Purpose Multiple randomized controlled trials have presented SGLT2 inhibitors (SGLT2i) as novel pharmacological therapy for patients with heart failure, resulting in reductions in hospitalization for heart failure and mortality. Given the absence of SGLT2 receptors in the heart, mechanisms of direct cardioprotective effects of SGLT2i are complex and remain to be investigated. In this study, we evaluated the direct biomechanical effects of SGLT2i empagliflozin on isolated myocardium from end-stage heart failure patients. Methods Ventricular tissue biopsies obtained from 7 patients undergoing heart transplantation or ventricular assist device implantation surgery were cut into 27 living myocardial slices (LMS) and mounted in custom-made cultivation chambers with mechanical preload and electrical stimulation, resulting in cardiac contractions. These 300 µm thick LMS were subjected to 10 µM empagliflozin and with continuous recording of biomechanical parameters. Results Empagliflozin did not affect the maximum contraction force of the slices, however, increased total contraction duration by 13% (p = 0.002) which was determined by prolonged time to peak and time to relaxation (p = 0.009 and p = 0.003, respectively). Conclusion The addition of empagliflozin to LMS from end-stage heart failure patients cultured in a biomimetic system improves contraction and relaxation kinetics by increasing total contraction duration without diminishing maximum force production. Therefore, we present convincing evidence that SGLT2i can directly act on the myocardium in absence of systemic influences from other organ systems.