Empagliflozin attenuates doxorubicin-impaired cardiac contractility by suppressing reactive oxygen species in isolated myocytes

Abstract

In animal studies, sodium-glucose co-transporter-2 inhibitors-such as empagliflozin-have been shown to improve heart failure and impaired cardiac contractility induced by anthracyclines-including doxorubicin-although the therapeutic mechanism remains unclear. Moreover, abnormalities in Ca²⁺ handling within ventricular myocytes are the predominant feature of heart failure. Accordingly, this study aimed to investigate whether empagliflozin can alleviate Ca²⁺ handling disorders induced by acute doxorubicin exposure and elucidate the underlying mechanisms. To this end, ventricular myocytes were isolated from C57BL/6 mice. Contraction function, Ca²⁺ handling, and mitochondrial reactive oxygen species (ROS) generation were then evaluated using IonOptix or confocal microscopy. Ca²⁺ handling proteins were detected by western blotting. Results show that incubation with 1 μmol/L of doxorubicin for 120-min impaired cardiac contractility in isolated myocytes, which was significantly alleviated by pretreatment with 1 μmol/L of empagliflozin. Doxorubicin also markedly induced Ca²⁺ handling disorders, including decreased Ca²⁺ transients, prolonged Ca²⁺ transient decay time, enhanced frequency of Ca²⁺ sparks, and decreased Ca²⁺ content in the sarcoplasmic reticulum. These dysregulations were improved by pretreatment with empagliflozin. Moreover, empagliflozin effectively inhibited doxorubicin-induced mitochondrial ROS production in isolated myocytes and rescued doxorubicin-induced oxidation of Ca²⁺/calmodulin-dependent protein kinase II (ox-CaMKII) and CaMKII-dependent phosphorylation of RyR2. Similarly, preincubation with 10 μmol/L Mito-TEMPO mimicked the protective effects of empagliflozin. Collectively, Empagliflozin ameliorated the doxorubicin-induced contraction malfunction and Ca²⁺-handling disorders. These findings suggest that empagliflozin alleviates Ca²⁺-handling disorders by improving ROS production in the mitochondria and alleviating the enhanced oxidative CaMKII signaling pathway induced by doxorubicin.

Keywords: Ca2+ handling; Doxorubicin; Empagliflozin; Heart failure; ROS.