Resveratrol in cardiovascular health and disease

Authors


Address for correspondence: Dipak K. Das, Ph.D., Sc.D., M.D. (hon), FAHA, Cardiovascular Research Center, University of Connecticut, School of Medicine, Farmington, CT 06030-1110. ddas@neuron.uchc.edu

Abstract

Resveratrol, initially used for cancer therapy, has shown beneficial effects against most degenerative and cardiovascular diseases from atherosclerosis, hypertension, ischemia/reperfusion, and heart failure to diabetes, obesity, and aging. The cardioprotective effects of resveratrol are associated with its preconditioning-like action potentiated by its adaptive response. During preconditioning, small doses of resveratrol can exert an adaptive stress response, forcing the expression of cardioprotective genes and proteins such as heat shock and antioxidant proteins. Similarly, resveratrol can induce autophagy, another form of stress adaptation for degrading damaged or long-lived proteins, as a first line of protection against oxidative stress. Resveratrol's interaction with multiple molecular targets of diverse intracellular pathways (e.g., action on sirtuins and FoxOs through multiple transcription factors and protein targets) intertwines with those of the autophagic pathway to give support in the modified redox environment after stem cell therapy, which leads to prolonged survival of cells. The successful application of resveratrol in therapy is based upon its hormetic action similar to any toxin: exerting beneficial effects at lower doses and cytotoxic effects at higher doses.

Introduction

Cardiovascular diseases (CVDs) are the major causes of morbidity and mortality in the developed nations, including the United States.1 About 100 million Americans suffer from some kind of CVD including high blood pressure, atherosclerosis, coronary heart disease, or stroke. Thirty-nine percent of all deaths—or one in every three deaths—is claimed to be caused by a heart disease. Although proper medicines are abundant in developing countries to combat CVDs, preventive measures appear to be necessary to lower the incidence of heart problems. Maintaining a healthy heart is particularly important for those with a family history of coronary heart disease including hypertension, heart attack, and atherosclerosis. Fortunately, a number of natural cures are available that appear to maintain cardiovascular health. For example, maintaining a healthy lifestyle, daily exercise, and choosing proper diets certainly help maintain a healthy heart.2,3 Certain mineral-rich foods including those high in potassium, calcium, and magnesium can control blood pressure. Low-fat milks are rich sources of calcium, while magnesium and potassium can be found in green vegetables and fish oil. Among the fatty foods, limiting saturated, polyunsaturated, monounsaturated, and trans- fats would help reduce cholesterol and associated forms of coronary heart diseases. Major sources of saturated fat include beef, butter, cheese, whole milk, and coconut and palm oils. In contrast, polyunsaturated fats like omega-3-fatty acids containing foods including fish oil, flax seed oil, and canola oil may reduce the risk of heart attack. Some of the examples for maintaining a healthy heart include garlic, olive oil, broccoli, capsicum, coenzyme Q10, Terminalia arjuna, Ashwagandha (Withania somnifera), cocoa, and fish oil, most of which are included in the Mediterranean diet. Numerous reports are available in the literature about heart healthy diet from Mediterranean foods.4,5

First published in 1992 as the French Paradox, the health benefits of daily drinking of wine in moderation has been proven by most of the scientists and clinicians all over the world.6 Wine contains 11–14% alcohol and many polyphenolic compounds, most of which possess antioxidant properties. Although alcohol possesses cardioprotective properties, it is universally believed that it is not the alcohol but some of the polyphenols present in wine that are responsible for the cardioprotective properties of wine.7 Among the polyphenols present in wine, especially red wine, resveratrol has drawn major attention. Resveratrol is present in the skins of grapes and thus in red wine. In addition to grapes, resveratrol is present in a large variety of fruits including cranberry, mulberry, lingonberry, bilberry, partridgeberry, sparkleberry, deerberry, blueberry, jackfruit, peanut, as well as in a wide variety of flowers and leaves including gnetum, butterfly orchid tree, white hellebore, scots pine, corn lily, eucalyptus, and spruce. Commercially, resveratrol is extracted from the dried roots of Polygonum cuspidatum, mainly found in Japan and China. Polygonum extract has been used in Japanese and Chinese traditional medicine to treat fungal infections, various skin inflammations, liver disease, and cardiovascular problems.8 Resveratrol is also an antifungal compound that is synthesized in response to environmental stressors including water deprivation, UV radiation, and especially fungal infection.9

A growing body of evidence now supports the notion that resveratrol is the major factor responsible for the cardioprotective effects of wine. In the heart, resveratrol blocks low-density lipoprotein (LDL) peroxidation, increases high-density lipoprotein (HDL) levels, induces vasorelaxation presumably through the induction of nitric oxide (NO) synthesis, inhibits endothelin (ET), modifies angiogenic response, reduces ventricular arrhythmias, possesses antithrombin activity and prevents platelet aggregation, inhibits formation of soluble adhesion molecules, reduces reactive oxygen species (ROS), reduces blood pressure, and ameliorates ischemic reperfusion injury (Fig. 1).8,10–12 It is believed that resveratrol-mediated cardioprotection is not due to its direct drug-like effect on the diseased heart, rather, it potentiates a preconditioning (PC)-like effect, a state-of-the art for cardioprotection. The PC effect results from an adaptive response, which receives further support from a recent discovery that resveratrol promotes autophagy (Fig. 2).13,14 More recently, resveratrol has been found to regenerate the infracted myocardium.11 Resveratrol appears to maintain a healthy heart in numerous ways, and can provide numerous health benefits, which will be the subject of discussion in this review.

Figure 1.

Effects of resveratrol on cardiovascular health and other diseases. Granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin (IL), low-density lipoprotein (LDL), nitric oxide (NO), and reactive oxygen species (ROS).

Figure 2.

Cardioprotective pathways of resveratrol.

Resveratrol and cardiovascular health

As mentioned earlier, CVDs—being the leading cause of death and illness worldwide—are greatly dependent not only on nonmodifiable predisposing factors (age, sex, and genetic composition), but also on a modifiable factor—the lifestyle. Indeed, risk factors other than increased plasma cholesterol15 and obesity,16 such as oxidative stress, inflammation, and endothelial damage, all have great relevance in the development of CVD. One of the most well-known benefits of resveratrol is improvement of cardiovascular health (Fig. 1).17 Resveratrol, in physiologic concentrations after red wine consumption, increases the expression of NO synthase, an enzyme responsible for synthesizing the potent vasodilator NO in human vascular endothelial cells.18 Resveratrol also decreases the expression of the potent vasoconstrictor ET.18 Cerebral ischemic damage in rat brains can be prevented via pretreatment with resveratrol19 by activating sirtuin (Sirt) 1, coupled with a decrease in mitochondrial uncoupling protein 2 and an increase in mitochondrial ATP synthesizing efficiency.19 Resveratrol can also exert cardioprotection by inhibiting platelet aggregation, similar to how aspirin works.17 In the following sections, the beneficial effects of resveratrol, as summarized in Figure 1, will be briefly discussed in relation to some major heart problems.

Atherosclerosis

Atherosclerosis is an inflammatory disease of the arterial wall caused by endothelial wall damage during continuous hemodynamic and redox stress conditions.20 Under such conditions, modified LDLs, lipids, and fibrous elements can accumulate in the wall. Its progression into plaque disruption can lead to local thrombosis and acute coronary syndrome.21,22 As previously mentioned, resveratrol can regulate the production of NO, a potent vasodilator, by counterbalancing the effect of the vasoconstrictor ET-1,18,23–25 thus providing thromboresistance and preventing atherogenesis.24 Resveratrol can potentially inhibit stress-induced ET-1 gene expression by interfering with its promoting extracellular-signal-regulated kinases (ERK) 1/2 pathway26 and therefore improve endothelial function.27

Inflammation mediates all stages of atherosclerosis from initiation to progression and, eventually, plaque rupture. Prostaglandin E2 (PGE2) plays a key role in inflammation—its synthesis is catalyzed by cyclooxygenase (COX) 2. Resveratrol can inhibit the atherosclerosis-associated inflammation via regulating the COX-2 activity at a transcriptional level, thereby inhibiting the production of PGE2.28 In cultured murine macrophages, resveratrol can suppress the proinflammatory interleukin (IL)-6 gene expression, protein synthesis, and secretion,29,30 and inhibit the release of both IL-8 and granulocyte-macrophage colony-stimulating factor.31 Oxidation of LDL and uptake into the vascular wall as a main cause of endothelial injury and inflammation can be prevented by resveratrol.32 Endothelial cell injury, platelet aggregation, and adherence to the injured cell surface can be blocked by this stilbene, inhibiting further the adhesion of collagen to platelets33 and ADP-induced platelet aggregation34 in a concentration-dependent manner.35 Low concentrations of resveratrol can significantly inhibit intracellular and extracellular ROS production36 by enhancing the intracellular free radical scavenger glutathione.37

Oxidized lipids, metabolic stress, and inflammation in atherosclerotic plaques can also induce autophagy as a safeguard against cellular distress by degrading damaged intracellular material. Basal autophagy can be enhanced by certain drugs for cardioprotection, which can reach damaging and autophagic cell death levels under excessive stimulation.38 Loss of smooth muscle cells may lead to plaque destabilization and rupture under excessive cell death and autophagy conditions.39 However, therapeutic induction of autophagy in atherosclerotic plaque resolution may have a counterbalancing effect on other survival or cell death pathways such as apoptosis and necrosis.

Hypertension

Higher systemic blood pressure (SBP) associates with increased heart weight, serum ET-1, angiotensin II (AngII) concentrations, and decreased serum NO. Resveratrol decreases SBP, heart weight, ET-1, and AngII concentrations while increases the vasodilator NO concentration, which protects against increased SBP and subsequent cardiac hypertrophy in mice.40,41 Mean arterial pressure after resveratrol treatment dropped significantly in spontaneously hypertensive rats.42 In another study on rats treated with resveratrol, echocardiographic analysis of cardiac structure and function after hypertensive stress showed improvement in the interventricular septal wall thickness and left ventricular posterior wall thickness, as well as isovolumetric relaxation time at systole and diastole. Endothelial NO synthase (eNOS), inducible nitric oxide synthase (iNOS), and redox factor-1 (ref-1) were significantly decreased under these stress conditions. Resveratrol may therefore be beneficial against certain types of cardiac hypertrophy found in clinical settings of hypertension and aortic valve stenosis.

For the last 30 years it has been know that lysosomal pathways are involved in the different models of heart disease.43–47 More recently, advances in our understanding of autophagy have revealed that hemodynamic load-induced aggregation of intracellular proteins may have a protective role.48 However, excessive pressure overload may elicit a robust autophagic response in cardiomycytes that is maladaptive and contributes to disease progression.

Ischemia/reperfusion injury and PC

PC is a protective and adaptive phenomenon whereby brief episodes of ischemia and reperfusion (I/R) render the heart resistant to subsequent ischemic injury or stress. A large number of stimuli such as short cyclic episodes of I/R, a number of pharmacological agents (agonists of adenosine, bradykinin, adrenergic and muscarinic receptor, NO donors, and phosphodiesterase inhibitors), and various noxious stimuli (endotoxins, cytokines, and ROS) have been found to generate PC-like phenotype.49 Resveratrol can similarly precondition the heart in a NO-dependent manner (Fig. 3), inducing the expression of iNOS, eNOS, and vascular endothelial growth factor (VEGF) in rats.50 Resveratrol provided cardioprotection as evidenced by superior postischemic ventricular recovery, reduced myocardial infarct size, and decreased number of apoptotic cardiomyocytes. Resveratrol likely activates both adenosine A1 and A3 receptors that phosphorylate phosphatidylinositol-3 kinase (PI3K), which then phosphorylates protein kinase B (Akt) and thus preconditions the heart by producing NO, as well as by the activation of antioxidant Bcl-2.50,51 Activation of adenosine A3 receptors could also precondition the heart by a survival signal through the cAMP response element-binding protein (CREB) phosphorylation via PI3K/Akt and via MERK (mitogen-activated extracellular signal-regulated protein kinase)/CREB pathways.52 Recent studies have also demonstrated that NO can induce the expression of heme oxygenase-1. Tin protoporphyrin (SnPP), a heme oxygenase-1 (HO-1) inhibitor, abolished increased cardiac function parameters, reduced myocardial infarct size, and decreased cardiomyocyte apoptosis that characterize the cardioprotection with resveratrol.53 The HO-1–mediated mechanisms were related to the p38 MAP kinase and Akt survival signaling but independent of nuclear factor-kappaB (NF-κB) activation. A polyphenol, resveratrol, protects the heart by its antioxidative properties through various redox signaling mechanisms. The ability of resveratrol to modulate redox signaling has been extensively reviewed,54,55 but the role of Sirt activation in the heart is yet to be determined. White wine lacking polyphenols but containing antioxidant compounds such as caffeic acid and tyrosol mediated cardioprotection against I/R injury in rat hearts work via similar survival pathway involving Akt/FOXO3a/NF-κB.56 Our recent results also indicate that at lower doses, resveratrol-mediated survival of cardiac myoblasts is, in part, mediated through the induction of autophagy,57 which, along other enhanced survival signals, helps to recover the cells from injury.58

Figure 3.

(A) Typical electron spin resonance (ESR) spectra recorded from the reduction of nitroxide during ischemia of the isolated heart. The reduction in nitroxide resulted in a decrease in the ESR signal intensity over time during the ischemic phase. (B) The decrease in signal intensity over time was fit to determine the rate of decay of the ischemic heart in control and resveratrol-treated hearts. (C) The rate of decay of the TEMPO nitroxide (2, 2, 6, 6-tetramethyl-piperidine-1-oxyl) during ischemic challenge in control and resveratrol-pretreated isolated hearts. Results are shown as mean ± SEM of 4 hearts per group.

Angiogenesis

Resveratrol in general has angiosuppressive effects—rat gliomas significantly decrease in size upon resveratrol treatment.59 Proliferation and migration of vascular endothelial cells under resveratrol treatment activates eukaryotic elongation factor-2 kinase, which in turn inactivates elongation factor-2, an important factor in protein translation.60 In mice that develop abnormal angiogenesis in the retina after laser treatment, when given resveratrol, the abnormal blood vessels began to disappear. Similarly, resveratrol suppressed the growth of new blood vessels in animals by directly inhibiting capillary endothelial cell growth61—it blocked the VEGF and fibroblast growth factor (FGF) receptor-mediated angiogenic responses and significantly delayed angiogenesis-dependent wound healing. Exposure of human umbilical vein endothelial cells (HUVECs) to low doses of resveratrol significantly blocks VEGF-mediated migration and tube formation but not cell proliferation.26 Under the same concentrations, resveratrol fails to affect VEGF-stimulated activation of VEGF receptor, extracellular signal-regulated protein kinase 1/2, p38 mitogen-activated protein kinase, and Akt. Interestingly, at low micromolar doses, resveratrol effectively abrogates VEGF-mediated tyrosine phosphorylation of vascular endothelial cadherin and its complex partner, β-catenin.

Insufficient angiogenesis can result in nutritional and metabolic deficiency, igniting prosurvival pathways of autophagy in cells and tissues. In this case as well, the double-edged sword of autophagy is evident, since low-level induction of this pathway may be prosurviving for the deficiently supplied tissue, but excessive autophagy in cells can be destructive and possibly proinflammatory.62,63

Metabolic syndrome: diabetes and obesity

Although most of the research done on the positive effect of resveratrol and its antidiabetic potential has been conducted on animals, the results from our own laboratory show that injecting streptozotosin diabetic rats with this compound results in a significant decrease in the levels of blood glucose.64 Similarly, direct injection of resveratrol into the brains of diabetic rats resulted in reduction of insulin levels like it does in hyperinsulinemic rats. A recent study reported that long-term intracerebroventricular infusion of resveratrol-normalized hyperglycemia and greatly improved hyperinsulinemia in diet-induced obese and diabetic mice65 independent of changes in body weight, food intake, and circulating leptin levels. In addition, central nervous system resveratrol delivery improves hypothalamic NF-κB inflammatory signaling by reducing acetylated-RelA/p65 and total RelA/p65 protein contents and inhibitor of NF-κB alpha and beta mRNA levels. Furthermore, this treatment leads to reduced hepatic phosphoenolpyruvate carboxykinase 1 mRNA and protein levels and ameliorates pyruvate-induced hyperglycemia in this mouse model of type 2 diabetes. The mechanism of resveratrol's action is by far complex and still evolving. Obesity studies in our laboratory demonstrate that resveratrol renders the hearts of ob/ob mice resistant to ischemic reperfusion injury.66 The antidiabetic potential of resveratrol appears to warrant further promise of imposing the same effect on humans.

Insulin signaling, which enhances protein synthesis, also inhibits autophagy in insulin target tissues.67 Thus, there is the potential for alterations in this control in insulin-resistant states such as diabetes, and defects in autophagy may adversely affect cell function in diabetes. Such defects may lead to inappropriate accumulation of dysfunctional mitochondria, insulin granules, or activation of apoptotic pathways. Furthermore, various cellular stresses can induce dysfunctional protein folding, which can cause the accumulation of misfolded and aggregated proteins in cells.68,69 Inability of the autophagy pathway to clear protein aggregates may be intimately related to cell death in diabetes.

Longevity

The first revolutionary discovery that resveratrol can activate the antiaging gene SIRT170 has been followed by recent studies supporting this antiaging function. However, animal studies of the dose effect of resveratrol would mean that one needs to consume several bottles of red wine per day to reach such a dose.71 More recently, low dose of resveratrol has been found to activate not only SIRT1, but also SIRT 3 and SIRT 4 as well as FoxOs and pre-B cell colony-enhancing factor (PBEF)—all being proteins linked or associated with longevity.72 In addition, white wine and its components tyrosol and hydroxytyrosol have been found to activate same longevity genes, suggesting that activation of SIRT1 could be a nonspecific phenomenon. Resveratrol prevented age-related and obesity-related cardiovascular functional decline in mice, but did not affect the overall survival or maximum life span for mice on a standard diet, compared to mice on the same diet with resveratrol.73 Different species appear to increase their life span upon resveratrol, from yeast and nematodes to flies (Drosophila melanogaster), as well as mice kept on a high fat diet. In another related study, SIRT1 overexpression enhanced the autophagic flux in cancer cells, and this effect was blocked by inhibitors of its catalytic activity. Resveratrol induced autophagy in Caenorhabditis elegans also via activation of SIRT2; this effect was abolished by siRNA mediated depletion of SIRT2.74 Autophagy, in turn, increases life expectancy of the worm.

A substantial amount of evidence suggests that resveratrol mimics calorie restriction and increases the life span. Ongoing research to identify an ideal antiaging compound has not been successful. To date, calorie restriction remains the only effective mean to increase the life span,75 although the mechanisms whereby calorie restriction increases life span remain speculative. Several reports can be found in the literature supporting the role of exercise and certain chemicals such as rapamycin and resveratrol in expanding life span.76 A study using microarray revealed that out of 6,347 modified genes by calorie restriction, about 58 genes displayed at least twofold alterations in gene expression.77 There is a striking similarity between resveratrol and calorie restriction on the alteration of metabolic pathways. For example, both resveratrol and calorie restriction improve insulin sensitivity thereby reducing the insulin and glucose levels in the body,78 which in turn reduce the life-threatening cardiovascular risk factors. Both resveratrol and calorie restriction can potentiate the expression of Glucose transporter type 4 (GLUT4).66 As mentioned earlier, the most important antiaging gene that is upregulated by calorie restriction and resveratrol is SIRT1, although it works in coordination with the FoxOs. Indeed, resveratrol and red wine containing significant amount of resveratrol that can inactivate the FoxOs by phosphorylation.72

In a recent study, life extension of fish by resveratrol was associated with the change in the slope of mortality trajectory,79 indicating lowering of the time-dependent increase in death risk with resveratrol. In the same study, the authors noticed an initial increase in death rate after resveratrol treatment, suggesting that the weak toxic effect of resveratrol was associated with a stress response that ultimately increased the expression of longevity genes, thereby increasing the life span. A similar effect was observed in Drosophila, when hypothermia was associated with a reduction in the accumulation of age-dependent irreversible injury.80 This effect was quite different from the life extension induced by calorie restriction, which is reflected in time-dependent reduction in acute risk of death without changing the slope of mortality rate.80

Resveratrol and autophagy

On the basis of our previous observation that cardioprotection induced by ischemic PC induces autophagy and that resveratrol induces PC-like effects, we sought to determine if resveratrol could induce autophagy. Macroautophagy, commonly known as autophagy, involves a bulk degradation process clearing organelles, long-lived proteins, and protein complexes. Cytosolic constituents destined to be degraded become enclosed by double membrane structures, known as autophagosomes or autophagic vacuoles, which are fused with lysosomes followed by the degradation of its contents. In fact, the name explains the term autophagy, a housekeeping process through self-cannibalism or eating inside-out.

Three types of autophagy are known: (i) microautophagy involving “dumping” of cytosolic constituents into the lysosome by direct invagination of the lysosomal membrane followed by budding of vesicles into the lysosomal lumen; (ii) macroautophagy, or autophagy involving the formation of a double membrane structure known as an autophagosome, which sequesters cytosolic constituents to be delivered into the lysosomes for digestion; and (iii) chaperone-mediated autophagy characterized by selectivity of cytosolic proteins to be degraded. Any autophagic process undergoes four stages: (i) induction by external stress, such as environmental stress (e.g., oxidative stress), nutritional stress (e.g., nutritional deprivation), and physical stress (e.g., ischemia or hypoxia)—the gatekeeper for induction is mTOR, which regulates transcriptional activation of downstream target genes; (ii) autophagosome formation, as described earlier, where a number of autophagy (Atg) genes participate and recruit beclin-1 and microtubule-associated protein light-chain 3 (LC3); (iii) autophagosomes undergo docking and fusion with the lysosome; and finally (iv) autophagic vesicles are broken down by lysosomal proteases, where lysosomal-associated membrane protein 2 (LAMP-2) plays a crucial role in the degradation process.

In a recent publication, we demonstrated that resveratrol at lower doses (0.1 and 1 μM in H9c2 cardiac myoblast cells and 2.5 mg/kg/day in rats) induced cardiac autophagy shown by enhanced formation of autophagosomes and its component LC3-II after hypoxia-reoxygenation or ischaemia-reperfusion.11 Autophagy was attenuated with the higher dose of resveratrol, suggesting hormetic action of resveratrol, and the induction of autophagy was correlated with enhanced cell survival and decreased apoptosis. Treatment with rapamycin (100 nM), a known inducer of autophagy, did not further increase autophagy compared with resveratrol alone.11 Autophagic inhibitors, wortmannin (2 μM), and 3-methyladenine (10 mM) significantly attenuated resveratrol-induced autophagy and induced cell death.11 The activation of mammalian target of rapamycin (mTOR) was differentially regulated by low-dose resveratrol, i.e., the phosphorylation of mTOR at serine 2448 was inhibited, whereas the phosphorylation of mTOR at serine 2481 was increased, which was attenuated with a higher dose of resveratrol.11 Although resveratrol attenuated the activation of mTOR complex 1, low-dose resveratrol significantly induced the expression of rictor, a component of mTOR complex 2, and activated its downstream survival kinase Akt (Ser 473).11 Resveratrol-induced rictor was found to bind with mTOR. Furthermore, treatment with rictor siRNA attenuated the resveratrol-induced autophagy. These results indicate that at lower doses, resveratrol-mediated cell survival is, in part, mediated through the induction of autophagy involving the mTOR-rictor survival pathway.11

It appears that resveratrol can generate a survival signal through autophagy. The important question is, “can we use such autophagy clinically for the health benefits?” The simple answer is yes, if the amount of stress can be controlled. There is no doubt that a small amount of stress induced by resveratrol can induce autophagy generating a survival signal, while the same autophagy (induced by higher doses of resveratrol) will potentiate a death signal if the amount of such stress is large and becomes cytotoxic to the cells. Thus, a “therapeutic amount of stress” (5–10 mM resveratrol) must be defined for the induction of autophagy, the same amount that has been known to induce an “adaptive response” for the cells subjected to hostile environment.

Resveratrol in regeneration of infracted myocardium

Another recent study from our laboratory using cardiac stem cells demonstrated that resveratrol can potentiate the regeneration of infracted myocardium.57 A major problem in the effectiveness of stem cell therapy is the death of stem cells due to the oxidative environment present in the normal tissue. Reduction of oxidative stress or maintaining a reduced environment in the target tissue can enhance stem cell survival and cardiac regeneration after stem cell therapy. In the study mentioned,57 we pretreated rats with resveratrol (2.5 mg/kg/day gavaged for 2 weeks) after which a left anterior descending coronary artery (LAD) occlusion was carried, followed by direct injection of adult cardiac stem cells stably expressing enhanced green-fluorescent protein (EGFP) on the border zone of the myocardium through survival surgery. The prevalence of cardiac-reduced environment was seen in resveratrol-treated rat hearts via significantly enhanced redox signaling observed through the nuclear factor-E2-related factor-2 (Nrf2), stromal cell-derived factor-1 (SDF1), and NF-κB, as well as ref-1, seven days after LAD occlusion.57 Significantly improved cardiac functional parameters (left ventricular ejection fraction and fractional shortening), enhanced stem cell survival and proliferation (expression of cell proliferation marker Ki67), and differentiation of stem cells toward the regeneration of the myocardium (expression of EGFP) was evident 28 days after LAD occlusion in rats treated with resveratrol, compared to control rats.57 Our study clearly demonstrated that resveratrol can modify the physiological redox environment within the myocardium. Maintaining a reduced tissue environment by treatment with resveratrol in rats enhanced the cardiac regeneration by adult cardiac stem cells via improved cell survival, proliferation, and differentiation, leading to improved cardiac function. This study is the first demonstrating that nutritional modification of the redox environment with resveratrol can prolong the regeneration of the infracted myocardium.57

In addition cardiac stem cells were modified with resveratrol by preincubating the cells stably expressing EGFP with 10 mM resveratrol for 60 min followed by washing the cells with buffer to get rid of any free resveratrol.57 Rats were anesthetized, their hearts were opened, and LAD was occluded to induce a heart attack. The animals were divided into two groups: one group was treated with resveratrol-modified stem cells, while the other group was treated with stem cells alone. One week after the LAD occlusion, the cardiac-reduced environment was confirmed in resveratrol-treated rat hearts by the enhanced expression of Nrf2 and Ref-1. M-mode echocardiography was performed to determine cardiac function up to three months after the stem cell therapy. Initially (after 72 h), both groups revealed improvement in cardiac function, but only the resveratrol-modified stem cell group revealed improvement in cardiac function (left ventricular ejection fraction, fractional shortening, and cardiac output) at the end of the 1, 2, and 4 months period. The improvement in cardiac function was accompanied by the enhanced stem cell survival and proliferation as evidenced by the expression of cell proliferation marker Ki67 and differentiation of stem cells toward the regeneration of the myocardium as evidenced by the expression of EGFP up to six months after LAD occlusion in the resveratrol-treated stem cell group of hearts.57 Again, our results demonstrate that resveratrol maintained a reduced tissue environment by overexpressing Nrf2 and Ref-1 in rat hearts up to four months, resulting in an enhancement of the regeneration of the adult cardiac stem cells as evidenced by increased cell survival and differentiation leading to improved cardiac function.

Resveratrol and hormones

Several recent studies implicate that resveratrol displays hormetic action, protecting the cells at lower doses while killing them at relatively higher doses. Since such a hormetic behavior might have significant impact on epidemiological and clinical studies, we sought to determine the dose–response curve for resveratrol action. We fed by gavaging up to 30 days a group of rats three different doses of resveratrol 2.5, and 100 mg/kg while the control group was given vehicles only. Our results indeed showed hormesis for resveratrol, being cardioprotective at lower doses only and detrimental for higher doses (D. Das, J. Clin. Exp. Cardiol., in press). At 100 mg/kg dose, 100% of the hearts died in case of resveratrol.a The results clearly demonstrate that resveratrol is beneficial to the heart only at low doses and is detrimental at higher doses. Also, the action of resveratrol is quickly realized, in most cases within 14 days; up to 30 days, resveratrol does not add any additional benefit. Such hormesis has been known for more than 100 years and frequently observed among the toxins. As mentioned earlier, resveratrol is a phytoalexin, whose growth is stimulated by environmental stress such as fungal infection, ultraviolet (UV) radiation, and water deprivation.81 Cardioprotective effects of resveratrol are exerted through its ability to precondition a heart through adaptive response, which causes the development of intracellular stress leading to the upregulation of intracellular defense system such as antioxidants and heat shock protein.82 Preconditioning is another example of hormesis, which is potentiated by subjecting an organ like heart to cyclic episodes of short durations of ischemia, each followed by another short durations of reperfusion.83 Such small but therapeutic amount of stress renders the heart resistant to subsequent lethal ischemic injury. Such an adaptive response is commonly observed with aging. Autophagy also constitutes another form of adaptive response. Consistent with this idea, resveratrol has been found to stimulate longevity genes, and at least in prokaryotic species extend the life span. In this respect, resveratrol may fulfill the definition of a hormetin.84 There is no doubt that alcohol, wine, and wine-derived resveratrol all display hormesis. It has been known for quite some time that cardioprotective effects of alcohol or wine intake follow a J-shaped curve.85 This study (D. Das, J. Clin. Exp. Cardiol. in press) echoed this concept that cardioprotective effects of resveratrol follow a J-shaped curve and display hormesis (Fig. 4). At lower doses, resveratrol acts as an antiapoptotic agent, providing cardioprotection as evidenced by increased expression in cell survival proteins, improved postischemic ventricular recovery, and reduction of myocardial infarct size and cardiomyocyte apoptosis by maintaining a stable redox environment compared to control. At higher doses, however, resveratrol depresses cardiac function, elevates levels of apoptotic protein expressions, results in an unstable redox environment, and increases myocardial infarct size and number of apoptotic cells. A significant number of reports are available in the literature to show that at a high dose, resveratrol not only hinders tumor growth, but also inhibits the synthesis of RNA, DNA, and protein; causes structural chromosome abberrations, chromatin breaks, chromatin exchanges, weak aneuploidy, and higher S-phase arrest; blocks cell proliferation; and decreases wound healing, endothelial cell growth by VEGF and FGF-2, and angiogenesis in healthy tissue cells leading to cell death.86–89

Figure 4.

Hormetic action of resveratrol. Dose of resveratrol [X-axis] is plotted against the values of cardiac function, infarct size, and apoptosis. Resveratrol at a dose of 2.5 mg/kg provided maximum protection [peak], which then progressively and steadily declined.

Summary and conclusion

Resveratrol, a grape- and wine-derived phytoalexin polyphenol, provides diverse health benefits, the most prominent being the best natural medicine to cure diverse CVDs including atherosclerosis, hypertension, ischemia/reperfusion, heart failure, diabetes, obesity, and aging. Resveratrol-mediated cardioprotection is not due to its drug-like action neutralizing the toxins introduced by the disease; rather, the protection is realized from its PC-like effects potentiated by its adaptive response. During the PC, a small amount of resveratrol (defined as therapeutic dose) exerts adaptive stress response thereby enabling the cardiac cells to make cardioprotective genes and proteins such as heat shock and antioxidant proteins. The ability of resveratrol to exert adaptive response is further supported by its ability to induce autophagy, another form of adaptation to stress. In that context, autophagy may be viewed as “cry for survival,” and such survival is a result of adaptive response to fight against stress. If the stress—such as oxidative stress—is mild, it generates an adaptive response to survive against it. Autophagy results in a survival signal by inducing a number of genes and transcription factors that alter the stress-induced death signal into a survival signal, leading to production of antiapoptotic or antideath proteins. On the other hand, if the stress is overwhelming, the adaptive response fails and the cells die due to induction of apoptotic signals. It is important to remember that to survive, cells must get rid of damaged, detrimental, and unwanted components, and they do so through autophagy. Thus, resveratrol may be used clinically to induce autophagy leading to cardioprotection. Finally, resveratrol can modify the redox environment within the cells through its ability to induce redox protein thioredoxin, which can pave the way for stem cell survival. Recent studies documented its ability to regenerate the infracted myocardium up to four months after the cell therapy. An important thing to remember is that for successful application of resveratrol in therapy, one must recognize the fact that resveratrol possesses hormetic action similar to any toxins, exerts beneficial action at lower doses, and becomes cytotoxic at higher doses, generating a J-shaped or inverted U-shape curve like wine and alcohol.

Further research and more clinical studies are necessary in order to ensure the safety of resveratrol and also for ascertaining the optimum doses and ways of inducing low-level autophagy for prevention and treatment. The findings that resveratrol can potentiate diverse health benefits from chemoprevention to cardioprotection—through its ability to induce autophagy and cardiac regeneration through prolonging the stem cell survival—certainly warrants an opening of a new era of pharmaceutical intervention in cardiovascular medicine.

Footnotes

  1. aDudley et al. 2009, J. Nutr. Biochem., 20(6), 2009, 443–452.

Conflicts of interest

The authors declare no conflicts of interest.

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