More Energy to Relax

Corresponding Author

Relaxation of sarcomeres requires adenosine triphosphate (ATP), which binds to the myosin head to trigger its detachment from actin following the power stroke that generates force. This releases the thick filament from the thin filament and permits myosin to reattach to the actin molecule, followed by ATP hydrolysis that drives the next power stroke. This cross-bridge cycling drives mechanical shortening of the sarcomere, to generate a contraction. ATP is also required to actively pump calcium ions (that initiate contraction by unmasking myosin binding sites on actin) out of the cytosol into the subcellular organelles or into the extracellular space, and regulates allosteric effects on cardiac contractile proteins. Relaxation is therefore an “active,” energy-requiring process and inherently slower than contraction, which is “passive” and rapidly triggered by membrane depolarization that drives a sudden influx of calcium ions into the cytosol to induce actin–myosin cross-bridge formation. Impaired myocardial relaxation underlies diastolic dysfunction, a sine qua non of heart failure, which is a fatal condition and increasing in prevalence worldwide. In this issue of JACC: Basic to Translational Science, Soranno et al. (1) implicate decline in myocardial ATP levels as the underlying mechanism for development of cardiac diastolic dysfunction in the long term following acute kidney injury (AKI) in mice. Treatment with ITF2357, a nonspecific inhibitor of histone deacetylases (HDACs) currently in human clinical trials for noncardiac indications, prevented both the decline in energy stores and development of diastolic dysfunction. These studies are further supported by metabolomic analyses of cardiac tissues and plasma that indicate accumulation of ATP precursors pointing to impaired flux in ATP generation pathways. Indeed, HDAC inhibitor (HDACi) treatment reverses these abnormalities, thus providing “more energy (ATP) for relaxation” as a mechanism for preventing diastolic dysfunction.

As discussed by the investigators, a growing body of epidemiological data indicate a strong association between heart failure and renal dysfunction, often referred to as the cardiorenal axis; and each condition has been documented to increase the future incidence of the other, with worsening morbidity and mortality. To examine the mechanisms for development of heart failure after AKI, the investigators modeled AKI in male mice with careful serial assessment of multiple outcome measures pertinent to both cardiac and renal function, up to a year following the renal injury. The resultant cardiac phenotype of diastolic dysfunction and preserved ejection fraction recapitulates pathophysiological features observed in humans with heart failure with preserved ejection fraction (HFpEF), which portends an equally poor prognosis as heart failure with reduced ejection fraction, with limited proven therapeutic options. Indeed, patients with HFpEF demonstrate reduced myocardial energy stores by ³¹P nuclear magnetic resonance spectroscopy techniques, mimicking the current observations (2). A novel finding of the current study is the observation of myocardial diastolic dysfunction before the development of hypertension following renal injury. Interestingly, HDACi treatment (with ITF2357) initiated 3 days after AKI prevented myocardial dysfunction, as well as the development of hypertension, suggesting common underlying pathogenic mechanisms that independently provoke both conditions. Inflammation is one such mechanism, and the current study revealed early induction in cytokine levels and persistent TNF elevation, pointing to the need to rigorously examine cellular inflammatory pathways as a target to prevent cardiovascular modeling post-AKI. It is also intriguing to note that HDACi did not improve renal outcomes in the current study and was associated with worsening renal fibrosis post-AKI, an aspect that will require further follow-up to define the safety profile of this approach with regard to renal (and cardiac) outcomes.

Multiple pathological mechanisms are implicated in inducing impaired relaxation in both animal models and tissues from failing human hearts. In cardiac myocytes, increased myofibril calcium sensitivity and alteration in titin isoforms—or its post-translational modifications—and other myofilament proteins, such as myosin heavy chain, tropomyosin, myosin binding protein C or troponin I, result in altered relaxation. Recent studies have demonstrated impaired cardiac myocyte sarcomere relaxation in 2 rodent models of HFpEF, and treatment with HDACi ITF2357 was demonstrated to improve myocyte relaxation ex vivo (3). Interestingly, in this study, HDACi improved diastolic function in aging female mice without affecting hypertension (3), whereby sex-based differences may explain the preventive effects on HDACi on hypertension post-renal injury observed in experiments by Soranno et al. (1), which focused on male mice (1). An analysis of the myocardial proteome in HDACi-treated aging female mice revealed altered acetylation; and experimental manipulation of protein acetylation by exposure of isolated myofibrils to purified enzymes altered relaxation parameters (3). The current study suggests that another mechanism may be operative in parallel whereby HDACi preserves myocardial ATP stores. ATP turnover rates in cardiac myocytes per beat are estimated at ∼10% of the entire ATP pool and are buffered by high-energy phosphate store depots, which exist as phosphocreatine in cardiac myocytes. A decline in steady-state ATP levels likely reflects a global decline in these high-energy phosphate stores, which requires further investigation in the model presented by Soranno et al. (1). Also, given that ∼90% of cardiac myocyte ATP generation is estimated to be from mitochondrial oxidative phosphorylation, the data presented in the current study point to long-term adverse effects of acute kidney injury on cardiac mitochondrial function and to a preventive role of HDACi therein. Indeed, this group had previously reported observations of increased oxidative stress that accompanied reduced cardiac ATP levels in the subacute period (7 days) after AKI, alluding to mitochondrial dysfunction as a potential source for increased generation of reactive oxygen species.

Unraveling the signaling pathways that transduce the cardiac effects of ITF2357, a pan-HDACi that inhibits both Class I and II HDACs, will be complicated by the divergent effects of these classes of HDACs on cardiac hypertrophy signaling and function. Moreover, HDACs demonstrate pleiotropic mechanisms of action that involve derepression of gene expression by an increased abundance of acetylated histones, and downstream effects on signaling kinases such as mTOR (mammalian target of rapamycin), a serine-threonine kinase that functions as a master regular of metabolism, growth, and autophagy signaling. Prior studies also implicate a role for lysine acetylation of transcription factor EB (TFEB, a master regulator of the autophagy-lysosome pathway) in regulating its transcriptional activity (4). In this study, pan-HDACi with suberoylanilide hydroxamic acid (SAHA) induced TFEB activation, which has been previously demonstrated to facilitate mitochondrial quality control in cardiac myocytes by simultaneously facilitating removal of damaged mitochondria and stimulating biogenesis of normal mitochondria. Interestingly, ITF2357 does not inhibit Class III HDACs, which require nicotinamide adenine dinucleotide (NAD+), levels. of which are observed to decline in renal tissues and in the circulation after AKI as noted by the investigators. Although NAD+ levels did not change in the metabolomic analyses presented, the nicotinamide metabolism pathway was significantly altered, suggesting the need for careful examination of NAD+ signaling in the heart in this model.

Modeling HFpEF in animals remains a challenge, given the complexity of various risk factors such as age, diabetes, hypertension, obesity, renal dysfunction, and inflammation that are implicated in the pathophysiology; and the model reported by Soranno et al. (1) does not faithfully recapitulate various aspects of HFpEF in humans. Indeed, AKI resulted in reduced left ventricular mass at 1 year despite development of hypertension, which is expected to increase afterload and trigger pathological myocardial hypertrophy. Moreover, myocardial fibrosis, a common pathological change in HFpEF patients, was not observed. In this model, the observed enrichment of amino acid metabolism pathways in both the heart and serum metabolite profiles is intriguing in light of the recent observations that demonstrate increased amino acid catabolism in patients with reduced ejection fraction and cardiomyopathy, as compared with individuals without heart failure (5). Indeed, although AKI is known to induce systemic protein catabolism; the data presented herein suggest that AKI induces cardiac protein catabolism in the long term in the setting of energetic insufficiency, analogous to the observed net release of amino acids from the heart via protein catabolism in the fasted state (5). The observed decline in left ventricular mass with AKI in the current study, along with a decline in body weight and a suggestion of lower skeletal muscle mass (potentially underpowered to reveal a statistical significant difference), indicates generalized augmentation of catabolic pathways in this model. A further decline in body weight and skeletal muscle (gastrocnemius) weight with HDACi treatment was noted by the investigators in this model, raising a potential red flag for activation of atrophic signaling (and potentially excess autophagy) that would require close monitoring with clinical translational of this approach. Another major limitation of the current study is the lack of data on lipid fluxes in the myocardium, examining which should be a priority, given their overwhelming contribution as substrates to cardiac energy generation in the normal heart and the observed decline with development of cardiomyopathy (5).

The major contribution of the current study is the recognition of long-term effects of renal injury on the myocardium that may portend development of HFpEF. It provides early insights into how AKI may regulate cardiac energy generation and the role of HDACs in remodeling the cardiac metabolome. Like any robust investigation, the data raise more questions, addressing which may realize the potential to develop targeted HDACi as a strategy to prevent HFpEF.

Funding Support and Author Disclosures

Dr. Diwan is supported by Department of Veterans Affairs grant I01BX004235 and National Institutes of Health grants HL143431 and HL107594. Dr. Diwan reports that he provides consulting services to ERT Systems for interpretation of echocardiograms in clinical trials. This did not affect the work reported in this paper.