ZINC AND THE PROOXIDANT HEART FAILURE PHENOTYPE

Abstract

Neurohormonal activation with attendant aldosteronism contributes to the clinical appearance of congestive heart failure (CHF). Aldosteronism is intrinsically coupled to Zn²⁺ and Ca²⁺ dyshomeostasis, in which consequent hypozincemia compromises Zn²⁺ homeostasis and Zn²⁺-based antioxidant defenses that contribute to the CHF prooxidant phenotype. Ionized hypocalcemia leads to secondary hyperparathyroidism with parathyroid hormone-mediated Ca²⁺ overloading of diverse cells, including cardiomyocytes. When mitochondrial Ca²⁺ overload exceeds a threshold, myocyte necrosis follows. The reciprocal regulation involving cytosolic free [Zn²⁺]_i as antioxidant and [Ca²⁺]_i as prooxidant that can be uncoupled in favor of Zn²⁺-based antioxidant defenses. Increased [Zn²⁺]_i acts as a multifaceted antioxidant by: i) inhibiting Ca²⁺ entry via L-type channels and hence cardioprotectant from the Ca²⁺-driven mitochondriocentric signal-transducer-effector pathway to nonischemic necrosis; ii) serving as catalytic regulator of Cu/Zn-superoxide dismutase; and iii) activating its cytosolic sensor, metal-responsive transcription factor that regulates the expression of relevant antioxidant defense genes. Albeit present in subnanomolar range, increased cytosolic free [Zn²⁺]_i enhances antioxidant capacity that confers cardioprotection. It can be achieved exogenously by ZnSO₄ supplementation or endogenously, using a β₃ receptor agonist, (e.g., nebivolol) that enhances NO generation to release inactive cytosolic Zn²⁺ bound to metallothionein. By recognizing the pathophysiologic relevance of Zn²⁺ dyshomeostasis in the prooxidant CHF phenotype and by exploiting the pharmacophysiologic potential of [Zn²⁺]_i as antioxidant, vulnerable cardiomyocytes under assault from neurohormonal activation can be protected and the myocardium spared from adverse structural remodeling.

Introduction

Zinc is an essential micronutrient with crucial biologic functions in both contractile and noncontractile cells. Zn²⁺-based regulatory actions can be categorized as catalytic, structural and regulatory (1). For example, Zn²⁺ catalyzes the activity of matrix metalloproteinases and angiotensin-converting enzyme, metalloenzymes expressed by inflammatory cells and myofibroblasts at sites of tissue repair. Zn²⁺ finger proteins serve as coordination complexes essential for the control of transcriptional and translational responses in which diminished [Zn²⁺]_i leads to protein misfolding and impaired function. Cytosolic free [Zn²⁺]_i in cardiomyocytes regulates the expression of its intracellular sensor, metal-responsive transcription factor (MTF)1.

Hypozincemia of varying severity, based on normal control values (75–140 μg/dL), has been widely reported in patients with congestive heart failure (CHF) irrespective of its etiologic origins (2–5). The most relevant underlying pathophysiologic mechanism for hypozincemia is renin-angiotensin-aldosterone system (RAAS) activation with renin-dependent aldosteronism, where a marked increase in fecal and urinary Zn excretion occurs and which can be blocked by an aldosterone receptor antagonist (6). Other factors contributing to hypozincemia in CHF include urinary Zn²⁺ losses associated with angiotensin-converting enzyme (ACE) inhibitor treatment (7) and reduced dietary Zn²⁺ intake and/or impaired small intestine absorption of Zn²⁺ (8). Resultant Zn²⁺ deficiency contributes to the systemic illness of CHF and its prooxidant phenotype and where increasing cardiomyocyte [Zn²⁺]_i functions as a multifaceted antioxidant to confer cardioprotection.

[Zn²⁺]_i contributes to the regulation of antioxidant defense genes by activating MTF1. Upon its translocation to the nucleus, MTF1 upregulates: metallothionein (MT)1, a Zn²⁺ binding protein and oxygen radical scavenger; glutathione synthase; and Cu/Zn-superoxide dismutase (SOD), a metalloenzyme and antioxidant (9, 10). [Zn²⁺]_i also plays a critical role as an endogenous competitive inhibitor to Ca²⁺ entry via L-type Ca²⁺ channels. Other contributions of Zn²⁺ in maintaining cardiomyocyte redox equilibrium include its catalytic regulation of Cu/Zn-SOD and its pool of inactive cytosolic Zn²⁺, bound to MT1 cysteine clusters, which upon its release by eNOS-mediated action of nitric oxide provides for [Zn²⁺]_i-based defenses (11–13).

This review focuses on the hitherto less well-recognized aspects of Zn²⁺ dyshomeostasis in the CHF prooxidant phenotype and where increasing [Zn²⁺]_i serves as antioxidant and cardioprotectant. The role of [Zn²⁺]_i as an antioxidant in preventing ischemia/reperfusion injury and diabetic cardiomyopathy has been reported elsewhere (14–16).

PATHOPHYSIOLOGY OF CHF

CHF: A Prooxidant Phenotype

Neurohormonal Activation

The symptoms and signs of the CHF syndrome have their origins rooted in neurohormonal activation, where endocrine properties of circulating effector hormones of the RAAS and adrenergic nervous (ANS) systems orchestrate an unwanted salt-avid state. Another deleterious outcome to such inappropriate hormonal activation is a systemic illness, in which persistent circulating hormones affect such diverse tissues as skeletal muscle, bone and peripheral blood mononuclear cells (i.e., lymphocytes and monocytes) to adversely influence their behavior. It is the concerted endocrine actions of RAAS and ANS hormones that account for this systemic illness, one of whose major features is a prooxidant phenotype (17) in which the rate of reactive oxygen species generation in diverse tissues overwhelms their rate of detoxification at these sites by endogenous antioxidant defenses that include crucial Zn²⁺-dependent enzymes.

Secondary Hyperparathyroidism (SHPT)

A contemporaneous dyshomeostasis of Zn²⁺, Ca²⁺ and Mg²⁺ is present on admission in patients hospitalized with decompensated biventricular failure having a dilated cardiomyopathy of ischemic or nonischemic origins (3–5, 18–20). This includes hypozincemia, ionized hypocalcemia and hypomagnesemia, together with abnormal elevations (>65 pg/mL) in plasma parathyroid hormone (PTH). SHPT accompanies the chronic aldosteronism of CHF, where PTH becomes a crucial mediator of pathologic remodeling. SHPT arises because of ionized hypocalcemia and hypomagnesemia, thus providing major stimuli to the parathyroids’ elaborated secretion of PTH (see Figure 1).

Figure 1.

Acute and chronic stressor states with excess catecholamines and aldosteronism, respectively, lead to hypozincemia, ionized hypocalcemia and hypomagnesemia with secondary hyperparathyroidism (SHPT), that provoke bone resorption and enhanced absorption of these cations from gut and kidney. Paradoxically, elevations in plasma parathyroid hormone (PTH) raise intracellular Ca²⁺ to induce oxidative stress and promote cardiomyocyte necrosis. See text. Adapted from Kamalov G, et al. J Cardiovasc Pharmacol. 2010;56:320–328.

SHPT is invoked to restore extracellular cation homeostasis through PTH-mediated resorption of bone Ca²⁺, Mg²⁺ and Zn²⁺ (21), while nephron-derived 1,25(OH)₂D₃ promotes their absorption from gut and kidneys (see Figure 1). PTH with its receptor-ligand binding property paradoxically leads to increased L-type Ca²⁺ channel activity and intracellular Ca²⁺ overloading involving various tissues (22–24). In the case of cardiomyocytes, adverse consequences of excessive intracellular Ca²⁺ accumulation include both cytosolic and mitochondrial domains. There follows the induction of oxidative stress by these organelles and ultimate necrosis of parenchymal cells with consequent tissue repair. Lost parenchyma and the appearance of microscopic scarring further compromise ventricular diastolic and systolic dysfunctions (25–29).

Oxidative Stress and Cardiomyocyte Necrosis

The failing heart is not spared the onslaught of circulating hormones (25–30). The myocardium undergoes a hormone-regulated structural and biochemical remodeling with progressive decline in contractile and diastolic functions (31, 32). This includes the ongoing prooxidant-based loss of cardiomyocytes and consequent appearance of reparative fibrosis that follows each bout of “dirty cell death,” or necrosis, to account for widely scattered foci of scarring found in the right and left heart. This contrasts to “sterile cell death,” or apoptosis, which does not evoke inflammatory cell-mediated tissue repair and, therefore, does not result in myofibroblast-based fibrous tissue formation (33).

Several sources account for the generation of reactive oxygen species in cardiomyocytes that includes superoxide and H₂O₂: membrane-bound NADPH oxidase; cytosolic-based xanthine oxidase; and subcellular organelles with the all-important role of subsarcolemmal mitochondria accounting for the altered redox state of the failing heart (9, 10, 34). Increased production of superoxide in the explanted failing heart, either ischemic or nonischemic origins, is coupled to increased activities of antioxidant defenses that include metalloenzymes: Cu/Zn- or Mn-based superoxide dismutases; and Se-dependent glutathione peroxidase (GPX) (35–37).

A Mitochondriocentric Pathway to Necrosis

As conceptually depicted in Figure 2, the early PTH-mediated Ca²⁺ overloading seen during wks 1–3 of aldosteronism is coupled to an induction of oxidative stress, expressed as H₂O₂ production, by subsarcolemmal mitochondria. Oxidative stress appears as the rate of reactive oxygen species (ROS) generation and overwhelms their rate of detoxification by the cumulative capacity of antioxidant defenses. Mitochondrial Ca²⁺ overloading is a determinant of irreversible cell injury in particular when it exceeds a threshold (38) and as depicted in Figure 2. Together with membrane depolarization and ATP depletion, there ensues a nonphysiologic cyclophilin D-based opening of mitochondrial inner membrane permeability transition pore (mPTP) and osmotic swelling with structural and functional degeneration. This pathophysiologic scenario triggers the pronecrotic phenotype with cell death. It represents a mitochondriocentric signal-transducer-effector pathway to nonischemic cardiomyocyte necrosis operative during either acute hyperadrenergic or chronic stressor states (see Figure 3) (39).

Figure 2.

The putative association between mitochondrial (mt) total [Ca²⁺]_m and their production of H₂O₂ during preclinical and pathologic stages of aldosteronism/salt treatment (ALDOST) when cardiomyocytes cross from prosurvival to pronecrotic phenotype. Necrosis occurs when a threshold in [Ca²⁺]_m has been surpassed. Actual data points are shown as closed circles.

Figure 3.

A mitochondriocentric signal-transducer-effector pathway to nonischemic cardiomyocyte necrosis. Aldosterone/salt treatment (ALDOST) and associated elevations in plasma PTH promote cytosolic free [Ca²⁺]_i and mitochondrial [Ca²⁺]_m overloading and ensuing oxidative stress. Together with lost membrane potential and ATP synthesis, a pathologic opening of the subsarcolemmal (SSM) mitochondrial permeability transition pore (mPTP) follows leading to degeneration of these organelles and cardiomyocyte necrosis with leakage of troponins. Adapted from Rutledge MR, et al. Cardiovasc Drugs Ther. 2013;27:161–170.

Necrosis is followed by the spillage of cell contents, including the leakage of troponins, which appear in the circulation as a biomarker confirmatory of necrosis. Proteins released by necrotic cardiomyocytes serve as “danger signals” activating the immune system (33).

A series of site-directed, sequential pharmacologic interventions targeted along cellular-subcellular cascades were carried out to block downstream events leading to PTH-mediated intracellular Ca²⁺ overloading, oxidative stress and mPTP opening to salvage cardiomyocytes (reviewed in 40).

Zn²⁺ DYSHOMEOSTASIS IN STRESSOR STATES

Zinc, the body’s sixth most abundant cation, is an essential micronutrient predominantly derived from dietary sources; it is present in most tissues, but stored at greater proportion in liver and skeletal muscle. Reduced antioxidant defenses can occur when dietary Zn intake is inadequate, excretory Zn²⁺ losses are excessive relative to intake, Cu/Zn-SOD is utilized in the dismutation of superoxide without adequate replenishment, or a combination of these scenarios (41).

Acute Stressor States

The importance of Zn²⁺ and Zn²⁺ deficiency was recognized by Prasad 50 years ago (42). In response to dietary Zn²⁺ deficiency plasma Zn²⁺ levels and tissue Zn²⁺ stores both decline and are associated with reduced tissue MT1, a cysteine-rich Zn²⁺-binding protein involved in Zn²⁺ homeostasis and free radical scavenging, together with reduced serum Cu/Zn-SOD activity. This contrasts to Zn²⁺ insufficiency seen with stressor states (43, 44). Rapid reductions in circulating Zn²⁺ occur in response to activation of the hypothalamic-pituitary-adrenal axis which accompanies acute stressor states, such as bodily injury, hyper- or hypothermia, acute myocardial infarction, or coronary artery bypass surgery (45–47). Hypozincemia can be reproduced by adrenocorticotropin hormone (ACTH) infusion. The degree of hypozincemia correlates with the severity of injury. Zn²⁺ is translocated to sites of injury and to liver for storage, facilitated by upregulated expression of MT1 (6, 21). In response to tissue injury involving heart, kidney or lung, where circulating Zn²⁺ levels decline while Zn²⁺ is redistributed to injured tissues based, in part, on MT1 upregulation, and where tissue Cu/Zn-SOD expression and activity are increased (48–51). At the site of injury, Zn²⁺ participates in wound healing, including: initiation of gene transcription and protein synthesis; immune cell replication and functions; and activation of metalloproteinases (6, 52). The hypokalemia and ionized hypocalcemia and hypomagnesemia seen on admission with acute hyperadrenergic states is based on catecholamine-mediated translocation of these cations to skeletal muscle and adipose tissue, respectively (reviewed in 53).

Chronic Stressor State

CHF is a chronic stressor state. The underlying pathophysiologic basis for hypozincemia in CHF is multifactorial. Most importantly is renin-dependent (or secondary) aldosteronism that accompanies RAAS activation, in which a marked increment in fecal and urinary Zn²⁺ excretion occurs (see Figure 1) and can be blocked by an aldosterone receptor antagonist (6). There is also marked excretory losses of Ca²⁺ and Mg²⁺ with ultimate appearance of ionized hypocalcemia and hypomagnesemia leading to SHPT (see Figure 1) (23, 54). Hyperparathyroidism itself raises urinary Zn²⁺ excretion (55, 56).

Other factors contributing to hypozincemia in CHF include: i) urinary Zn²⁺ losses associated with ACE inhibitor treatment (7); ii) reduced dietary Zn²⁺ intake and/or impaired small intestinal Zn²⁺ absorption (8); and iii) Zn²⁺ translocation for storage within the liver, muscle and bone with subsequent release for purposes of tissue repair, e.g., during recurrent episodes of myocyte necrosis (21, 52). Resultant Zn²⁺ deficiency contributes to the systemic illness of CHF by causing: i) prooxidant phenotype with oxidative stress-induced damage to cardiomyocytes, tissue DNA, proteins and lipids (57); and ii) an immunostimulatory state with activated immune cells elaborating proinflammatory cytokines.

Zn²⁺ TRAFICKING IN CARDIOMYOCYTES

Nano level quantitation and intracellular Zn²⁺ trafficking studies have made possible the advent of fluorescent proteins and sensor fluorescence energy transfer spectroscopy to monitor cytosolic [Zn²⁺]_i and nuclear [Zn²⁺]_i, together with total tissue Zn content by atomic absorption spectroscopy (54, 58, 59). Using a fluorescent dye, FluoZin-3, LeWinter et al. (60) identified Zn²⁺ entry via sarcolemmal Ca²⁺ channels to further implicate [Zn²⁺]_i as an endogenous competitive inhibitor of Ca²⁺ entry. Both extra- and intracellular cardiomyocyte Zn²⁺ are inhibitory of ATP-induced Ca²⁺ entry, including that which accompanies β-adrenergic receptor stimulation by catecholamines (61, 62). Pathophysiologic responses in [Zn²⁺]_i, as antioxidant, and [Ca²⁺]_i, as prooxidant, are depicted in Figure 4.

Figure 4.

An evolving schema of the pathophysiologic paradigm contributing to the dyshomeostasis of cardiomyocyte cytosolic free [Ca²⁺]_i and subsarcolemmal mitochondria [Ca²⁺]_m as prooxidant, together with [Zn²⁺]_i and [Zn²⁺]_m as antioxidant, during chronic aldosteronism with SHPT. Elevations in plasma PTH cause intracellular Ca²⁺ overloading via L-type Ca²⁺ channels (LTCC), and the induction of mitochondrial reactive oxygen species (mtROS) generation contributing to mitochondrial permeability transition pore (mPTP) opening and ensuing cardiomyocyte necrosis with scarring. Antioxidant defenses are activated to counteract the prooxidant state. These include: ROS-mediated or nebivolol β₃ adrenergic receptors (β₃R) binding-induced generation of nitric oxide (NO) from endothelial NOS (eNOS) producing cyclic guanosine monophosphate (cGMP) via guanylyl cyclase (GC) which inhibits LTCC Ca²⁺ entry. NO also promotes release of inactive cytosolic Zn²⁺ bound to metallothionein (Zn/MT) raising both [Zn²⁺]_i and [Zn²⁺]_m. The rise in [Zn²⁺]_i activates its sensor, metal-responsive transcription factor (MTF)-1 which, in turn, upregulates transcription of antioxidant defense genes. Furthermore, Zn²⁺ (Zip) transporters, induced by ROS, contribute to increased Zn²⁺ entry and consequent rise in [Zn²⁺]_i and [Zn²⁺]_m. Zn²⁺ supplement or Zn²⁺ ionophore will raise [Zn²⁺]_i, which regulates Cu/Zn-superoxide dismutase (SOD) activity. Adapted from Cheema Y, et al. J Cardiovasc Pharmacol. 2011;58:80–86.

More than 99% of total cardiomyocyte [Zn] is inactive and bound to MT1; upon its release from MT1 [Zn²⁺]_i rises (60). Nitric oxide (NO) mobilizes cardiomyocytes Zn²⁺ from MT1-thiol clusters raising [Zn²⁺]_i as an antioxidant in the attenuation of cardiac mitochondrial oxidative damage following reperfusion (14). The multifaceted antioxidant profile of nebivolol, a β₁ blocker, relates to its dual action as agonist of β₃ adrenergic receptors, which, in turn, activates eNOS to enhance NO generation and raise [Zn²⁺]_i, via its mobilization from the inactive MT-bound store (63).

Total Zn content of normal human heart is reported to be 20–30 mg/g wet tissue (64). This approximates our findings in normal rat and hamster myocardial tissue and reported as 82±3 and 79±5 ng/mg fat-free dried tissue, respectively (9, 59). The overall Zn content of cardiomyocytes is predominantly regulated by membrane-bound Zn²⁺ transporters: i) ZnT-based pumps which modulate Zn²⁺ efflux; and ii) Zip transporters that regulate Zn²⁺ influx (reviewed in 65). A lesser amount (≈30%) enters via L-type Ca²⁺ channels (9). Cytosolic [Zn²⁺]_i is further regulated by its sensor, MTF1. The rise in [Zn²⁺]_i seen with aldosteronism is facilitated by the upregulation of membranous Zn²⁺ importers and exporters (9). Because of this tight regulation of [Zn²⁺]_i, Zn²⁺ cytotoxicity is relatively uncommon.

Intrinsically Coupled Zn²⁺ and Ca²⁺ Dyshomeostasis

The dyshomeostasis of extra- and intracellular Zn²⁺ and Ca²⁺ that accompanies aldosteronism contributes to a deleterious but reversible dysequilibrium between them. Intrinsically coupled dyshomeostasis of intracellular Zn²⁺ and Ca²⁺ occurs spontaneously to regulate the integrated redox state of cardiomyocytes and mitochondria. These cations were monitored in their cytosolic free state using relevant dyes and fluorescence spectroscopy with ALDOST alone or in combination with either spironolactone or amlodipine cotreatment. As shown in Figure 5, aldosteronism was accompanied by increased total Zn and Ca in myocardial tissue (top panel), cardiomyocytes (middle panel), and in subsarcolemmal mitochondria (lower panel). Increased [Ca²⁺]_i and [Zn²⁺]_i, together with elevated subsarcolemmal mitochondrial [Ca²⁺]_m and [Zn²⁺]_m, could be prevented by spironolactone, acting to prevent SHPT, and significantly attenuated by amlodipine (9). These salutary iterations in divalent cation composition corroborated well with reduced biomarkers of oxidative stress in mitochondria and were coincident with increased activities of Cu/Zn-SOD and GPX (9, 39, 54). Adaptive modulation in [Zn²⁺]_i was accompanied by the contemporaneous upregulation of MT1, both a Zn²⁺ importer (Zip1) and exporter (ZnT-1), and MTF1.

Figure 5.

The responses to intrinsically coupled Zn²⁺ and Ca²⁺ dyshomeostasis that occur in chronic stressor state invoked by 4 wks ALDOST raise myocardial total Ca²⁺ and Zn²⁺ (top panel), cytosolic free cardiomyocyte [Ca²⁺]_i and [Zn²⁺]_i (middle panel), as well as mitochondrial total [Ca²⁺]_m and [Zn²⁺]_m (lower panel). *p<0.05 vs. control. Adapted from Kamalov G, et al. J Cardiovasc Pharmacol. 2009;53:414–423.

The temporal responses in coupled Ca²⁺ and Zn²⁺ dyshomeostasis, reflecting the tenuous prooxidant:antioxidant equilibrium, indicate elevations in [Ca²⁺]_i and [Ca²⁺]_m are coupled with [Zn²⁺]_i and [Zn²⁺]_m and accompanied by increased mitochondrial H₂O₂ production, cardiomyocyte xanthine oxidase activity, and cardiac and mitochondrial 8-isoprostane levels (9). This prooxidant state is initially counterbalanced by increased activity of antioxidant proteins, enzymes, and the nonenzymatic antioxidants, or cumulative antioxidant capacity, as well as by the augmented antioxidant defenses. However, these defenses ultimately prove inadequate in combating the persistent intracellular Ca²⁺ overloading and the consequential marked rise in cardiac tissue 8-isoprostane (39).

Uncoupling Zn²⁺ and Ca²⁺ Dyshomeostasis Confers Cardioprotection

Whether intrinsically coupled Ca²⁺ and Zn²⁺ dyshomeostasis and prooxidant:antioxidant dysequilibrium (9) seen with aldosteronism could be uncoupled in favor of antioxidants was examined using cotreatment with a ZnSO₄ supplement, pyrrolidine dithiocarbamate (PDTC), a Zn²⁺ ionophore (see Figure 4), or ZnSO₄ in combination with a Ca²⁺ channel blocker, amlodipine (10). Cotreatment with ZnSO₄ alone, PDTC alone, or ZnSO₄ plus amlodipine augmented the rise in cardiomyocyte [Zn²⁺]_i beyond that seen with aldosteronism, while attenuating the rise in [Ca²⁺]_i which together served to reduce biomarkers of oxidative stress in cardiomyocytes and their mitochondria that proved to be cardioprotective (10, 30, 54). ZnSO₄ supplement alone, however, does not reverse hypocalcemia and hypomagnesemia (54). Hence there is a need for a polycation supplement to correct the deficits of Zn²⁺, Ca²⁺ and Mg²⁺ that arise because of their excessive excretory losses.

The extent to which the coupled dyshomeostasis between Zn²⁺ and Ca²⁺ of aldosteronism could be uncoupled by ZnSO₄ supplement or Zn²⁺ ionophore was far exceeded by nebivolol cotreatment (63, 66). The reciprocal regulation between [Zn²⁺]_i and [Ca²⁺]_i seen in ALDOST alone, ALDOST plus ZnSO₄, and ALDOST plus nebivolol cotreatment is depicted in Figure 6. The increment in [Zn²⁺]_i seen with nebivolol related to its binding to β₃ receptors with activation of eNOS and NO generation that releases intracellular Zn²⁺ bound to MT1. This marked rise in [Zn²⁺]_i seen with nebivolol was associated with a nearly complete abrogation to the rise in [Ca²⁺]_i and accompanied by marked reduction in H₂O₂ production by mitochondria and abrogation in biomarkers of lipid peroxidation by these organelles and in cardiac tissue (63). The degree of cardioprotection seen with nebivolol was not found with another β₁ receptor blocker, atenolol, which is devoid of β₃ receptor binding (63, 67).

Figure 6.

An integrated reciprocal regulation between cytosolic free [Zn²⁺]_i and [Ca²⁺]_i is revealed by their percent increase in cardiomyocytes at: 4 wks ALDOST; ALDOST+ZnSO₄ supplement (10); and ALDOST+nebivolol (Neb) (63). Reprinted from Khan MU, et al. J Cardiovasc Pharmacol. 2013;62:445–451.

Zinc-Finger Transcription Factors

In addition to NO-based release of metallothionein-bound cytosolic Zn²⁺, it will also induce the release of Zn²⁺ residing within cell nuclei (13). This would suggest zinc-finger proteins are NO targets and that NO modulates zinc-finger transcription factors and thereby gene regulation (13).

Zinc-finger proteins recognize DNA sequences. They may therefore serve in drug discovery and therapeutics through their DNA binding properties to zinc-finger transcription factors (68). Engineered zinc-finger proteins could be used to regress disease-related genes in heart failure (69).

Clinical Correlates

SHPT has now been recognized as an independent predictor of decompensated heart failure and the need for hospitalization (70, 71). Moreover, PTH is emerging as an independent risk factor for mortality and cardiovascular events in community-dwelling individuals (72–74). The prognostic significance of PTH is independent of glomerular filtration rate (75, 76).

Hypozincemia has been widely reported in patients with CHF irrespective of whether they had an ischemic or idiopathic cardiomyopathy, or had been in sinus rhythm or atrial fibrillation (2–5). In patients who had gastric bypass surgery for the treatment of morbid obesity, chronic deficiencies of Zn²⁺ and Se²⁺ are common, together with the appearance of a dilated cardiomyopathy with reduced cations and activity of glutathione peroxidase (GPX) in the myopathic heart (77). Six months’ treatment with supplemental Zn²⁺ and Se²⁺, together with standard of care, resulted in normalized tissue cations and GPX activity and improved ventricular function. Supplemental Zn²⁺ reduces oxidative stress and chronic inflammation in chronic diseases (42). It prevents myocardial injury that accompanies: a hyperadrenergic state (78, 79); streptozocin-induced diabetes (80); and ischemia/reperfusion (15, 81). Likewise, supplemental Se²⁺ reverses a dilated cardiomyopathy known to be associated with dietary Se²⁺ deficiency (82). The minimal daily requirement for Zn²⁺ is 8–10 mg. As a supplement for acute or chronic Zn²⁺ deficiency, 50–100 mg/day ZnSO₄ is suggested (83).

SUMMARY AND CONCLUSIONS

The neurohormonal activation that accompanies CHF includes chronic aldosteronism with secondary hyperparathyroidism leading to intrinsically coupled Zn²⁺ and Ca²⁺ dyshomeostasis. This includes their excessive excretory losses that account for hypozincemia and ionized hypocalcemia. Zn²⁺ losses are also potentiated by ACE inhibitor treatment. Coupled with reduced dietary Zn²⁺ intake, a Zn²⁺ deficiency arises to compromise antioxidant defenses and predispose to the mitochondriocentric signal-transducer-effector pathway with PTH-mediated Ca²⁺ overloading and oxidative stress-induced cardiomyocyte necrosis.

Cardiomyocyte cytosolic free [Zn²⁺]_i has multiple biologic functions, none of which are more evident than its enhancement of antioxidant capacity thereby to confer cardioprotection in preventing nonischemic necrotic cell death and consequent microscopic scarring of myocardium. These antioxidant properties include: endogenous inhibitor to entry of prooxidant Ca²⁺; catalytic element to Cu/Zn-SOD activity; and [Zn²⁺]_i-based activation of its sensor, MTF1, which regulates antioxidant defense genes, including MT1.

In the failing heart, compromised antioxidant reserves can be reversed by exogenous Zn²⁺, such as with a nutriceutical like ZnSO₄, and from endogenous inactive cytosolic Zn²⁺ pool, using nebivolol, a β₃ receptor agonist, wherein inactive cytosolic Zn²⁺ bound to MT1 is released by nitric oxide. Thus, the spectrum of [Zn²⁺]_i as multifaceted antioxidant can be preferentially exploited promoting cardioprotection and counteracting the prooxidant heart failure phenotype.