Oxidative stress plays a significant role in the development and progression of various pathologies. In heart failure, a leading cause of death worldwide, oxidative stress is associated with multiple risk factors, such as diabetes and metabolic syndrome. The progression of heart disease is further linked to profound metabolic remodelling, mitochondrial dysfunction and disturbed redox homeostasis. These intercon-nected changes may lead to energy depletion, impaired cardiac function and ultimately heart failure. Based on preliminary data in the tissue of failing hearts, we aimed to design an experimental setup further focusing on cellular changes in vitro. The deployed cardiac models, including human stem cell-derived cardiac organoids (""cardioids"") as well as 2D human cardiomyocyte cell lines (AC16; HCM), were differentially treated to environments reflecting pathological aspects of heart disease. To address the crosstalk of aberrant metabolism and alterations of the myocardial redox state correlated with oxi-dative stress and heart failure, cellular samples were subjected to parallel mass spectrometry-based redox metabolomic and proteomic analyses. Preservative two-step alkylation allowed for the accurate assessment of cellular glutathione redox status as a proxy for oxidative stress and, additionally, the redox status of individual redox-sensitive (cysteine-containing) peptides. Label-free quantitative prote-omics, redox proteomics and untargeted metabolomics were carried out after corresponding chromato-graphic separation on a coupled timsTOF Pro mass spectrometer (Bruker) operated in positive or neg-ative mode, respectively, with enabled trapped ion mobility spectrometry and the scan mode set to par-allel accumulation–serial fragmentation. Targeted redox metabolomics is based on a LCMS-8060 sys-tem (Shimadzu), working in positive multiple reaction monitoring mode for glutathione and its derivatives. Results provide insights into molecular mechanisms of the cellular stress response and metabolism. This includes the adaptive and specific regulation of antioxidative enzymes matching glycolytic and mi-tochondrial flux to preserve redox homeostasis. The experimental setup was further validated by multi-ple signatures matching prior findings in the tissue of failing hearts, including distinct changes in the expression of proteins involved in collagen synthesis and modification, stress-mediated extracellular matrix remodelling, lipid metabolism and ion transport. Further efforts to mature and thereby improve the translational ability of the cardiac in vitro models to drive our investigation of nutrient-driven redox signalling have been productive. Follow-up experiments on potential key mediators between metabolism and oxidative stress, as identi-fied by redox proteomics, are being carried out. In conclusion, elucidation of the intricate pathophysio-logical events in heart failure requires profound analytical methods and the utilization of cardiac in vitro models appropriate to their respective translational ability to drive the discovery and validation of novel strategies in diagnosis and treatment.