Abstract 3302: ROS-mediated Dysregulation of Mitochondrial Permeability Transition Pore Underlies Increased Susceptibility of Hypertensive Hypertrophied Hearts to Ischemia/reperfusion Injury
Objective We examined the hypothesis that reduced tolerance of the myocardium of hypertensive hypertrophied hearts to ischemia/reperfusion injury is attributable to modified regulation of the mitochondrial permeability transition pore (mPTP).
Methods Ventricular tissues were sampled from stroke-prone spontaneously hypertensive rats (SHR-SP) and Wistar-Kyoto rats (WKY) at the age of 12~16 weeks, and protein levels of mPTP subunits and their interactions in the mitochondrial fractions were examined by immunoblotting and immunoprecipitation. As an index of oxidant stress-induced protein modification, carbonylation of proteins was determined. To induce cytoprotective signaling upstream of mPTP, erythro-poietin (EPO) receptor was activated by EPO at a dose of 10 units/ml in vitro or 5000 IU/kg in vivo.
Results Heart weight-to-body weight ratio in SHR-SP was 25% higher than in WKY. There was no significant difference between SHR-SP and WKY in levels of voltage dependent anion channel (VDAC), adenine nucleotide translocase (ANT), cyclophilin-D (Cyp-D) in the mitochondrial fraction. Levels of phosphorylation of Akt, ERK and GSK-3β before and after EPO receptor activation in vitro were also similar in the two groups. However, level of carbonylation of mitochondrial proteins less than 43kDa after 25-min ischemia/5-min reperfusion was significantly higher by 20% in SHR-SP than in WKY. Furthermore, Cyp-D level in the ANT immunoprecipitates was higher by 100% in SHR-SP than in WKY. This augmented Cyp-D-ANT interaction in SHR-SP was almost normalized by treatment with N-(2-mercaptopropionyl)-glycine (MPG, 1 mM), a scavenger of reactive oxygen species (ROS), commenced 10 min before ischemia. Infarct size after 25-min ischemia/2-hr reperfusion in vivo was significantly larger in SHR-SP than in WKY (71.2±3.1% of risk area vs. 54.8±3.7%), and MPG treatment reduced infarct size in SHR-SP to a level equivalent to that in WKY.
Conclusion Although pro-survival signaling upstream of mPTP is preserved, Cyp-D-ANT interaction, a trigger of mPTP opening, is augmented by a ROS-dependent mechanism and contributes to increased susceptibility to ischemia/reperfusion-induced necrosis in a model of hypertensive hypertrophied myocardium.