Stephen C. Kolwicz

Cardiac Metabolism and its Interactions With Contraction, Growth, and Survival of Cardiomyocytes

The network for cardiac fuel metabolism contains intricate sets of interacting pathways that result in both ATP-producing and non-ATP-producing end points for each class of energy substrates. The most salient feature of the network is the metabolic flexibility demonstrated in response to various stimuli, including developmental changes and nutritional status. The heart is also capable of remodeling the metabolic pathways in chronic pathophysiological conditions, which results in modulations of myocardial energetics and contractile function. In a quest to understand the complexity of the cardiac metabolic network, pharmacological and genetic tools have been engaged to manipulate cardiac metabolism in a variety of research models. In concert, a host of therapeutic interventions have been tested clinically to target substrate preference, insulin sensitivity, and mitochondrial function. In addition, the contribution of cellular metabolism to growth, survival, and other signaling pathways through the production of metabolic intermediates has been increasingly noted. In this review, we provide an overview of the cardiac metabolic network and highlight alterations observed in cardiac pathologies as well as strategies used as metabolic therapies in heart failure. Lastly, the ability of metabolic derivatives to intersect growth and survival are also discussed.

Mitochondrial Complex I Deficiency Increases Protein Acetylation and Accelerates Heart Failure

Mitochondrial respiratory dysfunction is linked to the pathogenesis of multiple diseases, including heart failure, but the specific mechanisms for this link remain largely elusive. We modeled the impairment of mitochondrial respiration by the inactivation of the Ndufs4 gene, a protein critical for complex I assembly, in the mouse heart (cKO). Although complex I-supported respiration decreased by >40%, the cKO mice maintained normal cardiac function in vivo and high-energy phosphate content in isolated perfused hearts. However, the cKO mice developed accelerated heart failure after pressure overload or repeated pregnancy. Decreased NAD(+)/NADH ratio by complex I deficiency inhibited Sirt3 activity, leading to an increase in protein acetylation and sensitization of the permeability transition in mitochondria (mPTP). NAD(+) precursor supplementation to cKO mice partially normalized the NAD(+)/NADH ratio, protein acetylation, and mPTP sensitivity. These findings describe a mechanism connecting mitochondrial dysfunction to the susceptibility to diseases and propose a potential therapeutic target.

Glucose metabolism and cardiac hypertrophy.

The most notable change in the metabolic profile of hypertrophied hearts is an increased reliance on glucose with an overall reduced oxidative metabolism, i.e. a reappearance of the foetal metabolic pattern. In animal models, this change is attributed to the down-regulation of the transcriptional cascades promoting gene expression for fatty acid oxidation and mitochondrial oxidative phosphorylation in adult hearts. Impaired myocardial energetics in cardiac hypertrophy also triggers AMP-activated protein kinase (AMPK), leading to increased glucose uptake and glycolysis. Aside from increased reliance on glucose as an energy source, changes in other glucose metabolism pathways, e.g. the pentose phosphate pathway, the glucosamine biosynthesis pathway, and anaplerosis, are also noted in the hypertrophied hearts. Studies using transgenic mouse models and pharmacological compounds to mimic or counter the switch of substrate preference in cardiac hypertrophy have demonstrated that increased glucose metabolism in adult heart is not harmful and can be beneficial when it provides sufficient fuel for oxidative metabolism. However, improvement in the oxidative capacity and efficiency rather than the selection of the substrate is likely the ultimate goal for metabolic therapies.

Cardiac-Specific Deletion of Acetyl CoA Carboxylase 2 Prevents Metabolic Remodeling During Pressure-Overload Hypertrophy

Rationale: Decreased fatty acid oxidation (FAO) with increased reliance on glucose are hallmarks of metabolic remodeling that occurs in pathological cardiac hypertrophy and is associated with decreased myocardial energetics and impaired cardiac function. To date, it has not been tested whether prevention of the metabolic switch that occurs during the development of cardiac hypertrophy has unequivocal benefits on cardiac function and energetics. Objective: Because malonyl CoA production via acetyl CoA carboxylase 2 (ACC2) inhibits the entry of long chain fatty acids into the mitochondria, we hypothesized that mice with a cardiac-specific deletion of ACC2 (ACC2H−/−) would maintain cardiac FAO and improve function and energetics during the development of pressure-overload hypertrophy. Methods and Results: ACC2 deletion led to a significant reduction in cardiac malonyl CoA levels. In isolated perfused heart experiments, left ventricular function and oxygen consumption were similar in ACC2H−/− mice despite an ≈60% increase in FAO compared with controls (CON). After 8 weeks of pressure overload via transverse aortic constriction (TAC), ACC2H−/− mice exhibited a substrate utilization profile similar to sham animals, whereas CON-TAC hearts had decreased FAO with increased glycolysis and anaplerosis. Myocardial energetics, assessed by 31P nuclear magnetic resonance spectroscopy, and cardiac function were maintained in ACC2H−/− after 8 weeks of TAC. Furthermore, ACC2H−/−-TAC demonstrated an attenuation of cardiac hypertrophy with a significant reduction in fibrosis relative to CON-TAC. Conclusions: These data suggest that reversion to the fetal metabolic profile in chronic pathological hypertrophy is associated with impaired myocardial function and energetics and maintenance of the inherent cardiac metabolic profile and mitochondrial oxidative capacity is a viable therapeutic strategy.

Metabolic Therapy at the Crossroad: How to Optimize Myocardial Substrate Utilization?

There has been growing interest in targeting myocardial substrate metabolism for the therapy of cardiovascular and metabolic diseases. This is largely based on the observation that cardiac metabolism undergoes significant changes during both physiologic and pathologic stresses. In search for an effective therapeutic strategy, recent studies have focused on the functional significance of the substrate switch in the heart during stress conditions, such as cardiac hypertrophy and failure, using both pharmacologic and genetic approaches. The results of these studies indicate that both the capacity and the flexibility of the cardiac metabolic network are essential for normal function; thus, their maintenance should be the primary goal for future metabolic therapy.

Mutation in the γ2-Subunit of AMP-activated protein kinase stimulates cardiomyocyte proliferation and hypertrophy independent of glycogen storage

Rationale: AMP-activated protein kinase is a master regulator of cell metabolism and an attractive drug target for cancer and metabolic and cardiovascular diseases. Point mutations in the regulatory &ggr;2-subunit of AMP-activated protein kinase (encoded by Prkag2 gene) caused a unique form of human cardiomyopathy characterized by cardiac hypertrophy, ventricular preexcitation, and glycogen storage. Understanding the disease mechanisms of Prkag2 cardiomyopathy is not only beneficial for the patients but also critical to the use of AMP-activated protein kinase as a drug target. Objective: We sought to identify the pro–growth-signaling pathway(s) triggered by Prkag2 mutation and to distinguish it from the secondary response to glycogen storage. Methods and Results: In a mouse model of N488I mutation of the Prkag2 gene (R2M), we rescued the glycogen storage phenotype by genetic inhibition of glucose-6-phosphate–stimulated glycogen synthase activity. Ablation of glycogen storage eliminated the ventricular preexcitation but did not affect the excessive cardiac growth in R2M mice. The progrowth effect in R2M hearts was mediated via increased insulin sensitivity and hyperactivity of Akt, resulting in activation of mammalian target of rapamycin and inactivation of forkhead box O transcription factor–signaling pathways. Consequently, cardiac myocyte proliferation during the postnatal period was enhanced in R2M hearts followed by hypertrophic growth in adult hearts. Inhibition of mammalian target of rapamycin activity by rapamycin or restoration of forkhead box O transcription factor activity by overexpressing forkhead box O transcription factor 1 rescued the abnormal cardiac growth. Conclusions: Our study reveals a novel mechanism for Prkag2 cardiomyopathy, independent of glycogen storage. The role of &ggr;2-AMP-activated protein kinase in cell growth also has broad implications in cardiac development, growth, and regeneration.

Substrain specific response to cardiac pressure overload in C57BL/6 mice

The C57BL/6 mouse strain is one of the most commonly used in experimental research. It is known to differ from other strains in baseline cardiovascular phenotypes as well as in response to pressure overload induced by aortic constriction. Since the generation of the C57BL/6 mouse line over a century ago, multiple substrains have been generated from the original. To identify potential substrain specific differences in response to pressure overload, we evaluated the effects of transverse aortic constriction (TAC) on survival, cardiac function, and expression of hypertrophic markers in three commonly used C57BL/6 substrains: C57BL/6J (JL), C57BL/6NCrl (CL), and C57BL/6NTac (TF). Survival and cardiac function were significantly lower in the CL and TF substrains compared with JL mice after TAC. Furthermore, the heart weight and lung weight as well as the expression of the hypertrophic marker Bnp were significantly greater in the CL mice compared with the JL. Histological assessment revealed marked left ventricular dilatation of CL and TF hearts while JL hearts showed increased wall thickness without dilatation. Our data demonstrate that cardiac response to pressure overload is distinct among the three commonly used C57BL/6 substrains of mice, which raises a cautionary note in study design and data interpretation.

Ketones Step to the Plate: A Game Changer for Metabolic Remodeling in Heart Failure?

It is increasingly recognized that metabolic remodeling is integral to heart failure (HF) development and progression.1,2 In particular, impairments in the ability of cardiac mitochondria to oxidize fatty acids have been noted, along with an increase in glycolysis that is uncoupled from glucose oxidation.3,4 This overall reduction in the myocardial oxidative capacity is purported to be the root cause of energy deficiency in the failing heart. Although past research has focused primarily on myocardial use of glucose and fatty acids, the heart is an omnivore and capable of oxidizing other substrates such as lactate, ketone bodies, and amino acids. The current understanding of the contribution of lactate, ketone bodies, and amino acids to cardiac metabolism is limited, particularly in the setting of HF. In this issue of Circulation , 2 independent studies provide new insights into the reliance of the failing heart on ketone bodies for energy supply. Proteomics analysis in mouse models of HF by Aubert et al5 and metabolomics analysis of end-stage human failing hearts by Bedi et al6 demonstrate strong and concordant evidence of increased ketone oxidation in the failing heart. Articles, see p 698 and 706 Ketone bodies, that is, acetoacetate, β-hydroxybutyrate (βOHB), and acetone (in low abundance), are produced in the liver and used in peripheral tissues as an energy source when glucose is not readily available because of either a limited exogenous supply or impaired insulin signaling or when fatty acids are in surplus, which occurs in marked activation of lipolysis. On entering the cell, they rapidly form acetyl CoA via a series of reactions catalyzed by βOHB dehydrogenase (BDH1), succinyl-CoA:3-oxoacid-CoA transferase (SCOT), and mitochondrial acetyl-CoA acetyltransferase 1 (also known as thiolase), as shown in the Figure. All enzymes are present in the heart, and …

Transgenic overexpression of ribonucleotide reductase improves cardiac performance

We previously demonstrated that cardiac myosin can use 2-deoxy-ATP (dATP) as an energy substrate, that it enhances contraction and relaxation with minimal effect on calcium-handling properties in vitro, and that contractile enhancement occurs with only minor elevation of cellular [dATP]. Here, we report the effect of chronically enhanced dATP concentration on cardiac function using a transgenic mouse that overexpresses the enzyme ribonucleotide reductase (TgRR), which catalyzes the rate-limiting step in de novo deoxyribonucleotide biosynthesis. Hearts from TgRR mice had elevated left ventricular systolic function compared with wild-type (WT) mice, both in vivo and in vitro, without signs of hypertrophy or altered diastolic function. Isolated cardiomyocytes from TgRR mice had enhanced contraction and relaxation, with no change in Ca2+ transients, suggesting targeted improvement of myofilament function. TgRR hearts had normal ATP and only slightly decreased phosphocreatine levels by 31P NMR spectroscopy, and they maintained rate responsiveness to dobutamine challenge. These data demonstrate long-term (at least 5-mo) elevation of cardiac [dATP] results in sustained elevation of basal left ventricular performance, with maintained β-adrenergic responsiveness and energetic reserves. Combined with results from previous studies, we conclude that this occurs primarily via enhanced myofilament activation and contraction, with similar or faster ability to relax. The data are sufficiently compelling to consider elevated cardiac [dATP] as a therapeutic option to treat systolic dysfunction.

Mutation in the γ2-subunit of AMPK Stimulates Cardiomyocyte Proliferation and Hypertrophy Independent of Glycogen Storage

Rationale: AMP-activated protein kinase is a master regulator of cell metabolism and an attractive drug target for cancer and metabolic and cardiovascular diseases. Point mutations in the regulatory &ggr;2-subunit of AMP-activated protein kinase (encoded by Prkag2 gene) caused a unique form of human cardiomyopathy characterized by cardiac hypertrophy, ventricular preexcitation, and glycogen storage. Understanding the disease mechanisms of Prkag2 cardiomyopathy is not only beneficial for the patients but also critical to the use of AMP-activated protein kinase as a drug target. Objective: We sought to identify the pro–growth-signaling pathway(s) triggered by Prkag2 mutation and to distinguish it from the secondary response to glycogen storage. Methods and Results: In a mouse model of N488I mutation of the Prkag2 gene (R2M), we rescued the glycogen storage phenotype by genetic inhibition of glucose-6-phosphate–stimulated glycogen synthase activity. Ablation of glycogen storage eliminated the ventricular preexcitation but did not affect the excessive cardiac growth in R2M mice. The progrowth effect in R2M hearts was mediated via increased insulin sensitivity and hyperactivity of Akt, resulting in activation of mammalian target of rapamycin and inactivation of forkhead box O transcription factor–signaling pathways. Consequently, cardiac myocyte proliferation during the postnatal period was enhanced in R2M hearts followed by hypertrophic growth in adult hearts. Inhibition of mammalian target of rapamycin activity by rapamycin or restoration of forkhead box O transcription factor activity by overexpressing forkhead box O transcription factor 1 rescued the abnormal cardiac growth. Conclusions: Our study reveals a novel mechanism for Prkag2 cardiomyopathy, independent of glycogen storage. The role of &ggr;2-AMP-activated protein kinase in cell growth also has broad implications in cardiac development, growth, and regeneration.