Prevenció de dany per reperfusió mitjançant inhibició reversible de la succinat deshidrogenasa mitocondrial. Mecanismes implicats.

Abstract

Cell death that occurs during an episode of coronary occlusion is directly proportional to the duration of ischemia. Therefore, the treatment of choice against myocardial infarction is the rapid reopening of the affected artery. However, the restoration of blood flow is associated with an additional damage, known as reperfusion injury, due to excessive production of reactive oxygen species (ROS). As a way to reduce deleterious effects of oxidative stress, different antioxidant molecules have been tested, but with contradictory results. A new approach could be to inhibit directly ROS production, by mitochondrial function modulation. Recent studies have suggested that some of ROS produced during reperfusion are due to reverse electron transfer from complex II (or succinate dehydrogenase) of the mitochondrial electron transport chain to complex I. The objective of this thesis has been to investigate the effects of reversible inhibition of mitochondrial succinate dehydrogenase with disodium malonate, administered at the moment of the restoration of the flow, on reperfusion injury, as well as to study the mechanisms involved. Administration of malonate, under normoxic conditions in isolated mice hearts perfused in a Langendorff system, induced a concentration-dependent reduction in left ventricular developed pressure, confirming an inhibitory action on mitochondrial respiration. Thereafter, additional isolated mice hearts were subjected to ischemia-reperfusion and were treated or not with 3 mM malonate, given during the first 15 minutes of reperfusion. Administration of malonate significantly reduced infarct size that was associated with a lower LDH release during reperfusion and with a better functional recovery. Nuclear magnetic resonance studies demonstrated the existence of differences in the metabolic profile between hearts treated with malonate, with increased succinate concentration. In addition, MitoSOX Red staining of myocardial tissue samples revealed that malonate treatment was associated with a reduction in ROS production. This decrease was also observed in isolated mitochondria obtained from mice hearts, treated in vitro with both succinate and malonate, that also depicted a reduction in mitochondrial permeability transition pore opening. Moving forward, we analyzed whether the protective effects of malonate persist in a more clinically relevant animal model, the porcine model of transient coronary occlusion. In this model we selectively administer malonate with intracoronary infusion. Under baseline conditions, only malonate of 50 mmol/L, but not lower concentrations, resulted in a significant reduction in systolic segment shortening in the myocardial region irrigated by the artery of interest, but not in distant regions. To analyze the effects of reversible inhibition of succinate dehydrogenase on reperfusion injury, animals were subjected to left anterior descending coronary artery occlusion (40 minutes), followed by reperfusion, and they were treated or not with malonate 10mmol/L, during the first 5 minutes of the same. Selective administration of malonate into the risk area significantly reduced infarct size, without changes in the incidence of malignant ventricular tachyarrhythmias during reperfusion, or in the contractility of remote myocardium. This cardioprotective effect was associated with a reduction in ROS production (MitoSOX Red). In conclusion, reversible inhibition of succinate dehydrogenase with disodium malonate at the onset of reperfusion has cardioprotective effects against myocardial infarction, both in isolated mice hearts and in a porcine model of transient coronary occlusion. This effect is due to a lower succinate accumulation in the tissue, resulting in a reduction of reverse electron transfer to complex I, reduced ROS production and mitochondrial permeability transition pore opening.

Date of Award	13 Nov 2019
Original language	Catalan
Supervisor	Antonio David Garcia Dorado Garcia (Director), Antonio Rodríguez Sinovas (Director) & Jaume Alijotas Reig (Tutor)