One of the major research efforts in Cardiology has been to salvage myocardium at risk for necrosis in patients with acute myocardial infarction. Two currently utilized interventions, namely the use of thrombolytic agents and percutaneous transluminal coronary angioplasty both involve reperfusion of acutely ischemic myocardium. Unfortunately, reperfusion may result in a form of myocardial damage which is caused by reflow itself, termed """"""""reflow injury"""""""". Free radical generation has been proposed as the central mechanism responsible for this reperfusion damage. Previous experimental studies of free radical generation in the heart have been indirect, documenting beneficial effects of the administration of free radical scavengers. There has been a great need for a technique which is capable of directly detecting and quantitating free radical generation during ischemia and during reperfusion of post-ischemic hearts. We have now demonstrated that Electron Paramagnetic Resonance, EPR, Spectroscopy can be used to directly measure and thus document radical generation in frozen samples obtained from ischemic and post-ischemic hearts. We have also designed and built a prototype resonator design suitable for in vivo EPR measurements in living isolated perfused hearts. In this grant application we propose to utilize these EPR techniques to definitively measure, quantitate and characterize free radical generation in the post-ischemic heart. Experiments will be performed to identify which free radicals are generated as well as the cellular mechanisms of their formation. In addition, optimal interventions for prevention of reperfusion injury will be determined. Free radical generation will be correlated with the observed reduction in cardiac contractile function, levels of high energy phosphates, and metabolic status of the heart. EPR measurements will be performed on frozen heart tissue at frequencies of 9 GHz and 35 GHz. We will also optimize resonator and microwave bridge design in order to perform in vivo free radical measurements in the heart at a frequency of 1-2 GHz. These experiments should provide new fundamental insight into the mechanisms of free radical generation in the heart on post-ischemic reperfusion, and elucidate the optimal approaches to prevent free radical induced reperfusion injury. Thus, this work will provide some of the crucial information which is clinically needed to determine how to save heart muscle at risk of infarction.
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