Succinate Accumulation Accelerates Oxidative Stress to Promote Pulmonary Epithelial Cell Apoptosis During Lung Ischemia–Reperfusion Injury

Abstract

During ischemia, succinate accumulates and leads to significant damage to the tissues. The specific role of succinate in lung ischemia–reperfusion injury (LIRI) remains unresolved. Differential metabolites in LIRI were identified through untargeted metabolomics using gas chromatography–mass spectrometry (GC–MS). Type II alveolar epithelial cells (AECs) were cultured and subjected to hypoxia/reoxygenation (H/R) in vitro, while an in vivo LIRI model was developed using C57BL/6 mice. Cytokine levels, lung oedema, histopathological alterations and lung functionality were evaluated. Protein levels were analysed through Western blotting. The mitochondrial membrane potential (Δψm) was measured using the JC-1 fluorescent dye, and mitochondrial morphology in Type II AECs following H/R damage was observed with a transmission electron microscope (TEM). Oxidative stress and apoptosis markers were detected in lung tissues and Type II AECs. Succinate was increased in the peripheral serum of LIRI patients and the C57BL/6 mices model. Succinate pre-treatment promotes Type II AEC cell apoptosis and oxidative stress, inhibits mitochondrial membrane potential and damages the alveolar epithelial cells' mitochondrial activity after H/R. Meanwhile, succinate may considerably reduce the amounts of acyl-CoA oxidase 1 (ACOX1) and isocitrate dehydrogenase 2 (IDH2) protein expression. Importantly, N-acetyl-L-cysteine (NAC) was observed to dramatically retard succinate-induced cell apoptosis, mitochondrial dysfunction and ROS levels in alveolar epithelial cells following H/R in vivo, with succinate-neutralising antibodies protecting LIRI in vitro. In conclusion, during ischemia, the build-up of succinate contributes to the advancement of LIRI by enhancing mitochondrial oxidative stress and promoting cell apoptosis, and blocking succinate may be a potential target for LIRI treatment.