Empagliflozin mitigates doxorubicin-induced cardiac dysfunction in association with preserved t-tubule integrity and improved Ca2+ handling: a structural mechanistic insight

Background Clinical and experimental studies have suggested that sodium-glucose cotransporter type-2 inhibitors (SGLT2i) may exert a preventive effect against doxorubicin (Dox)-induced heart failure; however, the underlying mechanisms remain incompletely understood. This study aimed to investigate whether empagliflozin (Emp) protects against Dox-induced cardiotoxicity by preserving transverse tubule (t-tubule) structure and improving Ca 2+ handling. Methods Adult male C57BL/6 mice were randomly assigned to four groups: Control, Emp-only, Dox-only, and Dox+ Emp. Emp was administered to the Emp-only and Dox+ Emp groups at 10 mg/kg/day, and Dox was administered to the Dox-only and Dox+ Emp groups at a cumulative dose of 15 mg/kg. Cardiac function was assessed by echocardiography. T-tubule ultrastructure and Ca 2+ handling were evaluated using confocal microscopy and IonOptix imaging. Protein expression was analyzed by western blotting. Results Dox-treated mice exhibited increased left ventricular dilatation and impaired cardiac function, both of which were attenuated in the Dox + Emp group. Dox induced blunted and dyssynchronous sarcoplasmic reticulum Ca 2+ release, characterized by reduced amplitude and delayed Ca 2+ transients, as well as increased Ca 2+ spark frequency. These abnormalities were significantly ameliorated by Emp treatment. In addition, Emp effectively prevented Dox-induced t-tubule loss and disorganization. Emp also ameliorated Dox-induced alterations in JPH2, BIN1, Ca 2+ /calmodulin-dependent protein kinase II (CaMKII), and CaMKII-dependent phosphorylation of RyR2, leading to improved synchronous Ca 2+ release, enhanced Ca 2+ transient amplitude, and reduced Ca 2+ sparks frequency. Conclusions Dox-induced cardiotoxicity is associated with severe t-tubule remodeling and impaired Ca 2+ -handling. Emp prevents Dox-induced structural and functional remodeling of t-tubule and improves Ca 2+ -handling, thereby exerting cardioprotective effects in mice. These findings provide structural-level mechanistic insight into Emp-mediated cardioprotection and support the potential of Emp as a preventive strategy against Dox-induced cardiotoxicity.