Spatial proteomics reveals cell-type-specific drivers in cardiomyopathies

Background Cardiomyopathies drive heart failure through complex cellular changes, yet we lack understanding of how individual cell types respond across the heterogeneous landscape of the diseased heart. Here, we use Deep Visual Proteomics (DVP) to systematically analyze cardiac cell types in their native tissue context to identify novel therapeutic targets with cell-type and region specificity. Purpose We aim to map the proteomic landscape of cardiac cell types across four cardiomyopathy subtypes and healthy controls, from remote myocardium to fibrotic scar. Additionally, we want to understand the molecular switch of individual cardiomyocytes in the immediate scar-border zone. Methods We applied DVP, combining immunofluorescence-guided cell identification, laser microdissection and ultra-sensitive mass spectrometry, to left ventricular tissue from 32 individuals (6 healthy controls; 26 patients with ischemic, non-ischemic, hypertrophic and arrhythmogenic cardiomyopathy). Cardiomyocytes (TNNT2+) and macrophages (CD68+) from remote, border and scar zones were dissected and analyzed by mass spectrometry. Additionally, we isolated individual cardiomyocytes along the immediate scar-to-border zone from ischemic cardiomyopathy (ICM) patients. Results We quantified up to 5000 protein groups per cell type with clear separation of cardiac cell populations. Cardiomyocytes displayed a progressive molecular shift from healthy through remote to scar zones, shared across cardiomyopathy subtypes. Diseased cardiomyocytes showed downregulation of oxidative phosphorylation and ATP synthesis, alongside activation of extracellular matrix remodeling, immune signaling and complement pathways. Region-specific and cell interaction analyses uncovered cardiomyopathy-subtype specific signatures, including differential regulation of inflammatory regulators and sarcomere proteins. Analysis of individual cardiomyocytes in the scar-border zone resolved proteomic transitions, including changes in metabolic enzymes and structural proteins. Conclusion Our workflow enables spatially resolved, cell-type-specific proteomic profiling of the failing human heart. We demonstrate that tissue location is a major determinant in cardiomyopathy. While cardiomyopathies share a common signature of metabolic collapse and extracellular matrix activation, disease-specific and region-specific programs emerge that represent candidate targets for therapeutic intervention. Single-cell proteomics applied to the scar-border zone pinpoints where cardiomyocytes transition from adaptive to maladaptive states, opening new avenues for targeted therapies in heart failure.