Pathophysiology of heart failure and an overview of therapies

Definition of heart failure and general features of the condition

Heart failure (HF) is formally defined as a complex clinical syndrome resulting from any structural and/or functional impairment of ventricular filling or ejection. This somewhat broad definition is necessary as HF represents the final common pathway of disparate cardiovascular pathologies that lead to an insufficient supply of oxygen to meet the body’s metabolic demands. These pathologies differ in the time of onset and progression, the systems responsible for driving failure, the types of injury/insult initiating dysfunction, and the resulting functional/anatomical cardiovascular alterations that define HF. This diverse set of precipitating etiologies has made prognostic measures for HF difficult and requires multiple criteria for making a diagnosis, including historical data on individual patients.

To appreciate the clinical approach to HF, the historical context in which this area of study has grown provides some insight into how HF has been managed clinically. Before an intimate understanding of the cardiovascular system, HF was considered an incurable terminal condition. The approach to clinical management has evolved out of various theories regarding the primary underlying causes, which has shifted from an initial focus on the heart purely as a failing pump, to a focus on the role of abnormal hemodynamics, to a more integrated approach dealing with neurohormonal response. Currently, a greater emphasis on targeting structural changes occurring in the failing heart has grown and efforts have expanded to therapeutically target the underlying cellular and molecular processes involved in the development of HF; these include energy metabolism, inflammation/immune response, protein turnover, and the regulation of gene expression, among others.

Prevalence and socioeconomic burden of heart failure

HF is a prominent health-care issue in industrialized countries, despite a high quality of life and improved health care. Increasing life span coupled with enhanced survival rates of various types of cardiovascular injury has resulted in substantial jumps in health-care costs related to treating patients with HF. The global impact of HF on total health-care costs (indirect and direct) is estimated over $108 billion annually, with the United States accounting for nearly a third of this spending . HF’s financial burden is the greatest of all diagnostic groups and will continue to lead as incidence rates are projected to increase . Globally, HF affects more than 20 million people. In the United States alone, nearly 6 million people have HF (roughly 2% of the general population) .

The predominant subpopulation affected by HF is the elderly where the prevalence increases to 11% in an ever-increasing population of individuals over 75 years of age . Historically, pediatric HF has a relatively low incidence compared to adults, with underlying causes linked to infectious disease or heritable cardiomyopathies; however, the growing population of children with obesity and diabetes mellitus has led to greater incidences of hypertension and subsequently HF in a younger patient population. Women have been historically underrepresented in clinical studies of HF, but some gender differences in HF have been defined. Despite a lower overall incidence of cardiovascular disease (CVD), women have a greater risk of HF development , and variation in the levels of diagnostic biomarkers (based on studies in male patients) suggests future work to better understand what gender means for HF is needed . Ethnicity influences HF prevalence as well; African Americans are more susceptible to HF, likely due to greater occurrence of hypertension and differences in socioeconomic status .

Long-term prognosis: morbidity and mortality

Despite advances in therapeutic strategies that have led to a significant decline in mortality from hypertension, acute myocardial infarction (MI), and valvular disease, there has been little change in the prognosis of HF precipitated by these conditions. Patient care is palliative in nature and progression to end-stage failure is often inevitable without surgical intervention. The average age for primary diagnosis of HF is 70 years with a survival rate of roughly 90%; however, this drops to 58% by 5 years post-HF onset .

Risk factors and underlying causes of heart failure

HF results from a diverse set of etiologies. Ultimately, HF reflects a severe decline in cardiomyocyte function or cell death, which can result from direct injury to the myocardium or stem from ongoing stress that overwhelms the heart’s ability to compensate. HF can be sudden in onset or progressive in nature. Multiple risk factors for HF are known, including ischemic heart disease and hypertension, among the most prominent. Despite being treatable, hypertension is often uncontrolled in patients. Hypertension afflicts nearly a quarter of the American population, carrying around a 40% lifetime risk of developing congestive HF . A history of MI increases the chances of developing HF. Established atherosclerotic disease in the coronary or any peripheral blood vessels also enhances the likelihood of HF. Obesity, insulin resistance, and dyslipidemia are considered part of the metabolic syndrome, which increases the risk of HF . Diabetes mellitus overlaps with these pathologies, with structural changes resulting in declining diastolic function, followed by an impact on systolic function, characterizing diabetic cardiomyopathy.

Genetic (familial) cardiomyopathies cover a range of gene mutations. Still, many clinically involve contractile apparatus protein mutations that result in structural abnormalities, culminating in HF . These cardiomyopathies are usually of early-onset and commonly account for the HF seen in children, as they manifest much earlier in life to affect both the structure and function of the myocardium negatively. However, the incidence of heredity in dilated and hypertrophic cardiomyopathy is also significant in the adult population and may account for nearly 30% of diagnosed cardiomyopathies, though exact numbers are difficult to define, as genetic screening is not routine .

Guidelines for the genetic evaluation of cardiomyopathy were established in 2009 by the Heart Failure Society of America (HFSA) and updated in collaboration with the American College of Medical Genetics and Genomics (ACMG) in 2018 . In recent years, genetic evaluation has become more feasible with high-throughput sequencing in clinical medicine and appreciated as specific interventions have emerged for a few genetic cardiac diseases. In contrast to nongenetic testing, a systemic approach to genetic diagnosis of HF is critical, including family history, genetic counseling, and specific guidance for drug and/or device therapies .

Genetic causes of cardiomyopathies are defined by clinical phenotypes, including hypertrophic cardiomyopathy (HCM), familial dilated cardiomyopathy (DCM), and arrhythmogenic right ventricular cardiomyopathy (ARVC). HCM is caused by mutations in the sarcomeric (and related) proteins. While DCM can have both nongenetic (e.g., coronary artery disease, primary valve/congenital causes, or exposure to drugs such as chemotherapies), and genetic causes, those with familial characteristics have many genes with diverse variants that are more extensive than other cardiomyopathies . ARVC, much less common than HCM and DCM, results from mutations in genes associated with desmosomal components. Restrictive cardiomyopathy shares genetic causes with HCM, although much rarer than the other types of cardiomyopathies. Guidelines are available for selecting whom to test and how to test, by a team of collaborators, commonly including cardiologists (adult or pediatric) with interest/training in cardiovascular genetics and genetic professionals, such as board-eligible/-certified counselors and/or clinical geneticists, to provide both patients and family of those with cardiomyopathy a comprehensive service .

Exposure of the heart to numerous compounds that are cardiotoxic also leads to HF. These include both medicinal drugs and drugs of abuse. Anthracyclines (commonly Adriamycin) are part of many chemotherapeutic regimens. Though efficacious in tumor regression, anthracyclines are potent cardiotoxic agents with acute effects on contractility that can be dose-limiting and lead to long-term defects linked to oxidative damage and permanent mitochondrial function defects . Alcohol (ethanol) consumption has a varied impact on the myocardium, with moderate habitual drinking linked to cardiovascular health benefits. However, heavy or “binge” drinking induces atrial fibrillation and can precipitate sudden cardiac death . Chronic excessive alcohol also increases HF risk in a dose-dependent manner by causing hypertension and atrial fibrillation. Drugs of abuse such as cocaine also present both acute and chronic HF. Cocaine-induced MI can result from increased heart rate, vasoconstriction leading to platelet activation, and arrhythmia generation, resulting in acute HF . Prolonged cocaine usage increases HF risk .

Diagnosing heart failure

Historically, HF is a condition characterized by systolic dysfunction evidenced most directly by a decline in ejection fraction (EF). However, there has been an increasing awareness of patients who suffer from HF with preserved ejection fraction (HFpEF) that historically was thought to be only diastolic dysfunction (DD). However, it is now recognized to be more complex and associated with a greater number of comorbidities . The criteria HF is diagnostically challenging as indicators and symptoms vary and rely heavily on observational findings. The multifactorial causes of HF make defining single, stand-alone endpoints of HF impossible. This is of special concern when dealing with HFpEF where functional findings may be absent or very mild and only evident during acute decompensation. HF diagnosis still commonly depends on comprehensive patient history and physical examination, chest radiography, electrocardiography, echocardiography, and laboratory tests.

Symptomatic presentation in patients

Suspected HF patients exhibit many characteristic signs and symptoms, though the combination of these findings and their intensity varies. For example, fatigue (generalized weakness) and exercise intolerance are common as the body is underserved in terms of its metabolic demand. Patients often present with dyspnea upon exertion or may experience shortness of breath even at rest in more severe cases. Examination of respiratory function may reveal rales and congestion manifested by wheezing and cough. Palpitations or tachycardia with weak pulse pressure is as likely as compensatory increases in HR counter a diminished stroke volume. With diminishing right ventricular filling, the onset of edema in the lower limbs and ascites often occurs along with weight gain. Venous jugular distension may also be present but is often difficult to assess.

Structural and functional

Chest X-rays are a primary means of noninvasive assessment of suspected HF, particularly in the acute setting. Imaging is useful for determining the presence of cardiomegaly and can show pulmonary venous congestion or interstitial edema to indicate HF due to the progressive increase of hydrostatic capillary pressure driving fluid accumulation . Two-dimensional echocardiography with Doppler is also a standard diagnostic tool for the assessment of structure and function and intracardiac pressures. Primary findings on echo are a lower cardiac output, evidenced by decreased EF (see Fig. 5.1 ), though in patients with HFpEF, including left ventricular (LV) filling pressures and ejection times should be assessed .

Criteria for classifying heart failure severity

Several schemes of classifying HF have been developed to standardize clinical diagnoses. The standard is the New York Heart Association (NYHA) system, which categorizes patients’ HF severity according to the physical limitations and physiological symptoms they experience as a result of HF. Despite this being an indirect system for classifying HF, it has been used as a standard since the 1920s and is still employed as the standard for clinical trials on HF. Table 5.1 outlines the categorical breakdown of both the long-standing NYHA and the more recently devised American Heart Association/American College of Cardiology Foundation (AHA/ACCF) system, which are ordered in terms of severity. The AHA scoring system uses “objective assessment” criteria based on measures of structural changes in the heart where stage A is used to document the presence of risk factors in the absence of disease, B implies modest changes in structure, C indicates structural changes with symptomatic HF of varying functional class, and D designates HF requiring surgical intervention and/or transplantation; this scoring complements the NYHA functional classifications based on subjective evaluation of patient symptoms. The combination of the NYHA and AHA classes is generally accepted as the standard for documenting HF severity and has become increasingly important with the recognition of previously underdiagnosed patients who present with HFpEF that may have mild changes but are functionally limited. Multiple scoring schemes have been created that provide a comprehensive diagnostic profile, which are varied, but incorporate additional clinical parameters as discussed above .

HF represents a significant health-care problem with an increasing population at risk, and end-stage failure is currently limited in therapeutic options other than surgical intervention. The expansion of treatment and management strategies for HF requires a better understanding of the events, both physiological and cellular, which precipitate and drive HF. This chapter aims to highlight contemporary topics in HF by exploring some of the underlying pathologies culminating in HF and discuss how these translate to potential therapeutic strategies representing the most promising direction in HF’s clinical management.

Structure of the failing heart

HF is a complex clinical syndrome that results in part from pathological alterations in the myocardium’s structure and function. As early as the 16th century, anatomists recognized characteristic structural abnormalities in patients’ hearts who died from HF . At autopsy, failing hearts were enlarged, with increased mass due to thickening of the ventricular walls and/or dilatation of the ventricular cavity. The former pattern came to be known as concentric hypertrophy and the latter as eccentric hypertrophy ( Fig. 5.2 ). Osler and others concluded that eccentric hypertrophy typically was associated with a decline in the heart’s contractile function and portended a worse prognosis. We now know that concentric hypertrophy develops as an initial adaptive (compensatory) response to chronic pressure loading, as demonstrated by the relationships between increased internal pressure and wall stress (tension) ( Fig. 5.3 ). Eccentric hypertrophy can arise from chronic volume loading, primary cardiomyopathy, or can evolve in a concentrically hypertrophied heart exposed to ongoing elevated wall tension. The complex molecular underpinnings of pathological hypertrophy have been elucidated in the century since Osler’s observation and have been invaluable in understanding HF.

Hemodynamic alterations in heart failure

Late nineteenth- and early 20th-century physiologists discovered that HF exhibited structural abnormalities that lead to alterations in basic hemodynamic parameters (e.g., preload, afterload, and contractility). Perhaps the most influential of these pioneers was Ernest Starling, who demonstrated that stroke volume increased proportionally with end-diastolic volume (preload) . This principle, now called the Frank-Starling Law, allows the heart to compensate for impaired contractility and forestall the development of HF’s clinical syndrome. This initially adaptive response is maintained, in part, by the retention and redistribution of fluid, resulting from neurohormonal responses to impaired cardiac performance. As HF progresses, fluid retention accounts for many of the signs and symptoms, which almost always is characterized by increased preload.

Afterload is defined as the ventricular wall tension during contraction and typically approximated blood pressure or systemic vascular resistance. The role of afterload in HF is multifactorial: chronically elevated afterload, as in hypertension or aortic stenosis, can cause hypertrophy and eventually lead to HF. Afterload can also increase as a compensatory response to maintain systemic perfusion in the setting of low cardiac output. Invasive hemodynamic evaluation, as pioneered by Andre Cournand and Dickinson Richards in the early 1940s , enabled the calculation of systemic vascular resistance critical to the current understanding that judicious pharmacologic reduction of afterload is beneficial in the management of HF.

Contractility is a measure of the force of myocardial contraction at any given preload and afterload and is determined at the cellular level by the degree of sarcomeric shortening. Historically, HF was understood primarily as a failure of contractility due to myocardial insult, though more recently, it has become clear that HF can develop in the setting of ostensibly normal contractility. In pathophysiological terms, HF in the setting of decreased contractility is called systolic HF, whereas HF develops in the setting of normal contractility and it is called diastolic HF. Clinically, these entities are known as heart failure with reduced ejection fraction (HFrEF) and HFpEF, where EF is a surrogate measure for contractility (stroke volume/end-diastolic volume). Surprisingly, roughly 50% of HF patients have HFpEF. The pathophysiology of HFpEF is poorly understood, but encompasses abnormalities in diastolic function, as well as pathological ventricular–vascular coupling due to arterial stiffness, inadequate HR response (chronotropic incompetence), and inadequate contractile reserve .

The degree of hemodynamic disturbance determines the clinical manifestations of HF. Most patients with symptomatic HF, either HFpEF or HFrEF, have markedly elevated preload due to fluid retention. HFrEF patients with severely impaired contractility may also develop hypoperfusion signs and symptoms. These symptoms are due to inadequate cardiac output, though these abnormalities develop only during the disease’s advanced stages.

Left and right heart failure

The pathophysiology and clinical manifestations of HF depend on which ventricle is affected (right or left), regardless of contractility (preserved or reduced). Left HF is associated with elevated LV filling pressure (preload) communicated passively back into the left atrium (LA) and subsequently through the pulmonary veins to the pulmonary vasculature. When the hydrostatic pressure in the pulmonary vessels exceeds the interstitial pressure, fluid extravasates to cause pulmonary edema. At markedly elevated pressures, fluid can move from pulmonary capillaries into the alveolar air space, leading to dyspnea and hypoxia. The development of left HF is caused by multiple etiologies: ischemic heart disease, dilated cardiomyopathy, abnormalities of the mitral or aortic valves, and systemic hypertension.

Right HF is associated with elevated central venous pressure, which serves as right ventricular preload. The resultant elevation in systemic venous pressure leads to liver edema, intestinal wall edema, and abdominal cavity ascites when markedly elevated. Collectively, these processes often cause patients with right HF to experience a sense of abdominal fullness, nausea, or early satiety. Fluid retention, elevated systemic venous pressure, and gravity conspire to cause lower extremity edema, the most typical right HF manifestation. The most common cause of right HF is left HF, wherein fluid retention leads to elevated preload and afterload in the right ventricle. However, right HF can also occur in the absence of left HF in conditions that cause elevated right ventricular afterload (pulmonary hypertension). Chronic hypoxic lung disease, collagen vascular disease, venous thromboembolic disease, and primary pulmonary arterial hypertension are causes of pulmonary hypertension that contribute to right-sided HF. While the vast majority of right-sided HF results indirectly from pressure increases, right HF can rarely be caused by a primary disease process in the right ventricular muscle itself, MI resulting from coronary artery blockage supplying blood to the right side of the heart, congenital heart disease, or arrhythmogenic right ventricular cardiomyopathy .

Myocardial remodeling in heart failure: cell death and regeneration

The anatomic basis for cardiac dysfunction and HF was originally described by Linzbach and Hort . As pathologists, they hypothesized that chronic cardiac stress leads to progressive cardiac hypertrophy (increase in cardiomyocyte size), which over time exceeds the capacity of the coronary vasculature to perfuse the increased cellular mass . They proposed that this led to the formation of ischemic foci to cause multifocal, subendocardial necrosis, slippage, and replacement fibrosis in addition to the stereotypical chamber dilation. The heart weight for this effect to manifest occurs at 500 g, corresponding to a critical LV weight of 200 g, according to Linzbach and Hort’s work . This myocardial remodeling, indicated by structural dilation, has more recently been recognized due to a significant number of complex processes . Linzbach found that the observed increase in cross-sectional diameter was less than predicted given the overall increases in heart mass, concluding that hyperplasia (increase in the number of cardiomyocytes) must occur in hearts with eccentric hypertrophy, exceeding the critical weight of 500 g. While the mitotic rate was not documented, histological assessment of myocardium indicated an increase in myocyte numbers, leading to the conclusion that cardiomyocyte hyperplasia was due to the longitudinal cleavage of cardiomyocytes . At the time, the concept of cardiomyocyte proliferation was speculative in mammals through a documented phenomenon in amphibians and teleost fish hearts.

Several reports suggested apoptosis as the mechanism responsible for the cell death occurring in failing hearts . However, the data suggested unrealistic rates of apoptosis that were later determined to be related to technical issues but gave rise to misleading publications . More recently, the application of more precise methods has found that between 0.001% and 0.002% of myocytes have apoptotic nuclei, in contrast to 0.08%–0.25% in various cardiac diseases . Additionally, types of cell death are reported with fewer apoptotic cells detected. For example, cardiomyocytes exhibit the following rates of occurrence in failing hearts: 0.002% apoptosis, 0.08% autophagy, and 0.06% oncosis . The dynamic nature of these processes precludes a precise quantification at a given point of the initiation of apoptosis (to completion), autophagy, and oncosis, which occur in parallel with dyssynchronous clearance of cells summarized in Table 5.2 . In the broader context, chronic pathological stimulus precipitate HF (detailed in sections below) results from an array of parallel processes that lead to the complex alterations in the shape of the failing LV (see Fig. 5.4 ) .

Importantly, parallel to cardiomyocyte hypertrophy, dysfunction, and cell death, there is emerging evidence that despite a long-standing view of cardiomyocytes as differentiated cells with a fixed number in the myocardium, there is a mechanism by which cell numbers may be partially replenished. Our understanding of regeneration has come largely from amphibian and fish models, including the newt and zebrafish, as summarized in Fig. 5.5 . These studies have extended into mammals, with mice demonstrating some regenerative capacity as neonates (see Fig. 5.5 ) . Recent studies have combined pulse-chase approaches, genetic fate mapping with stable isotope labeling, and multiisotope imaging mass spectrometry to quantify the low rate of division preexisting cardiomyocytes divide during normal aging, which increases in areas adjacent to myocardial injury . These fascinating new studies revealed that the preexisting cardiomyocytes are the dominant source of replaced cardiomyocytes in both myocardial homeostasis and after injury in mouse models .

The regenerative capacity of myocardial muscle has been a critical finding as recent breakthroughs have identified new ways in which de novo cardiomyocyte-like cells can be created from terminally differentiated nonmyocytes in the heart such as fibroblasts . Recent reviews consider the opportunities and challenges for this technology, where both transcription factors and micro-RNA (miR) can direct cardiac reprogramming in mouse embryos as depicted in Fig. 5.6 . Several important caveats should be made here, including the critical need to regenerate multiple cell types, including vessels via angiogenesis as outlined in Fig. 5.7 . Moreover, direct reprogramming of adult somatic cells in humans has proven more difficult than in nonmyocyte mouse heart cells as few cardiomyocytes with which sarcomere function, calcium transients, and action potentials were found after reprogramming human fibroblasts . Future work will require refinements to in vitro and potentially in vivo use to successfully exploit the heart’s limited ability to regenerate in cardiac disease.

Heart failure with preserved ejection fraction

Approximately 50% of all HF cases represent HFpEF. The high and rising prevalence of HFpEF is attributed to increased life expectancy, the growing aging population, and the epidemic of comorbidities such as hypertension, coronary artery disease, diabetes, obesity, metabolic syndrome, atrial fibrillation, chronic kidney disease, and chronic obstructive pulmonary disease (COPD) . The most common cause and common risk factors of HFpEF include hypertension , preclinical DD (DD with preserved left vetricular ejection fraction (LVEF) and no HF symptoms) , and LV hypertrophy. The major pathophysiologic abnormalities in HFpEF are DD, abnormal ventricular–arterial coupling, longitudinal systolic dysfunction in the face of a normal LVEF, pulmonary hypertension with HF, chronotropic incompetence, impaired vasodilator reserve, reduced LA function, impaired skeletal muscle function, and extracardiac causes of volume overload . While hypertension can trigger the development of LV hypertrophy and fibrosis , development of HFpEF in this scenario is highly variable. In the Framingham Heart study, isolated systolic hypertension leads to concentric LVH in females, but in males leads to eccentric hypertrophy . Animal studies have similarly found this relationship , suggesting that isolated systolic hypertension contributes to the increased HFpEF in females . Due to collagen cross-linking, the arterial stiffness associated with advanced age, geometric changes, and altered endothelial cell function may contribute to HFpEF . Borlaug et al. identified endothelial cell dysfunction in 42% of patients with HFpEF, compared to 28% of hypertensives with no HF. Furthermore, diabetes, obesity, and COPD interact to create a milieu that causes endothelial cell dysfunction, a prominent condition in HFpEF . Fig. 5.8 summarizes the interplay between the major risk factors that contribute to the development of HFpEF .

Neurohormonal paradigm

The understanding of HF began to shift away from the purely mechanical paradigm in the 1970s and 1980s, paralleling greater knowledge of neurohormones. Implicit in this recognition is the fact that HF is a systemic process, rather than simply a disease of the heart itself. Understanding the hemodynamic basis of HF remains essential to understanding the disease process, but the evolution of the neurohormonal paradigm has provided both the mechanistic underpinning for organ-level pathophysiology and the targets for clinically important HF therapies.

Sympathetic nervous system in heart failure

A decline in cardiac performance leads to activation of the sympathetic nervous system (SNS), and elevations in circulating catecholamines are central to the pathobiology of HF . In fact, plasma norepinephrine level is directly proportional to mortality in HF patients . Though acute stimulation of β1-adrenergic receptors (β1-ARs) on cardiac myocytes leads to increased contractility, persistent β1-AR activation results in pathological hypertrophy and cell death. Chronic adrenergic activation ultimately leads to a depletion of norepinephrine stores as well as downregulation and desensitization of myocardial β-ARs, further impairing the contractile and inotropic reserve of the failing heart. Activation of β2-ARs on cardiac fibroblasts leads to accelerated collagen deposition and differentiation of fibroblasts into myofibroblasts, contributing to myocardial fibrosis. Cardiac fibroblasts also secrete multiple paracrine signals that promote pathological changes in cardiac myocytes to include IL-6, angiotensin II, and endothelin. Collectively these direct myocardial effects cause pathological hypertrophy and remodeling, leading to perpetuation of HF.

Chronic catecholamine surge also exerts detrimental effects on other organs. Stimulation of ARs in the kidney leads to sodium and water retention, in addition to release of renin from the juxtaglomerular apparatus; these events collectively contribute to the elevated preload that is characteristic of symptomatic HF. Stimulation of vascular α1-ARs leads to smooth muscle cell proliferation and vascular remodeling that causes unfavorable changes in afterload. As such, persistent SNS activation contributes to all of the adverse hemodynamic changes that previous generations of physiologists described in patients with HF.

Renin–angiotensin–aldosterone system in the failing heart

Elevations in effector hormones of the renin–angiotensin–aldosterone system (RAAS) also contribute to the progression of HF . Decreased cardiac output, increased venous pressure, and increased renal arteriolar resistance during HF all conspire to increase renin release, which catalyzes the protease cascade that ultimately leads to formation of angiotensin II from angiotensinogen. Angiotensin II binds to AT1 receptors on vascular smooth muscle, vascular endothelium, cardiac myocytes, cardiac fibroblasts, and the adrenal cortex. Angiotensinogen, the precursor of angiotensin II, is synthesized primarily in the liver, though intrinsic production of angiotensin II has been identified in the heart, and likely plays important roles in the progression of HF .

Traditionally, angiotensin II was believed to exert its pathological effects primarily in the vasculature and kidneys. Indeed, activation of vascular AT1 receptors leads to vasoconstriction, adverse vascular remodeling, and endothelial dysfunction, collectively increasing afterload to the failing heart. Additionally, renal AT1 receptors in the proximal convoluted tubule promote retention of sodium and water. Angiotensin II also induces aldosterone release from the adrenal cortex, which independently promotes vasoconstriction and sodium retention through binding to mineralocorticoid receptors. Elevated angiotensin II levels also lead to activation of the SNS, through release of norepinephrine in cardiac nerve terminals as well as central activation of sympathetic nerves that target the heart. More recently, it has become clear that angiotensin II and aldosterone exert adverse effects through activation of receptors on cardiac myocytes and fibroblasts. Angiotensin II activates AT1 receptors on cardiac myocytes, leading to hypertrophy. Angiotensin II and aldosterone both bind their respective receptors on cardiac fibroblasts, leading to proliferation and fibrosis. Thus RAAS effectors cause cardiac hypertrophy both directly by activation of receptors in cells of the heart, and indirectly through their vascular effects.

Other neurohormonal abnormalities in heart failure

Many other neurohormones contribute to the pathophysiology of HF. Arginine vasopressin (AVP), a nine amino-acid peptide, is secreted from the posterior pituitary in response to high system blood osmolality, angiotensin II, catecholamines, and hypotension. In response to high osmolality and hypotension, AVP is released to increase arterial pressure (through vasoconstriction), blood volume (through renal water handling), and cardiac output. AVP then acts upon oxytocin receptors and vasopressin receptor subtypes V1a, V1b, and V2 found in vascular, pituitary, and renal, respectively . AVP binding to V1 receptors on vascular smooth muscle promotes calcium flux and vasoconstriction; AVP binding to V1 receptors on cardiomyocytes leads to the induction of the hypertrophic response. Activation of V2 receptors by AVP in the kidneys leads to reabsorption of water independent of sodium, lending vasopressin its other name, antidiuretic hormone. The renal effects of vasopressin contribute significantly to hyponatremia, a hallmark of advanced HF. In HF, AVP levels are increased ; increased AVP is related to an increased severity of HF , resulting in impaired release of free water (from the kidneys). The resulting water retention contributes to hyponatremia , inducing the pituitary release of more AVP and amplifying the congestion associated with HF due to volume overload. The hyponatremia itself contributes to an increased mortality in HF . AVP antagonists have been developed to treat HF. The Acute and Chronic Therapeutic Impact of a Vasopressin Antagonist in Congestive Heart Failure (ACTIV in CHF) trial investigated the short- and intermediate-term effects of tolvaptan in inpatient HF patients . When tolvaptan was administered to patients with congestive HF and LVEF% <40, the treatment groups had a net fluid loss, resulting in reduced body weight with no adverse effects . Similar results have been reported in the EVEREST and ECLIPSE trials with tolvaptan . An additional study examining the renal effects of lixivaptan illustrated an increase in urine volumes, with significant increases in solute-free water excretion, leaving serum sodium levels significantly higher , suggesting its utility as a therapy for HF. Conivaptan has demonstrated significant increases in urine output and decreased body weight, but this nonselective V1a/V2-receptor antagonist did not improve the clinical status of HF patients . While AVP contributes to the severity and progression of HF, studies blocking its effects have just begun to investigate the therapeutic utility of these drugs either as an add-on or monotherapy (as recently reviewed ).

Though most of the characteristic neurohormonal alterations in HF are maladaptive, NP synthesis is protective and counteracts many of the deleterious effects of SNS and RAAS activation . There are three recognized NPs: ANP, BNP, and CNP. ANP is released predominantly from cells in the atria in response to wall stress, angiotensin II, endothelin, or vasopressin. Similar stimuli promote release of BNP from ventricular cardiac myocytes. CNP is synthesized in endothelial cells and released in response to cytokines and other hormones, including acetylcholine. All three NPs bind membrane-bound guanylyl cyclases, GC-A and GC-B, leading to increased intracellular cyclic guanoside monophosphate (cGMP). Collectively, the NPs mediate physiological effects that are broadly beneficial in HF, including vasodilation, salt and water excretion, decreased renin release, and diminished SNS activity. GC receptors on cardiac fibroblasts and cardiac myocytes exert direct antifibrotic and antihypertrophic effects.

NP synthesis is used as an indicator of abnormal cardiac myocyte function, both experimentally and clinically. In cell culture and animal models, ANP (or ANF) is considered part of the “fetal gene program” that is reactivated in the cardiac myocyte response to hypertrophic stimuli. In clinical settings, elevated blood levels of BNP are detected in HF patients and the degree of elevation is inversely proportional to survival.

Therapeutic neurohormonal modulation

Drugs that antagonize the systemic effects of neurohormonal excess are the cornerstones of contemporary HF therapy. Beta-blockers, angiotensin-converting enzyme (ACE) inhibitors, angiotensin II receptor blockers, and aldosterone antagonists all decrease symptoms and prolong life in HF, reinforcing the importance of both the RAAS and SNS in HF pathobiology. Vasopressin receptor antagonists are approved and available but have no clear effect on survival and are used very rarely in practice. Recombinant BNP (nesiritide) improves symptoms in HF and was used extensively in the previous decade, though has fallen from favor due to expense and failure to improve survival in clinical trials .

Signaling responses to various stimuli during heart failure

The cells of the heart have a limited repertoire of responses to external stimuli. The heart comprises fibroblasts, endothelial cells, smooth muscle cells, adipocytes, and circulating blood cells, and cardiomyocytes. Since the cardiomyocytes (and their sarcomeric contractile apparatuses) are responsible for the primary pumping functions of the heart, their responses are particularly important. Since cardiomyocytes are terminally differentiated cells, their response repertoire is limited, with cardiac hypertrophy (the increase in cardiomyocyte size due to the addition of sarcomeric units) and atrophy (reduction in sarcomeric units) being a primary response. Cardiac hypertrophy occurs in response to stimuli such as IGF-1 and angiotensin II. Considering cardiomyocytes account for more than two-thirds of the normal cardiac mass and are mostly terminally differentiated, cardiac hypertrophy mostly occurs through enlargement of said cardiomyocytes. Cardiomyocytes will hypertrophy in an attempt to maintain pressures across the ventricular wall in the face of increased intraventricular pressures (Laplace’s law) .

Cardiac hypertrophy is divided into two fundamental categories based on functional and mechanistic properties: physiological and pathological hypertrophy. The most descriptive characteristic that differentiates physiological from pathological hypertrophy is that during physiological hypertrophy, cardiac function (both systolic and diastolic) remains normal or even improves (as in the case of exercise). Conversely, pathological hypertrophy is characterized by impaired ventricular function. Another major determinant is that physiological hypertrophy is reversible, which is not the case for pathological hypertrophy . Here we explore the underlying mechanisms and signaling pathways that elicit a growth response in the heart while differentiating why one form is beneficial and the other is detrimental.

Physiological hypertrophy

Enlargement of the heart, induced by exercise or pregnancy, is known as physiological hypertrophy. Pregnancy- and exercise-induced physiological hypertrophy is characterized by a 10%–20% increase in heart mass, compared to age-matched nonpregnant and sedentary individuals, respectively . These examples of physiological hypertrophy are reversible, one of the intriguing differences between pathological and physiological hypertrophy. Of course, there is physiological hypertrophy that occurs during the postnatal development period that is characterized by more than a twofold increase in cardiac mass, though this is not reversible . Many biomolecules (proteins, hormones, and cellular intermediates) play a role in the initiation and regulation of the hypertrophic response at the cellular level and the aim of the following section is to explore the function of these molecules and the impact they have on whole organ function in HF. This is made up primarily of signaling through the IGF-1 receptor complex and downstream PI3K/Akt/GSK-3β and mammalian target of rapamycin (mTOR) signaling pathways.

Insulin/IGF-1

Insulin is commonly discussed in terms of its regulation of glucose metabolism, but its intracellular role is much broader, and this peptide hormone is a known regulator of physiological hypertrophy . Binding of insulin to the insulin receptor (a transmembrane tyrosine kinase receptor) stimulates the recruitment and subsequent phosphorylation of insulin receptor substrate-1 (IRS1) and IRS2 ( Fig. 5.9 ) and activates the PI3K-Akt pathway (discussed in detail below). Insulin signaling in the heart regulates energy metabolism through regulation of glucose transport, glycolysis, glucose oxidation, glycogen synthesis, and fatty acid (FA) oxidation, and also protein synthesis . These processes all tie into cellular growth, and the broad role of downstream signaling from the insulin receptor is demonstrated in postnatal heart development that is known to be retarded by cardiac-specific genetic deletion of the insulin receptor where a 20%–30% decrease in heart size is observed . Additionally, cardiac-specific deletion of the insulin receptor sensitizes the heart to pressure overload, as exemplified by an expedited transition to dilated cardiomyopathy, depressed systolic function, and increased myocardial fibrosis in a rodent model . In a model of ischemic cardiac injury using MI, the mortality rate in cardiac-specific insulin receptor-deficient mice is decreased, in which a reduced mitochondrial FA oxidation capacity appears to be the underlying mechanism . Collectively, these results demonstrate the importance of insulin and its intracellular signaling pathway(s) in regulating postnatal physiological hypertrophy, along with compensatory mechanisms to deter and in compensating against pathological hypertrophy.

Similar to insulin, insulin-like growth factor-1 (IGF-1) is influential in initiating physiological hypertrophy. IGF-1 is synthesized and secreted by the liver in response to circulating growth hormone stimulation. Target and nearby tissues also synthesize IGF-1 as an autocrine or paracrine factor, respectively . IGF-1 binding to the insulin receptor or the IGF-1 receptor (also a tyrosine kinase receptor) activates the PI3K/Akt pathway and subsequently glycogen synthase kinase-3β (GSK-3β). IGF-1 null and IGF-1 receptor deficiency results in high postnatal mortality and severe growth retardation in surviving animals . Conversely, evidence suggests that IGF-1 receptor overexpression increases cardiomyocyte volume and systolic function with no change in myocardial fibrosis . However, this may be a time-dependent effect as IGF-1 overexpression in a mouse model also demonstrates physiological hypertrophy development early on but becomes pathological over time as evidenced by increased myocardial fibrosis and reduced cardiac function . In line with transient induction of IGF- evoking a beneficial physiological hypertrophic response, there are data showing systemic and cardiac concentrations of IGF-1 typically rise during exercise, postnatal cardiac growth, and physiological hypertrophy . These findings highlight the importance of IGF-1-mediated signaling during physiological hypertrophy.

PI3K/Akt/GSK-3β

Both insulin and IGF-1 transduce signals via the IGF-1 receptor at the cell surface to activate physiological hypertrophy via the PI3K/Akt pathway . The IGF receptor activates PI3K, a membrane-bound lipid kinase that is essential for physiological growth of the heart, which phosphorylates and thus activates Akt (also known as protein kinase B) and the signal is transmitted to numerous downstream elements of the physiologic hypertrophic response to affect protein activity and gene expression, along with the protein synthesis/degradation ratio.

First, Akt can phosphorylate and activate mTOR, which is discussed in detail below. Briefly, the mTOR pathway is key for enhancing protein synthesis, tilting the synthesis/degradation ratio toward growth and hypertrophy when activated . Akt also phosphorylates and deactivates GSK-3β ( Fig. 5.9 ). GSK-3β, which is a negative regulator of hypertrophy, suppresses hypertrophic transcriptional activators such as GATA4, β-catenin, c-Myc, and nuclear factor of activated T-cells (NFAT). GSK-3β can also phosphorylate NFAT, which leads to its exclusion from the nucleus and degradation , resulting in inhibition of the NFAT/calcineurin signaling cascade. GSK-3β also targets eukaryotic translation initiation factor 2B (eIF2B) . Activating eIF2B (via inhibiting its inhibitor, GSK-3β) will result in increased protein translation. In addition to GSK-3β, Akt can inactivate the transcription factor forkhead box protein O3, FOXO3 . The FOXO proteins regulate an atrophy gene program that regulates myocyte size.

The AKT signaling axis is not specific to physiological hypertrophy; however, how this pathway is regulated during HF differs from physiological hypertrophy. In the short term, activation of Akt results in adaptive growth; however, chronic stimulation of these pathways eventually progresses to pathological hypertrophy. For example, overexpression of Akt in mice for 2 weeks results in reversible physiological hypertrophy, but by 6 weeks develops into HF . This makes it exceedingly difficult to target these pathways pharmacologically as they are under tight regulation and the appropriate level of activation is both magnitude- and time-dependent in nature.

Mammalian target of rapamycin

mTOR is a protein kinase consisting of two distinct protein complexes, mTOR complex 1 (mTORC1) and mTORC2. Activation of mTOR influences cardiac hypertrophy by regulating protein translation and autophagy. mTORC1 is activated by Akt-mediated phosphorylation but can be inhibited by AMP-activated protein kinase (AMPK)-mediated phosphorylation at a separate site . Activated mTORC1 initiates translation by phosphorylating two key proteins: S6 kinase (S6K) and 4 eukaryotic binding protein 1 (4E-BP1) ( Fig. 5.9 ) .

Exercise-induced physiological cardiac hypertrophy involves mTOR regulation. Pharmacological inhibition of mTORC1 with rapamycin or its analogs effectively reduces cardiac hypertrophy, without fatally impairing the physiological actions of mTOR. The underlying mechanisms are not completely clear, but current thinking is that rapamycin analogs do not fully inhibit mTORC1 actions. Induction of physiological hypertrophy through overexpression of Akt is attenuated by mTOR inhibition . Genetic deletion of mTORC1 is associated with a high rate of embryological lethality and mice that survive experience cardiac dilation and dysfunction, cardiomyocyte apoptosis, mitochondrial derangement, and ultimately HF and death . A recent study illustrated how mTOR is activated during swimming-induced cardiac hypertrophy, which was correlated with stimulation of downstream mTOR targets .

The physiological cardiac hypertrophy response, most notably present in response to exercise, is mediated by a series of signaling pathways initiated by pituitary release of growth hormone release in response to exercise. Growth hormone then acts on hepatocytes to release IGF-1, which then mediates the effects on physiological cardiac hypertrophy by stimulating IGF-1 complexes and downstream PI3K/Akt/GSK-3β and mTOR signaling pathways. The compensatory increase in heart size, due to increased cardiomyocyte size, allows performance increases in heart function without detrimental effects with ongoing stimuli. The lack of long-term detriment in physiological hypertrophy differentiates it distinctly from the pathological hypertrophy response, discussed next.

Pathological hypertrophy

In response to pathological stress such as pressure or volume overload or ischemia, the heart will hypertrophy in an attempt to compensate for increased wall stress. Although beneficial in the short term, in which LV function is maintained, chronically, this compensation is maladaptive and will progress to HF. Despite overlap in the associated cell signaling pathways, even in the acute phase of the hypertrophic response, the signaling activated during physiological and pathological hypertrophy differs . Therefore pathological hypertrophy evokes distinctive events to give rise to the hallmarks of pathological hypertrophy, including decreased ventricular function, wall thickening, and a failure to regress from the hypertrophic state . This section discusses the causes and underlying mechanisms along with identifying possible therapeutic targets of pathological hypertrophy.

Hypertension

Hypertension affects nearly 40% of the population over 30 years of age in the United States, and chronic uncontrolled hypertension and the accompanied sustained cardiac pressure overload induces compensatory pathological cardiac hypertrophy culminating in HF. Left ventricular dysfunction results in a reduced stroke volume and cardiac output associated with increased end-systolic and end-diastolic volumes along with end-diastolic pressure. Collectively, these changes elicit increased LA pressure, with pressure backing up into the pulmonary circulation, which impairs pulmonary alveolar gas exchange manifesting in dyspnea . The exact cause of hypertension remains a mystery, making management of hypertensive patients essential.

Much of what we know about hypertension comes from studying mouse models and cells in culture. These have been invaluable for dissecting the signaling pathways underlying the hypertrophic response. In particular, they allow for genes to be turned on or off in a cell-specific manner (i.e., cardiomyocyte-specific) to monitor responses in more complex in vivo environments . Cultured cells can be easily manipulated through various mechanisms: drug treatments, gene transfer (via adenovirus, lentivirus, or plasmid), or knock-down experiments with silencing RNA. Other models exist and new models are continually being developed to better mimic human disease . Hypertension/cardiac pressure overload can be modeled in vivo by perfusing animals with angiotensin II, phenylephrine, or isoproterenol or by constricting the aorta between the left and right carotid arteries, referred to as transverse aortic constriction . Cardiac hypertrophy is modeled in vitro by treating cells with the aforementioned agents in addition to endothelin-1 .

Calcineurin

A common hallmark of pathological hypertrophy is induction of the calcineurin/NFAT pathway. Calcineurin, also referred to as protein phosphatase 2B, is a calcium/calmodulin-activated serine/threonine phosphatase that, once stimulated, dephosphorylates and thereby activates NFAT . Once dephosphorylated, NFAT translocates to the nucleus where it associates with the transcription factors Myocyte enhancer factor 2 (MEF2) and GATA4 (discussed in detail below) . These actions culminate in the transcription of hypertrophy-associated genes (the fetal/hypertrophic gene program) α-actin, endothelin-1, ANP, BNP, β-myosin heavy chain (β-MHC), transient receptor potential cation channel 6 (TRPC6), and regulator of calcineurin signaling 1 (RCAN1) . As its name implies, RCAN1 regulates calcineurin as part of a negative feedback loop to suppress calcineurin activation. The ubiquitin ligase, atrogin-1, also regulates calcineurin by ubiquitinating calcineurin, which targets it for proteasome degradation (discussed below) . NFAT activity is regulated by various protein kinases through phosphorylation to antagonize NFAT nuclear translocation allowing transcriptional regulation of target genes. These protein kinases include phosphatidylinositide 3-kinase (PI3K), Akt (protein kinase B), and GSK-3β .

Studies have revealed the impact the calcineurin/NFAT pathway has on pathological hypertrophy during conditions of pathological cardiac stress. Transgenic mice with cardiac-specific activation of the calcineurin/NFAT pathway develop cardiac hypertrophy, whereas inhibition of the calcineurin/NFAT pathway effectively attenuates cardiac hypertrophy . The clinical application of calcineurin inhibitors has long been sought after with promising preclinical findings; however, the lack of acceptable safety profiles currently hinders their application .

Small guanosine triphosphate-binding proteins Ras/Rho

Small guanosine triphosphate (GTP)-binding proteins are divided into five main subfamilies: Ras, Rho, Rab, Arf, and Ran. Also known as GTPases, these small GTP-binding proteins act as intracellular molecular switches, in conjunction with guanine nucleotide exchange factors (GEFs), by cycling between an active GTP-bound state and an inactive GDP-bound state . In response to various extracellular stimuli, the GEF will stimulate the release of GDP from the GTPase and the binding of GTP to the GTPase, and thus activation . Stimulation of cardiomyocyte receptors coupled to G _αq/11 and G _12/13 G-proteins with angiotensin II, endothlin-1, or phenylephrine activates Ras and Rho (G _αq/11 ) or Rho only (G _12/13 ) . In the heart, both Ras and Rho activation have been linked to the progression of pathological cardiac hypertrophy and remodeling . Downstream targets of Ras and Rho activation include the mitogen-activated protein kinases (MAPKs) and extracellular signal-regulated protein kinases (ERK), in addition to Rho-activating Rho kinase (ROCK). Activation of the MAPKs, ERK, and ROCK from Ras and Rho has been shown to induce transcription of the hypertrophic gene program, increased protein synthesis, and increased cardiomyocyte size. Furthermore, ROCK increases actin production and organization, both of which are hallmarks of cardiac hypertrophy . While Ras and Rho activation are associated with increased cardiac hypertrophy, their inhibition is antihypertrophic . A known inhibitor of Ras and Rho is guanine nucleotide dissociation inhibitors, which inhibit the dissociation of GDP from the GTPase, thus not allowing activation to occur . Activators of cGMP-dependent protein kinase (PKG) potently inhibit Ras and Rho, as part of PKG’s multifaceted antihypertrophic actions .

Protein kinase C

It is clear that protein kinase C (PKC) plays a key role in hypertrophic signaling. There are numerous studies showing PKC as a central player, and there is significant interest in generating therapeutics that target PKC signaling. However, the situation has turned out to be quite complicated, as there are at least 12 different isoforms of PKC, organized into three families based on their mechanisms of activation. Determining which isoform is the most important has been difficult, as many of the isoforms can compensate for each other. The prevailing opinion, however, is that PKCα, PKCβ, and PKCɛ are most directly responsible for hypertrophic signaling.

Neurohormonal signals such as angiotensin II, endothelin-1, and catecholamines bind to an alpha-adrenergic receptor on the surface of cardiomyocytes, which is coupled to a heterotrimeric G-protein named G _αq/11 . These G _αq/11 coupled receptors are associated with phospholipase Cβ (PLCβ). Activation of PLCβ results in the production of diacylglycerol (DAG) and inositol-1,4,5-trisphosphate (IP3) . IP3 can bind to an IP3 receptor on the sarcoplasmic reticulum (SR), stimulating the release of calcium . DAG and calcium can then bind to and activate PKC . Once activated, PKC will phosphorylate many intracellular proteins, including myofilament proteins (altering sensitivity to calcium ) and promote increased calcium release.

PKCα will phosphorylate and inhibit the antihypertrophic HDACs 4, 5, 7, and 9 (detailed description below), which enhances protein synthesis and cardiomyocyte growth ( Fig. 5.10 ) . PKCα inhibition in mice protected mice against cardiac hypertrophy and reduced cardiac remodeling . Furthermore, the cardiomyocyte expression of a dominant negative PKCα inhibited the progression to HF . Multiple independent transgenic mice strains have been used to implicate the PKCβ, and both support that the isoform is sufficient to produce hypertrophy, but not necessarily required. The most understood isoform, however, is PKCɛ, which is activated by both mechanical stress and Gq signaling. In hypertrophy, PKCɛ is upregulated and translocates within the cell. These mechanisms are compensatory, however, as if these mechanisms are blocked in HF, the result is disastrous, lethal dilated cardiomyopathy . Thus therapeutically, enhancement of PKCɛ translocation would be beneficial.

Serum response factor/GATA4

Serum response factor (SRF) and GATA4 (so named as it binds to the DNA sequence GATA) are transcription factors that regulate many cardiac genes both at baseline and in response to stress ( Fig. 5.11 ). SRF has been implicated in regulating the hypertrophic genes , cardiac α-actin, α-MHC, and β-MHC, and even other transcription factors such as GATA4 . Cardiac overexpression of SRF resulted in hypertrophy . Interestingly, mice with cardiac-specific deletion of SRF showed dilated cardiomyopathy and reduced cardiac contractility, along with defects in their cardiac structural proteins (regulated by SRF) and early-onset HF . SRF is regulated by the ubiquitin ligase, MuRF1. Genetic deletion of MuRF1 yielded mice with exaggerated hypertrophy and enhanced expression of the SRF-dependent hypertrophy genes . These findings suggest that SRF plays a pivotal role during the development of hypertrophy and that SRF is needed at baseline to maintain adequate cardiac function.

GATA4 belongs to the GATA family of transcription factors that regulate cardiac development, differentiation, proliferation, and survival . GATA4 mediates the induction of a select set of genes including α-MHC, myosin light chain 1/3 (MLC1/3), cardiac troponin C, cardiac troponin I, ANP, BNP, cardiac-restricted ankyrin repeat protein (CARP), cardiac sodium–calcium exchanger (NCX1), cardiac m2 muscarinic acetylcholine receptor, A1 adenosine receptor, and carnitine palmitoyl transferase Iβ, many during cardiac hypertrophy . Overexpression of GATA4 alone is sufficient to cause hypertrophy . GATA4 phosphorylation by ERK1/2 induces GATA4 DNA binding, thus promotion of hypertrophic gene expression . During hypertrophy, NFAT associates with GATA4 to induce expression of the hypertrophic genes . Genetic inhibition of GATA4 attenuated hypertrophy induced by pathological and physiological stimuli . GSK-3β can phosphorylate GATA4, which induces its nuclear export, eventual ubiquitination, and subsequent degradation by the 26S proteasome (discussed above) . Collectively, these studies indicate the pro-hypertrophic effects of GATA4 activation.

Beta-adrenergic/alpha-adrenergic/renin–angiotensin–aldosterone system

Beta-1-adrenergic receptors (β1-AR) are coupled to the G _s G-protein/adenyl cyclase signal transduction pathway, a central pathway to regulating cardiac function ( Fig. 5.12 ) . Downstream effects from this pathway are mediated by cAMP-dependent protein kinase (PKA), which phosphorylates its targets to increase HR and contractility . There is dysregulation and uncoupling of the β1–AR pathway during cardiac stress, such as increased B1 adrenergic signaling . Genetic overexpression of β1-AR alone was enough to induce cardiac hypertrophy, increased apoptosis, and eventually HF, which was attenuated by β1–AR inhibition . Notably, β1–AR blockade is one of the most prevalent and effective treatments of human HF patients .

The other main class of AR is the alpha 1 AR (α1-AR), which is coupled to a G _q /G ₁₁ G-protein/phospholipase Cβ1 pathway. Stimulation of this pathway generates IP3, subsequent release of intracellular calcium, and DAG, followed by PKC activation ( Fig. 5.10 ) . There is an increased stimulation of α1-AR during a cardiac stress, resulting in increased cardiac force of contraction, vasoconstriction, and protein synthesis. Cardiac hypertrophy is a common feature of α1-AR stimulation, whereas α1-AR inhibition effectively attenuated the onset of hypertrophy . Overexpression of α1-AR in mice yields hypertrophy, increased fibrosis, and early death. Unfortunately, α1-AR blockade has yielded mixed results in the clinical setting, likely due to differing etiologies, demonstrating the complexity of this system .

The renin angiotensin system (RAS) and its primary effector, angiotensin II (AngII), are principal mediators of cardiac hypertrophy. Once produced, AngII can bind to AngII type 1 (AT1) or AngII type 2 (AT2) receptors on cardiomyocytes. AngII binding to the AT1 receptor, which is coupled to a G _q /G ₁₁ G-protein, results in activation of ERK1/2, p38 MAPK, and protein synthesis . The RAS, through aldosterone, also promotes water retention that increases blood pressure and cardiac stress. Additionally, AngII can be cleaved by angiotensin-converting enzyme 2 (ACE2) to yield Ang1–7, which is thought to be cardioprotective by counterbalancing the harmful effects of AngII . Inhibition of RAS in mice exposed to pressure overload reduced hypertrophy, decreased fibrosis, and increased life span . Inhibition of the RAS, either with an ACE inhibitor, an AT1 receptor blocker, or an aldosterone inhibitor, has generally had beneficial effects although there have been some adverse effects reported with combinational therapy .

Calpains

Calpains are a family of ubiquitously expressed calcium-dependent, nonlysosomal cysteine proteases. Calpains are involved in apoptosis, cellular proliferation, and cell motility. While mostly calcium-dependent, calpains may also be activated through ERK-mediated phosphorylation. Commonly regarded as intracellular proteases, calpains can be secreted to regulate the extracellular milieu . Additionally, calpains can be targeted to specific intracellular compartments to cleave and/or degrade specific proteins . A study utilizing transgenic mice that overexpress an inhibitor of calpains and calpastatin found that calpain inhibition blunted pathological cardiac hypertrophy and the subsequent cardiomyocyte apoptosis induced by angiotensin II infusion . Calpains contribute to pathological hypertrophy through degradation of IκBα [inhibitor of nuclear factor-κB (NF-κB)] and calcineurin activation, which allows NF-κB and NFAT, respectively, to enter the nucleus. Once in the nucleus, NF-κB and NFAT will induce the transcription of genes involved in cardiac hypertrophy and fibrosis . As the calpastatin study noted, calpain inhibition may be a useful therapeutic strategy to treat pathological hypertrophy.

Protein synthesis/protein degradation

Cardiac hypertrophy, in the simplest sense, is an imbalance of the protein synthesis-to-protein degradation ratio, shifting toward a net increase in protein synthesis. Acutely, cardiac hypertrophy is characterized by an increase in protein synthesis and ubiquitin proteasome system (UPS)-mediated proteolysis. However, protein synthesis is increased . The sequence of events, altered protein synthesis versus degradation, was a mystery until one study observed that increased protein synthesis is responsible for acute hypertrophy (within 1 month), while reduced protein degradation is responsible for chronic hypertrophy .

Considering the shift toward increased protein synthesis that occurs during hypertrophy, there is growing interest in correcting this imbalance by enhancing protein degradation. Multiple lines of evidence support this hypothesis. Prolonged fasting or caloric restriction is known to tip the balance toward enhanced degradation and is associated with regression of cardiac hypertrophy or atrophy . Additionally, most, if not all forms of pathological cardiac hypertrophy present with an increase of ubiquitinated proteins, misfolded proteins, and toxic protein aggregates . Protein degradation primarily occurs through two mechanisms: UPS-mediated proteolysis and autophagy-lysosome (autophagy)-mediated degradation. Detailed in the following section is the current understanding of the role the UPS (the proteasome and ubiquitinating enzymes) has in cardiac hypertrophy. Autophagy is discussed in the following chapter.

Ubiquitin proteasome system

The UPS is responsible for degradation of most intracellular proteins (~80%) and therefore influences many cellular processes and maintains proteostasis. UPS-mediated protein degradation is divided into two main steps: targeting of the substrate protein through attachment of a polyubiquitin chain, termed ubiquitination, and the subsequent protein degradation by the proteasome ( Fig. 5.13 ) . Ubiquitination of a substrate protein occurs through a series of enzymatic reactions involving an ubiquitin activating enzyme (E1), ubiquitin conjugating enzyme (E2), and an ubiquitin ligase (E3) . The typical 26S proteasome consists of a 20S proteolytic core and one or two 19S lids or caps .

Increasingly, while implicated in the development of pathological hypertrophy, the exact role of the UPS in cardiac hypertrophy is complex and occasionally controversial, and thus not completely understood . An increase in proteasome activities is known to occur during the initial development of cardiac hypertrophy but decline in the transition to HF while there are increases in ubiquitinated proteins, misfolded proteins, and protein aggregates . These results suggest that while proteasome activity is increased, UPS-mediated degradation is impaired or at least inadequate to deal with the increased protein load during cardiac hypertrophy and failure. Cardiovascular disorders characterized by oxidative stress see an increase in an alternative lid for the 20S proteasome, named the proteasome activator 28, which when overexpressed is associated with enhanced protein degradation, reduced oxidative damage, and attenuated cardiac mass . Additionally, the proteasome can be regulated by posttranslational modifications , which is greatly expanding the ability of protein degradation to be targeted as a potential treatment . Interestingly, a few studies have illustrated that the proteasome is positively regulated by protein kinase A , protein kinase G , and calcium calmodulin kinase II , among others. For proteasome activators to be clinically applicable, there must be a druggable target. Recent studies have employed therapeutic treatments to enhance UPS-mediated degradation, which, in general, were associated with cardioprotection during a pathological stress . However, there were a few studies that reported an antihypertrophic response with proteasome inhibition . Proteasome inhibition and proteasome functional insufficiency are characterized by an increase in NFAT nuclear translocation, with subsequent transcription of hypertrophic genes . Notably, patients who received an LV-assist device saw a restoration of proteasome function . These studies highlight the important role that the proteasome has during the development of cardiac hypertrophy and transition to failure.

Not to be overlooked, much of the specificity of the UPS is due to the presence, activity, and/or substrate selection of the ubiquitin ligases. Atrogin-1 (also known as MAFbx1) is a striated muscle-specific ubiquitin ligase that attenuates cardiac hypertrophy, primarily by targeting calcineurin for proteasome-mediated degradation and thereby reducing NFAT-induced transcription . Expression of atrogin-1 is regulated by Forkhead family (FOXO) of transcription factors, mainly FOXO1 and FOXO3a. Interestingly, atrogin-1 controls its own expression by lysine-63 linked ubiquitination rather than the classical degradation-targeting lysine-48 linked ubiquitination, and this further increases FOXO activity . Thus a positive feedback loop is created, intriguingly though not just for atrogin-1 but also for another family of ubiquitin ligases, the muscle ring finger proteins (MuRFs) . MuRF1, 2, and 3 are all involved in myogenic responses and contractile regulation, but only MuRF1 appears to inhibit cardiac hypertrophy . Samples from human patients post-right ventricular assist device (RVAD) therapy revealed that MuRF1 is increased during cardiac atrophy. These findings were recapitulated in mouse studies where experimentally induced pressure overload was alleviated . MuRF1 elicits antihypertrophic properties by ubiquitinating sarcomeric troponin I for degradation; interacting with the receptor for activated PKC, which suppresses focal adhesion kinase and ERK1/2; and associating with the SRF transcription factor . Furthermore, mice lacking MuRF1 were unable to undergo cardiac atrophy and displayed enhanced expression of the SRF-dependent genes: BNP, smooth muscle actin, and β-MHC . Recent studies have highlighted the importance of another ubiquitin ligase, C-terminus of HSC70-interacting protein (CHIP), during pathological hypertrophy and lipid accumulation . CHIP-knockout mice exhibit robust cardiac hypertrophy in response to pressure overload . This group also uncovered the ability of CHIP to stabilize and enhance the activity of antihypertrophic AMP kinase . A recent study uncovered that CHIP is posttranslationally regulated by phosphorylation via protein kinase G, which strengthens the interaction of CHIP with heat shock cognate 70 to enhance cardiomyocyte proteostasis following an MI . A developing area of interest is how CHIP and MuRF1 not only regulate protein degradation to be antihypertrophic but also their newly revealed roles in cellular metabolism .

Mammalian target of rapamycin

The onset, development, and progression of cardiac hypertrophy, both physiological (discussed above) and pathological, are intricately regulated by mTOR . This atypical serine/threonine protein kinase interacts with the adaptor proteins mTORC1 and mTORC2. mTORC1 regulates protein synthesis, cell growth and proliferation, ribosomal and mitochondrial biogenesis, metabolism, and autophagy. mTORC2 is a complex that in general regulates cellular metabolism and the cytoskeleton and regulated by insulin, nutrient levels, and growth factors . mTORC2 role in pressure overload-induced cardiac hypertrophy has recently been described , but much less is understood about mTORC2 compared to mTORC1, which has been studied to a much greater extent.

mTORC1 has been more extensively studied and continues to be of great therapeutic interest, mostly for its ability to regulate protein synthesis and degradation. Activated Akt and ERK1/2 stimulate mTORC1, resulting in increased protein synthesis and reduced autophagy; while AMP kinase and protein kinase G inhibits mTORC1, which reduces protein synthesis and increases autophagy .

mTORC1 is activated during pathological hypertrophy for which, pharmacological inhibition of mTOR with rapamycin or rapamycin analogs generally elicits cardioprotection, while genetic deletion of mTOR exacerbates cardiac injury. As mentioned above, the reasoning behind this disparity is due to pharmacological therapies eliciting only partial inhibition of mTORC1, whereas genetic deletion ablates all activity, including actions necessary for homeostasis. The heart cannot adapt to increased stress, and rapidly develop dilation and HF . Partial mTORC1 inhibition with rapamycin attenuates and can regress cardiac hypertrophy, while reducing fibrosis and improving cardiac contractile function . The beneficial response elicited by rapamycin or rapamycin analogs likely results from decreased protein synthesis and increased autophagy, thereby shifting the balance toward net protein degradation . Collectively, these findings illustrate that mTORC1 inhibition represents a promising therapeutic target; however, clearly this approach needs further validation.

Micro-RNA

miRs are relative newcomers to our understanding of the players in hypertrophic signaling. First discovered in 1993 , miR was not studied in cardiac tissue until 2002 and only more recently have been appreciated as a component to the hypertrophic response . Essentially, miRs are small (usually around 22 nucleotides long) nonprotein-coding RNAs that bind to their complementary sequence in the 3-prime untranslated region of target mRNA sequences. Once bound, the miR either inhibits translation or marks it for degradation; either way, the result is repression of protein expression ( Fig. 5.14 ).

While there are many miRs (currently estimated around 2000) and each can target hundreds of transcripts, only a handful of miRs have been shown to be involved in cardiac hypertrophy. One of these is miR-133, the most abundant miR in myocardial tissue . Targets of mIR-133 include the hypertrophic regulator calcineurin , and members of the beta-receptor cascade (see above). Interestingly, miR-133 expression is also regulated by a feed-forward mechanism via calcineurin/NFAT signaling .

Another key cardiac miR is miR-1, the highest expressed miR in cardiac myocytes, which targets important hypertrophic signaling molecules calmodulin, Mef2a, GATA4, and IGF-1 (all discussed above). As a clinically relevant approach, miR-1 delivered to rats with cardiac hypertrophy using gene therapy was found to reverse maladaptive remodeling, including an improvement in calcium handling and apoptosis . Another key miR, miR 208, is embedded in the myosin heavy chain gene and regulates β-MHC expression and can induce cardiac hypertrophy . Finally, there are several other miRs that correlate with hypertrophic response, including miR29, miR26, miR378, miR22, miR23, miR24, and miR199.

The anti-miRNA therapy targeting mIR-122 was one of the first miRNA-based therapies tested in patients with hepatitis C virus, which significantly reduced liver viral titers . To date, at least 18 clinical studies using eight different miRNA-based therapies have been reported, although none have targeted the cardiovascular system . That said, dozens of clinical trials are in process to quantify miRNAs in heart disease, including coronary heart disease (e.g., clinicaltrials.gov studies NCT03635255 , NCT03855891 ), cardiac allograft rejection (e.g., clinicaltrials.gov study NCT02672683 ), cardiotoxicity in breast cancer therapy (e.g., clinicaltrials.gov study NCT02065908 ), acute coronary syndrome (e.g., clinicaltrials.gov study NCT02755207 ), and HF (e.g., clinicaltrials.gov study NCT02541773 ).

The proof of concept of targeting miRNAs in heart disease remains in the animal model stages. The delivery of an anti-miR-21 in a pig model of HF confirms that catheter-based delivery of miRNAs can be delivered to the myocardium . Catheter delivery overcomes the issues with intracardiac injection delivery which exposes the heart to high concentrations of the drug. Anti-miR-21 treatment in pigs reduced macrophage infiltration and downregulated fibroblast proliferation post-MI, leading to a sustained dose-dependent restoration of cardiac functions and modification of the myocardial transcriptome .

Diabetic cardiomyopathy

Diabetic patients have a higher incidence of CVD than nondiabetic patients and CVD is the leading cause of morbidity and mortality in this population . Furthermore, the progression of CVD seems to be exacerbated by the presence of diabetes . As the incidence of diabetes continues to increase, primarily type 2, so does the occurrence of CVD . Diabetic cardiomyopathy involves both structural and functional changes of the myocardium . Hallmarks of diabetic cardiomyopathy include myocardial energy metabolism derangement, increased production of reactive oxygen species (ROS), inflammation, interstitial fibrosis, and cardiomyocyte apoptosis. Ultimately, these changes contribute to the development of HF . A major underlying mechanism of diabetic cardiomyopathy is lipotoxicity resulting from a substantial increase in FA uptake, which is accompanied by alterations in the transcription factor network that regulates sugar (glucose) and lipid (FA) metabolism .

Nuclear factor-κB

NF-κB belongs to a family of transcription factors involved in the regulation of cell proliferation, differentiation, inflammation, immune response, apoptosis, and cardiac hypertrophy. Normally, NF-κB is bound by the inhibitor of κB (IκB), which prevents nuclear translocation, and subsequent DNA-IκB can be phosphorylated by IκB kinase (IKK), which tags IκB for ubiquitination and degradation to abolish NF-κB .

Both the protein expression and activity of NF-κB are increased in diabetic cardiomyopathy. Persistent NF-κB activation in cardiomyocytes is associated with increased cardiac fibrosis and apoptosis, thereby exacerbating HF progression . Increased NF-κB is also believed to promote insulin resistance hinting at overlapping effects on the insulin signaling pathway . Both NFAT and NF-κB are transcription factors that are known to interact to promote cardiac hypertrophy and remodeling. Furthermore, cardiomyocyte-specific deletion of NF-κB is associated with a reduction in NFAT activity and reductions in hypertrophic response and cardiac remodeling, which was associated with improved contractile function during pressure overload stimulation . An intriguing possibility for treatment of diabetic cardiomyopathy is resveratrol, which activates sirtuin 1 (SIRT1) activator, that as a histone deacetylase, regulates the transcription of NF-κB through lysine310 deacetylation of the p65 subunit of NF-κB . It should be noted that resveratrol does modulate other pathways than NF-κB. Preliminary findings are encouraging; however, a lot of work still remains.

Peroxisome proliferator-activated receptor

Peroxisome proliferator-activated receptors (PPARs) are transcription factors that belong to the nuclear receptor superfamily and are important regulators of cardiomyocyte metabolism, particularly as they relate to FAs. Three isoforms are present in cardiomyocytes: PPARα, PPARγ, and PPARδ, which differ by their localization, function, and ligand specificity . Under normal physiological conditions, the heart prefers to utilize FAs for energy metabolism, due to the higher yield of energy. However, under stress conditions, where oxygen supply is likely compromised, the heart reverts to utilizing glucose for energy metabolism . Diabetic cardiomyopathy is characterized by an increase in FA metabolism and insulin resistance of the cardiomyocytes. This limits the cardiomyocytes’ ability to shift to glucose metabolism, therefore increasing the reliance on FA metabolism, leading to increased myocardial oxygen consumption and cardiac dysfunction .

PPARα is the most abundant isoform in the heart and regulates metabolism by modulating expression of enzymes involved in mitochondrial FA oxidation . Reduced PPARα activity shifts the cardiomyocyte substrate utilization from FA to glucose metabolism . Mice with cardiac-specific overexpression of PPARα exhibit increased myocardial FA oxidation with reduced glucose uptake and oxidation, which coincides with cardiac dysfunction and ventricular hypertrophy . Notably, treatment of diabetic rats with a PPARα agonist, fenofibrate, reduces cardiomyocyte apoptosis and pathological hypertrophy, suggesting that enhancing PPAR activity may be protective against diabetic cardiomyopathy .

Diabetes also induces increased PPARγ expression in rodent models. In the heart, PPARγ stimulates an increased production of intracellular triglycerides, which may contribute to cardiac lipotoxicity that is often observed as lipid accumulation . Mice with overexpression of PPARγ in the heart exhibit increased expression of FA oxidation genes, increased triglyceride uptake and storage, increased glycogen stores, and alterations in mitochondrial morphology in addition to developing dilated cardiomyopathy . Treatment of diabetic rats with a PPARγ agonist, rosiglitazone, is associated with a reduction in cardiomyocyte apoptosis, cardiac fibrosis, and DD .

Compared to PPARα and PPARγ, there is much less known regarding PPARδ. PPARδ is reduced in rats with diabetic cardiomyopathy that also exhibits an increase in ROS production, increased protein synthesis, and cell size . While PPARδ levels are reduced, the activities of the inflammatory cytokines tumor necrosis factor α (TNF-α), IL-6, and nicotinamide-adenine dinucleotide phosphate are increased in the rat diabetic heart . Collectively, these studies highlight the advances that have been made in the fight against diabetic cardiomyopathy.

Heart failure-related alterations in cellular function and biochemical pathways

HF can result from a diversity of injuries that include (1) MI, (2) chronically increased hemodynamic load that can be caused by pressure (aortic stenosis) and volume (valvular regurgitation) overload, (3) viruses (myocarditis), (4) cardiomyopathy (idiopathic or genetic-based), and (5) cardiotoxic drugs (e.g., doxorubicin) . The initiation and development of HF is accompanied by maladaptive changes in myocardial architecture and molecular responses that are defined as ventricular remodeling. Remodeling is associated with a diversity of cellular changes within the myocardium, including (1) loss of cardiac muscle cells due to apoptosis and necrosis , (2) compensatory effects on remaining cardiomyocytes resulting in cardiomyocyte hypertrophy , and (3) recruitment and activation of other cell types such as fibroblast proliferation and fibrosis , as well as inflammatory cell infiltration , which all lead to the (4) loss of cardiac muscle cell contractility ( Fig. 5.15 ).