Date of Award

10-1-2005

Degree Type

Restricted Access Dissertation

Degree Name

Doctor of Philosophy

Field of Study

Integrative Physiology

Department

Graduate School of Biomedical Sciences

First Advisor

Robert T. Mallet

Second Advisor

James Caffrey

Third Advisor

Jerry Simecka

Abstract

Knott, E. Marty, The Effects of Pyruvate on Oxidative Stress and Myocardial Energetics During Cardioplegic Arrest and Reperfusion Doctor of Philosophy (Integrative Physiology), October 2005, 118 pp, 1 tables, 17 figures, references, 130 titles. Cardioplegic arrest for bypass surgery imposes global ischemia on the myocardium generating oxyradicals which contribute to post surgical cardiac dysfunction. Early clinical trials have demonstrated that pyruvate-fortified cardioplegia reduces myocardial energy and improves mechanical recovery in patients undergoing elective cardiopulmonary bypass for coronary artery bypass grafting. This study was designed to determine the effects of the natural carbohydrate, pyruvate, on oxidative stress, myocardial energy state, and activities of myocardial metabolic enzymes during and immediately following cardiopulmonary bypass. In the first set of experiments, in situ swine hearts were arrested for 60 min with a 4:1 mixture of blood and crystalloid cardioplegia solution containing 188 mM glucose alone (control) or with additional 23.8 mM lactate or 23.8 mM pyruvate, then reperfused for 3 min with cardioplegia-free blood. Glutathione redox state (GSH/GSSG) and phosphocreatine phosphorylation potential were determine from measurements of myocardial metabolites in left ventricular heart tissue snap frozen at 45 min arrest and 3 min reperfusion. Coronary sinus 8-isoprostane indexed oxidative stress. Pyruvate-fortified cardioplegia decreased oxidative stress, lowering 8-isoporstane content accumulate during arrest and reperfusion. Phosphorylation potential was maintained in all groups during arrest but fell upon reperfusion in the control and lactate and cardioplegia groups. Use of pyruvate cardioplegia during arrest prevented the decline in phosphorylation potential during reperfusion. Pyruvate cardioplegia doubled GSH/GSSG during arrest as compared to lactate, but GSH/GSSG fell during reperfusion all 3 groups. Pyruvate proved to be an effective antioxidant and energy yielding fuel in the setting of carioplegic arrest and reperfusion. From these data, we hypothesized that pyruvate would protect oxidant-sensitive enzymes from inactivation. To test this hypothesis, in situ swine hearts were arrested for 60 min with control cardioplegia and reperfused for 3 min with cardioplegia-free blood alone or with co-infusion c. 12 mM pyruvate. Activities of oxidant-sensitive enzymes, 8-isoprostane content, and energy and antioxidant metabolites were measured in left ventricular myocardium snap-frozen at 45 min arrest and 3 min reperfusion. At 3 min reperfusion, glutathione redox state fell by 70% while 8-isoprostane content increased 75%. Pyruvate administration during reperfusion suppressed oxidative stress, maintained glutathione redox state, and enhanced phosphocreatine phosphorylation potential. Aconitase and glucose 6-phosphate dehydrogenase activities fell during arrest; creatine kinase and phosphofructokinase were inactivated upon reperfusion. Pyruvate protected creatine kinase and reactivated aconitase, which are at least partially mitochondrial enzymes, but did not modify the cytosolic enzymes glucose 6-phosphofructokinase. We conclude that 1) pyruvate-fortified cardioplegia and administration of pyruvate during early perfusion increase the antioxidant state of the heart and reduce oxidative stress occurring as a result of cardioplegic arrest and reperfusion; 2) pyruvate bolsters the myocardial energy state during early reperfusion when administered during cardioplegic arrest or during reperfusion; 3) Cardioplegic arrest and reperfusion inactivated several key metabolic enzymes. Pyruvate administration during reperfusion, the period of most intense oxidative stress, increases the activity of two mitochondrial enzymes during early reperfusion when compared to control. These investigations provide likely mechanisms for the ability of pyruvate-fortified cardioplegia to reduce myocardial injury and improve post-surgical cardiac performance in patients undergoing CPB. More research must be done to solidify pyruvate’s role as a cardioprotective intervention during CPB.

Comments

Knott, E. Marty, The Effects of Pyruvate on Oxidative Stress and Myocardial Energetics During Cardioplegic Arrest and Reperfusion Doctor of Philosophy (Integrative Physiology), October 2005, 118 pp, 1 tables, 17 figures, references, 130 titles W 4 K72E 2005

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