Malonate given at reperfusion prevents post-myocardial infarction heart failure by decreasing ischemia/reperfusion injury

Abstract

The mitochondrial metabolite succinate is a key driver of ischemia/reperfusion injury (IRI). Targeting succinate metabolism by inhibiting succinate dehydrogenase (SDH) upon reperfusion using malonate is an effective therapeutic strategy to achieve cardioprotection in the short term (< 24 h reperfusion) in mouse and pig in vivo myocardial infarction (MI) models. We aimed to assess whether inhibiting IRI with malonate given upon reperfusion could prevent post-MI heart failure (HF) assessed after 28 days. Male C57BL/6 J mice were subjected to 30 min left anterior coronary artery (LAD) occlusion, before reperfusion for 28 days. Malonate or without-malonate control was infused as a single dose upon reperfusion. Cardiac function was assessed by echocardiography and fibrosis by Masson's trichrome staining. Reperfusion without malonate significantly reduced ejection fraction (~ 47%), fractional shortening (~ 23%) and elevated collagen deposition 28 days post-MI. Malonate, administered as a single infusion (16 mg/kg/min for 10 min) upon reperfusion, gave a significant cardioprotective effect, with ejection fraction (~ 60%) and fractional shortening (~ 30%) preserved and less collagen deposition. Using an acidified malonate formulation, to enhance its uptake into cardiomyocytes via the monocarboxylate transporter 1, both 1.6 and 16 mg/kg/min 10 min infusion led to robust long-term cardioprotection with preserved ejection fraction (> 60%) and fractional shortening (~ 30%), as well as significantly less collagen deposition than control hearts. Malonate administration upon reperfusion prevents post-MI HF. Acidification of malonate enables lower doses of malonate to also achieve long-term cardioprotection post-MI. Therefore, the administration of acidified malonate upon reperfusion is a promising therapeutic strategy to prevent IRI and post-MI HF.

Keywords: Heart failure with reduced ejection fraction; Ischemia/reperfusion injury; Malonate; Mitochondria; Reactive oxygen species; Succinate.

Fig. 1

Schematic of malonate inhibition of IRI in MI. Ischemia-accumulated succinate is rapidly oxidized on reperfusion by SDH upon reperfusion. This drives the ROS burst at reperfusion via RET at complex I, which subsequently primes cardiomyocytes for cell death and leads to post-MI HF. Malonate inhibits SDH, slowing down succinate oxidation and ROS production on reperfusion, reducing cell death. Malonate entry into cardiomyocytes can be enhanced by ischemia or low pH, driving the entry of monocarboxylate malonate via MCT1. Whether malonate can prevent post-MI HF by preventing RET-ROS on reperfusion is currently unknown. MCT1 monocarboxylate transporter 1, DIC dicarboxylate carrier, SDH succinate dehydrogenase, CxI complex I, RET reverse electron transport, ROS reactive oxygen species, TCA tricarboxylic acid cycle, EF ejection fraction

Fig. 2

Pharmacokinetics of malonate in vivo. A Malonate action is short-lived in vivo. C57BL/6 J mice were IP injected malonate (160 mg/kg as disodium malonate (DSM); 100 µl bolus in neutral pH saline) and serial blood samples taken at 0, 15, 30, 60, 120 min and 24 h after injection. Plasma malonate was assessed by LC–MS/MS. B Malonate does not accumulate in tissues. C57BL/6J mice were IP injected daily (160 mg/kg as DSM; 100 µl bolus in neutral pH saline) for 7 days before harvesting tissues 24 h after the last dose and measuring tissue malonate by LC–MS/MS. Data presented as mean ± S.D. from N = 4 biological replicates

Fig. 3

Malonate infusion on reperfusion prevents post-MI HF. C57BL/6 J mice were subjected to 30 min LAD ligation prior to 28 day reperfusion ± 16 mg/kg/min 10 min infusion of malonate (pH 7.4), 1.6 or 16 mg/kg/min 10 min infusion of acidified malonate (pH 6) or saline control, experimental outline depicted in (A). Cardiac function was monitored for ejection fraction (B) and fractional shortening (C) by echocardiography at baseline, 3 and 28 days post-MI. Quantification of scar size as a percentage of LV area (D) and fibrosis (E) of heart Sections 28 days post-MI and representative stained heart images (E). Data presented as mean ± S.D. of N = 5–6 biological replicates. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 versus without malonate-treated animals