Basal characteristic and risk factors.
Radiofrequency ablation (RFA) in Brugada syndrome (BrS) has been performed by both endocardial and epicardial. The substrate in BrS is not completely understood. We investigate the functional endocardial substrate and its correlation with clinical, electrophysiological and ECG findings in order to guide an endocardial ablation. Two patients agreed to undergo an endocardial biopsy and the samples were examined with transmission electron microscopy (TEM) to investigate the correlation between functional and ultrastructural alterations. About 13 patients (38.7 ± 12.3 years old) with spontaneous type 1 ECG BrS pattern, inducible VF with programmed ventricular stimulation (PVS) and syncope without prodromes were enrolled. Before endocardial mapping, the patients underwent flecainide testing with the purpose of measuring the greatest ST-segment elevation for to be correlated with the size and location of substrate in the electro-anatomic map. Patients underwent endocardial bipolar and electro-anatomic mapping with the purpose of identify areas of abnormal electrograms (EGMs) as target for RFA and determine the location and size of the substrate. When the greatest ST-segment elevation was in the third intercostal space (ICS), the substrate was located upper in the longitudinal plane of the right ventricular outflow tract (RVOT) and a greatest ST-segment elevation in fourth ICS correspond with a location of substrate in lower region of longitudinal plane of RVOT. A QRS complex widening on its initial and final part, with prolonged transmural and regional depolarization time of RVOT corresponded to the substrate located in the anterior-lateral region of RVOT. A QRS complex widening rightwards and only prolonged transmural depolarization time corresponded with a substrate located in the anterior, anterior-septal or septal region of RVOT. RFA of endocardial substrate suppressed the inducibility and ECG BrS pattern during 34.7 ± 15.5 months. After RFA, flecainide testing confirmed elimination of the ECG BrS pattern. Endocardial biopsy showed a correlation between functional and ultrastructural alterations. Endocardial RFA can eliminate the BrS phenotype and inducibility during programmed ventricular stimulation (PVS).
- Brugada syndrome
- radiofrequency catheter ablation
Since the original publication in 1992 , many researchers have tried to explain the mechanisms and substrate that causes an abnormal electrocardiographic (ECG) pattern and ventricular arrhythmias in Brugada syndrome (BrS) and few therapeutic options have been found. Initially three hypotheses were proposed for explain the mechanism and arrhythmias in BrS, the abnormal repolarization theory , the abnormal depolarization theory  and the abnormal expression of neural crest cells during cardiac development .
BrS is characterized by an elevated ST segment in the right precordial leads (V1–3) on the ECG and risk of sudden cardiac death (SCD) [1, 5]. The ECG 1 pattern is frequently intermittent and can be unmasked by the administration of a sodium channel blocker (Figure 1). The incidence of SCD in subjects with Brugada type 1 ECG pattern and no previous cardiac arrest is 2 per 1000 patients per year [6, 7].
At present, there are just two therapeutic strategies, which include implantable cardioverter-defibrillator (ICD) and/or chronic quinidine therapy [6, 7]. However, quinidine is not effective in many patients and its use is frequently associated with intolerable adverse effects. ICD implantation may be effective in preventing SCD, and is currently recommended as a class I indication for symptomatic patients with type 1 Brugada ECG pattern. Unfortunately, ICD therapy in many patients is associated with inappropriate shocks (overall ICD complication rate is 9.1% and inappropriate shocks in BrS occur in 13.7%), lead fractures/failure, device infections and frequent ICD discharges by electric storms [6, 8, 9].
As an autosomal dominant disease with incomplete penetrance, BrS was initially linked to mutations in the SCN5A gene . Currently, more than 450 pathogenic variants have been identified in 24 genes encoding sodium, potassium, and calcium channels or associated proteins [10, 11]. Known BrS-susceptibility genes can only partially explain the clinically diagnosed cases; therefore, many patients (65–70%) remain “genetically unresolved” [8, 9]. For many years, BrS has been considered a purely electric disease even if, more recently some authors have shown the presence of morphological and functional abnormalities (regional conduction slow in the endocardium and epicardium), predominantly located in the right ventricle outflow tract (RVOT) [12–14].
Radiofrequency ablation (RFA) has recently emerged as a therapeutic option in BrS patients of high risk. RFA in two previous reports was effective in preventing ventricular fibrillation (VF) in BrS [15, 16]. Two studies have recently shown fractionated systolic electrograms (EGMs) in epicardium of RVOT and RF normalized the ECG pattern and prevented ventricular fibrillation and ventricular tachycardia (VT/VF) occurrence in a short follow-up [17, 18].
2. Study population and risk factors
The arrhythmic events occur in patients who presented spontaneous type 1 ECG BrS pattern and syncope of presumed arrhythmic origin, so both are considered high-risk factors . The risk of SCD in patients without ICD is 2 per 1000 patients per year [6, 7]. But unfortunately, the possibility of survival out of hospital is low if the first symptom is the SCD.
We in a prospective single-center study consecutively included 13 caucasian patients when they presented all three high-risk criteria: (1) documented spontaneous type 1 BrS ECG pattern, (2) syncope of probable arrhythmic cause (syncope was defined as a no traumatic and reversible loss of consciousness, and was considered of arrhythmic origin in the absence of a prodrome or triggering circumstances), (3) inducible VF with PVS . These were associated with at least one of the following conditions: family history of SCD at age < 45 years, type 1 BrS ECG pattern in family members, early repolarization pattern, and/or nocturnal agonal respiration [6, 9]. Structural heart disease, systemic diseases and phenocopies was ruled out in each case on the basis of clinical history and extensive evaluation with 2D Echocardiography, tilt test, brain computed tomography, 24-hour ambulatory ECG monitoring, HIV test, coxsackie and parvovirus B19 test, Chagas disease test, myocardial perfusion and cardiac nuclear magnetic resonance. All patients had 5 points according to the risk score model currently proposed by Sieira et al.  (spontaneous type 1 ECG pattern =1 point, inducible VF =2 points and syncope =2 points). None had a history of SCD or documented spontaneous VT/VF and did not receive antiarrhythmic drugs. Patients were submitted to endocardial bipolar and electroanatomic mapping and RFA. One month before of mapping and RFA, 10 patients accepted the implant of an ICD with class IIa indication .
Syncope constitutes an important diagnostic and therapeutic challenge in BrS. Approximately one-third of BrS patients present syncope. Some cases of syncope may be related to VF that terminates spontaneously. Vagal syncope is probably the most frequent cause of syncope in the BrS  and vagal hypertony may facilitate the onset of spontaneous VF in BrS . Also symptoms suggesting of vagal syncope may also be observed in syncope of cardiac origin . In our study, two patients (15%) after RFA had near-syncope with vaso-vagal prodrome and without arrhythmias in the ICD interrogation .
BrS is eight times more prevalent in males, probably for higher testosterone levels and a more prominent transient outward current (Ito). Males are at increased risk for developing a spontaneous type 1 ECG BrS pattern and VF during PVS. Nevertheless, because the majority of the asymptomatic patients are also male the gender is not an independent predictor of arrhythmic events [9, 24]. It is striking that eight of our patients (61.5%) were females. Five of these had between 38 and 58 years of age and menopausal symptoms, so we might suspect that lack of estrogens could induce the expression of phenotype .
3. Electrophysiological study and mapping: identification of functional substrate
Nademanee et al., found prolonged and fractionated late potentials in the anterior zone of epicardium of RVOT . Recently, Brugada et al. in 14 inducible patients reported abnormal EGMs only in epicardium of the anterior free wall of right ventricle and in RVOT . However, consistently we find a substrate in the endocardium of RVOT and RFA eliminates abnormal EGMs, ECG BrS pattern and inducibility during a median follow-up of 44.7 ± 15.5 months in 13 patients. We saw the substrate not only in the anterior zone of RVOT but also in septal and lateral regions, but never in the posterior region . Sunsaneewitayakul et al. reported late depolarization zones on the endocardial of RVOT. Endocardial RFA about these zones modified the ECG BrS pattern and suppress the VF storm . Similarly, we reported areas with late depolarization, diastolic electrical activity and abnormal systolic EGMs. In our study high-density detailed endocardial electroanatomical and bipolar voltage mapping of right ventricle and RVOT was performed, using 3-dimensional (3D) mapping system En Site NavXTM under local anesthesia and sedation, during stable sinus rhythm . AH and HV interval to DI and V2 lead and ventricular refractory period (VRP) were measured. Bipolar EGMs were filtered from 10 to 400 Hz and displayed at 100–200 mm/s speeds. Systolic EGMs with an amplitude ≤1.5 Mv, split or fractionated whit a duration >80 ms and delayed components extending beyond the end of QRS complex and accompanied by late potentials (LPs) were defined as abnormal. The EGMs found in the diastole were referred as “diastolic electrical activity”. The number of diastolic EGMs (separated by isoelectric line) in two successive sinus cycles was counted. PVS of RVOT with 3 cycle lengths (600, 500 and 400 ms) and up to two premature extrastimuli was performed. Premature extrastimuli was decreased in 20 ms step until a coupling interval of 200 ms or the VRP was reached or VF lasting >10 seconds was induced. The induction with PVS up to two premature beats is independent predictors of poor prognosis with a high negative predictive value and was associated with increased risk, but has a controversial prognostic value. The lack of induction does not necessarily portend a low risk and hence clinical factors are the most important determinants [6, 20, 25, 26]. VF inducibility rate is highest in patients with BrS and syncope of unknown origin (80%), the lowest in asymptomatic patients (61.5%), and intermediate in patients with vasovagal syncope (70.5%) [26–29]. However, it is important to note that these observations correspond to the pre-ablation era of BrS and therefore, the PVS could be a good predictor of outcome after RFA . In our patients the endocardial RFA of diastolic electrical activity and abnormal systolic EGMs suppressed the type 1 ECG BrS pattern and inducibility with PVS, making the patients asymptomatic, as was previously reported for the epicardial RFA [17, 18].
We identified three zones of substrate according to amplitude of the systolic EGMs: central very low voltage zone <0.5 mV, peripheral low voltage zone of 0.5–1.5 mV (border) and normal voltage zone >1.5 mV . The central zone of substrate using filling scaling was measured in mm2 and located in the RVOT. Areas showing low-amplitude signals were mapped with greater point density. Abnormal endocardial electroanatomic voltage maps, characterized by very low-voltage EGMs with clean diastoles in the central area of substrate were found. Only three patients (23%) during bipolar mapping showed fragmented systolic EGMs of low-voltage (≤ 1.5 mV) with duration greater of 80 ms between central and peripheral zone (border zone) of voltage mapping. As shown in Figure 3, only in the peripheral zone of substrate the endocardial diastolic EGMs (mean 6.7 ± 1.4 EGMs in two successive sinus cycles) were present. Overall, the median baseline very low voltage or central area of substrate (≤ 0.5 mV) was 14.2 mm2 (SD = 10.5) (Table 1).
|Sex||M||F||F||M||F||M||M||F||F||M||F||F||F||F = 8 M = 5||F = 61.5%-M = 38.5%|
|Spontaneous type 1 ECG pattern||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||13||100%|
|Syncope without prodromes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||Yes||13||100%|
|Nocturnal agonal respiration||No||No||No||No||Yes||No||Yes||No||Yes||No||No||Yes||No||4||31%|
|Family history SCD||No||No||Yes||Yes||Yes||No||No||Yes||Yes||Yes||Yes||No||No||7||54%|
|Family history BrS||No||Yes||No||No||No||No||No||No||Yes||Yes||Yes||No||No||4||31%|
|VRP ≤ 200 ms||180||180||160||200||160||180||200||180||160||180||180||200||180||13||100%||180 ms||13.6|
|QRS duration in V2 lead (> 120 ms)||160||105||120||180||120||120||140||100||90||140||170||140||100||6||46%||129.6 ms||27|
|R wave in aVR lead (≥ 3 mm) (#)||3||4.5||2||2.5||1.5||1||4||4||1.5||6||3||4||1||7||54%||3 mm||1.4|
|HV interval to DI lead (>55 ms)||60||57||40||120||57||40||46||35||45||45||55||35||59||5||38.5%||53.4 ms||21|
|HV interval to V2 lead (>55 ms)||40||40||35||80||45||40||46||35||40||45||55||35||46||1||7.7%||44.8 ms||11.5|
|Very low-voltage AREA(≤ 0.5 mV) in (mm2)||23||8||19||42||5||4||8||10||19||10||8||25||4||13||100%||14.2 mm2||10.5|
diastolic EGMs IN 2
successive cycles sinus (≥4 EGMs)
|Procedure time (min)||120||110||100||90||120||95||105||95||100||100||160||90||170||112||24.5|
|Fluoroscopy time (min)||15||10||10||10||15||10||12||13||12||10||27||10||25||13.7||5.5|
It is important to note, that epicardial mapping may not recognized a substrate located in the septal zone of RVOT. In addition, during epicardial mapping the interposition of fat tissue between the epicardium and the exploratory catheter could decrease the amplitude of the potentials recorded, giving false areas of low voltage. Our study only inclusion 13 patients and may be considered a small size sample . However, our results are consistent allowing us to reach reliable conclusions. We not performed genetic studies. However, it is unlikely that this could affect our observations because mutations have been described in over 24 genes and BrS-susceptibility genes can only partially explain the clinically diagnosed cases. Although the results of mapping and RFA were good, we do not perform epicardial mapping and do not ignore the possibility that a portion of the substrate can remain present after ablation.
In addition, we found pre-systolic potentials as was previously reported by Haissaguerre et al. . We showed with TEM Purkinje fibers in RVOT (Figures 2 and 3) . These could be involved in the origin of pre-systolic potentials and genesis of early-onset PVCs that can trigger VT or VF, by spontaneous depolarization or micro-reentry circuit in the Purkinje network .
4. Electrocardiographic analysis
4.1. Size of substrate and ST-segment elevation
In the BrS the mechanism of ST segment elevation has been explained by the repolarization theory (decreased of Na + and/or Ca++ channel function with a prominent Ito current in epicardium of RVOT generates a transmural voltage gradient), or the depolarization theory (disturbances in depolarization of RVOT can be cause of delayed conduction and ST segment elevation) [18, 20]. Nevertheless, the ECG changes are actually explained by the theorem of solid angle, where a substrate larger increases the magnitude of the ST segment elevation . In 13 patients of high risk we have analyzed QRS complex duration, R wave amplitude in aVR lead, presence of fragmented QRS (f-QRS) and end-QRS slur or notch in DI, aVL, DII, DIII and aVF leads with J point peak ≥0.2 mv with descending ST segment, corresponding to an early repolarization or “J wave” . Before of endocardial mapping the patients were underwent flecainide testing (400 mg, orally) with the purpose of measuring the greatest ST-segment elevation. The partial greatest ST-segment elevation in millimeters (ST-segment elevation in V1 + V2 leads in the third ICS and ST-segment elevation in V1 + V2 + V3 leads in fourth ICS), and total sum of greatest ST-segment elevation (ST segment elevation in V1 + V2 leads in the third ICS plus ST-segment elevation in V1 + V2 + V3 leads in fourth ICS) were measured. (Figure 4-A). Correlation between size and location of substrate in the electro-anatomic map and the ST-segment elevation were analyzed. As shown in Figure 4 and Table 2, with a cut-off ≥13 mm in the total sum of ST segment elevation two variants were found: (1) A total sum of ST-segment elevation <13 mm (n = 8, 61.5%, mean 9.6 ± 1.3 mm) corresponded to a central area of substrate of 7.7 ± 1.8 mm2; (2) A total sum of ST segment elevation ≥13 mm (n = 5, 38.5%, mean 15 ± 1.1 mm) corresponded to a central area of substrate of 28.2 ± 9.2 mm2 (p < 0.001 for correlation of ST segment elevation and correlation of central area of substrate, with a value >0.90 of ROC curve).
|Patient||ST-segment with||Total sum of ST-segment||Size of low voltage||Location of substrate|
|>elevation (mm)||Elevation (mm)||Area (mm2)||In the longitudinal and|
|V1-V2 leads V1-V2-V3 leads|
Third ICS 4TH ICS
|V1 = 2 V1 = 2|
|V2 = 4 V2 = 7||16||36||Bottom|
|V3 = 1||Anterior-lateral|
|V1 = 3 V1 = 1|
|V2 = 4 V2 = 2||10||8||Top|
|V3 = 0||Anterior-lateral|
|V1 = 2 V1 = 3|
|V2 = 3 V2 = 5||14||19||Bottom|
|V3 = 1||Anterior-lateral|
|V1 = 2 V1 = 1|
|V2 = 6 V2 = 5||16||42||Intermediate|
|V3 = 2||Anterior-lateral|
|V1 = 2 V1 = 2|
|V2 = 1 V2 = 2||8||5||Bottom|
|V3 = 1||Anterior-lateral|
|V1 = 2 V1 = 0|
|V2 = 2 V2 = 2||8||5||Intermediate|
|V3 = 2||Anterior|
|V1 = 1 V1 = 1|
|V2 = 3 V2 = 4||10||8||Bottom|
|V3 = 1||Septal|
|V1 = 3 V1 = 1|
|V2 = 5 V2 = 2||11||10||Top|
|V3 = 0||Anterior|
|V1 = 2 V1 = 3|
|V2 = 3 V2 = 4||13||19||Bottom|
|V3 = 1||Anterior-septal|
|V1 = 5 V1 = 1|
|V2 = 3 V2 = 2||11||10||Top|
|V3 = 0||Anterior|
|V1 = 2 V1 = 1|
|V2 = 6 V2 = 2||11||8||Top|
|V3 = 0||Septal|
|V1 = 2 V1 = 1|
|V2 = 3 V2 = 2||8||8||Top|
|V3 = 0||Anterior-septal|
|V1 = 3 V1 = 3|
|V2 = 4 V2 = 5||15||25||Intermediate|
|V3 = 0||Anterior|
|3 ± 1.3 2.3 ± 1.5||<13 (n = 8, 61.5%) 9.6 ± 1.3||7.7 ± 1.8|
|≥13 (n = 5, 38.5%) 15 ± 1.1||28.2 ± 9.2|
Brugada et al. during electro-anatomic mapping with administration of a sodium channel blocker showed an increase in the size of the low voltage . In concordance, our observations suggest that the total sum of ST segment elevation during flecainide testing would approximately determine the substrate size. In addition, the ECG leads with greater ST segment elevation would locate the substrate in the RVOT .
Morita et al. (145 patients who experienced syncope or had VF events) proposes ECG risk markers for the initial and recurrent episodes of VF in symptomatic patients with BrS. The f-QRS, inferolateral early repolarization and complete RBBB were associated with occurrence of ventricular tachyarrhythmia in the symptomatic patients . Our population show in three patients a QRS fragmentation (23%) and in two patients a J wave (15.4%) .
4.2. Location of substrate in the longitudinal plane
Nademanee et al. found functional substrate in the anterior zone of epicardium of RVOT . Brugada et al. in 14 inducible patients reported a functional substrate in epicardium of RVOT and anterior free wall of right ventricle .
In our study a high-density endocardial electroanatomical mapping of right ventricle and RVOT was performed . Taking the pulmonary valve as upper limit and the supraventricular crest as lower limit in the longitudinal plane the RVOT was divided in top and bottom. As shown in Figure 4-A and Table II, we have found three variants: (1) When the substrate was located in the top region of RVOT, it corresponded with the greater sum of ST-segment elevation in V1-V2 leads in the 3rd ICS (n = 5, 38.5%); (2) When the substrate was located in the bottom region of RVOT (n = 5, 38.5%), it corresponded with the greater sum of ST-segment elevation in V1-V2-V3 leads in the 4th ICS; (3) When the substrate was located in the intermediate region of RVOT (n = 3, 23%), the sum of ST-segment elevation was of equal magnitude in the 3rd and 4th ICS (n = 3, 23%).
4.3. Location of substrate in the transverse plane
In our study during endocardial electroanatomical mapping, in the transverse plane an anterior, lateral, posterior and septal areas were identified . During endocardial bipolar mapping, the regional depolarization time (RDT) from the endocardial EGM of right ventricular inflow tract (RVIT) recorder by the catheter located at the site of His, until the beginning of endocardial EGM of RVOT, recorded by the catheter located in the RVOT was measured. A value from 0 to 10 ms was considered normal. Moreover, the trans-mural depolarization time (TDT) of RVOT from the beginning of endocardial EGM of RVOT until the end of QRS complex in V2 lead was measured. The TDT measured at DI was considered as normal value. As shown in Figure 5, we have found two variants were found: (1) When the substrate was located in the anterior lateral region of RVOT (n = 5, 38.5%), the HV interval measured to DI lead (mean 70.6 ± 24 ms) was longer than the HV interval to V2 lead (mean 50 ± 15 ms). This was accompanied with widening of the QRS complex in its initial and final parts (widening of QRS left and right) in V1 and V2 lead. We defined this as “mixed delay of depolarization of RVOT”. Simultaneously, TDT and RDT were prolonged, because early depolarization of RVOT occurs. (2) When the substrate was located only in the anterior part (30.8%), or anterior-septal (15.4%) or exclusively in the septal region (15.4%) of RVOT, the HV interval measured to DI and V2 leads showed no increase in its duration (mean 42.6 ± 6 ms and 41 ± 6.5 ms, respectively) and the widening of QRS was only rightward (widening QRS rightward). Moreover, the endocardial EGM of RVIT and RVOT, and beginning of QRS complex in DI-V1-V2 lead they were activated simultaneously, indicating that there is no RDT delay, while TDT of RVOT was prolonged of dynamic manner. We defined this as “end delay depolarization of RVOT”.
Was report that 11% of patients with BrS have early repolarization pattern in the inferior-lateral leads and a more severe phenotype . Interestingly, as shown in Figure 6 we found in two patients who had a substrate of exclusively septal location, showed end-QRS notching or slurring pattern. When the substrate was located in the bottom-septal zone of RVOT (patient N°7) only end-QRS notch in aVL lead and slurred S-wave in DII, DIII and a VF leads was observed (Figure 6-A). Whereas, when the substrate was located in the top-septal zone of RVOT (patient N°11) an end-QRS slur in DI and aVL leads was observed (Figure 6-B). Our observations suggest that a location of substrate in septal region of RVOT, with beginning of the depolarization at the endocardium correlated with the presence of end-QRS notching or slurring pattern in inferior or/and lateral leads by slow conduction. As shown in Figure 6 A and B the endocardial RFA of substrate produced their disappearance . Normally the epicardium is electropositive with respect to electronegative endocardium creating a current flow of endocardium to epicardium. When activation spreads from endocardium to epicardium, in a context of slow conduction, the J wave coincides with the notch in the epicardial AP mediated by the Ito current and it is recorded by ECG. Conversely, when the activation begins in the epicardium, the J wave disappears hidden by the QRS complex . Consequently, our observations lead us to think that the J wave depends more of late depolarization that early repolarization as was suggested by other authors.
5. Effects of RFA on the ECG BrS pattern and substrate
Endocardial and epicardial RFA has been proposed as a new strategy to prevent SCD and VT/VF in BrS patients of high risk. Nademanee et al. found what RFA on prolonged and fractionated late potentials in the anterior zone of epicardium of RVOT normalized the ECG BrS pattern and prevented VT/VF in all but one patient during a follow-up of 20 ± 6 months . Recently, Brugada et al. in 14 inducible patients reported abnormal EGMs only in epicardium of the anterior free wall of right ventricle and in RVOT. RFA eliminated both ECG BrS pattern and inducibility, with a median follow-up of 5 months .
In our study, the endocardial RFA in 13 patients resulted in normalization of the ECG BrS pattern, disappearance of end-QRS notching or slurring and suppression of inducibility in all patients during a mean follow-up of 47.7 ± 15.5 months (Table 1) . About 30 days after RFA a flecainide testing did not develop ECG BrS pattern. Seven patients who entered to procedure with spontaneous type 1 ECG pattern showed ECG normalization at the end of the procedure. Immediately after RFA was applied, activity and varying degrees of changes in ST segment were observed. With following applications of RFA the ECG pattern progressively decreased (Figure 7). After RFA local abnormal diastolic EGMs completely disappear and systolic EGMs were replaced by residual very low voltage areas. The mean time of procedure and fluoroscopy were 112 ± 24.5 and 13.7 ± 5.5 minutes, respectively (Table I). Postprocedure, predischarge, and follow-up 12-lead ECG confirmed the absence of BrS ECG pattern (Figure 6C). The patients were asymptomatic and free of arrhythmic events in the 24-hour ambulatory ECG monitoring and in follow-up the ICD interrogation. Two patients (15%) had a near-syncope with prodrome at 24 of 46 months and at 18 of 36 months of follow-up respectively, without arrhythmias in ICD interrogation.
Sunsaneewitayakul et al. reported that endocardial RFA on the late depolarization zones modified the ECG BrS pattern in three patients and suppress the VF storm in four patients, during follow-up of 12–30 months . Similarly, we obtained suppression of inducibility, normalization of BrS ECG pattern and early repolarization pattern with endocardial RFA of areas with late depolarization, diastolic electrical activity and abnormal systolic EGMs ; probably by substrate homogenization and transmural lesion of the thin wall of RVOT (mean 3 mm). In addition, as show Figures 2 and 3 we found pre-systolic potentials as was previously reported by Haissaguerre et al. . In our patients the endocardial RFA of these potentials suppress PVCs and inducibility on PVS .
6. Correlation between functional and ultrastructural substrate
Coronel et al. show in a heart explanted of a BrS patient, fibrosis with epicardial fat infiltration as well as conduction slow without transmural repolarization differences . Furthermore, interstitial fibrosis, fat tissue and myocyte disorganization with reduced gap junction expression in the presence of fractionated and unfractionated low voltage systolic EGMs in the endocardium and epicardium of RVOT was reported with optical microscopy [36–38].
In two patients, before RFA and after right internal jugular venous access through a steerable sheath we advanced a bioptome to RVOT and connected to 3-dimensional (3D) mapping system . Guiding by electroanatomic and voltage map two samples of endo-myocardial biopsies of the three previously defined zones of substrate was obtained. Samples were fixed in 4% glutaraldehyde and 0.1% sodium phosphate (pH 7.4) for transmission electron microscopy (TEM) study as was described previously . As show Figure 8 the ultrastructural substrate and functional substrate were correlated . In the Figure 8A, the patient N°13 in (a), (d) and (g) shows electro-anatomic and voltage map (functional substrate) with a central area of substrate of 25 mm2 located in the intermediate-anterior zone of the RVOT and the bioptome connected to the navigation system. In (b) and (c) on the right side can be seen the ultrastructural substrate which corresponds to normal zone with mitochondria, myofibrils, and a Purkinje cell of normal characteristics. In contrast, in the peripheral zone of substrate (e and f) note that when approaching to pathological area, cytoplasmic vacuolization, myofibrillar and mitochondrial disorganization with myofibrillar residue can be observed. The (h) and (i) corresponds to the central zone, which depicts strong vacuolization and cell destruction with intense cytoplasmic disorganization and myofibrillar residue. In the Figure 8B, the patient N° 11 in (a), (d) and (g) shows the voltage and electro-anatomic map (functional substrate) with central area of substrate of 8mm2 located in the top-septal zone of the RVOT. In (b) and (c) on the right side can be seen the ultrastructural substrate which corresponds to normal zone with normal characteristics of mitochondria, myofibrils, and a Purkinje cell. The approaching to pathological area in the peripheral zone of substrate (e and f), myofibrillar disorganization, citoplasmic vacuolization, swelling and disappearance of mitochondrial crests, myofibrillar rests and remains of erythrocytes were observed. The (h) and (i) corresponds to the central zone showing strong vacuolization and cell destruction and myofibrillar residue. Fat replacement, lymphocytic infiltration, Chagasic myocarditis, collagen tissue or apoptotic bodies were not observed.
It is important to note that when we approach to pathological areas progressive cell damage was observed. In the central zone of substrate low voltage systolic EGMs coincided with strong cell destruction and cytoplasmic disorganization. The peripheral zone of substrate with cell damage, mitochondrial swelling and myofibrillar residue without apoptotic bodies coincided with late potentials, diastolic and/or presystolic activity (Figures 2 and 3). These findings support mitochondrial energy loss as possible non-apoptotic progressive tissue damage and death cell. Our results suggest the interesting possibility that substrate could be generated by an abnormal expression of neural crest cells localized in RVOT during cardiac development. Because an epicardial and endocardial substrate was demonstrated, our findings together with those of other researchers support the probability of a transmural substrate.
7. Mechanisms of arrhythmias
Two theories suggest that an abnormal repolarization (local re-excitation by phase 2 reentry in the epicardium) or a defect of the depolarization (disturbances in depolarization of RVOT can cause conduction delay) may be responsible of phenotype and VF in BrS [40–43]. However, in BrS the arrhythmias are usually polymorphic VT or VF, and these cannot be supported by macro-reentry mechanisms. VF depends of a firing focus initiated by early or delayed after-depolarization or a micro-re-entry . Surviving cells surrounded by fibrosis has demonstrated to be responsible of slow conduction and reentry in inhomogeneous scars . The residual electrical activity within scar was reported as delayed or isolated EGMs, late potentials or diastolic EGMs and their elimination during sinus rhythm was effective to prevent VT/VF . In peripheral zone of substrate when a sufficient degree of cell damage was reached such as we found with TEM and resting potentials are reduced the polymorphic VT/VF may occur. This event could be originated through a firing focus or by multiple wavelets from a reentrant microcircuit, and would explain the “diastolic electrical activity” observed in our patients.
In addition, we found pre-systolic potentials as was previously reported by Haissaguerre et al. . As show Figure 8, the Purkinje fibers in RVOT could be involved in the origin of pre-systolic potentials and in genesis of early-onset PVCs that can trigger VT or VF, by spontaneous depolarization or micro-reentry circuit in the Purkinje network .
In patients with BrS of high risk the substrate RFA may be a potential option of treatment. We successfully ablated the substrate of BrS from the endocardium based on the electrophysiological and ultrastructural findings. Our data together with the observations of other researchers suggest a transmural substrate, contributing to future definition of the arrhythmogenic substrate in BrS. As many phenotypes are involved in BrS, it is not unthinkable that different substrates may exist in BrS. ECG analysis during administration of a sodium channel blocker allows approximately determined the size and location of substrate. Careful endocardial mapping allows identify late potentials, presystolic and diastolic EGMs as a new risk marker to guide an endocardial substrate RFA, probably with the same results what a more complex epicardial ablation. A comparative study between endocardial and epicardial RFA should be performed.
This research was supported by funding sources from the Electrophysiology Division (Model Heart Center) to PET, and PIP 2015 No.183 (CONICET Argentina) and PICT 2013 No. 1949 (ANPCyT, Argentina) grants to SSS and SMH. We thank to Abbott-Argentina for providing financial support to publication.
Conflict of interest
The authors declare none conflict of interest.
|ICD||implantable cardioverter defibrillator|
|PVC||premature ventricular contractions|
|PVS||programmed ventricular stimulation|
|RVIT||right ventricular inflow tract|
|RVOT||right ventricular outflow tract|
|SCD||sudden cardiac death|
|TEM||transmission electron microscopy|
|VRP||ventricular refractory period|