Abstract 162: Liproxstatin-1 Treatment Protects the Myocardium Against I-R Injury By Decreasing VDAC1 Levels and Increasing GPX4 Activity

Introduction: Ferroptosis is a non-apoptotic form of cell death characterized by iron-dependent accumulation of lethal lipid reactive oxygen species (ROS) and liproxstatin-1 is a potent inhibitor of ferroptosis. We investigated whether Liproxstatin-1 can protect the heart against ischemia/reperfusion (I/R) injury and determined the mechanism underlying cardioprotection. Methods: Isolated hearts from C57BL/6J mice were subjected to 35 min ischemia followed 120 min reperfusion and treated with Liproxstatin-1 (200nM). Myocardial infarct size was assessed by TTC staining at the end of reperfusion. Mitochondrial integrity was measured by analyzing cristae morphology and ROS production was assessed using Amplex red method. The calcium retention capacity (CRC) required to induce mitochondrial permeability transition pore (mPTP) opening was measured using calcium green dye. The levels of mitochondrial proteins cyclophilin D (CypD), Glutathione peroxidase 4 (GPX4), VDAC1, 2 and 3 were assessed by Western blot. And GPX4 activity was measured using a kit. Results: We found that liproxstatin-1 treatment decreased myocardial infarction size, ROS production, and VDAC1 expression as well as preserved mitochondrial structural integrity by protecting cristae from damage. These mitochondrial effects of liproxstatin-1 were associated with the increase in GPX4 levels and activity. But, we also found that liproxstatin-1 did not affect the CypD, VDAC2, and VDAC3 expression as well as mitochondrial CRC. Conclusion: Together, these results indicate that liproxstatin-1 treatment protects myocardium against I/R injury by increasing GPX4 activity resulting in the decrease in mitochondrial ROS production; preserving mitochondrial structure as well as a decrease in VDAC1 (but not VDAC 2 and 3) expression. Liproxstain-1 cardioprotective action does not involve the regulation of the mPTP opening. These results point to the cardioprotective role of this anti-ferroptosis compound after I/R.