Carlos De Diego

Muscleblind‐like 2 (Mbnl2) ‐deficient mice as a model for myotonic dystrophy

Myotonic dystrophy (DM), the most common adult‐onset muscular dystrophy, is caused by CTG or CCTG microsatellite repeat expansions. Expanded DM mRNA microsatellite repeats are thought to accumulate in the nucleus, sequester Muscleblind proteins, and interfere with alternative mRNA splicing. Muscleblind2 (Mbnl2) is a member of the family of Muscleblind RNA binding proteins (that also include Mbnl1 and Mbnl3) that are known to bind CTG/CCTG RNA repeats. Recently, it was demonstrated that Mbnl1‐deficient mice have characteristic features of human DM, including myotonia and defective chloride channel expression. Here, we demonstrate that Mbnl2‐deficient mice also develop myotonia and have skeletal muscle pathology consistent with human DM. We also find defective expression and mRNA splicing of the chloride channel (Clcn1) in skeletal muscle that likely contributes to the myotonia phenotype. Our results support the hypothesis that Muscleblind proteins and specifically MBNL2 contribute to the pathogenesis of human DM. Developmental Dynamics 237:403–410, 2008.

Spatially discordant alternans in cardiomyocyte monolayers

Repolarization alternans is a harbinger of sudden cardiac death, particularly when it becomes spatially discordant. Alternans, a beat-to-beat alternation in the action potential duration (APD) and intracellular Ca (Cai), can arise from either tissue heterogeneities or dynamic factors. Distinguishing between these mechanisms in normal cardiac tissue is difficult because of inherent complex three-dimensional tissue heterogeneities. To evaluate repolarization alternans in a simpler two-dimensional cardiac substrate, we optically recorded voltage and/or Cai in monolayers of cultured neonatal rat ventricular myocytes during rapid pacing, before and after exposure to BAY K 8644 to enhance dynamic factors promoting alternans. Under control conditions (n = 37), rapid pacing caused detectable APD alternans in 81% of monolayers, and Cai transient alternans in all monolayers, becoming spatially discordant in 62%. After BAY K 8644 (n = 28), conduction velocity restitution became more prominent, and APD and Cai alternans developed and became spatially discordant in all monolayers, with an increased number of nodal lines separating out-of-phase alternating regions. Nodal lines moved closer to the pacing site with faster pacing rates and changed orientation when the pacing site was moved, as predicted for the dynamically generated, but not heterogeneity-based, alternans. Spatial APD gradients during spatially discordant alternans were sufficiently steep to induce conduction block and reentry. These findings indicate that spatially discordant alternans severe enough to initiate reentry can be readily induced by pacing in two-dimensional cardiac tissue and behaves according to predictions for a predominantly dynamically generated mechanism.

Electrophysiological Consequences of Acute Regional Ischemia/Reperfusion in Neonatal Rat Ventricular Myocyte Monolayers

Background— Electrophysiological changes promoting arrhythmias during acute regional ischemia/reperfusion are challenging to study in intact cardiac tissue because of complex 3-dimensional myocardial and vascular geometry. We characterized electrophysiological alterations and arrhythmias during regional ischemia/reperfusion in a simpler 2-dimensional geometry of cultured neonatal rat ventricular myocyte monolayers. Methods and Results— Optical mapping of intracellular Ca (Cai) and voltage was performed with the use of Rhod 2-AM and Rh-237, respectively. Regional ischemia was mimicked by covering the central portion of monolayer with a glass coverslip, and reperfusion was mimicked by removing the coverslip. Monolayers were stained with fluorescent antibodies to detect total and dephosphorylated connexin-43 at various time points. During coverslip ischemia, action potential duration shortened, Cai transient duration was prolonged, and local conduction velocity (CV) slowed progressively, with loss of excitability after 10.6±3.6 minutes. CV slowing was accompanied by connexin-43 dephosphorylation. During ischemia, spontaneous reentry occurred in 5 of 11 monolayers, initiated by extrasystoles arising from the border zone or unidirectional conduction block of paced beats. On reperfusion, excitability recovered within 1.0±0.8 minutes, but CV remained depressed for 9.0±3.0 minutes, promoting reentry in the reperfused zone. As connexin-43 phosphorylation recovered in the reperfused zone, CV normalized, and arrhythmias resolved. Conclusions— Acute regional ischemia/reperfusion in neonatal rat ventricular myocyte monolayers recapitulates electrophysiological alterations and arrhythmias similar to those observed during acute coronary occlusion/reperfusion in intact hearts. During early reperfusion, slow recovery from connexin-43 dephosphorylation leads to persistent CV slowing, creating a highly arrhythmogenic substrate.

Cardiac resynchronization therapy response is associated with shorter duration of atrial fibrillation.

Background: Atrial fibrillation (AF) is commonly associated with heart failure. The benefit of cardiac resynchronization therapy (CRT) on atrial remodeling has been demonstrated. However, biventricular pacing did not reduce the global incidence of AF. We evaluated the relationship between CRT response and AF duration.

Intrapericardial balloon placement for prevention of collateral injury during catheter ablation of the left atrium in a porcine model

BACKGROUND Catheter ablation of the left atrium (LA) is associated with potential collateral injury to surrounding structures, especially the esophagus and the right phrenic nerve (PN). OBJECTIVES The purpose of this study was to evaluate the efficacy and feasibility of intrapericardial balloon placement (IPBP) for the protection of collateral structures adjacent to the LA. METHODS Electroanatomic mapping was performed in porcine hearts using a transseptal endocardial approach in eight swine weighing 40-50 kg. An intrapericardial balloon was inflated in the oblique sinus, via percutaneous epicardial access, to displace the esophagus. Similarly, with the balloon positioned in the transverse sinus, IPBP was used to displace the right PN. Esophageal temperature was monitored while endocardial radiofrequency (RF) energy was delivered to the distal inferior PV. RESULTS In all cases, balloon placement was successful with no significant effects on hemodynamic function. Balloon inflation increased the distance between the esophagus and posterior LA by 12.3 +/- 4.0 mm. IPBP significantly attenuated increases in luminal esophageal temperature during endocardial RF application (6.1 +/- 2.4 degrees C vs. 1.2 +/- 1.1 degrees C; P<.0001). High-output endocardial pacing from the right superior pulmonary vein ostium stimulated PN activity. After displacement of the right PN with IPBP, PN capture was abolished in 30 (91%) of 33 sites. CONCLUSIONS These findings demonstrate that in an animal model, IPBP is feasible in the setting of catheter ablation procedures and has the potential to decrease the risk of collateral damage to the esophagus and PN during LA ablation.

Atrioventricular conduction and arrhythmias at the initiation of beating in embryonic mouse hearts.

To investigate cardiac physiology at the onset of heart beating in embryonic mouse hearts, we performed optical imaging of membrane potential (Vm) and/or intracellular calcium (Cai). Action potentials and Cai transients were detected in ∼50% of mouse embryo hearts at E8.5, but in all hearts at E9.0, indicating that beating typically starts between E8–E9. Beating was eliminated by Ca channel blocker nifedipine and the If blocker ZD7288, unaffected by tetrodotoxin and only mildly depressed by disabling sarcoplasmic (SR) and endoplasmic (ER) reticulum Ca cycling. From E8.5 to E10, conduction velocity increased from 0.2–1 mm/s to >5 mm/s in first ventricular and then atrial tissue, while remaining slow in other areas. Arrhythmias included atrioventricular reentry induced by adenosine. In summary, at the onset of beating, If‐dependent pacemaking drives both AP propagation and Cai transient generation through activation of voltage‐dependent Ca channels. Na channels and intracellular Ca cycling have minor roles. Developmental Dynamics 239:1941–1949, 2010.

Relationship between mechanical and electrical remodelling in patients with cardiac resynchronization implanted defibrillators

AIMS Cardiac resynchronization therapy (CRT) is associated with reverse left ventricular (LV) remodelling. However, the effects of CRT-induced mechanical remodelling on electrical remodelling, and the occurrence of ventricular arrhythmias have not been clearly established. We studied the relationship between mechanical remodelling, electrical remodelling, and the occurrence of appropriate implantable cardioverter-defibrillator (ICD) therapy 1 year after CRT. METHODS AND RESULTS We analysed data from 45 patients who underwent ICD-CRT implantation at our centre. Significant LV reverse remodelling was defined by a minimum 10% decrease in the LV end-diastolic diameter (LVEDd) at 1 year of follow-up. Electrocardiographic indices of dispersion of repolarization [QTc, Tpeak-Tend (Tp-e) and their dispersion] were measured immediately and 1 year post-CRT implantation. The occurrence of appropriate ICD therapy was noted for each patient. Patients with (n= 21) and without (n= 24) significant LV reverse remodelling had similar baseline characteristics. At 1 year of follow-up, patients with mechanical reverse LV remodelling exhibited a significant decrease in QTc (505 ± 42 vs. 485 ± 52 ms, P < 0.05) and Tp-e (107 ± 26 vs. 92 ± 22 ms, P < 0.0001). However, patients without mechanical LV reverse remodelling exhibited a significant increase in QT dispersion (29 ± 43 vs. 98 ± 47 ms, P = 0.002) and Tp-e dispersion (22 ± 21 vs. 54 ± 36 ms, P = 0.0001). Finally patients with mechanical LV reverse remodelling experienced a lower rate of ICD therapy (P = 0.0025) after a mean follow-up of 19 months. CONCLUSION Reverse LV mechanical remodelling is associated with reversal of electrical remodelling and a lower rate of appropriate ICD therapy following CRT.

Effects of angiotensin-neprilysin inhibition compared to angiotensin inhibition on ventricular arrhythmias in reduced ejection fraction patients under continuous remote monitoring of implantable defibrillator devices

BACKGROUND Angiotensin-neprilysin inhibition compared to angiotensin inhibition decreased sudden cardiac death in patients with reduced ejection fraction heart failure (rEFHF). The precise mechanism remains unclear. OBJECTIVE The purpose of this study was to explore the effect of angiotensin-neprilysin inhibition on ventricular arrhythmias compared to angiotensin inhibition in rEFHF patients with an implantable cardioverter-defibrillator (ICD) and remote monitoring. METHODS We prospectively included 120 patients with ICD and (1) New York Heart Association functional class ≥II; (2) left ventricular ejection fraction ≤40%; and (3) remote monitoring. For 9 months, patients received 100% angiotensin inhibition with angiotensin-converting enzyme inhibitor (ACEi) or angiotensin receptor blocker (ARB), beta-blockers, and mineraloid antagonist. Subsequently, ACEi or ARB was changed to sacubitril-valsartan in all patients, who were followed for 9 months. Appropriate shocks, nonsustained ventricular tachycardia (NSVT), premature ventricular contraction (PVC) burden, and biventricular pacing percentage were analyzed. RESULTS Patients were an average age of 69 ± 8 years and had mean left ventricular ejection fraction of 30.4% ± 4% (82% ischemic). Use of beta-blockers (98%), mineraloid antagonist (97%) and antiarrhythmic drugs was similar before and after sacubitril-valsartan. Sacubitril-valsartan significantly decreased NSVT episodes (5.4 ± 0.5 vs 15 ± 1.7 in angiotensin inhibition; P <.002), sustained ventricular tachycardia, and appropriate ICD shocks (0.8% vs 6.7% in angiotensin inhibition; P <.02). PVCs per hour decreased after sacubitril-valsartan (33 ± 12 vs 78 ± 15 in angiotensin inhibition; P <.0003) and was associated with increased biventricular pacing percentage (from 95% ± 6% to 98.8% ± 1.3%; P <.02). CONCLUSION Angiotensin-neprilysin inhibition decreased ventricular arrhythmias and appropriate ICD shocks in rEFHF patients under home monitoring compared to angiotensin inhibition.

A simple pacing method to diagnose postero-anterior (clockwise) cavo-tricuspid isthmus block after radiofrequency ablation.

AIMS Bidirectional block of the cavo-tricuspid isthmus (CTI) is a widely accepted endpoint for typical atrial flutter ablation, but its evaluation may be difficult, especially in the postero-anterior (clockwise) direction. The main goal was to evaluate pacing at the septal edge of the ablation line as an indicator of clockwise CTI block and as a predictor for flutter recurrence. METHODS AND RESULTS In 94 patients undergoing flutter ablation, CTI block in the antero-posterior (counterclockwise) direction was determined by differential pacing from several levels of the anterior right atrial (RA). CTI block in the clockwise direction was evaluated by analysing electrograms (EGM) at the ablation line during differential pacing of the septal RA (differential septal pacing) or by anterior sequence of RA during pacing septal isthmus, next to the ablation line (septal CTI pacing). Ablation produced bidirectional block in 78% of the patients, unidirectional counterclockwise block in 9% and bidirectional conduction persisted in 13%. After follow-up (37 +/- 23 months), flutter recurrence occurred in 13% (48% if persistent conduction vs. 3% if bidirectional block, P < 0.001). During differential septal pacing, EGMs were difficult to interpret in 36% of the patients; in these cases, the diagnosis of CTI block or conduction in the clockwise direction was clearly established by using septal CTI pacing. CONCLUSION Activation sequence of anterior RA during septal CTI pacing, next to the ablation line, is a reliable and simple method to diagnose clockwise CTI block and is associated with a low flutter recurrence.