Mechanical unloading via LVAD reduced phospholamban aggregate burden and altered p62 compartmentalization in human myocardium with PLN-R14del cardiomyopathy.
Human myocardium and cultured slices reveal a pronounced PLN aggregate phenotype that is reduced under mechanical unloading, while iPSC-CM models capture stress-dependent functional defects preceding overt aggregate formation.
Abstract Background/Introduction The PLN-R14del mutation is associated with arrhythmogenic and dilated cardiomyopathy and marked structural remodeling. Human myocardium frequently displays phospholamban (PLN) aggregates, interstitial fibrosis, and lipid accumulation, reflecting disrupted protein handling. The mechanisms driving these features remain insufficiently defined. Diverse cardiac models offer complementary approaches to investigate structural signatures of PLN-R14del cardiomyopathy. Purpose To characterize structural and autophagy-related features of PLN-R14del cardiomyopathy across human myocardial tissue, cultured tissue slices, 2D iPSC-derived cardiomyocytes, and iPSC-CM spheroids using immunofluorescence-based analysis. Methods Multiple PLN-R14del models were examined. Fresh human myocardium and patient-derived tissue slices provided direct readouts of disease-associated pathology and, when cultured, enabled controlled experimental manipulation. iPSC-CMs and cardiac spheroids allowed investigation of phenotype development and responses to pharmacological stressors. Across all systems, immunofluorescence staining for PLN, SERCA, connexin-43, Troponin-I3, and p62 was performed to assess PLN localization and aggregate formation. Results Non-LVAD myocardium showed abundant PLN aggregates with consistent PLN–SERCA–p62 co-localization and marked structural disorganization. In contrast, tissue and slices from a mechanically unloaded heart supported by an LVAD displayed almost no detectable aggregates and a more preserved architecture. In LVAD samples, p62 localized both within the few remaining aggregates and at connexin-43–positive intercalated discs, whereas in non-LVAD samples p62 was confined to aggregates. These observations indicate unloading is associated with reduced aggregate burden and altered p62 compartmentalization. In iPSC-derived PLN-R14del cardiomyocytes, PLN distribution was uniform under baseline conditions and progressively redistributed following exposure to isoproterenol, caffeine, or epinephrine, indicating that pharmacological stress unmasks phenotypic features absent at baseline. Calcium handling was impaired under stress despite the absence of structural aggregates in iPSC-CMs suggesting functional alterations can precede overt aggregate formation. Spheroids showed preserved PLN organization without aggregate formation under the examined conditions. Conclusion Human myocardium and cultured slices reveal a pronounced PLN aggregate phenotype that is reduced under mechanical unloading and accompanied by shifts in p62 localization toward intercalated discs. In contrast, 2D iPSC-CMs and spheroids do not form aggregates at baseline and instead display stress-dependent PLN redistribution and impaired calcium handling without structural pathology. These complementary models capture distinct aspects of PLN-R14del cardiomyopathy and support the investigation of relationships between aggregate biology, autophagy, and functional defects.
Berg et al. (Fri,) conducted a other in PLN-R14del induced cardiomyopathy. Mechanical unloading (LVAD) and pharmacological stressors vs. Non-LVAD myocardium and baseline conditions was evaluated on PLN localization and aggregate formation. Mechanical unloading via LVAD reduced phospholamban aggregate burden and altered p62 compartmentalization in human myocardium with PLN-R14del cardiomyopathy.