Greta Dockenfuss was supported by an Otto-Hess stipend of the DGK
Background: Ischemic heart disease represents a major public health problem. The number of fatal myocardial infarctions (MI) has declined; however, MI-surviving patients develop heart failure as consequence of an irreversible loss of cardiomyocytes. The remaining myocardium functionally compensates the loss in part for a certain period of time, but fibrotic and metabolic remodeling occurs and results in a decline of heart function and ultimately death of the patient. The present project investigates the hypothesis that contractile arrest alters mitochondrial complexome composition in a way that promotes cardiomyocyte proliferation. Therefore, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were cultured in engineered heart tissue format (EHT) and subjected to contractile arrest.
Methods and Results: We used genetically engineered PSAM-GlyR hiPSC to generate CMs and EHTs. In these PSAM-GlyR-EHTs, the chemical compound PSEM-89-S induced fast and reversible arrest of contraction (“stopped EHTs”). Transmission electron microscopy analysis revealed disassembly of sarcomeric structure. Proteomic analysis demonstrated an enrichment of pathways associated with cell cycle activity in stopped EHTs. Furthermore, mitochondrial respiration was significantly reduced under coupling conditions (control: 12.28 pmol/s/mg O₂flux vs. PSEM-89-S: 5.47 pmol/s/mg O₂ flux) in stopped EHTs. The reduced OCR was accompanied by a relative increase in extracellular acidification rate (ECAR), resulting in an elevated ECAR/OCR ratio (control: 0.20 vs. PSEM-89-S: 0.27), indicative of a shift toward glycolytic metabolism. Dynamic mitochondrial complexome analysis using Stable Isotope Labeling by Amino Acids in Cell Culture (SILAC), followed by Blue Native Gel Electrophoresis and mass spectrometry, revealed a selective reduction of respiratory supercomplexes, and ATP synthase-containing assemblies. Notably, contractile arrest was associated with a pronounced accumulation of ATP-Synthase Inhibitor 1 at ATP synthase complexes in stopped EHTs, exerting an off-switch on mitochondrial respiration.
Our data connect for the first time cardiomyocyte contractility with distinct functional alterations in mitochondrial complexome composition with impact on proliferation that could be modulated for therapeutic purposes in the future.
