Molecular Determinants and Systems-Level Modulation of Cardiac Excitation–Contraction Coupling: Integrative Mechanisms Governing Calcium Handling, Sarcomeric Dynamics, and the Regulation of Myocardial Contractile Force in Health and Heart Failure

RESEARCH PROPOSAL JCTC, CardioCare 2026 Title Molecular Determinants and Systems-Level Modulation of Cardiac Excitation–Contraction Coupling: Integrative Mechanisms Governing Calcium Handling, Sarcomeric Dynamics, and the Regulation of Myocardial Contractile Force in Health and Heart Failure Principal Investigator Camilo Fernández Bravo, MD/PhD/FACC ORCID ID: 0009-0004-6848-7115 MD, Family Medicine – UCM Carlos J Finlay, Cuba Diplomate in Cardiovascular Diseases, General Cardiology Fellow of the American College of Cardiology (FACC), USA Postdoctoral Research Fellow, Harvard Medical School Associate Professor, European Society of Preventive Cardiology Medical Office Manager Professional Certificate, Johns Hopkins University, USA Emergency & Critical Care Fellowship, Stanford University UN Global Surgery Learning Hub Abstract (≈250 words) Cardiac excitation–contraction coupling (ECC) is the fundamental process linking electrical excitation to mechanical contraction in cardiomyocytes, primarily orchestrated by precise calcium (Ca²⁺) handling, sarcomeric protein interactions, and regulatory signaling pathways. In heart failure (HF), dysregulation of these mechanisms— including altered L-type Ca²⁺ channel function, ryanodine receptor (RyR2) leak, reduced SERCA2a activity, phospholamban dysregulation, and sarcomeric myofilament Ca²⁺ sensitivity—leads to impaired systolic force generation, diastolic dysfunction, and arrhythmogenesis. This proposal aims to elucidate the molecular and systems-level determinants of ECC in health and HF using an integrative approach combining advanced imaging, multi-omics analyses, computational modeling, and functional assays. We hypothesize that HF-specific remodeling disrupts the spatiotemporal coordination between Ca²⁺ microdomains, sarcomeric dynamics, and contractile force, and that targeted modulation of key nodes (e.g., SERCA-phospholamban axis, RyR2 stabilization) can restore contractility. Specific aims include: (1) mapping molecular alterations in Ca²⁺ handling proteins and sarcomeric components in human and animal HF models; (2) quantifying systems-level impacts on ECC using high-resolution live-cell imaging and force measurements; and (3) testing therapeutic modulation strategies to rescue contractile function. Expected outcomes will provide novel insights into ECC pathophysiology and identify translational targets for HF therapy, aligning with CardioCare 2026 priorities in precision cardiovascular medicine. Specific Aims Aim 1: Characterize molecular determinants of ECC dysregulation in heart failure. • Profile expression, post-translational modifications, and localization of key Ca²⁺ handling proteins (LTCC, RyR2, SERCA2a, phospholamban, NCX) and sarcomeric regulators (troponin complex, titin, myosin-binding protein C) in healthy vs. failing myocardium using proteomics, transcriptomics, and super-resolution microscopy. Aim 2: Investigate integrative mechanisms governing Ca²⁺ handling, sarcomeric dynamics, and contractile force at the systems level. • Employ live-cell Ca²⁺ imaging, T-tubule structural analysis, and simultaneous sarcomere length/force measurements in isolated cardiomyocytes and engineered heart tissues to dissect spatiotemporal ECC impairments in HF models. Aim 3: Evaluate systems-level modulation strategies to restore myocardial contractility. • Test pharmacological and genetic interventions (e.g., SERCA activators, RyR stabilizers, myofilament Ca²⁺ sensitizers) for their ability to normalize ECC parameters and improve force generation in failing models, with validation in translational platforms. Significance Heart failure remains a leading cause of morbidity and mortality worldwide, with excitation–contraction coupling defects central to contractile dysfunction. Despite advances, current therapies (e.g., beta-blockers, SGLT2 inhibitors) do not fully address underlying Ca²⁺ and sarcomeric abnormalities. This project bridges molecular insights with systems physiology to uncover integrative failure mechanisms, potentially revealing novel therapeutic nodes. It holds high translational relevance for developing precision interventions in HFrEF and HFpEF. Innovation • Integrative multi-scale approach: combining molecular profiling, real-time ECC imaging, computational modeling of Ca²⁺-myofilament coupling, and force dynamics—not typically combined in prior studies. • Focus on human-relevant models alongside animal systems for enhanced translatability. • Exploration of novel crosstalk between Ca²⁺ microdomains, T-tubule remodeling, and sarcomeric compliance in HF progression. Approach (Overview) Preliminary Considerations / Feasibility The PI brings extensive clinical cardiology expertise (FACC), postdoctoral training at Harvard Medical School, and interdisciplinary fellowships, ensuring robust interpretation of basic and translational findings. Access to advanced facilities (via Harvard/Stanford collaborations) includes confocal/super-resolution microscopy, patch-clamp electrophysiology, high-throughput proteomics, and engineered tissue platforms. Experimental Design (per Aim) Aim 1: Human failing hearts (from transplant/biopsy repositories) and rodent TAC/ischemic HF models. Techniques: LC-MS/MS proteomics, RNA-seq, STED microscopy for protein localization. Statistical power: n≥8/group, ANOVA/Machine learning clustering for pattern detection. Aim 2: Isolated adult cardiomyocytes and 3D engineered heart tissues. Methods: Fluo-4/Indo-1 Ca²⁺ imaging, line-scan confocal for spark/wave analysis, optical coherence tomography or traction force microscopy for sarcomere/force dynamics. Metrics: Ca²⁺ transient amplitude, decay kinetics, ECC gain, myofilament Ca²⁺ sensitivity (via force-Ca²⁺ relationships). Aim 3: Interventions: small-molecule SERCA activators (e.g., istaroxime analogs), JTV519/RyR stabilizers, omecamtiv mecarbil (myosin activator). Endpoints: restoration of Ca²⁺ transients, reduced diastolic leak, normalized force-frequency response. Validation: in vivo echocardiography in small-animal HF models. Timeline & Milestones (2–3 year proposal) Year 1: Aim 1 completion + preliminary Aim 2 data. Year 2: Full Aim 2 + intervention screening (Aim 3). Year 3: Mechanistic validation, data integration, manuscript preparation. Ethical Considerations All studies will adhere to institutional IACUC/IRB guidelines, Helsinki Declaration, and CIOMS standards. Human tissues will use de-identified samples with prior ethics approval. Budget Overview (high-level) Personnel, supplies (reagents/imaging), animal costs, sequencing/proteomics core fees, travel for CardioCare 2026 presentation. References (selected; full list available upon request) • Bers DM. Calcium cycling in heart failure: the arrhythmia connection. J Mol Cell Cardiol. 2000. • Zima AV et al. Ca handling during ECC in heart failure. Pflugers Arch. 2014.