Abstract 217: Hexokinase II Binding to Mitochondria Suppresses Irreversible Ischemia Reperfusion Injury in the Beating Heart by Respiratory Inhibition and Reduced ROS Levels
Introduction: The glycolytic enzyme hexokinase II (HKII) can be mitochondrial-bound (mtHK) or free in the cytosol. Increased mtHKII protects the heart against ischemia-reperfusion (I/R) injury. The mechanism of protection is not yet elucidated.
Hypothesis: We hypothesized that mtHKII protects the heart against I/R injury by respiratory inhibition and reducing reactive oxygen species (ROS).
Methods: Langendorff perfused rat hearts were exposed for 20min to 1 μ M TAT-only, 1 μ M TAT-HK or 200 nM TAT-HK, followed by 15 min ischemia and 30 min reperfusion. TAT-HK contains the binding motif of HKII and dislodges HKII from mitochondria. ROS was measured using DHE fluorescence (n=6/group). Lactate was measured after peptide treatment, necrosis (LDH release) was determined during reperfusion and oxygen consumption (MVO2) and heart function (rate pressure product (RPP)) were monitored (n=12-15/group).
Results: Disruption of mtHKII binding resulted in increased MVO2/RPP values during both baseline and reperfusion in the 1 μ M TAT-HK group. Effluent lactate increased from (mean±SEM) 0.024±0.0041 μ mol/ml effluent in the TAT-only group to 0.042±0.0072 and 0.091±0.016 μ mol/ml effluent in the 200 nM and 1 μ M TAT-HK treated hearts, respectively. TAT-HK treatment had no effect on ROS production during baseline conditions. However, during both ischemia (normalized values (mean±SEM) 1.14±0.05 and 1.17±0.08 versus 1.09±0.06) and reperfusion (1.58±0.24 and 1.84±0.15 versus 1.30±0.17) TAT-HK significantly and dose dependently increased ROS when compared to TAT-only treatment. This was accompanied by an increased LDH release during reperfusion of (mean±SEM) 27.1±4.1 and 42.9±4.2 μ mol/30min/g heart for the 200 nM and 1 μ M TAT-HK treated hearts, respectively. There was no cell necrosis in the TAT-only group.
Conclusion: >Our results show for the first time that disruption of the mtHK binding affects cardiac MVO2 and causes an increase in ROS production during ischemia and reperfusion in the intact heart. These data indicate that mtHKII is a master switch turning reversible ischemia into irreversible ischemia. Our data also suggest that mtHKII is a determinant of glycolysis/glucose oxidation coupling. Supported by the Dutch Heart Foundation (NHS 2010B011)
- © 2012 by American Heart Association, Inc.