Recent reports indicate that elevating
DNA glycosylase/
AP lyase repair
enzyme activity offers marked cytoprotection in cultured cells and a variety of injury models. In this study, we measured the effect of EndoIII, a fusion
protein construct that traffics
Endonuclease III,
a DNA glycosylase/
AP lyase, to the mitochondria, on
infarct size in a rat model of
myocardial ischemia/reperfusion. Open-chest, anesthetized rats were subjected to 30 min of occlusion of a coronary artery followed by 2 h of reperfusion. An intravenous bolus of EndoIII, 8 mg/kg, just prior to reperfusion reduced
infarct size from 43.8 ± 1.4% of the risk zone in control animals to 24.0 ± 1.3% with no detectable hemodynamic effect. Neither EndoIII's vehicle nor an enzymatically inactive EndoIII mutant (K120Q) offered any protection. The magnitude of EndoIII's protection was comparable to that seen with the
platelet aggregation inhibitor cangrelor (25.0 ± 1.8%
infarction of risk zone). Because loading with a P2Y12 receptor blocker to inhibit platelets is currently the standard of care for treatment of acute
myocardial infarction, we tested whether EndoIII could further reduce
infarct size in rats treated with a maximally protective dose of
cangrelor. The combination reduced
infarct size to 15.1 ± 0.9% which was significantly smaller than that seen with either
cangrelor or EndoIII alone. Protection from
cangrelor but not EndoIII was abrogated by pharmacologic blockade of phosphatidylinositol-3
kinase or
adenosine receptors indicating differing cellular mechanisms. We hypothesized that EndoIII protected the heart from spreading
necrosis by preventing the release of proinflammatory fragments of
mitochondrial DNA (
mtDNA) into the heart tissue. In support of this hypothesis, an intravenous bolus at reperfusion of
deoxyribonuclease I (
DNase I) which should degrade any
DNA fragments escaping into the extracellular space was as protective as EndoIII. Furthermore, the combination of EndoIII and
DNase I produced additive protection. While EndoIII would maintain mitochondrial integrity in many of the ischemic cardiomyocytes,
DNase I would further prevent
mtDNA released from those cells that EndoIII could not save from propagating further
necrosis. Thus, our
mtDNA hypothesis would predict additive protection. Finally to demonstrate the toxicity of
mtDNA, isolated hearts were subjected to 15 min of global
ischemia.
Infarct size doubled when the coronary vasculature was filled with
mtDNA fragments during the period of global
ischemia. To our knowledge, EndoIII and
DNase are the first agents that can both be given at reperfusion and add to the protection of a P2Y12 blocker, and thus should be effective in today's patient with acute
myocardial infarction.