Enhanced pyruvate dehydrogenase activity improves cardiac outcomes in a murine model of cardiac arrest

Abstract

Rationale: Post-ischemic changes in cellular metabolism alter myocardial and neurological function. Pyruvate dehydrogenase (PDH), the limiting step in mitochondrial glucose oxidation, is inhibited by increased expression of PDH kinase (PDK) during ischemia/reperfusion injury. This results in decreased utilization of glucose to generate cellular ATP. Post-cardiac arrest (CA) hypothermia improves outcomes and alters metabolism, but its influence on PDH and PDK activity following CA are unknown. We hypothesized that therapeutic hypothermia (TH) following CA is associated with the inhibition of PDK activity and increased PDH activity. We further hypothesized that an inhibitor of PDK activity, dichloroacetate (DCA), would improve PDH activity and post-CA outcomes.

Methods and results: Anesthetized and ventilated adult female C57BL/6 wild-type mice underwent a 12-minute KCl-induced CA followed by cardiopulmonary resuscitation. Compared to normothermic (37°C) CA controls, administering TH (30°C) improved overall survival (72-hour survival rate: 62.5% vs. 28.6%, P<0.001), post-resuscitation myocardial function (ejection fraction: 50.9±3.1% vs. 27.2±2.0%, P<0.001; aorta systolic pressure: 132.7±7.3 vs. 72.3±3.0 mmHg, P<0.001), and neurological scores at 72-hour post CA (9.5±1.3 vs. 5.4±1.3, P<0.05). In both heart and brain, CA increased lactate concentrations (1.9-fold and 3.1-fold increase, respectively, P<0.01), decreased PDH enzyme activity (24% and 50% reduction, respectively, P<0.01), and increased PDK protein expressions (1.2-fold and 1.9-fold, respectively, P<0.01). In contrast, post-CA treatment with TH normalized lactate concentrations (P<0.01 and P<0.05) and PDK expressions (P<0.001 and P<0.05), while increasing PDH activity (P<0.01 and P<0.01) in both the heart and brain. Additionally, treatment with DCA (0.2 mg/g body weight) 30 min prior to CA improved both myocardial hemodynamics 2 hours post-CA (aortic systolic pressure: 123±3 vs. 96±4 mmHg, P<0.001) and 72-hour survival rates (50% vs. 19%, P<0.05) in normothermic animals.

Conclusions: Enhanced PDH activity in the setting of TH or DCA administration is associated with improved post-CA resuscitation outcomes. PDH is a promising therapeutic target for improving post-CA outcomes.

Fig 1. TH improves post-CA myocardial function, neurological outcomes and survival.

Following twelve-minute asystolic CA, CPR was performed. Echocardiography showed fractional shortening (A, FS%) was reduced in the CA group and was improved by therapeutic hypothermia (TH). Aortic systolic pressure (B, ASP) was deceased following CA, but was elevated with TH. Neurological scores (C), and survival (D) were decreased following CA, but were improved with TH.

Fig 2. Post-CA inhibition of pyruvate dehydrogenase (PDH) in the brain is improved by TH.

A. Brain lactate concentrations increased following CA, but not in mice treated with TH. B. PDH enzyme activity decreased following CA, whereas TH increased PDH enzyme activity as demonstrated by PDH dipstick activity assay. C. Phosphorylated PDH (P-PDH) increased following CA, but not in the TH group. D. Pyruvate dehydrogenase kinase (PDK) in the brain, PDK2, increased following CA, but not in mice treated with TH.

Fig 3. Lactate increases despite elevations in Glut1 expression in the post-CA myocardium.

A. Lactate concentrations in the heart were elevated following CA, but normalized with TH treatment. B&C. Western blot and immunostaining showed that Glut1 was increased following CA, but not affected by TH.

Fig 4. Post-CA inhibition of PDH in heart is improved by TH.

A. PDH enzyme activity in the heart was decreased following CA, but was improved with TH. B. P-PDH in the heart was elevated following CA but was normalized with TH. C&D. PDK4, the primary PDK isoform in the heart, was elevated following CA but not in TH treated animals.

Fig 5. Dichloroacetate (DCA) treatment improves post-CA myocardial function and survival.

A. PDH enzyme activity in the heart post-CA was improved by DCA administration. B&C. P-PDH and PDK4 in the heart post CA were both reduced by DCA treatment. D&E. FS% and ASP post-CA were both improved by DCA administration. F&G. ROSC rate and survival rate post-CA were both improved by DCA.

Fig 6. Metabolic alterations in PDH activity signaling in the heart and brain post-CA.

There are metabolic imbalances in both brain and heart post-CA. GO is depressed evidence by decreased pyruvate, whereas glycosis is increased evidenced by increased lactate concentration. These are direct effects of PDK activation and PDH inhibition, which result in further cardiac and neurological dysfunction. Both TH and DCA, a PDK inhibitor, improve post-CA resuscitation outcomes, including cardiac output, neurological scores and survival by decreasing lactate concentrations, reducing PDK expressions, and elevating PDH activity.