Overexpression of SOD2 in Myocytes Protects Mitochondrial Function from Myocardial Ischemia and Reperfusion Injury

Abstract

Mitochondrial dysfunction in myocardial ischemia and reperfusion (IR) is caused by oxidative stress. Mitochondrial electron transport chain (ETC) is the major source of superoxide and superoxide‐derived reactive oxygen species during I/R injury. Ischemic defect and reperfusion‐induced injury to ETC and Krebs cycle components are critical in mitochondrial dysfunction and the related disease pathogenesis of post‐ischemic injury. SOD2 is the primary antioxidant enzyme in a network of detoxification enzymes that neutralizes superoxide to H2O2 in the mitochondrial matrix. Mouse model with transgenic overexpression of human SOD2 (SOD2‐tg) in myocytes develops supernormal cardiac function through enhanced bioenergetic efficiency and myocardial blood flow. To test the therapeutic potential of using human SOD2 for cardioprotection and the effect of human SOD2 overexpression in the mouse heart on the post‐ischemic injury, isolated mouse heart of the SOD2‐tg mouse model was subjected to global ischemia for 30 min followed by reperfusion for 45‐min. TCC (2,3,5‐triphenyltetrazolium chloride) staining indicated that transgenic overexpression of SOD2 in myocytes significantly decreased the infarct area of the post‐ischemic heart. Analysis of oxygen consumption rates (OCR) by the isolated mitochondria indicated that ADP‐dependent and uncoupling OCRs were preserved in the mouse heart of SOD2‐tg after I/R injury, and mitochondrial integrity was further protected from IR impairment due to transgenic overexpression of SOD2 in the mouse heart. Assessment of the electron transport activity (ETA) from the alamathicin‐permeabilized mitochondria further indicated that SOD2 overexpression significantly preserved the ETA of complexes I, II, III, and IV in the post‐ischemic heart. For the components of Krebs cycle in the isolated mitochondria, it was measured that the activities of aconitase and isocitrate dehydrogenase were impaired by IR in wild type heart, but preserved in the mouse heart of SOD2‐tg. However, the enzymatic activity of malate dehydrogenase and citrate synthase in wild type mouse heart was not affected after IR injury, suggesting accumulation of citrate for cytosolic fatty acid biosynthesis and malate for cytosolic gluconeogenesis under the physiological conditions of hypoxia and reoxygenation. Immunoblotting analysis indicated a downregulation of ETC components and aconitase was induced by IR heart in the wild type heart, but protein expression of ETC components and aconitase was preserved in the post‐ischemic heart of SOD2‐tg mice, suggesting that SOD2 overexpression maintains protein quality control in the mitochondria of post‐ischemic heart. Overexpression of SOD2 in myocytes results in a more reduced redox setting of mouse heart as determined by the redox activity of hydroxylamine oxidation using EPR measurement. However, SOD2 overexpression did not prevent myocardium from IR‐mediated increased redox activity of hydroxylamine oxidation, suggesting that IR mediated a more oxidized redox setting in myocardium independently of superoxide generation by mitochondria. In conclusion, transgenic overexpression of human SOD2 in the mouse myocyte protects myocardium from IR injury by preserving mitochondrial function via maintaining protein quality control of ETC and Krebs cycle components.Support or Funding InformationNIH/HL083237