Keywords

1 Introduction

Increased hemodynamic load on the left ventricle initiates a cascade of events that leads to ventricular remodeling, cardiac hypertrophy, and eventually heart failure [1]. This ventricular remodeling is a myocardial response to various stresses and involves gradual and progressive changes in chamber architecture, cardiomyocyte phenotype, as well as non-myocytic components [2, 3]. Clinical conditions that produce the pressure-overload stimulus include essential hypertension, aortic stenosis, aortic coarctation, and drug-induced hypertension. The pattern of ventricular remodeling and hypertrophy is dependent upon the type of cardiac stress. In the case of pressure overload, the left ventricle responds with concentric hypertrophy in contrast to the eccentric hypertrophy which occurs in response to volume overload. The process of hypertrophy is initially considered to be an adaptive response because it serves to normalize wall stress and helps to mechanically compensate heart function. However, with progression of the stimulus, the initial compensatory hypertrophy can lead to maladaptive responses that can eventually lead to heart failure. Although the exact point at which hypertrophy becomes maladaptive is difficult to identify, there may be a clear delineation of compensated and decompensated phases of hypertrophy [4–6]. Alternatively, it has also been proposed that the initial hypertrophic process involves a combination of both adaptive and maladaptive cardiac processes and that initiation of any pathological hypertrophy will eventually compromise heart function [7].

Left ventricular hypertrophy is recognized as a major independent risk factor for cardiovascular morbidity and mortality from sudden cardiac death, dysrhythmia, heart failure, ventricular ischemia, and coronary heart disease [6, 8–11]. A growing body of evidence suggests that oxidative stress, which occurs when the generation of reactive oxygen species (ROS) exceeds the cell’s intrinsic antioxidant defenses, is an important contributor to the development and progression of cardiac hypertrophy, in part by activating multiple intracellular signaling pathways [12]. Therefore, ameliorating oxidative stress has been proposed as a potential strategy for preventing and/or treating pathological cardiac hypertrophy and heart failure [13, 14]. Our current understanding of the pathophysiology of cardiac hypertrophy as well as potential new therapies targeting oxidative stress will be reviewed herein.

2 Physiological Process of Ventricular Hypertrophy

It is generally accepted that mechanical stress begins the cascade of events that leads to ventricular remodeling. However, it is important to note that in vivo, there is rarely, if ever, an isolated increase in pressure independent of any neurohormonal activation. As a result, pressure overload involves mechanical as well as neurohormonal activation that then modulates the ventricular response to stress. Left ventricular hypertrophy begins with normal ventricle and myocyte function; ­however, with prolonged hypertrophy this situation eventually decompensates into a hypertrophied ventricle with corresponding myocyte dysfunction. These changes influence both structural and mechanical properties of the myocardial tissue and coronary circulation [1].

At the cellular level, concentric left ventricular hypertrophy is identified by an increase in cardiomyocyte cell size, parallel addition of new sarcomeres, and enhanced protein synthesis arising from alterations in gene transcription and translation [15]. In addition to changes to the cardiomyocyte, left ventricular hypertrophy is also characterized by proliferation of fibroblasts, increased generation of extracellular matrix proteins, and fibrosis [8, 16, 17]. This increased fibroblast growth in combination with the deposition of collagen and fibrosis leads to myocardial stiffening, which on its own is an important contributor to cardiac dysfunction [18, 19]. In the adult mammalian heart, α-myosin heavy chain (MHC) is the predominant isoform expressed and possesses a high ATPase activity and faster rate of contractility than the fetal β-MHC isoform. During hypertrophy, α-MHC expression is reduced, and β-MHC expression is increased which is characteristic of the re-induction of the fetal gene program [20]. Indeed, the transition from α- to β-MHC isoforms that occurs during pathological cardiac hypertrophy may play a role in the development of cardiac dysfunction by reducing myocardial contractility [19].

Generally there are three broad steps regarding the molecular mechanism of ventricular hypertrophy. Initially, heart remodeling is activated by biomechanical stress induced by increased pressure. The left ventricle is stretched during diastole and overloaded during systole which initiates both local myocardial and systemic neurohormonal responses. The mechanical stretch is sensed by myocyte membrane and sarcomere-coupled stretch receptors [21], whereas neurohormonal signals [i.e., endothelin-1 (ET-1), angiotensin-II (Ang-II), catecholamines] are sensed by G protein-coupled receptors that lead to activation of Gαq/11 proteins [22]. In the second stage of the hypertrophic process, these mechanical and neurohormonal stimuli activate intracellular signaling pathways which eventually transmit a signal to the myocyte nucleus [7, 21]. Finally these signaling pathways activate transcription factors to induce expression of particular genes in the cardiac myocyte which ultimately causes phenotypic changes in the myocytes as well as altered gene transcription that includes re-expression of the fetal gene profile [i.e., atrial natriuretic peptide (ANP), B-type natriuretic peptide (BNP)] and new cellular growth [21, 23]. Activation of specific pathways leads to beneficial and adaptive remodeling, while activation of other pathways eventually has maladaptive and harmful results [21, 24]. Ultimately the imbalance between maladaptive and beneficial signaling results in the development of pathological hypertrophy and the progression to heart failure. Since multiple molecular signaling pathways have been shown to be involved in the development of pathological hypertrophy, these pathways are becoming ­potential targets for cardiovascular treatments [25].

3 Transition from Compensated Hypertrophy to Heart Failure

As mentioned, the initial stimulus for hypertrophy may also stimulate maladaptive events in the cardiac tissue [2, 18, 26]. Several of the initial changes occurring at the level of the cardiomyocyte may be detrimental in left ventricular hypertrophy. For instance, the aforementioned re-expression of the fetal gene profile also induces a change in energy metabolism that results in enhanced glycolysis and a reduction in overall oxidative metabolism [27]. Rates of glycolysis are accelerated in the hypertrophied heart likely as an initial adaptive mechanism to increase ATP production in the presence of impaired oxidative metabolism [28–31]. As a result of this metabolic reprogramming, there is a decrease in myocardial ATP production, leading to energy deprivation [23, 32]. In addition, when glucose oxidation is not correspondingly increased along with glycolysis, there is an accumulation of protons and ROS [28, 33, 34]. Moreover, the reduction in fatty acid utilization itself may lead to lipid accumulation within cardiomyocytes, which has been shown to result in contractile dysfunction [35]. In addition to metabolic changes, apoptosis of cardiomyocytes is augmented in hypertensive heart disease, which leads to a reduction in contractile mass and affects contractility [36]. Other events which occur during hypertrophy such as collagen deposition may promote ventricular stiffness and diastolic dysfunction and heart failure [37–39]. As a result of these and a myriad of other changes that occur [1], it is apparent that myocardial hypertrophy in the setting of pressure overload is ultimately a maladaptive process that should be prevented or reversed in order to prevent the development of impaired cardiac contractile function and heart failure.

4 Oxidative Stress in Cardiac Hypertrophy and Heart Failure

Although the fundamental mechanisms underlying the development and ­progression of cardiac hypertrophy and heart failure are not fully understood, evidence from experimental models of heart failure and clinical studies demonstrates increased oxidative stress in the failing myocardium and strongly supports the concept that it plays a major role in the development of cardiac hypertrophy and progression to heart failure [40–43]. Oxidative stress is defined as the excessive production of ROS relative to the body’s natural antioxidant systems. ROS are a group of molecules including oxygen and its derivatives that are produced in all aerobic cells; this includes highly reactive free radicals such as superoxide anion (•O2 −) and hydroxyl radical (OH•), as well as compounds such as hydrogen peroxide (H2O2) that can be converted to reactive radical species [13]. Nitric oxide (NO) is an important molecule that regulates normal cardiac function by activating soluble guanylyl cyclase and synthesis of intracellular cGMP to inhibit activation of hypertrophic signaling. In the setting of oxidative stress, •O2 − can react with NO to form highly reactive ­species such as peroxynitrite, which has cytotoxic effects and further reduces NO bioavailability.

There are several enzymatic sources of ROS in mammalian cells including mitochondrial respiration, arachidonic acid pathway enzymes (lipoxygenase and cyclooxygenase), cytochrome p450s, xanthine oxidase, NADH/NADPH oxidases, nitric oxide synthase (NOS), peroxidases, and other hemoproteins [12]. The three key producers of ROS in the cardiovascular system are xanthine oxidase, NADH/NADPH oxidase, and NOS [12]. To combat ROS, the body has many specific and nonspecific antioxidant defense mechanisms that scavenge and degrade ROS to nontoxic molecules, including scavenging enzymes such as superoxide dismutase (SOD), glutathione peroxidase, and catalase as well as nonenzymatic antioxidants, such as vitamins E and C [44]. Under normal conditions, SOD converts •O2 − to H2O2, which is then further detoxified by catalase and glutathione peroxidase to H2O [13]. Excessive production of ROS causes cell dysfunction, protein and lipid peroxidation, and DNA damage—all of which lead to cell damage and death [12]. In addition, ROS appear to participate in several steps involved in the development and progression of myocardial remodeling and failure, including modification of proteins critical for proper contractile function [45], activation of hypertrophy signaling kinases or transcription factors [46] and apoptosis [47], as well as stimulation of cardiac fibroblast proliferation and activation of matrix metalloproteinases leading to extracellular matrix remodeling [48–50].

4.1 Cardiac Sources of Reactive Oxygen Species

Increased oxidative stress in the failing heart may be reflective of an increase in the production of ROS from many sources and also a deficiency in antioxidant capacity. The mechanisms responsible for increased oxidative stress in cardiac hypertrophy are unknown, and little is known about the mechanism by which stimuli such as mechanical strain, neurohormones, or cytokines lead to increased ROS. However, cardiac sources of ROS include cardiac myocytes, endothelial cells, and neutrophils. Within cardiac myocytes, ROS can be produced by mitochondria due to leakage of electrons from the respiratory chain which results in the conversion of oxygen to •O2 − [51]. Mitochondria from the failing heart produce more superoxide than normal mitochondria, suggesting that mitochondrial electron transport could be a predominant source of •O2 − production [52]. In addition, these mitochondria were associated with decreased complex enzyme activity, making mitochondria an important source of ROS in failing hearts and indicating a link between mitochondrial dysfunction and oxidative stress [52].

Regarding ROS production via oxidative enzymes, NADPH oxidase activity was found to be increased by stimuli relevant to the development of heart failure (i.e., mechanical stretch, Ang II, ET-1, tumor necrosis factor-α) [53]. This increased cardiac NADPH oxidase activity has also been observed in human heart failure [54]; increased xanthine oxidase expression and activity have been similarly noted [55]. In addition to these sources of ROS, uncoupled NOS has also been shown to produce ROS in a variety of pathological conditions [56, 57]. Under normal conditions, endothelial NOS (eNOS) also known as NOS3 catalyzes the formation of NO from l-arginine and oxygen. Uncoupling of NOS due to absence of the necessary cofactor tetrahydrobiopterin (BH4), for example, produces •O2 − which is known to contribute to the progression of atherosclerosis and hypertension [58, 59]. eNOS is also uncoupled and figures importantly in pathological remodeling and progression to heart failure in response to chronic pressure overload in mice [60].

In addition to being a potential source of ROS during cardiac hypertrophy, ­mitochondria themselves can also be damaged by ROS. As ROS damage mitochondrial components, the •O2 − produced within the mitochondria is unable to pass through the membranes, causing still more damage to this organelle, thus compromising mitochondrial function [61]. Increased ROS generation in failing hearts was associated with mitochondrial damage and dysfunction with increased lipid peroxidation in the mitochondria and reduced oxidative capacity [62]. This establishes a vicious cycle in which mitochondrial functional decline promotes further ROS generation, more mitochondrial damage, and cellular injury.

5 Hypertrophic Signaling Pathways

A number of complex signaling cascades have been identified that regulate the hypertrophic process in the heart. There appear to be distinct signaling cascades that distinguish physiological versus pathological hypertrophy (see [63] for review of physiological hypertrophy). At a molecular level, pathological cardiac hypertrophy is mediated by several molecular growth signals and induction of fetal gene expression programs (for proteins involved in cardiac contractility and calcium handling) [22, 63]. Some of the better characterized signaling pathways involved in the development of pathological cardiac hypertrophy will be discussed in the section below, with a focus on those pathways where ROS have been shown to be important in activation of these signaling pathways. Indeed, ROS are known to regulate the activity of a variety of signaling kinases and transcription factors to mediate the cardiac hypertrophic response, including calcineurin–NFAT, mitogen-activated protein kinases (MAPKs), and AMP-activated protein kinases (AMPK) among others [63].

5.1 Calcineurin–NFAT Signaling

Calcineurin is a ubiquitously expressed calcium (Ca2+)–calmodulin-regulated serine/threonine phosphatase that consists of a catalytic A and regulatory B subunit. In response to sustained increases in intracellular Ca2+ levels, Ca2+/calmodulin complexes bind to the B subunit, inducing a conformational change allowing calcineurin to dephosphorylate downstream effector proteins, most notably the members of the nuclear factor of activated T-cells (NFAT) family of transcription factors [64]. NFAT transcription factors are normally sequestered in the cytoplasm; however, upon calcineurin-mediated dephosphorylation of NFAT, these proteins rapidly translocate into the nucleus and activate gene expression [65, 66]. Calcineurin activation is observed in hypertrophied hearts following pressure overload induced by constriction of the abdominal aorta [67]. Moreover, partial inhibition of calcineurin by nonspecific immunosuppressive drugs cyclosporin A (CsA) and FK506 has been shown to prevent hypertrophy in some studies [67, 68].

Using a more targeted genetic approach, several transgenic mice have been generated and characterized in order to better define the role of calcineurin in the development and progression of cardiac hypertrophy. Mice expressing a constitutively active form of calcineurin in the heart are sufficient to induce cardiac hypertrophy (two- to threefold increase in heart size) and the expression cardiac fetal hypertrophic genes (i.e., BNP, β-MHC), a phenotype which then rapidly progresses to dilated cardiomyopathy, interstitial fibrosis, and heart failure [66]. Furthermore, targeted inhibition of calcineurin by cardiac overexpression of the inhibitory domains of Cain/Cabin-1 and A-kinase anchoring protein (AKAP) 79, which are endogenous calcineurin inhibitors, reduced cardiac hypertrophy in response to both isoproterenol infusion and pressure overload [69].

Calcineurin activity is negatively regulated by myocyte-enriched calcineurin-interacting protein (MCIP1), which is highly expressed in striated muscle and capable of binding and inhibiting the activity of calcineurin [70]. The cardiac hypertrophic response induced by aortic banding was significantly blunted in transgenic mice overexpressing the calcineurin inhibitory domain from MCIP1 [71]. Mice expressing either a cardiac-specific dominant-negative mutant of calcineurin [72] or a deletion of calcineurin Aβ [73] both displayed significantly less cardiac hypertrophy following aortic banding. Lastly, similar protection from pressure-overload-induced hypertrophy is observed in mice with targeted disruption of NFATc3, a downstream transcription factor effector of calcineurin-mediated hypertrophy [74]. Collectively these different animal models provide strong evidence that the calcineurin–NFAT pathway plays a fundamental role in regulating sustained hypertrophic growth of the heart. Moreover, it is thought that activation of the calcineurin–NFAT pathway is specific to pathological hypertrophy, such as that produced by hypertension or aortic constriction, and does not contribute to the development of physiological cardiac hypertrophy (i.e., exercise), which does not lead to deleterious consequences, thus creating a distinction between signaling pathways responsible for the two different types of cardiac hypertrophy [64].

Alterations in calcium homeostasis, as a result of changes in expression and/or activity of Ca2+ handling proteins, are a common phenomenon observed in the setting of cardiac hypertrophy and heart failure [75]. Since cardiac contractile function is highly dependent upon intracellular Ca2+ concentration, modifications in Ca2+ handling proteins can have a profound effect on cardiac contractility and function [76]. Oxidative stress plays a major role in the indirect regulation of intracellular Ca2+ homeostasis and in general is thought to lead to a rapid increase in cytosolic Ca2+ ­levels. Both the sarcoplasmic reticulum ATPase (SERCA2), an ATP-dependent Ca2+ pump that transports Ca2+ from the cytoplasm into the endoplasmic/sarcoplasmic reticulum, and the ryanodine receptor (RyR) SR Ca2+ release channels are redox sensitive [45]. ROS have been shown to reduce expression and/or activity of the SERCA2A [77] and increase the activation of the RyR [78]. Indeed, SERCA protein expression and activity are found to be decreased in pressure-overload-induced cardiac hypertrophy [79]. As well, oxidants have been shown to stimulate reverse mode of the sarcolemmal sodium–calcium exchanger (NCX) to facilitate Ca2+ entry into the cell [45]. Therefore, together these mechanisms may lead to an overall increase in intracellular Ca2+ levels that then may result in activation of the calcineurin–NFAT signaling pathway to trigger hypertrophy of the myocardium [80].

Since the cardiac myocyte is constantly beating with cyclic Ca2+ transients mediating excitation–contraction coupling, it is likely that Ca2+-dependent activation of the calcineurin–NFAT pathway is more complex than in other cell types where sustained increases in total intracellular Ca2+ are sufficient to trigger activation of this pathway [81]. More recently, it has been proposed the existence of specialized cellular microdomains where Ca2+ concentration is locally regulated and sensed by Ca2+-dependent proteins localized within these microdomains [81]. Transient receptor potential (TRP) channels, which are G protein-coupled receptor (GPCR)-operated Ca2+ channels that are located on the sarcolemma and membrane of the SR/ER, are now emerging as potential key regulators of Ca2+ entry during hypertrophy [82]. Indeed, TRP channel expression and activity are upregulated in pathological hypertrophy and heart failure [83–85]. Transgenic mice overexpressing various forms of TRP channels (i.e., TRPC3 and TRPC6) display marked ventricular hypertrophy at baseline and are more sensitive to pressure-overload-induced hypertrophy, which is associated with increased calcineurin–NFAT activity [85, 86]. Moreover, targeted disruption of the calcineurin Aβ gene blocked the exaggerated hypertrophic response in TRPC3 transgenic mice when subjected to TAC [86], supporting a link between TRPC and activation of the calcineurin–NFAT pathway. ROS generated during pressure-overload hypertrophy may also contribute to activation of TRP channels in this pathological setting [87]. Collectively these data suggest that TRP channels may play an important role in mediating pathological cardiac hypertrophy, likely by increasing local Ca2+ concentrations in microdomains resulting in activation of the calcineurin–NFAT signaling pathway and subsequent gene expression.

5.2 Mitogen-Activated Protein Kinases (MAPKs)

In general, mitogen-activated protein kinases (MAPKs) are divided into three classes based on the terminal kinase in the pathway: the extracellular signal-­regulated kinases (ERKs), the c-Jun amino-terminal kinase (JNKs), and the p38 MAPKs [88–90]. There is evidence for activation of all three families of MAPKs in cultured cardiac myocytes in response to hypertrophic stimuli (i.e., mechanical stress, GPCR agonists, Ca2+) [91–93], in the experimental pressure-overloaded heart [94, 95], and in the human failing heart [96]. Although these MAPKs have been largely considered to be pro-hypertrophic, the precise role of each of these MAPKs in the pathophysiology of cardiac hypertrophy and whether they are necessary mediators and/or modulators of the hypertrophic process remains to be clearly identified. However, early during the development of cardiac hypertrophy, it has been thought that ROS may contribute to activation of each these MAPK signaling pathways [97]. As these kinases are considered pro-hypertrophic, antioxidant therapies that target these signaling pathways may also prove to be advantageous in the treatment of pathological cardiac hypertrophy and the progression to heart failure.

5.3 Extracellular Signal-Regulated Kinases (ERKs)

ERK1/2 are ubiquitously expressed protein kinases that are activated in the setting of pathological cardiac hypertrophy and heart failure [63, 90, 98, 99]. In cultured cardiac myocytes, ERK1/2 are activated in response to pro-hypertrophic agonist stimulation, oxidative stress, or mechanical loading [91, 97, 100, 101]. Furthermore, activation of ERK 1/2 is required for increased protein synthesis in isolated cardiac myocytes in response to hypertrophic stimuli, such as ET-1 and α-adrenergic agonists [102]. Consistent with these in vitro findings, expression of a dominant-negative mutant of Raf-1, an upstream kinase of Erk1/2, inhibited the activation of ERK1/2 and blunted development of pressure-overload-induced cardiac hypertrophy [103]. Interestingly, mice with expression of cardiac-specific constitutively active MEK1, an immediate upstream MAPK kinase of ERK1/2, led to the development concentric physiological hypertrophy associated with enhanced cardiac systolic function without decompensation over time [104]. Furthermore, neither a global deletion of ERK1 or ERK1 −/− and ERK2 +/− mice led to a reduction in cardiac hypertrophy in mice subjected to TAC [105], suggesting that ERK1/2 may be sufficient, but not critical for mediating pathological pressure-overload-induced cardiac hypertrophy.

5.4 c-Jun Amino-Terminal Kinase (JNKs)

The JNK family consists of at least ten isoforms derived from three mammalian genes: JNK1, JNK2, and JNK3 [106]. JNK is rapidly activated in vitro in response to multiple cellular stresses, including mechanical stress, oxidative stress, and proinflammatory cytokines [106]. Activation of JNK is mediated via phosphorylation by MAPK kinase 4 (MEK4) and MEK7, which in turn are activated by MAPK kinase kinase (MEKK1) [107, 108]. Although several in vitro studies suggest that JNKs may contribute to the regulation of pathological hypertrophy [109, 110], data from in vivo studies have proven inconclusive. Studies in transgenic mouse models with blunted JNK activation have provided conflicting results with some showing that cardiac hypertrophy in response to pressure overload is either attenuated [111] or enhanced [112]. Moreover, TAC in mice with selective deletions of JNK1, JNK2, or JNK3 show that the cardiac hypertrophy developed to a similar extent as in wild-type mice [113], suggesting that either JNK isoforms perform similar functions and are redundant or that JNK activation may not be required for cardiac growth. At present, the exact role of JNK in the development of pathological cardiac hypertrophy remains unclear.

5.5 p38 MAPKs

The p38 MAPK family is an important regulator of a diverse array of biological functions including cell growth, cell proliferation, metabolism, and cell death [108, 114]. Of the four p38 isoforms, it appears that only p38α and p38β are expressed in the heart. Similar to JNK, p38 is a stress-activated protein kinase that is activated by a multitude of external stimuli including cytokines, oxidative stress, osmotic stress, and growth factors among others (see [63] for review). In vitro studies have largely supported a key role for p38 in promoting cardiac growth and hypertrophy with small molecular inhibitors of p38 and dominant-negative p38 adenovirus inhibiting hypertrophic growth [115]. Similar to the MAPKs described above, myocardial p38 activity is found to be increased by pressure overload [95] and ET-1/phenylephrine stimulation [116, 117]. Despite these findings, targeted activation of p38 in the heart by transgenic overexpression of upstream kinases MKK3 or MKK6 did not produce a significant degree of cardiac hypertrophy in the basal state. However, these mice did have increased interstitial fibrosis and ventricular wall thinning and died prematurely at 7–9 weeks as a consequence of heart failure [118], suggesting that p38 may be important in mediating late-stage ventricular remodeling as opposed to hypertrophy. Furthermore, studies in cardiac-specific p38 dominant-negative transgenic mice (DN-p38) showed that loss of p38 activity either had no effect on the development of cardiac hypertrophy [119] or enhanced the hypertrophic response [120] to pressure overload by aortic banding. In support of the latter finding, p38 MAPK has been shown to inhibit NFAT-transcriptional activity and nuclear translocation by directly phosphorylating NFAT [120, 121], suggesting that p38 may in fact be a negative regulator of hypertrophy. As several studies performed by different groups even in the same transgenic mice have given contradictory results, the precise role of p38 in the development and progression of cardiac hypertrophy and heart failure remains unclear.

5.6 AMP-Activated Protein Kinase (AMPK)

AMPK is a key metabolic sensing serine/threonine kinase that is activated by cellular and metabolic stresses that deplete ATP (i.e., ischemia). AMPK responds to increases in the AMP/ATP ratio by activating energy-producing metabolic pathways and inhibiting energy-consuming pathways [122]. The exact role of AMPK in cardiac hypertrophy has not yet been clearly defined. Protein translation and synthesis are known to be necessary mediators of increased myocardial cell size, and pharmacological activation of AMPK (i.e., AICAR, resveratrol) has been shown to inhibit protein synthesis associated with cardiac hypertrophy via numerous molecular pathways (see [122, 123] for review). Thus, while AMPK may act as a negative regulator of cardiac hypertrophy [124–127], it is not certain if inactivation of AMPK is a necessary step of the hypertrophic process or if it simply creates a permissive environment for cardiac growth. Supporting this idea, recent evidence has shown that impaired AMPK activity makes the heart more susceptible to pro-hypertrophic stimuli, such as hemodynamic overload [128–130]. In the SHR, increased oxidative stress [specifically the lipid peroxidation by-product, 4-hydroxy-2-nonenal (HNE)] leads to a reduction in AMPK activity via inhibition of its upstream activating kinase LKB1 [128], and this was associated with the development of cardiac hypertrophy. Consistent with studies in isolated myocytes, impaired AMPK activity was associated with activation of the mTOR/p70S6 kinase pathway that regulates protein synthesis in the heart [128]. Furthermore, cardiomyocyte-specific deletion of LKB1 results in left ventricular hypertrophy [131], suggesting that modification of LKB1 activity may contribute to the hypertrophic process. Although some studies have shown that the activation of AMPK is associated with the development of pressure-overload-induced cardiac hypertrophy [132], this is likely due to the heart becoming energetically compromised in the later stages of the disease and requiring AMPK to restore depleted ATP levels. Therefore, AMPK may play dual roles in the cardiac hypertrophic process whereby early in the disease, inactivation of AMPK may be necessary for cardiac growth, whereas activation of AMPK occurs during the later stages in order to maintain adequate ATP supply to the heart (see [122] for review).

5.7 JAK–STAT Pathway

Janus kinase (JAK) proteins are a family of cytosolic tyrosine kinases associated with the intracellular domain of membrane-bound receptors, which act to transduce signals from extracellular ligands such as cytokines, growth factors, and hormones to the nucleus to elicit cellular responses [133]. The JAK family of proteins rapidly transduces signals by recruitment of signal transducers and activators of transcription pathway (STAT) transcription factors. The JAK–STAT pathway plays a critical role in cardiac hypertrophy and the transition from hypertrophy to failure as well as mediates signal transduction from gp130 cytokine receptor [134] and GPCR [135] to the nucleus. Gp130 is a promiscuous receptor for several cytokines including interleukin 6/11 and transduces its signal mainly through induction of STAT3. Specifically, STAT3 is translocated to the nucleus in response to gp130 activation, which results in the induction of genes involved in hypertrophy [136]. Overexpression of STAT3 is sufficient to induce cardiomyocyte hypertrophy under both in vitro [137] and in vivo [138] settings. Transgenic mice with a deletion of gp130 in the myocardium have reduced STAT3 activity and display normal cardiac structure and function at baseline; however, these mice fail to develop compensatory hypertrophy following acute pressure overload and develop heart failure [139]. Therefore, the JAK–STAT pathway may be a novel therapeutic target for developing agents that prevent the development and/or progression of pathological cardiac hypertrophy.

The signaling pathways involved in cardiac hypertrophy are numerous and complex. The generation and characterization of transgenic rodent models have allowed investigators to better delineate the potential molecular mechanisms responsible for mediating distinct forms of cardiac growth. However, future research is still needed to clearly identify the pathways that are most critical for the development of pathological cardiac hypertrophy, and in doing so this may provide new targets for drug discovery in the management and treatment of cardiac hypertrophy and heart failure.

6 Antioxidants: Implications for Therapy

Since oxidative stress has been strongly implicated in the pathophysiology of cardiac hypertrophy induced by pressure overload, antioxidant therapies have received much attention as a potential therapeutic strategy to prevent and/or regress cardiac hypertrophy and/or prevent the transition to heart failure. To date, this therapeutic approach using antioxidants such as vitamin E and n-acetylcysteine, while having had some success in experimental animal models, has not successfully translated to the clinic for use in humans [140]. However, the natural polyphenol resveratrol that is found in a number of dietary food sources is emerging as a potential novel new therapy that may be a treatment option for those with cardiovascular disease, in particular cardiac hypertrophy.

6.1 Vitamin E

The antioxidant vitamin E is a naturally occurring lipid-soluble vitamin found in a variety of food sources, including nuts and sunflower seeds, and is a nonenzymatic part of the cell’s intrinsic antioxidant system to counter the accumulation of ROS [141]. Indeed, symptoms in humans with a vitamin E deficiency further support that vitamin E plays an important role in protecting membranes and the nervous system from oxidative stress [142]. Of the various forms of vitamin E, α-tocopherol is the most abundant and biologically active form of vitamin E, with the acetate and synthetic forms of α-tocopherol being the primary constituents of vitamin E supplements [141, 143]. Vitamin E is a potent peroxyl radical scavenger that breaks radical-propagated chain reactions and due to its lipid solubility is present in cell membranes and plays an integral role in protecting cell membranes and plasma lipoproteins from lipid peroxidation [142, 143]. Indeed, peroxyl radicals (ROO−) react at a rate 1,000 times faster with vitamin E than with polyunsaturated fatty acids [144].

In experimental models, treatment with vitamin E has been shown to reduce oxidative stress and prevent the development of cardiac hypertrophy and heart failure [145]. In the guinea pig model of pressure overload, chronic vitamin E treatment improved cardiac function and blunted the progression of heart failure over 20 weeks. Interestingly, however, this occurred in the absence of improvements in hypertrophy [145]. In contrast, clinical studies in humans have produced conflicting results regarding the cardiovascular benefit of vitamin E supplementation with some studies showing a significant reduction in cardiovascular risk [146–151], as well as a number of studies having failed to show a benefit to vitamin E supplementation [152–155]. Therefore, simply supplementing the antioxidant defenses of the heart using vitamin E may be insufficient to prevent cardiac hypertrophy and cardiovascular disease.

6.2 N-Acetylcysteine

N-acetylcysteine is a widely used thiol-containing antioxidant that is a pharmacological precursor of l-cysteine [156]. Reduced thiols are molecules with a sulfhydryl group whose biological properties include scavenging of oxygen free radicals, acting as cofactors for enzymatic reactions and potentiating the half-life and activity of NO by forming NO adducts (S-nitrosothiols) which are more stable than NO itself [157, 158]. Oral treatment with N-acetylcysteine (500 mg/kg/day) administered in the drinking water for 7 days led to a marked reduction in left ventricular weight induced by 2 weeks of abdominal aortic constriction in mice [159], which is consistent with a major role for ROS in the generation of pressure-overload-induced left ventricular hypertrophy in vivo. Interestingly, N-acetylcysteine administration (1.5 g/kg/day) to adult spontaneously hypertensive rats with established hypertension is both unable to reduce blood pressure and cardiac hypertrophy [160, 161]. Due to a lack of large-scale clinical trials, it is not clear whether N-acetylcysteine is effective in reducing cardiac hypertrophy in humans. However, N-acetylcysteine may be beneficial following myocardial infarction to limit infarct size [162, 163]. As well, N-acetylcysteine treatment has been shown to improve endothelial function and reduce systolic blood pressure in hypertensive diabetic patients via a reduction in oxidative stress and increased NO bioavailability [164]. This suggests that N-acetylcysteine can reduce oxidative stress in vivo and may be a useful antioxidant strategy to supplement current therapies in a variety of cardiovascular conditions.

7 Resveratrol as an Antioxidant

Resveratrol (3,5,4′-trihydroxystilbene) is a polyphenol that is found in a variety of berries, grapes, peanuts, and medicinal plants. Scientific interest in resveratrol has increased over the past 15 years since it has been shown to be a calorie-restriction mimetic and increase overall health in mammals [165]. Of relevance, resveratrol also appears to prevent the development of pressure-overload-induced cardiac hypertrophy [128, 166, 167] as well as show benefit in a variety of other cardiovascular diseases [168]. Indeed, resveratrol has many properties which are considered the basis of its cardiovascular effects and protection. Some of these include antioxidant, •O2 − scavenging, ischemic preconditioning, and angiogenic actions [169]. The exact mechanism of action of resveratrol in vivo has not been unequivocally determined; however, many cell components and processes are thought to be involved such as surface receptors, signaling pathways, metabolic pathways, nuclear receptors, and gene transcription and translation [170]. The antioxidant and •O2 − scavenging properties of resveratrol are of interest as they directly relate to the potential therapeutic benefits of this compound to reduce oxidative stress in the setting of cardiac hypertrophy. Some experiments suggest that resveratrol can directly scavenge •O2 − in potassium superoxide and xanthine oxidase-based antioxidant systems [171], with added evidence of direct inhibition of xanthine oxidase. Resveratrol has also been shown to inhibit NADPH oxidase and subsequently reduce •O2 − production as a mechanism for vasorelaxation in isolated rat aorta [172]. Additional evidence supports direct scavenging of ROS by resveratrol [173, 174] as well as increased expression of antioxidant enzymes such as SOD, catalase, and glutathione peroxidase [174–176]. Thus a key protective mechanism of resveratrol appears to be an augmentation of endogenous antioxidant systems in addition to direct inhibition/scavenging of ROS to counter the characteristic increase in oxidative stress during the development of cardiac hypertrophy.

In vivo experiments suggest that resveratrol is indeed a potent antioxidant via upregulation of NOS and increased NO production or by modulation of thioredoxin and heme oxygenase systems [177]. In fact, NO may be an important component in resveratrol-induced changes in intracellular redox state because NO has a greater affinity for •O2 − radicals compared to superoxide dismutase and thus can rapidly lower •O2 − concentrations [178]. In support of this concept, platelet eNOS was activated at physiologically achievable doses of resveratrol and blunted the proinflammatory pathway linked to p38 MAPK to inhibit production of ROS [179]. The proposed mechanism for increased NO production by resveratrol is thought to involve activation of the sirtuin SIRT1 protein deacetylase [180] which in turn has been shown to deacetylate eNOS at lysine residues to stimulate NO production [181, 182]. Related to this, resveratrol also activates AMPK [165, 183], which can then directly phosphorylate eNOS to stimulate NO production [184, 185]. As NO possesses vasodilatory, anti-inflammatory, anti-hypertrophic properties [186], increasing NO production may be an important pathway by which resveratrol ameliorates pathological cardiac hypertrophy.

As mentioned, mitochondria contribute importantly to the increased oxidative stress in cardiac hypertrophy. Interestingly, resveratrol has also been shown to attenuate mitochondrial oxidative damage [187], which could be another potential mechanism by which this compound is cardioprotective. Resveratrol-induced overexpression of SIRT1 attenuates mitochondrial oxidative damage in endothelial cells [188, 189], suggesting that prevention of mitochondrial oxidative damage alone could be an indirect mechanism for reduction of ROS by resveratrol. Furthermore, resveratrol also reduces mitochondrial •O2 − production in many cell types, including human coronary arterial endothelial cells which have been attributed to direct stimulation of mitochondrial antioxidant systems [189].

8 Modulation of Cell Signaling by Resveratrol

Although a currently accepted mode of action of resveratrol is as an antioxidant, this polyphenol also triggers other mechanistic pathways, which include modulation of cell signaling, apoptosis, and gene expression [190]. For example, resveratrol modulates phorbol-ester-induced signal transduction pathways, which leads to elevated COX2 expression [191] as well as other signals such as NFkB, MAP kinases, transcription factor activating protein-1 (AP-1), and ERK [192]. In addition, resveratrol interferes with many intracellular signaling pathways that regulate cell survival and apoptosis [193].

With specific relevance to cardiac pathophysiology, it is known that resveratrol reduces phenylephrine-induced hypertrophy of rat cardiac myocytes through the activation of AMPK in an AMPK kinase, LKB1-dependent manner [194]. Resveratrol acts as a negative regulator of cardiac hypertrophy via inhibition of the mTOR–p70S6 kinase protein synthesis pathway [195, 196]. Furthermore, resveratrol has been shown to inhibit calcineurin activity, NFAT translocation to the nucleus, and NFAT-dependent transcription in phenylephrine-induced hypertrophy of isolated rat cardiac myocytes [194]. As the calcineurin–NFAT signaling pathway is a major contributor to the development of pathological hypertrophy, resveratrol’s ability to suppress activation of this pathway may be an important mechanism by which resveratrol mediates its anti-hypertrophic effects.

As discussed, it is widely accepted that oxidative stress due to excessive production of ROS is a prominent factor in triggering the events that result in cardiomyocyte death. Cardiac SIRT1 is the mammalian ortholog of the silent information regulator 2 (Sir 2) family and is upregulated in response to oxidative stress [197]. Resveratrol has also been shown to be an important activator of SIRT1 [198], and in neonatal rat ventricular cardiomyocytes with simulated ischemia–reperfusion injury, resveratrol-induced SIRT1 activation and overexpression protected cardiomyocytes from oxidative injury, mitochondrial dysfunction, and cell death [198]. Resveratrol-induced SIRT1 overexpression in turn affects the MAPK pathway by reducing p38 and JNK phosphorylation [198]. In a porcine coronary artery preparation, resveratrol treatment countered ET-1 enhancement of MAPK activity by directly inhibiting MAPK activity and similarly reducing JNK-1 and p38 phosphorylation, as well as phosphorylation of ERK 1/2 was also reduced [199]. In support of this finding, resveratrol pretreatment protected against oxidative stress (H2O2)-induced cell proliferation and ERK 1/2 activation in human coronary smooth muscle cells [200]. In addition, resveratrol strongly inhibited Ang-II-induced hypertrophy and produced dose-dependent reductions in Akt 1 protein kinase and ERK 1/2 phosphorylation (two mediators thought to be essentially involved in Ang-II-mediated hypertrophy) [201].

Although activation of the p38 MAPK pathway is thought to contribute to the development of cardiac hypertrophy, recent but limited studies support a beneficial role for resveratrol-mediated activation of this pathway [202]. In H9c2 embryonic rat heart-derived cells treated with H2O2 to induce oxidative stress, resveratrol protected cells from oxidative damage, increased autophagy, and reduced peroxide-induced apoptosis [202]. All of these protective effects of resveratrol were prevented by prior treatment with a p38 MAPK inhibitor [202]. Overall resveratrol protected these cells from oxidative stress by upregulating autophagy via the p38 MAPK pathway [202]. Altogether these studies emphasize that the cellular actions of resveratrol are varied and dependent on the particular tissue or system under study (Fig. 14.1).

Fig. 14.1
figure 00141

Schematic representation of multiple cell signaling events modulated by resveratrol during the development of cardiac hypertrophy. Resveratrol affects several signaling events, many of which lead to a reduction in ROS and ultimately ameliorate cardiac hypertrophy. NFAT nuclear factor of activated T-cells, mTOR mammalian target of rapamycin, AMPK AMP-activated protein kinase, eNOS endothelial nitric oxide synthase, NOS nitric oxide synthase, NO nitric oxide, ROS reactive oxygen species, SIRT1 sirtuin (silent mating type information regulation 2 homolog) 1, p38: p38 mitogen-activated kinase, JNK: c-Jun N-terminal kinases, ERK extracellular signal-­regulated kinase

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9 Therapeutic Applications for Resveratrol in Cardiac Hypertrophy

To complement direct antioxidant effects which could attenuate oxidative stress and the progression of hypertrophy, resveratrol also modulates other aspects of the pathophysiology of cardiac hypertrophy. As increased arterial pressure is often an initiating factor for cardiac remodeling, the ability of resveratrol to prevent increases in blood pressure in vivo is protective against left ventricular hypertrophy [203]. In several rat models of hypertension, resveratrol prevented left ventricular hypertrophy due to reduced systolic blood pressure [204–206]. Resveratrol also appears to have direct effects on cardiac myocyte growth since it can prevent cardiac hypertrophy in the absence of changes in blood pressure in spontaneously hypertensive rats or rats subjected to abdominal aortic banding as a model of pressure overload [128, 166, 167, 207]. This direct effect of resveratrol on cardiomyocytes is also supported by the modulation of signaling pathways that regulate growth and protein synthesis [194, 195]. Resveratrol also protects against other hypertrophy-induced events such as preventing the reduction in eNOS and iNOS [207] and presumably preventing a subsequent reduction in NO production. In addition, resveratrol inhibits critical steps in cardiac collagen deposition such as cardiac fibroblast proliferation which is thought to involve the cardiac NO–cGMP signaling pathway [196, 208]. Overall, evidence demonstrates that resveratrol protects cardiomyocytes from oxidative stress and death via ROS reduction, increased expression of antioxidant enzymes, and mitochondrial protection [209, 210], with ultimate reduction of fibrosis and preservation of cardiac function and survival in rodent models of heart failure [211].

10 Clinical Uses of Resveratrol

Although experimental models suggest that resveratrol may be an effective therapy for cardiac hypertrophy, to date there is a paucity of published human trials in this area. One study in healthy humans showed that a supplement which included 100 mg of resveratrol reduced the oxidative and inflammatory responses normally induced by a high-fat, high-carbohydrate meal [212]. A recent study focused on patients after myocardial infarction who were given a 10 mg/day resveratrol supplement for 3 months. In this study, resveratrol appeared to improve left ventricular diastolic function and endothelial function as measured by flow-mediated dilation of the brachial artery with no change in blood pressure [213]. However, despite the limited published evidence in humans, there are several clinical trials underway with this compound, and resveratrol may be a promising new therapeutic strategy for the treatment of cardiac hypertrophy and heart failure.

11 Conclusion

A thorough grasp of the pathophysiological mechanisms underlying the ­development of cardiac hypertrophy and the progression to heart failure is essential considering that left ventricular hypertrophy is a significant independent risk factor for sudden cardiac death, dysrhythmia, heart failure, ventricular ischemia, and coronary heart disease [214]. Although the specific steps that lead to hypertrophy are not completely clear, it appears that oxidative stress does play an important role in this condition. It is also promising that a treatment approach involving antioxidant ­therapies, including the natural polyphenol resveratrol, appears to target several signaling and pathogenic events in the etiology of this condition and may well prove to be effective in the prevention and treatment of cardiac hypertrophy alone or in combination with other drugs. Although we have a great deal of evidence supporting resveratrol treatment of hypertrophy, this is largely in cell-based and animal models with limited evidence from human studies—particularly focused on oxidative stress and cardiac function. Therefore, this is a critical and undoubtedly fruitful avenue of research in the future that may have significant clinical implications.