-band M3S transducer (2.5 MHz). Lead II was used for ECG recording. Color tissue Doppler myocardial images of the RV free wall and the LV lateral wall were recorded in the apical 4-chamber view with a median frame rate of 144.1 FPS (interquartile range [IQR] 94.1–203.3 FPS). Images and loops were digitally stored for offline analysis (EchoPac version 113; GE Vingmed Ultrasound).

Offline Tissue Velocity Imaging Analysis and Reproducibility of Measurements

As reported previously,¹⁴ a sample area was placed at the basal portions of both the RV free wall and the lateral LV wall in the color tissue Velocity imaging (TVI)-coded apical 4-chamber view. We used a 30 ms temporal smoothing filter and the QRS gain was increased to ensure correct assessment of the beginning of the QRS. In the absence of a Q wave, the onset of the QRS was determined at the beginning of the R wave. RVED and left ventricular ejection delay (LVED) were measured as the time from QRS onset to the onset of the systolic ejection wave at the end of the isovolumetric contraction (Figure 1).¹⁴ All timings were corrected for heart rate (HR) using Bazett’s formula. TTE analyses were performed separately and blinded with regard to other patient data. Intra- and interobserver variability was assessed and reported elsewhere.¹⁴

Figure 1.

Myocardial velocities measured by color tissue velocity imaging in (A) a carrier of a truncation (T) variant, (B) a carrier of an inactive missense mutation with >90% I_Na reduction (M_inactive), and (C) a carrier of an active missense mutation with <90% I_Na reduction (M_active). Color Doppler sample areas were placed at the basal portions of the lateral left ventricular (LV) wall (blue sample/curve) and the free right ventricular (RV) wall (yellow sample/curve). The onset of QRS onset (red arrowhead) was determined at the beginning of the R-wave if a Q-wave was absent (A). The ejection delay of the RV and LV (RVED and LVED, respectively) was measured as the time from QRS onset to the onset of the systolic ejection wave (open blue/yellow arrows) at the end of the isovolumetric contraction (asterisk). These ejection delays represent both electromechanical coupling and the conduction delay at each wall. Note the longer RVED in the patient carrying the T-mutation (A) and the M_inactive carrier (B) compared with the M_active carrier (C).

Electrocardiography

A standard 12-lead ECG was acquired before every TTE using a GE Healthcare MAC 5500 ECG Diagnosis System. The data assessed on the ECG were the PQ interval, QRS duration, the QTc interval, and the type of BrS repolarization pattern based on the second consensus document published in 2005 (Table 1).² Any previous ECGs (e.g., from medical records) that patients had undergone were also obtained.

Table 1. Patient Characteristics and Clinical Presentation of SCN5A Variant Carriers and Non-Carriers

	T variants (nall=13; nClassIV/V=13)	Mall variants (nall=21) (nClassIV/V=15)	Minactive variants (nall=6; nClassIV/V=6)	Mactive variants (nall=5; nClassIV/V=4)	Muncl variants (nall=10; nClassIV/V=5)	Noncarriers (n=66)	P value
							T vs. Mall	T vs. Minactive	T vs. Mactive	Minactive vs. Mactive
Males						37 (56.1)
All carriers	8 (61.5)	10 (47.6)	1 (16.7)	2 (40.0)	7 (70.0)		NS	NS	NS	NS
Carriers of Class IV/V mutations	8 (61.5)	4 (26.7)	1 (16.7)	1 (25.0)	3 (60.0)		NS	NS	NS	NS
Age (years)						44.1±15.1
All carriers	46.4±11.3	41.4±13.7	34.7±12.8	44.6±10.5	43.9±15.3		NS	NS	NS	NS
Carriers of Class IV/V mutations	46.4±11.3	39.9±12.8	34.7±12.8	44.0±12.0	42.8±13.9		NS	NS	NS	NS
BMI (kg/m2)						24.2±3.2
All carriers	25.6±3.5	25.2±3.7	26.3±3.4	26.5±3.1	24.3±4.1		NS	NS	NS	NS
Carriers of Class IV/V mutations	25.6±3.5	25.9±3.8	26.3±3.4	26.8±3.5	25.4±0.31		NS	NS	NS	NS
SBP (mmHg)						120.7±14.5
All carriers	119.6±10.5	119.6±14.5	121.0±13.1	129.8±12.7	111.6±13.1		NS	NS	NS	NS
Carriers of Class IV/V mutations	119.6±10.5	121.2±16.8	121.0±13.1	131.0±14.3	108.3±20.2		NS	NS	NS	NS
DBP (mmHg)						75.0 [70.0–80.0]
All carriers	75.0 [70.0–80.0]	75.0 [60.0–80.0]	80.0 [65.0–80.0]	80.0 [60.0–82.5]	70.0 [60.0–80.0]		NS	NS	NS	NS
Carriers of Class IV/V mutations	75.0 [70.0–80.0]	80 [60.0–80.0]	80.0 [65.0–80.0]	70.0 [60.0–83.8]	70.0 [50.0–/]		NS	NS	NS	NS
HR (beats/min)						68.1±9.5
All carriers	65.6±11.4	63.5±8.7	66.1±5.9	63.2±6.2	61.6±11.1		NS	NS	NS	NS
Carriers of Class IV/V mutations	65.6±11.4	65.0±9.0	66.1±5.9	65.0±5.5	63.6±14.6		NS	NS	NS	NS
Ethnic group
Asian
All carriers	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)	1 (1.5)	NS	NS	NS	NS
Carriers of Class IV/V mutations	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)		NS	NS	NS	NS
African						1 (1.5)
All carriers	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)		NS	NS	NS	NS
Carriers of Class IV/V mutations	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)		NS	NS	NS	NS
Caucasian						64 (97.0)
All carriers	13 (100.0)	21 (100.0)	6 (100.0)	5 (100.0)	10 (100.0)		–	–	–	–
Carriers of Class IV/V mutations	13 (100.0)	15 (100.0)	6 (100.0)	4 (100.0)	5 (100.0)		–	–	–	–
Clinical presentation
Syncope						25 (37.9)
All carriers	5 (38.5)	8 (38.1)	2 (33.3)	2 (40.0)	4 (40.0)		NS	NS	NS	NS
Carriers of Class IV/V mutations	5 (38.5)	4 (26.7)	2 (33.3)	1 (25.0)	1 (20.0)		NS	NS	NS	NS
Spontaneous VT/VF						12 (18.2)
All carriers	1 (7.7)	2 (9.5)	0 (0.0)	1 (20.0)	0 (0.0)		NS	NS	NS	NS
Carriers of Class IV/V mutations	1 (7.7)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)		NS	NS	NS	–
ACA						5 (7.6)
All carriers	2 (15.4)	1 (4.8)	0 (0.0)	0 (0.0)	1 (10.0)		NS	NS	NS	–
Carriers of Class IV/V mutations	2 (15.4)	1 (6.7)	0 (0.0)	0 (0.0)	1 (25.0)		NS	NS	NS	–
ECG features
PQ (ms)						149.9±22.7
All carriers	221.8±42.4*	183.9±34.4*	184.0±24.2*	156.8±33.0	197.4±34.8*		0.010	NS	0.009	NS
Carriers of Class IV/V mutations	221.8±42.4*	190.0±27.0*	184.0±24.2*	170.0±17.0	213.2±21.8*		0.048	NS	0.013	NS
QRS (ms)						102.5±22.7
All carriers	129.6±24.6*	114.6±15.6*	109.0±11.6	100.0±5.8	125.2±13.7*		0.042	NS	0.020	NS
Carriers of Class IV/V mutations	129.6±24.6*	113.2±17.1*	109.0±11.6	99.0±6.2	129.6±16.3*		0.035	0.030	0.005	NS
Type 1 at TTE						10 (15.2)
All carriers	2 (14.3)	2 (9.5)	0 (0.0)	0 (0.0)	2 (20.0)		NS	NS	NS	–
Carriers of Class IV/V mutations	2 (14.3)	0 (0.0)	0 (0.0)	0 (0.0)	0 (0.0)		NS	NS	NS	–
Intermittent Type 1						10 (15.2)
All carriers	6 (46.2)*	9 (42.8)*	1 (16.7)	0 (0.0)	8 (80.0)*		NS	NS	NS	–
Carriers of Class IV/V mutations	6 (46.2)*	5 (33.3)	1 (16.7)	0 (0.0)	4 (80.0)*		NS	NS	NS	–
Dynamic ECG						17 (35.4)
All carriers	5 (38.5)	8 (38.1)*	2 (33.3)	1 (20.0)*	1 (20.0)*		NS	NS	NS	NS
Carriers of Class IV/V mutations	5 (38.5)	6 (40.0)	2 (33.3)	1 (25.0)	3 (60.0)*		NS	NS	NS	NS
General TTE measurements
TAPSE (mm)						24.4±4.5
All carriers	25.5±4.1	23.2±6.3	22.2±5.6	23.0±7.1	23.8±6.8		NS	NS	NS	NS
Carriers of Class IV/V mutations	25.5±4.1	23.1±1.4	22.2±5.6	25.5±4.9	22.2±6.7		NS	NS	NS	NS
LVEF (%)						72.4±7.7
All carriers	75.2±8.7	73.9±7.5	71.2±10.1	72.9±5.7	76.0±6.5		NS	NS	NS	NS
Carriers of Class IV/V mutations	75.2±8.7	72.4±2.0	71.2±10.1	70.8±3.9	75.1±8.1		NS	NS	NS	NS
Genetic variants
Class I	0 (0.0)	3 (14.3)	0 (0.0)	0 (0.0)	3 (30.0)
Class II	0 (0.0)	1 (4.8)	0 (0.0)	1 (20.0)	0 (0.0)
Class III	0 (0.0)	2 (9.5)	0 (0.0)	0 (0.0)	2 (20.0)
Class IV	11 (84.6)	13 (61.9)	6 (100.0)	4 (80.0)	3 (30.0)
Class V	2 (15.4)	2 (9.5)	0 (0.0)	0 (0.0)	2 (20.0)

Variables with a normal distribution are presented as the mean±SD, variables with an abnormal distribution are presented as the median [interquartile range]. Other data are presented as n (%). *P<0.05 compared with non- SCN5A variant carriers (P values are provided in the text). Variants were categorized as not pathogenic (Class I), unlikely pathogenic (Class II), unknown pathogenicity (Class III), likely pathogenic (Class IV), and (putative) pathogenic (Class V). ACA, aborted cardiac arrest; BMI, body mass index; DBP, diastolic blood pressure; ECG, electrocardiogram; HR, heart rate; LVEF, left ventricular ejection fraction (determined by the Teichholz method); M_all, all missense variants, including M_inactive, M_active, and M_uncl; M_active, active missense mutation with <90% I_Na reduction; M_inactive, inactive missense mutation with >90% I_Na reduction; M_uncl, unclassified missense mutation with unknown I_Na reduction; n_all, number of all SCN5A carriers (Class I–V); n_ClassIV/V, number of Class IV and Class V SCN5A carriers; SBP, systolic blood pressure; SCN5A, gene encoding the α-subunit of the cardiac voltage-gated sodium channel; T, truncation; TAPSE, tricuspid annular plane systolic excursion; TTE, transthoracic echocardiography.

Genetic Analysis

Total genomic DNA was isolated from whole blood samples using standard techniques (Chemagen; PerkinElmer, Zaventem, Belgium). SCN5A mutation analysis of all 28 exons and flanking intron-exon boundaries was performed by high resolution melting curve analysis (HRMCA) as a first-line mutation detection assay using a Lightcycler 480 reverse transcription-polymerase chain reaction (RT-PCR) instrument (Roche Applied Science). In regions with aberrant HRMCA profiles, the exons and flanking intronic sequences were amplified using PCR and Sanger sequenced on an ABI 3130xl DNA Sequence Detection System (Applied Biosystems, Foster City, CA, USA). Variants were classified according to the system suggested by Hofman et al.¹⁷ This classification system is based on the results of various pathogenicity prediction tools, databases that store variants from healthy individuals, databases containing variants already associated with a disease, and information from familial variant segregation analysis and functional studies described in literature. Variants were categorized into five classes: not pathogenic (Class I), unlikely pathogenic (Class II), unknown pathogenicity (Class III), likely pathogenic (Class IV), and (putative) pathogenic (Class V).

T and M Variants

SCN5A variants were divided in 2 groups as described previously.¹² The first group consisted of variants leading to premature protein truncation (T variants) such as splice site, nonsense, and frameshift variants. These variants result in a truncated, incomplete protein with a significant change in channel gating properties and a decreased number of functional channels expressed in the membrane due to folding or trafficking anomalies. Consequently, T variants are believed to result in complete loss of I_Na or a more pronounced reduction in I_Na compared with M variants. M variants, defined by a single amino acid exchange, constitute the second group of SCN5A variants in the present study. All M variants (M_all) were divided into 3 subgroups based on previously published biophysical properties (Table 2). Partially active M variants (M_active) were defined by a <90% peak reduction in I_Na (or >10% I_Na availability) in patch-clamp studies; functionally inactive M variants (M_inactive) were defined by a >90% peak reduction in I_Na (or <10% I_Na availability); and unclassified M variants (M_uncl) were defined as those with an unknown peak reduction in I_Na. The 90% cut-off value is based on the dynamic multicellular Luo-Rudy fiber model, which incorporates kinetic properties of I_Na and I_Ca(L) and states that I_Na availability <11.25% results in a difficulty to reach the cell membrane threshold.¹⁶ Variant characteristics are listed in Table 2.

Table 2. SCN5A Variants

Nucleotide change	Amino acid change	Type	Location	Classification	No. patients	No. families	Segregation within families	Type 1 ECG	Symptoms	SCD/ACA in family	INa peak reduction (%)
T variants
c.[2466G>A];[=]26–28	p.(W822*)	N	E16, transmembrane helical DII-S4	IV	1	1	–	No	Dynamic ECG, palpitations	No	100
c.[4813+3_4813+ 6dupGGGT];[=]7,29,30	–	Sp	I27	IV	4	2	5/8, 21/23	Yes (3, 4)	None (0/1, 1/3), syncope (1/1, 1/3), EPS+ (0/1, 2/3)	Yes	100
c.[2092G>T];[=]7	p.(E698*)	N	E14, cytoplasmic DI/DII	IV	1	1	–	Yes	Inducible VT	No	100A
c.[5356_5357delCT];[=]7,31	p.(L1786Efs*2)	F	E28, cytoplasmic C-terminal	V	2	1	3/3	No	None (1/2), syncope, ACA, EPS+ (1/2)	Yes	100
c.[4300–2A>T];[=] (novel)	–	Sp	I24	V	2	1	8/8	Yes (1/2)	None (2/2)	Yes	100A
c.[3840+1G>A];[=]7,15,32,33	–	Sp	I21	IV	1	1	–	No	Syncope, spontaneous VT	Yes	100
c.[−53+1G>A];[=] (novel)	–	Sp	I1	IV	1	1	4/5	Yes	Syncope, ACA, AFl	No	100A
c.[4083delG];[=] (novel)	p.(R1362Gfs*12)	F	E23, extracellular DIII-S5/S6	IV	1	1	1/2	No	None	Yes	100A
Minactive variants
c.[2632C>T];[=]7,34–36	p.(R878C)	M	E16, extracellular DII-S5/S6	IV	6	1	27/30	Yes (1/6)	None (3/6), EPS+ (1/6), syncope (2/6)	Yes	100
Mactive variants
c.[1715C>A];[=]37–40	p.(A572D)	M	E12, cytoplasmic DI/DII	II	1	1	1/6	No	Syncope, spontaneous VT/VF	No	0
c.[4895G>A];[=]34,41	p.(R1632H)	M	E28, transmembrane helical, voltage sensor, DIV-S4	IV	4	1	9/10	No	None (3/4), syncope, AFl (1/4)	No	0–10
Muncl variants
c.[4346A>G];[=]7,42	p.(Y1449C)	M	E25, transmembrane helical, DIII-S6	IV	1	1	1/2	Yes	Syncope, AFl, VT after flecainide	No	–
c.[3673G>A];[=]7,22,43	p.(E1225K)	M	E21, extracellular, DIII-S1/S2	III	1	1	–	Yes	Syncope	No	–
c.[4981G>A];[=]7	p.(G1661R)	M	E28, transmembrane helical, DIV-S5	V	2	1	7/7	Yes (1/2)	None (2/2)	Yes	–
c.[1003T>C];[=] (novel)	p.(C335R)	M	E9, extracellular, DI-S5/S6	IV	1	1	–	Yes	SSS, AF, AFl, TA UC	Yes	–
c.[4283C>T];[=]7	p.(A1428V)	M	E24, extracellular, DIII-S5/S6	III	1	1	4/6	Yes	Syncope	Yes	–
c.[274–24C>T];[=]7,44,45	–	I	I2	I	3	3	2/5, 1/1,1/1	Yes (2/3)	None (2/3), syncope (1/3)	Yes	–
c.[5189C>A];[=] (novel)	p.(P1730H)	M	E28, extracellular, DIV-S5/S6	IV	2	2	2/5	Yes	ACA, spontaneous VF	No	–

Variants were categorized as not pathogenic (Class I), unlikely pathogenic (Class II), unknown pathogenicity (Class III), likely pathogenic (Class IV) and (putative) pathogenic (Class V). ^ASodium current (I_Na) reduction is predicted to be 100% (based on the type of mutation; i.e., T and/or F). AF, atrial fibrillation; AFl, atrial flutter; D, transmembrane domain; EPS+, positive electrophysiological study with inducible sustained ventricular tachycardia (VT) or ventricular fibrillation (VF); E, exon; F, frameshift; I, intronic; M, missense; N, nonsense; S, segment; SCD, sudden cardiac death; Sp, splice site; SSS, sick sinus syndrome; TA UC, traffic accident of unknown cause at 36 years of age. Other abbreviations as in Table 1.

Statistical Analysis

Normality of distribution was assessed using the Shapiro-Wilk test. Descriptive statistics are presented as the mean±SD for continuous variables if normally distributed, or as the median and IQR for variables with a non-normal distribution. Where appropriate, continuous data were compared using the Kruskal-Wallis test, Mann-Whitney test, and paired Student’s t-test. Categorical data are expressed as percentages and were compared using the Chi-squared test. Receiver operating characteristic (ROC) curves, the area under the curve (AUC) of the ROC, sensitivity, and specificity were calculated to determine the value of RVED that best differentiates BrS patients with T variants from BrS patients with M_all, M_active, and M_inactive SCN5A variants. Analyses were performed using SPSS version 21 (IBM Corp., Armonk, NY, USA) and 2-sided P<0.05 was considered statistically significant.

Results

Patient Characteristics and Clinical Presentation

Thirty-four BrS patients with an SCN5A variant (mean age 43.3±12.9 years; 52.9% male) and 66 non-carriers (mean age 44.1±15.1 years; 56.1% male) were enrolled in the present cross-sectional study. The mean follow-up period from diagnosis until inclusion in the study was 81.9±73.9 months for SCN5A variant carriers and 52.5±51.3 months for non-carriers. Of the SCN5A variants, 13 (38.2%) were T, 6 (17.6%) were M_inactive, 5 (14.7%) were M_active, and 10 (29.4%) were M_uncl. Of the 13 T variants, 2 were nonsense, 3 were frameshifts, and 8 were splice site variants (Table 2). The clinical and genotypic characteristics of these groups are listed in Table 1.

No significant differences between subgroups were found regarding malignant (arrhythmic) events or the presence of a Type 1 ECG during TTE. However, compared with non-carriers, more intermittent Type 1 ECG changes were seen during follow-up in T- and M_all-carriers (P=0.011 for both), and more dynamic ECG changes were observed in M_all-carriers (P=0.001). T, M_all, and M_uncl variants had longer PQ (P<0.001, P<0.001, and P=0.002, respectively) and QRS (P<0.001 for all) intervals than non-carriers. M_inactive variants only had longer PQ values than non-carriers (P=0.001; Table 1).

RV Ejection Delay

Ejection delays in relation to the nature of SCN5A variants are shown in Figure 2. T-carriers had longer RVEDs than M_all-carriers (139.3±15.1 vs. 124.8±11.9 ms, respectively; P=0.008), M_active-carriers (125.5±19.0 ms; P=0.035), and M_uncl-carriers (125.3±9.7 ms; P=0.019). In contrast, no significant difference regarding RVED was found between T- and M_inactive-carriers, and only T-carriers had longer RVEDs than non-carriers (127.7±17.3 ms; P=0.027). No differences were found in LVED between any subgroups (Figure 2). ROC analysis revealed that an RVED of 128.9 ms had 76.2% specificity and 76.9% sensitivity to separate patients with T variants from patients with M_all variants (AUC 0.810; 95% CI 0.650–0.969; P=0.003), with a negative predictive value (NPV) of 84.2% and a positive predictive value of (PPV) of 66.7%. An RVED of 128.2 ms had 80.0% specificity and 76.9% sensitivity to separate patients with T variants from patients with M_active variants (AUC 0.846; 95% CI 0.663–1.000; P=0.027), with an NPV of 57.1% and a PPV of 90.9%. No significant cut-off value was determined to differentiate patients with T variants from patients with M_inactive variants (P=0.066).

Figure 2.

Ejection delays in relation to the type of SCN5A variant in all carriers and non-carriers. RVED, right ventricular ejection delay; LVED, left ventricular ejection delay; T, truncated variant (nonsense, splice site, frameshift); M_all, missense variants; M_inactive, inactive missense variant with >90% I_Na reduction; M_active, active missense variant with <90% I_Na reduction; M_uncl, unclassified missense variant; SCN5A–, non-SCN5A variant carrier. Data are presented as the mean±SD. *P<0.050 compared with T; **P<0.050 compared with SCN5A–.

All SCN5A Variants vs. Only Class IV/V Variants

When looking only at values of BrS patients carrying a Class IV or V variant, RVED remained significantly longer in T-carriers than M_all-carriers (139.3±15.1 vs. 124.8±13.8 ms, respectively; P=0.013), M_active-carriers (121.9±6.6 ms; P=0.044), and non-carriers (127.7±17.3 ms; P=0.027). Although RVED values of M_uncl-carriers remained shorter (126.2±13.0 ms), they did not differ significantly from those of T-carriers. Again, no differences in LVED were observed between any subgroups.

Discussion

RV conduction delay in BrS has been demonstrated ‘purely electrically’ on standard and signal-averaged ECG recordings,^{12,15,19–21} as well as on body surface mapping.^21,22 The reduced I_Na decreases action potential upstroke velocity and results in prolongation of PQ and QRS intervals, and more PQ and QRS widening following challenge with Na channel blockers, especially in those carrying a more severe T SCN5A variant.^12,15,19 The data in the present study confirm the presence of the most prolonged baseline PQ and QRS intervals in carriers of T SCN5A variants (Table 1).

In addition, RV conduction delay has also been confirmed ‘electromechanically’ by using TVI to determine the RVED.^14,18 The present study confirms the hypothesis that RVED differs significantly among patients carrying different SCN5A variants. A longer RVED in T- than M-carriers reflects the increased effect of the variant subtype on Nav1.5 sodium channel dysfunction, and consequently on conduction delay. As anticipated, a complete or more pronounced loss of I_Na in T-carriers thus results in an increased ‘electromechanical’ delay (from QRS onset to onset of RV ejection) in this particular BrS subpopulation, whereas M-carriers exhibit shorter electromechanical delays equal to those of non-carriers.

These findings provide 2 major explanations for the absence of a significant RVED prolongation in BrS patients with any SCN5A variant compared with non-carriers in our previous study.¹⁴ First, M_all-carriers exhibit a shorter RVED than T-carriers and this difference is driven primarily by the shortest RVED of the M_active-carriers. Mall-variants thus shorten the mean RVED of all SCN5A variant-carriers and neutralize the impact of T-carriers in the mixed group of all SCN5A variant-carriers. Second, non-carriers may carry variants in other genes with differing modulatory effects on cardiac I_Na that could consequently also prolong RVED. These genes may involve regulatory subunits (i.e., β-subunits, glycerol-3-phosphate dehydrogenase 1-like gene [GPD1-L], sarcolemma-associated protein [SLMAP]) or associated proteins (i.e., Connexin 43) interacting with Nav1.5.^4,23–25

In conclusion, merely comparing SCN5A variant carriers with non-carriers is not sufficient any more to predict the phenotypic effect of an SCN5A variant on parameters of electromechanical coupling such as RVED. Even more, future analyses on SCN5A variant-specific RV fibrosis or Connexin 43 expression, or SCN5A variant-based functional genetic studies could further clarify the delayed ‘electromechanical coupling process’ seen in T-carriers vs. M- or non-carriers.

Study Limitations

It is important to realize that BrS is an orphan disease with a very low incidence, and that only 20–30% of BrS patients carry a (likely) pathogenic SCN5A variant.⁷ Moreover, only carriers with adequate RV tissue velocity images and not taking any BB or AAD were included in the present study in order to obtain reliable data. A study analyzing 34 SCN5A variant carriers to assess the link between genetics and parameters of ‘electromechanical’ coupling is thus quite unique. As in previous studies,¹² when analyzing data between carriers of different SCN5A variants, some clinical differences did not reach statistical significance because of the relatively small overall and subgroup numbers of patients and families. Therefore, and despite the fact that their biophysical data are not available for reclassification as M_active- or M_inactive-carriers, M_uncl-carriers were not excluded from the present study because they added valuable information to the entire M_all group.

Future Perspectives

T-carriers have been associated with increased conduction delay, proven both electrically¹² and now electromechanically. Therefore, additional studies with a larger number of SCN5A variants should determine the potential role of genetic testing, especially differentiating between different types of (likely) pathogenic SCN5A variants in future diagnostic work-up and risk stratification.

Conclusions

Using the simple, non-invasive echocardiographic parameter RVED, the present study links BrS patients carrying T variants to a more pronounced electromechanical delay. In addition, deformation imaging again confirmed its pivotal role in clarifying and understanding complex cardiac pathophysiology.

Disclosures

There were no relationships with the industry, and no grants, contracts or other forms of financial support in relation to this study.

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