HMC ameliorates myocardial ischaemia–reperfusion injury through suppressing cuproptosis via the de‐ubiquitination and stabilization of ATP7A

BACKGROUND AND PURPOSE: Myocardial ischaemia-reperfusion injury (MI/RI) is a major contributor to poor prognosis following revascularization in myocardial infarction patients, yet targeted therapies remain elusive. While hesperidin methyl chalcone (HMC) demonstrates antioxidant and vasoprotective properties, its role in modulating regulated cell death pathways during MI/RI is undefined. This study investigates the cardioprotective effects of HMC against MI/RI and its underlying mechanisms. EXPERIMENTAL APPROACH: Hypoxia/reoxygenation (H/R) models in H9c2 cardiomyocytes and a model of MI/RI in mice were employed. The effects of HMC on cardiac performance, copper flux analysis, ubiquitination assays and molecular docking, mitochondrial integrity and cell death modalities in response to MI/RI were examined. KEY RESULTS: HMC pretreatment exhibited significant cardioprotection, reducing infarct size and improving cardiac function in MI/RI mice. Mechanistically, HMC specifically attenuated cuproptosis, as evidenced by decreased copper overload and downregulation of cuproptosis effectors, mimicking the action of the copper chelator ammonium tetrathiomolybdate (ATTM). Crucially, the cuproptosis inducer elesclomol-Cu (Es-Cu) abolished the cardioprotective effects of HMC. Furthermore, HMC interacted with the E3 ubiquitin ligase MARCHF7, and disrupted the complex of MARCHF7/ATP7A, thereby reducing ubiquitination and proteasomal degradation of the copper exporter ATP7A. The stabilization of ATP7A enhanced copper efflux, alleviating cuproptosis, oxidative stress, inflammation and mitochondrial damage induced by MI/RI. CONCLUSIONS AND IMPLICATIONS: Our work unveils a ubiquitin-regulated copper homeostasis axis in MI/RI. By targeting MARCHF7-ATP7A interaction, HMC sustains copper export machinery to combat cuproptosis-driven injury. These findings position HMC as a novel therapeutic candidate for MI/RI through regulating ubiquitination-dependent regulation of copper homeostasis.