JACC: Clinical Electrophysiology

Background: Characterizing cardiac activation by its site of origin, propagation, and conduction velocity underlies arrhythmia diagnosis and management, but invasive electrophysiology (EP) mapping requires vascular access, fluoroscopy, and sedation. Magnetocardiography (MCG) enables contactless mapping, and recent solid-state sensors remove the cost, cryogenic, and shielding barriers of legacy systems. We assessed the feasibility of a novel solid-state MCG system for noninvasive arrhythmia site-of-origin (SOO) localization and activation reconstruction, benchmarked against electrocardiographic imaging (ECGi). Methods: In nine swine implanted with right atrial and right ventricular pacing leads, we recorded MCG and ECGi simultaneously during atrial and ventricular pacing. Invasive epicardial contact EP mapping provided the activation-time reference and MRI-derived lead-tip location the SOO reference. Local activation time (LAT), conduction velocity (CV), and SOO were compared on a co-registered chamber mesh. SOO error was the Euclidean distance to the MRI lead tip; LAT and CV agreement with EP were quantified by Pearson r and compared using Wilcoxon signed-rank tests. Results: Across 17 datasets (8 atrial, 9 ventricular), median SOO error was lower for MCG than ECGi in the atrium (19.6 vs 31.2 mm; p=0.023) and ventricle (12.0 vs 26.1 mm; p=0.074). LAT agreement with EP was comparable between modalities and higher in the ventricle (MCG r=0.63; ECGi r=0.68) compared with the atrium (MCG r=0.40; ECGi r=0.53), each correlating with invasive EP mapping above chance. CV agreement was modest and numerically higher for MCG in the ventricle. Conclusions: Solid-state MCG was feasible for noninvasive site-of-origin localization and activation mapping, with accuracy comparable to ECGi, motivating larger prospective studies to define its clinical role in noninvasive mapping.

Background and Aims: Machine learning has shown potential in predicting ablation targets for ventricular tachycardia (VT) in an animal model. This study progresses to externally validating deep learning approaches for human data. Methods: The development and external validation dataset included 21 and 13 patients, respectively, with structural VT undergoing catheter ablation. In the development datasets, electrophysiological studies were conducted using the AdvisorTM HD grid (EnsiteTM X), while both CARTO and Ensite Precision were used in the validation dataset. In each patient, VT ablation targets were defined as mapping points within 8 mm of VT isthmuses. Three advanced machine learning models were trained using cardiac mapping data acquired in both omnipolar and unipolar configurations during sinus rhythm and ventricular pacing. Discrimination was evaluated using nested leave-one-out cross-validation at patient level. Results: Overall, graph convolutional networks (GCNs), which integrate intracardiac signal waveforms with three-dimensional electroanatomical geometries, achieved the highest performance, with optimal results obtained from unipolar electrograms acquired in sinus rhythm (median AUC 0.793, sensitivity 83.6%, specificity 69.0%). This may be partly explained by the inclusion of repolarization dynamics in unipolar electrograms and the higher point density of sinus rhythm maps. Comparable performance was observed in the external dataset. Conclusion: This study demonstrates that graph convolutional networks applied to sinus rhythm EGM waveforms collected during substrate mapping can localise critical components of VT re-entry circuits. This approach has potential to provide fast and accurate ablation guidance without the need to induce and map VT, improving safety and efficacy of VT catheter ablation.

Background: Pulsed-field ablation (PFA) systems increasingly provide impedance-based contact indicators, such as tissue proximity indication (TPI), derived from local impedance changes relative to a blood-pool baseline. These indicators are largely binary and do not quantify post-application catheter?tissue energy coupling. We evaluated Bipolar Local Impedance Delta (BiLID), the peri-application bipolar local impedance drop, as a complementary impedance-based index of delivered energy coupling. Methods: We retrospectively analyzed 1,556 VARIPULSE applications in 23 patients undergoing pulmonary vein isolation. BiLID was derived from 29,822 paired pre-/post-ablation impedance measurements obtained from numeric local impedance readouts displayed by the mapping system, without proprietary data export or waveform estimation. Reproducibility was assessed by intraclass correlation. Associations with TPI status, vein anatomy, application order, and peak creatine kinase-MB (CK-MB) were examined using linear mixed-effects models with within-patient clustering and exploratory patient-level analyses. Results: BiLID showed excellent interobserver reproducibility and increased stepwise with the number of TPI-positive electrodes per pair (0?2; P < 0.0001), while varying widely among TPI-positive signals. BiLID differed by electrode position and was lower during right than left pulmonary vein ablation (both P < 0.0001). Total BiLID correlated with peak CK-MB (r = 0.71; 95% CI, 0.42?0.87; P = 0.0001), whereas application count (r = 0.16, P = 0.4711) and TPI-positive signals (r = 0.26, P = 0.2334) did not. Lower CK-MB elevation was associated with larger left atrial volume index, female sex, and heart failure, suggesting substrate-modulated biomarker release. Conclusions: BiLID is a reproducible, continuous index of catheter?tissue energy coupling that complements pre-delivery binary contact indicators by quantifying the response after PFA delivery. It captures graded contact quality and anatomical heterogeneity and may inform individualized, coupling-guided PFA titration.

Background: Three pulsed-field ablation (PFA) systems are FDA-approved for atrial fibrillation (AF), yet whether their safety profiles differ from each other and from radiofrequency (RF) ablation has not been systematically characterized using real-world adverse event data. We compared platform-specific complication profiles across three FDA-approved PFA systems and contemporary RF catheters in MAUDE. Methods: We analyzed 2,262 manually adjudicated MAUDE adverse event reports (760 PFA, 1,502 RF) through July 2025. Neurologic events underwent independent adjudication into five tiers by three auditors. Disproportionality was assessed using Reporting Odds Ratios (ROR) with Benjamini-Hochberg (BH) correction. Results: Pooled PFA had significantly lower BH-adjusted ROR for tamponade (0.52, 95% CI 0.41-0.67) and esophageal injury (0.09, 0.01-0.66), consistent with a tissue-selective reporting profile across platforms. Platform-level analysis, however, revealed substantial heterogeneity: the stroke signal was driven by Varipulse (ROR 16.41, 8.61-31.28) and was not observed with Farapulse (ROR 1.26, NS). Pooled PFA had higher ROR for imaging-confirmed stroke (3.84, 2.27-6.49) and arrhythmia (2.57, 1.91-3.45). Coronary vasospasm (24 vs. 0 events) and hemolysis (15 vs. 1 events) were PFA-specific. Composite serious adverse events were similar. In a pre-specified extension period analysis (August-December 2025), the pooled PFA stroke signal attenuated to non-significance (ROR 1.60, 0.89-2.85), consistent with notoriety bias following the FDA Safety Communication. Conclusions: PFA adverse-event reporting shows substantial platform heterogeneity across approved systems. Varipulse was associated with a disproportionate neurologic reporting signal, while all PFA platforms show tissue-selective reporting patterns relative to RF. These findings support platform-aware clinical decision-making and post-market surveillance.

Background: Iatrogenic atrial septal defect (iASD) is a known consequence of transseptal catheterization. Left atrial intracardiac echocardiography (LA ICE) requires additional septal instrumentation, yet data on persistent iASD after pulsed field ablation (PFA) with an LA ICE workflow remain limited. We evaluated the incidence, predictors, and one-year clinical significance of persistent iASD in this setting. Methods: Consecutive patients undergoing PFA for atrial fibrillation with LA ICE were prospectively evaluated with transthoracic echocardiography before ablation and at one year, including systematic agitated saline contrast. Persistent iASD was defined as residual interatrial shunting on color Doppler at follow-up, classified as small (<3 mm), moderate (3-5 mm), or large (>5 mm). Groups were compared by t-test and chi-square test. Results: Among 850 patients, persistent iASD was identified in 153 (18.0%) at one year; 97 (63.4%) were small and 56 (36.6%) moderate, with no large defects. All shunts were left-to-right. No stroke or transient ischemic attack, paradoxical embolism, hypoxemia, right-heart enlargement, or septal closure occurred. Persistent iASD was associated with female sex (64.7% vs 48.1%), longer septal dwell time (52{+/-}12 vs 31{+/-}11 min), higher left atrial pressure (28{+/-}4 vs 12{+/-}3 mmHg), lower LVEF (32{+/-}11% vs 54{+/-}14%), and larger-caliber sheaths ([&ge;]17 Fr; 80.4% vs 48.2%; all p<0.001). Conclusions: Persistent iASD following PFA with LA ICE occurs in approximately one in five patients but is predominantly small, exclusively left-to-right, and clinically benign at one year. Persistence is associated with mechanical and hemodynamic factors, particularly sheath caliber, rather than the ablation energy source.

Background. Durable mitral isthmus (MI) block remains challenging in persistent atrial fibrillation (PeAF) ablation. Recent epicardial vein of Marshall (VoM) recordings have shown incomplete MI transmurality and time-dependent conduction recovery after pulsed field ablation (PFA). Whether systematic VoM ethanol infusion (VoM-EI) followed by focal PFA provides stable acute MI block remains unknown. **Objectives.** To assess the incidence, timing, and procedural implications of early MI conduction recovery after systematic VoM-EI followed by focal Sphere-9 PFA. Methods.In this prospective single-center study, 55 consecutive patients undergoing first ablation for symptomatic PeAF with planned MI ablation were screened. VoM-EI was systematically attempted before left atrial access and successfully performed in 51 (92.7%), who constituted the study cohort. Pulmonary vein isolation, roof-line, and MI ablation were performed with the Sphere-9? lattice-tip catheter. After bidirectional MI block, conduction was systematically reassessed during a standardized 30-minute waiting period. Results.Mean age was 70.3 {+/-} 8.2 years, and 36 patients (70.6%) were men. Initial bidirectional MI block was achieved in 50/51 patients (98.0%). During the waiting period, conduction recovered in 9/50 (18.0%; 95% CI, 9.8%-30.8%), at a median of 16 minutes (IQR, 10-20; range, 8?23). Six of 9 patients with recovery (66.7%) required targeted coronary sinus (CS) ablation. Block was restored in all 9, yielding a final block rate of 50/51 (98.0%). Median procedure duration was 82 minutes (IQR, 73-95), with no major complications. Conclusions. Immediate bidirectional MI block was not synonymous with stable block. Despite systematic VoM-EI followed by focal Sphere-9 PFA, conduction recovered in approximately one in five patients, including beyond 20 minutes, and two thirds required targeted CS ablation. These findings support standardized 30-minute reassessment and targeted CS interrogation rather than reliance on immediate block. Chronic invasive remapping is required to determine whether this strategy improves long-term MI block durability.

BackgroundEmerging evidence in humans has linked Xp22.31 genetic deletions to an increased risk of stress-induced arrhythmias, and association analysis across Xp22.31 has shown enrichment for atrial fibrillation genetic risk variants around STS (steroid sulfatase). MethodsWe compared heart rhythm in homozygous STS-knockout (STS-KO) adult mice (n=19) to wildtype (WT) mice (n=11) under baseline (Tyrodes solution), and {beta}-adrenergic stimulation (stress), conditions using ex vivo perfused-heart electrocardiography (ECG). Ventricular ectopic beats (VEBs) were manually-identified, and rhythm abnormalities were quantified using a semi-automated approach. ResultsAt baseline, the groups displayed comparable sinus cycle length and ECG intervals; all WT hearts showed stable sinus rhythm, whereas [~]30% of STS-KO hearts developed spontaneous VEBs. WT hearts typically maintained steady rhythm under {beta}-adrenergic stimulation; in contrast, [~]60% of STS-KO hearts displayed VEBs. Under baseline and stimulated conditions the QRS interval was greater during VEBs than normal beats in STS-KO hearts, consistent with a ventricular origin. We identified a higher frequency of any abnormal beats in STS-KO hearts than in WT hearts under baseline (1.9{+/-}0.7% vs. 0.3{+/-}0.1%, p=0.038) and stimulated (5.3{+/-}2.2% vs. 0.5{+/-}0.3%, p=0.047) conditions. Under stimulated conditions, abnormal beats only occurred singly in WT hearts, whereas in STS-KO hearts, [~]50% of the time they occurred in runs of two or more. ConclusionSTS deficiency in mice predisposes to arrhythmias, and the STS-KO mouse represents a tractable model for mechanistic investigation. These data support the contention that STS activity influences arrhythmic vulnerability in humans and that STS should be considered for inclusion in arrhythmia-related gene panels.

BACKGROUND: Atrial fibrillation (AF) remains difficult to explain using a single focal driver or rotor-centered mechanism across disease stages. We tested whether progressive atrial substrate remodeling can drive a critical transition toward turbulence-like, decentralized multi wavelet electrical activity. METHODS: We constructed a controlled two-dimensional atrial reaction-diffusion model with six graded substrate-remodeling stages. We evaluated effective wavelength, theoretical wavelet capacity, AF inducibility, vulnerable-window dynamics, spatial randomness, temporal memory, spectral dispersion, nonlinear indices, virtual ablation response and ERP-prolongation reverse mechanistic testing. RESULTS: Progressive remodeling shortened effective wavelength from 12.0 to 2.4 cm and increased theoretical wavelet capacity from 0.69 to 17.36. Inducibility rose sigmoidally as wavelength shortened, with a model-derived transition near lambda50=4.5 cm. Advanced substrates showed increased wavebreak, spatial randomness, short-memory dynamics, broad spectral dispersion, positive nonlinear indices and resistance to random local ablation. Culprit atrial premature beats within the vulnerable window efficiently triggered AF, whereas counter pacing at 20 to 35 ms reduced inducibility from 52% to 11% in stage 2. CONCLUSIONS: In this controlled model, AF initiation and maintenance were linked to substrate-dependent wavelength, wavelet capacity and vulnerable-window triggering. The model-derived transition provides a testable framework for future high-density mapping, patient30 specific modeling and device-based studies. Key Words atrial fibrillation; turbulence-like electrical activity; substrate remodeling; critical wavelength; multi-wavelet re-entry; vulnerable window; culprit premature atrial beat; counter pacing

Background: Mitral regurgitation (MR) is a major risk factor for the development of atrial fibrillation (AF), yet the molecular mechanisms linking volume overload to arrhythmogenic remodeling remain poorly understood. Although fibrosis has long been considered the primary substrate for AF, increasing evidence suggests that fibroblast heterogeneity and cell-cell interactions may play important roles in disease progression. Methods: MR was created endovascularly by chordal avulsion in 12 dogs with 6 controls. AF inducibility was assessed by transvenous burst pacing, left atrial volume by echocardiography, and collagen content by Masson trichrome and picrosirius red staining. Single-nucleus RNA sequencing (snRNA-seq) was performed on left atrial posterior wall tissue from control, 4-week, and 6-month MR animals. Fibroblast subpopulations and fibroblast-cardiomyocyte communication were analyzed and markers validated by RNA in situ hybridization in all 18 animals. Results: MR resulted in progressive left atrial dilation, but neither the change in left atrial volume from baseline nor total collagen burden correlated with the inducibility of AF (n=6 each). SnRNA-seq resolved seven major cardiac cell populations and identified four transcriptionally distinct fibroblast populations (NOX4/GRIA4, PCOLCE2, ADRB2/HCN1, PTX3/ICAM1). Fibroblast composition shifted markedly: matrix-associated PCOLCE2 fibroblasts starkly declined by 6 months, whereas inflammatory-associated PTX3/ICAM1 fibroblasts expanded stepwise over time. Cardiomyocyte-to-fibroblast signaling, dominated by PTPRM and LAMA2, was progressively redirected toward PTX3/ICAM1 fibroblasts. RNAscope confirmed a stepwise rise in ICAM1 transcripts and higher ICAM1 in AF-inducible than non-inducible animals. Conclusions: In a canine model of MR, the inducibility of AF was associated with fibroblast state remodeling rather than with atrial dilation or collagen burden. Progressive expansion of inflammatory-associated PTX3/ICAM1 fibroblasts, together with reorganized fibroblast-cardiomyocyte signaling, defines a candidate arrhythmogenic mechanism and therapeutic target in MR.

Background Substrate mapping using multielectrode catheters is increasingly used for post-myocardial infarction (MI) ventricular tachycardia (VT) avoiding repeated VT induction and mapping during VT. However, these catheters may mechanically induce ventricular arrhythmias with hemodynamic compromise. This study compares pro-arrhythmogenicity between single-tip and multi-spline catheters during functional substrate mapping. Methods Thirty post-MI patients (age 68{+/-}8 years, 97% male, LVEF 40% [IQR 33-46]) referred for VT ablation at two centers (2021-2024) underwent endocardial mapping during baseline rhythm in random order with both a multi-spline catheter (Octaray, n=4; Pentaray, n=26) and a single-tip QDOT catheter. The protocol was prematurely terminated if (i) two mechanically induced VTs required ECV, (ii) recurrent ATP-treated mechanical VTs caused hemodynamic compromise, or (iii) excessive mechanically induced ectopy impaired catheter contact. Mapping time, point density, and mechanically induced arrhythmias were assessed. Results Multi-spline catheters enabled faster mapping (26{+/-}9 vs 60{+/-}16 minutes, p<0.001) with more acquired points (p<0.001). VTs were more frequently mechanically induced with multi-spline catheters (median 2 [IQR 1-4] vs 0 [0-3], p<0.05) and these VTs were faster (304ms, IQR 292-320] vs 373ms, IQR [316-405], p=0.01) and degenerated more often into VF (3 vs. 0). Overall, 17 patients (57%) experienced at least one mechanically induced VT; seven (23%) required cardioversion, and mapping was prematurely terminated in eight (27%), all while using multi-spline catheters. Conclusion Multi-spline catheters allow rapid substrate mapping but with substantial risk of mechanically induced arrhythmias, requiring premature termination of substrate mapping because of safety concerns. Their use in post-MI VT ablation warrants careful risk?benefit assessment.

Background: Vagally mediated syncope and functional bradyarrhythmias can cause recurrent symptoms, injury and a need for permanent pacing, yet standard therapies do not address the underlying autonomic reflex. Cardioneuroablation (CNA) targets this mechanism, but adoption has been limited by uncertainty about patient selection, procedural endpoints and durable outcomes. Methods: We studied 95 consecutive patients with vagally mediated syncope (n=78) or functional bradyarrhythmia (n=17) who underwent CNA at a single center by a single lead operator under a uniform, prospectively maintained protocol. Extracardiac vagal stimulation (ECVS) was performed before and after ablation to confirm vagally mediated sinus or atrioventricular (AV) nodal responses and to verify their attenuation. Outcomes through 1 year included clinical recurrence, quality of life, pacing burden and pacemaker extraction. Results: Before ablation, ECVS provoked sinus pauses in 95.8% of patients and AV block in 91.6%; after ablation, sinus pauses were abolished in every patient and residual AV block was present in 4.2%. One-year freedom from recurrent syncope and from bradyarrhythmia-related events was 93.8% and 94.1%. Syncope burden fell from a median of 2.7 episodes per year to none, and disease-specific quality of life improved substantially (P<0.001). Among patients with pre-existing devices, atrial pacing burden fell from a median of 22% to 0%. There were no strokes, deaths or myocardial infarctions. Conclusions: In this single-center cohort, physiology-guided CNA was associated with durable symptom control, improved quality of life and reduced pacing across vagally mediated bradyarrhythmia syndromes. Multicenter controlled studies are needed to confirm these findings.

Background: Abnormal ventricular repolarisation in hypertrophic cardiomyopathy (HCM) may predispose patients to lethal arrhythmias, but repolarisation in HCM remains poorly spatially characterised. Integrating spatial cardiac magnetic resonance with ECG data has the potential to map electrical function throughout the ventricles. This study applied novel digital twin inverse ECG methods to map repolarisation patterns underlying abnormal T-waves in HCM. Methods: Patient-specific full ventricular electrophysiological models were iteratively refined to match the patient 12-lead ECG. Data from 32 healthy volunteers and 69 HCM patients were analysed, with inferred substrates incorporating activation times, repolarisation times, and rate-corrected action potential durations (APDcs). Patients were stratified by T-wave phenotype to identify distinct spatial repolarisation signatures associated with different ECG presentations. Results: Clinical 12-lead ECGs were accurately reproduced by the inferred ventricular models in 95 of 101 cases. Healthy volunteers (N=30) and HCM patients with normal T-waves (N=33) were characterised by apex-to-base APDc gradients of 60 ms (40?80) and 60 ms (30?80), respectively. HCM patients with V1-V3 T-wave abnormalities (N=6) had attenuated apex-to-base APDc gradients of 30 ms (-20?40) driven by apical-to-mid anterior APDc prolongation, greatest at the apical segment (?APDc vs. healthy: 54 ms; 95% CI: 23?84 ms). HCM patients with V4-V6 T wave abnormalities (N=21) had reversed apex-to-base APDc gradients of -20 ms (-40?0) driven by apical-to-mid APDc prolongation, most severe at the apical segment (?APDc vs. healthy: 94 ms; 95% CI: 74?120 ms). Despite significant APDc prolongation, only 4 of 69 HCM patients had QTc > 480 ms, due to masking by the intrinsically healthy longer APDcs at the ventricular base. Conclusions: Distinct spatial distributions of APDc prolongation, not necessarily mirroring the distribution of hypertrophy, underlie different ECG repolarisation phenotypes in HCM and may be missed by the QTc interval.

Background: Cardiac resynchronization therapy (CRT) fails in 30% of patients, often due to suboptimal left ventricular pacing site (LVPS) selection. Current practice lacks tools for pre-procedural, patient-specific LVPS optimization within the accessible coronary sinus (CS) tributaries. This study aimed to develop a digital twin and an explainable ML-based clinical decision support framework to address this issue. Methods: Personalized 3D cardiac models incorporating ventricular anatomy, myocardial fibrosis, and CS anatomy were constructed from CT and LGE-MRI for 74 CRT candidates. Finite-element Eikonal simulations of biventricular pacing generated patient-specific electrophysiological features at candidate LVPS. A Machine Learning (ML) classifier was trained on a hybrid feature set of pre-procedural clinical variables and model-derived indices, validated by leave-one-out cross-validation. SHAP analysis provided a physiologically interpretable rationale for each prediction. The framework was applied to a pilot cohort of 19 patients with reconstructed 3D CS anatomy to generate a spatial likelihood map of CRT response across all clinically implantable pacing sites within each patient's CS. Results: The ML classifier outperformed the reference Feeny clinical calculator under LOO-CV (accuracy 0.78 vs 0.58; F1-score 0.75 vs 0.43), AUC=0.78, sensitivity=0.80, specificity=0.77. Bootstrap analysis yielded mean AUC=0.85 (95% CI 0.70-0.95). In the pilot CS cohort, the framework identified that 8 of 13 clinical non-responders had no accessible CS site predicted to yield a positive response, supporting redirection towards alternative pacing strategies. In the remaining 5, alternative implantable sites with high predicted response probability were identified. SHAP analysis confirmed that dominant predictors were patient-specific in their relative contributions, supporting individualized over heuristic-based LVPS selection. Conclusion: This pilot study demonstrates the feasibility of a digital twin and explainable ML framework as a pre-procedural clinical decision support tool for CRT planning, stratifying patients and identifying optimal implantable sites with transparent anatomical rationale. Prospective validation and regulatory evaluation are required before clinical deployment.

Three-dimensional genome organization stabilizes cell-type-specific gene expression, yet the tissue-restricted factors that maintain chromatin insulation remain poorly understood. Here, we identify the muscle-specific ribosomal protein Rpl3l as an unexpected nuclear regulator of genome architecture in atrial cardiomyocytes. Rpl3l is enriched in the nucleus and nucleolus, where it binds its own genomic locus and stabilizes a CTCF-anchored chromatin boundary that represses the T-type calcium channel gene Cacna1h. Loss of Rpl3l weakens local chromatin insulation, increases long-range contacts across the Rpl3l-Cacna1h locus, derepresses Cacna1h, and increases susceptibility to atrial fibrillation (AF), which is suppressed by pharmacological inhibition of T-type calcium channels. Furthermore, AF-associated RPL3L variants exhibit impaired nucleolar localization, reduced rRNA binding, and defective repression of CACNA1H in human iPSC-derived atrial cardiomyocytes. Together, these findings reveal a ribosomal protein-chromatin axis linking genome insulation to ion-channel dosage control and cardiac rhythm stability, expanding the repertoire of cell-type-specific genome architecture regulators.

Background: Left atrial volume index (LAVI) reflects atrial remodeling in heart failure with preserved ejection fraction (HFpEF), but its nonlinear, hemodynamic, and time-varying associations with atrial fibrillation (AF) remain uncertain. Objectives: To assess the association of baseline LAVI with documented incident AF in HFpEF. Methods: We studied 764 patients with HFpEF without documented AF in a Hong Kong registry. LAVI was analyzed continuously, by tertiles, and with restricted cubic splines. Incident AF was the first qualifying AF diagnosis or electrocardiographic record after echocardiography. Cause-specific Cox and Fine-Gray models were used, with death before AF as a competing event. A hemodynamic overlap-adjusted model additionally included E/e' ratio and pulmonary artery systolic pressure. Results: During a median follow-up of 5.81 years, 360 patients developed documented incident AF and 272 died before AF was documented. In the primary clinical model, each 10-mL/m2 increase in LAVI was associated with incident AF in cause-specific Cox regression (HR, 1.08; 95% CI, 1.06-1.11) and Fine-Gray regression (sHR, 1.06; 95% CI, 1.04-1.09). After hemodynamic overlap adjustment, the continuous association was attenuated. However, the highest LAVI tertile remained associated with incident AF (HR, 1.78; 95% CI, 1.28-2.49; sHR, 1.47; 95% CI, 1.05-2.04). Splines showed nonlinear excess risk at higher LAVI, and period-specific analyses showed the strongest association during the first year. Conclusions: Marked left atrial enlargement identified a structural-hemodynamic phenotype associated with early documented incident AF and may support risk-enriched rhythm surveillance.

Approximately half of the adult sinoatrial node (SAN) consists of non-myocyte populations, indicating that cardiac pacemaking depends on interactions within a multicellular tissue rather than on pacemaker cardiomyocytes alone. Among these, an S100B-associated cell population has been implicated in pacemaker function, yet its identity and physiological roles remain poorly understood. These cells are rare and dispersed throughout the small, structurally complex SAN, making them difficult to observe repeatedly while preserving the native multicellular environment. Here, we established a dissociated multicellular culture of adult mouse SAN tissue on soft collagen-gelatin hydrogels that retains spontaneous electrical activity and permits longitudinal live imaging of S100B-associated cells. Using an S100B-EGFP+ reporter, we identified at least six reproducible morphological and behavioral phenotypes, including migration, proliferation, phagocytic behavior, and spontaneous self-organization into three-dimensional clusters. Cultures remained spontaneously electrically active for more than 10 days in vitro, with peak activity around day 10. This multicellular culture model bridges the gap between intact SAN preparations and isolated-cell cultures, allowing repeated observation of rare S100B-associated cells within a spontaneously active multicellular environment. HighlightsO_LIThe platform enables longitudinal live imaging of rare S100B-associated cells within an diverse multicellular SAN culture. C_LIO_LILive imaging reveals at least six reproducible morphological and behavioral phenotypes of S100B-associated cells. C_LIO_LIDissociated multicellular SAN cultures remain spontaneously electrically active for more than 10 days in vitro. C_LI

Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia and a major cause of cardioembolic stroke. Although polygenic risk scores (PRS) are well characterized to quantify inherited susceptibility for AF, they provide limited insight into the pathways and tissues underlying genetic risk, which are critical to uncover for individual risk prediction. In this study, we develop a pathway-level multi-omics representation learning framework that converts individual genetic profiles into interpretable biological features by integrating GWAS-derived pathway burden scores with tissue-specific transcriptomic pathway signals. We constructed machine learning models to assess population-level AF risk prediction performance across genomic and transcriptomic tissue contexts; the pathway-based global attention models substantially improved risk prediction performance over PRS and other baselines (AUROC improved from 0.601 to 0.738). Transformer and graph neural network frameworks then assessed individual-level pathway interpretability, revealing heterogeneous contributions from electrical signaling, cardiac development, and DNA repair pathways to AF risk. This added interpretability highlights the potential of this pathway approach to enable more mechanistically informed risk stratification than static PRS by capturing underlying heterogeneity. To independently assess whether prioritized pathways reflected cardiac regulatory biology, we compared pathway rankings with transcriptional effects predicted by the AlphaGenome foundation model. Variants in highly ranked pathways showed significantly greater predicted effects on expression in atrial and ventricular tissues (FDR = 0.032) relative to controls, providing orthogonal evidence that the model identifies biologically relevant mechanisms. Overall, this work reframes polygenic risk from a single measure of susceptibility to tissue-informed pathway mechanisms, providing a framework for interpretable genomic stratification in complex diseases.

RationaleSCN1B encodes the {beta}-subunits of the main cardiac voltage-gated sodium channel, NaV1.5. Variants are linked to cardiac conduction disease, often with concealed phenotypes. Whether {beta}1-subunits regulate conduction through nanoscale intercalated disc (ID) structures, e.g. perinexi, and ephaptic coupling remains unresolved. ObjectiveTest whether Scn1b haploinsufficiency induces latent conduction abnormalities that are unmasked by perturbations in extracellular nanodomains. Methods and ResultsAdult Scn1b+/- mice and wild-type (WT) littermates underwent multiscale phenotyping (qRT-PCR, Western blot, patch clamp, transmission electron microscopy (TEM), ex vivo optical mapping, in vivo ECG). Scn1b+/- hearts showed [~]50% reductions in Scn1b mRNA and {beta}1 protein without changes in canonical conduction proteins. Peak sodium current, baseline conduction velocity ex vivo, and baseline QRS duration in vivo were unchanged. However, TEM revealed increased baseline perinexal width in Scn1b+/- hearts. Osmotic expansion of the perinexus with mannitol slowed conduction to a greater extent in Scn1b+/- hearts and prolonged QRS duration in vivo. In contrast, perinexal narrowing with dextran 2MDa selectively increased conduction velocity in Scn1b+/- hearts. ConclusionsScn1b haploinsufficiency preserves baseline excitability and conduction but structurally remodels the ID at the nanoscale, increasing sensitivity to extracellular nanodomain perturbations. These data support a structural role for {beta}1-subunits in ephaptic coupling, and that conduction is maintained over a range of perinexal widths with pathological conduction slowing occurring beyond a critical width. Importantly, osmotic stress unmasks a concealed conduction phenotype, identifying extracellular nanodomain stability as a potential therapeutic target to mitigate arrhythmia risk in SCN1B-associated disease.

Background: The His-Purkinje network drives rapid ventricular activation and is a major substrate for ventricular arrhythmias, yet it is among the least molecularly characterized cardiac compartments. Markers validated in rodents transfer poorly across species, few are confirmed at the protein level in large mammals or humans, and most lack the stability and surface accessibility that demanding applications require. Methods: We combined histology-guided laser-capture microdissection with low-input, cell-type-resolved RNA-sequencing to profile Purkinje fibers, left-ventricular cardiomyocytes and peri-Purkinje stroma from adult sheep. Differentially expressed genes were ranked by a transparent composite framework weighting expression specificity, cross-individual stability and predicted subcellular accessibility; leading candidates were validated by RT-qPCR and immunolabelling in sheep and by RT-qPCR in human myocardium. Results: RNA-sequencing resolved a Purkinje transcriptome distinct from cardiomyocytes and stroma and defined 331 concordantly enriched genes, which the composite framework ranked into stable, specific candidates spanning intracellular and cell-surface compartments. By RT-qPCR, the canonical conduction markers connexin-40/GJA5, HCN4, NEFM and MYL4 were strongly enriched in Purkinje fibers, whereas the rodent gold-standard contactin-2 was not, underscoring species divergence. Thirteen of sixteen prioritized candidates were confirmed by RT-qPCR, and immunolabelling localized MYL4, CNN1, TAGLN and DKK3 to Purkinje fibers; contactin-5 emerged as a novel transcript- and protein-validated Purkinje marker. In human myocardium, a defined subset - MYL4, connexin-40/GJA5, contactin-5 and TAGLN - was conserved, while several markers proved species-restricted. Conclusions: We provide the first genome-wide, cell-type-resolved molecular portrait of the Purkinje fiber in a large-animal model and a generalizable strategy that selects markers for specificity, stability and accessibility. The resulting resource - including the cross-species marker contactin-5 and compartment-matched candidates - supplies validated tools to identify, isolate and target Purkinje cells and demonstrates the necessity of cross-species validation.

Background Postoperative atrial fibrillation (POAF) is a common complication of cardiac surgery with POAF susceptibility thought to be primarily the result of pre-existing structural atrial disease, specifically fibrosis. We tested this hypothesis directly by combining preoperative imaging of atrial fibrosis with continuous physiological monitoring before POAF onset. Methods This prospective single center study (2023?2025) comprised 6,697 adults without prior atrial fibrillation (AF) undergoing elective cardiac surgery. Subjects were enrolled into three prespecified, non-overlapping cohorts: a mechanistic imaging cohort (n=52) to test whether preoperative atrial fibroblast activation predicts POAF; a physiological monitoring cohort (n=3,183) to characterize peri-event autonomic dynamics via time-resolved HRV analysis; and an independent prospective observability cohort (n=3,451) for validation. The prespecified primary analyses assessed time-domain and frequency-domain HRV across six consecutive 10-minute intervals during the 60 minutes preceding POAF onset. Generalized estimating equations models were applied. Results Preoperative atrial fibroblast activation did not differ significantly between patients with (n=22) or without POAF (n=52). By contrast, in a physiological monitoring cohort (n=3,183), time-resolved heart rate variability analysis revealed progressive autonomic destabilization beginning approximately 20 minutes before POAF onset, with significant divergence in heart-rate-corrected SDNN in the final 10-minute pre-event interval (marginal mean difference 0.0139, 95% CI 0.0113?0.0164; P<0.001; Cohen's d=0.728). This signal was independently validated in a prospective observability cohort (n=3,451), achieving fragment-level sensitivity of 69.6% and specificity of 97.5% at the 10-minute horizon. Conclusions POAF is more closely associated with immediately preceding detectable autonomic destabilization than with preoperative structural substrate. These findings challenge the hypothesis that POAF susceptibility is structurally determined(structural-determinism) and reframe this frequent complication as a dynamic, state-dependent process that may be a target for active prevention.