Epoxylipids and Soluble Epoxide Hydrolase in Heart Diseases

Abstract

Cardiovascular and heart diseases are leading causes of morbidity and mortality. Coronary artery endothelial and vascular dysfunction, inflammation, and mitochondrial dysfunction contribute to progression of heart diseases such as arrhythmias, congestive heart failure, and heart attacks. Classes of fatty acid epoxylipids and their enzymatic regulation by soluble epoxide hydrolase (sEH) have been implicated in coronary artery dysfunction, inflammation, and mitochondrial dysfunction in heart diseases. Likewise, genetic and pharmacological manipulations of epoxylipids have been demonstrated to have therapeutic benefits for heart diseases. Increasing epoxylipids reduce cardiac hypertrophy and fibrosis and improve cardiac function. Beneficial actions for epoxylipids have been demonstrated in cardiac ischemia reperfusion injury, electrical conductance abnormalities and arrhythmias, and ventricular tachycardia. This review discusses past and recent findings on the contribution of epoxylipids in heart diseases and the potential for their manipulation to treat heart attacks, arrhythmias, ventricular tachycardia, and heart failure.

Graphical Abstract

1. Introduction

Cardiovascular diseases are estimated to result in 18 million deaths globally each year which is projected to rise to 24 million deaths per year by 2030 [1,2]. This represents one third of all global deaths with over 80% of cardiovascular deaths being due to heart attacks and strokes [1,2]. Risk factors for cardiovascular diseases include a sedentary lifestyle, inappropriate diet, tobacco use, and alcohol intake [3,4]. These risk factors can result in an elevated blood pressure, hyperglycemia, increased plasma lipid levels, and obesity [3,4]. There have been advances in patient care that have led to improvements in cardiovascular outcomes [3,5]. Despite the progress, the decreases in cardiovascular mortality are dissipating [1,2]. Unlike other diseases areas such as cancer and neurological disorders, the pipeline for cardiovascular disease drugs is weak [6]. Therefore, identifying targets for cardiovascular drugs is desperately needed.

The pathology of cardiovascular diseases includes changes in vascular and cardiac function that can damage multiple organs resulting in heart failure [7,8]. Vascular endothelium dysfunction is a major contributor to cardiovascular diseases such as atherosclerosis, coronary artery disease, and heart disease [9,10]. Heart attacks are a consequence of these cardiovascular diseases [3,9]. Damage to the heart is due to inflammation, oxidative stress, apoptosis, necrosis, ionic disturbances, and alterations in mitochondrial bioenergetics [11,12]. Extensive deterioration of cardiac function occurs due to atrial or ventricular arrhythmias, myocardial fibrosis, and ventricular hypertrophy [7,8,9]. Eventually cardiac function declines and progresses to heart failure [7,8].

Although drugs for cardiovascular diseases can combat hypertension, dyslipidemia, and diabetes, there is an urgent need to identify therapeutic targets that will further decrease mortality. Manipulation of fatty acids provides opportunities to develop cardiovascular drugs [13–16]. Epoxy fatty acids and soluble epoxide hydrolase (sEH) are promising targets treating cardiovascular diseases [13,15,17,18]. It is well recognized that polyunsaturated fatty acids have vascular and cardiac biological activities that could be beneficial in combating cardiovascular diseases [14,15,16]. In addition, there is significant evidence that regulation of enzymes that regulate polyunsaturated fatty acids can impact cardiovascular diseases [14,15,16]. Epoxy fatty acids of arachidonic acid, eicosapentaenoic acid (EPA), and docosohexaenoic acid (DHA) are epoxylipids that can improve mitochondrial function, decrease lipidemia, decrease inflammation, improve insulin sensitivity, lower blood pressure, and improve endothelial dysfunction [14,16]. These epoxy fatty acids can be converted to diols by sEH that decreases beneficial vascular and cardiac biological activities (Figure 1) [14,15,16]. These findings have led to development of drugs that can inhibit sEH to increase epoxy fatty acid levels or drugs that act as epoxy fatty acid mimetics [14,15,16].

Figure 1 – Epoxylipid Metabolites.

Fatty acids arachidonic acid, eicosapentaenoic acid, and docosohexaenoic acid are metabolized by CYP2C and CYP2J epoxygenase enzymes to epoxyeicosatrienoic acids (EET), epoxyeicosatetraenoic acids (EEQ), and epoxydocosapentaenoic acids (EDP). Epoxylipids are then metabolized to their corresponding diols, dihydroxyeicosatetraenoic acids (diHETE) and dihydroxydocosapentaenoic acids (diHDPA) by soluble epoxide hydrolase (sEH).

This review will focus on the contribution of epoxy fatty acids and sEH to myocardial infarction, arrythmias, ventricular tachycardia, and chronic heart failure. The protective actions and cell signaling mechanisms by which arachidonic acid, EPA, and DHA epoxy fatty acids improve cardiac function will be detailed. Finally, the development and testing of sEH inhibitors and epoxylipid mimics in cardiac experimental models will be provided. The progress to date demonstrates the potential for epoxylipid based drugs to treat heart attacks, arrhythmias, ventricular tachycardia, and heart failure.

2. Epoxylipids and Soluble Epoxide Hydrolase in Heart Diseases

Cardiovascular diseases have been associated with a decrease in arachidonic acid epoxylipids, epoxyeicosatrienoic acids (EETs) and an increased degradation by sEH to their corresponding diols [19,20]. Biological actions demonstrated by EETs can combat cardiovascular diseases. EETs demonstrate vasodilation, anti-inflammatory, anti-apoptotic, and anti-smooth muscle cell migration actions [15,17]. Human genetic studies have found associations between cytochrome P450 (CYP) epoxygenase and sEH enzymes with increased risk for cardiovascular diseases [19,21–24]. In most of these studies genetic polymorphisms that result in decreased EETs or increased sEH activity associate with increased cardiovascular disease risk or progression [19,24]. Associations between genetic variations that decrease CYP epoxygenases or plasma EET levels and hypertension have been demonstrated in different cohorts [19,24,25,26]. On the other hand, increased plasma EET levels were found in patients with stable angiographically confirmed coronary artery disease [23]. Increased plasma EET levels in patients after stroke and decreased expression of EPHX2, the gene for sEH, in heart failure patients have also been observed [27,28]. These finding demonstrate that mechanisms to upregulate EETs can act to oppose cardiovascular disease in certain patients.

Experimental studies in animal disease models have provide greater mechanistic insight into epoxylipids and their ability to combat cardiovascular diseases [14,15,16]. Hypertension animal models have been the most extensively studied regarding EETs and sEH to blood pressure control and progression of end organ damage and heart disease [13,15]. Decreased EET levels and increased sEH expression were found to contribute to increased vascular resistance and electrolyte homeostasis in hypertension [29,30,31]. Genetic manipulation in mice that increased or decreased EET levels further demonstrated a contribution to regulation of blood pressure and progression of heart disease in hypertension [32–36]. In addition to effects on vascular resistance and electrolyte homeostasis, EETs regulate inflammation and oxidative stress in hypertension [13,15]. These findings led to evaluating sEH inhibitors and EET analogs as therapeutics for hypertension [13,18,37].

Myocardial infarction studies in rodents have provided additional mechanistic evidence for EETs and other epoxylipids to cardiovascular diseases [15,16,38]. EETs have been demonstrated to improve cardiac mitochondrial function, decrease inflammation, and oppose apoptosis to reduce cardiac fibrosis and hypertrophy (Figure 2) [14,38]. Cardiomyocyte mitochondrial function is regulated by EETs which improve heart function following ischemia [39,40,41]. Cardioprotective mitochondrial actions were found in mice overexpressing the CYP2J2 epoxygenase or lacking sEH [38,39,41]. EET actions on vascular smooth muscle cell K⁺ channels vasodilate coronary arteries and EET actions on cardiomyocyte K⁺ channels oppose metabolic stress to decrease heart damage following ischemia [42]. EETs have also been clearly demonstrated to reduce cardiac cell death [14,38]. Cardiomyocytes exposed to hypoxic conditions and treated with EETs demonstrated increased cell viability, mitochondrial membrane stability, and decreased caspase-3 activity [43,44]. EET actions on phosphoinositide 3-kinase (PI3K) and K_ATP channels are critical signaling mechanisms for the anti-apoptotic cardiomyocyte effects. Lastly, EET anti-inflammatory actions on the heart provide protection from adverse cardiac remodeling and fibrosis that contribute to left ventricular dysfunction [14,15]. EETs can oppose vascular and heart monocyte and neutrophil infiltration and can inhibit nuclear factor-κB (NF-κB) activation to reduce proliferation of cardiac fibroblasts and development of heart fibrosis [34,35]. Cardiac dysfunction following lipopolysaccharide injection in mice is decreased by EET actions on peroxisome proliferator-activated receptor gamma (PPARγ) and heme oxygenase-1 (HO-1) pathways to reduce M1 macrophage polarization and infiltration resulting in decreased inflammatory cytokine production [45]. These cardioprotective EET actions ultimately protect from ischemic and non-ischemic heart diseases.

Figure 2 – Cardiomyocyte Actions for Epoxyeicosatrienoic Acids (EET), Epoxyeicosatetraenoic Acids (EEQ), and Epoxydocosapentaenoic Acids (EDP).

Arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosohexaenoic acid (DHA) are metabolized by CYP2C and CYP2J epoxygenase enzymes to EET, EEQ, and EDP. EET, EEQ, and EDP decrease endoplasmic reticulum (ER) stress, combat mitochondrial dysfunction, inhibit transforming growth factor β1 (TGF-β1), reduce the NLRP3 inflammasome, and inhibits nuclear factor-κB (NF-κB) activation to reduce proinflammatory cytokines, chemokines, and adhesion molecules. These cardiomyocyte EET, EEQ, and EDP actions reduce apoptosis and inflammation to decrease fibrosis and heart remodeling.

The cardioprotective actions for n-3 polyunsaturated fatty acids diets have been long recognized [16,46,47]. EPA and DHA rich diets result in increased plasma and tissue levels of 17,18-epoxyeicosatetraenoic acid (17,18-EEQ) and 19,20-epoxydocosapentaenoic acid (19,20-EDP) [16]. These experimental studies have demonstrated cardiomyocyte cell signaling mechanisms responsible for improved heart function (Figure 2) [16]. Increased dietary EPA and DHA can improve cardiac mitochondrial function and decrease apoptosis [46,48,49]. In addition, cardiomyocytes in cell culture have demonstrated direct actions for EPA and DHA to decrease mitochondrial dysfunction, decrease oxidative stress, and oppose apoptosis [46,49]. EPA can attenuate H92C cardiac cell oxidative stress and apoptosis through activating an adaptive autophagic response [46,49]. EPA treatment protected against apoptosis via ERK activation in cultured cardiomyocytes exposed to hypoxia [50]. Likewise, the DHA epoxygenase metabolite 19,20-EDP can protect against lipopolysaccharide-induced cardiac cell toxicity by activation of sirtuin 1 (SIRT1) to improve mitochondrial function resulting in cell survival [51]. DHA also decreased cardiac damage in isolated heart rats exposed to ischemic reperfusion injury [52]. An experimental study that compared EPA, DHA, 17,18-EEQ, and 19,20-EDP on isolated mouse hearts exposed to ischemia reperfusion revealed that DHA and 19,20-EDP but not EPA and 17,18-EEQ provided cardioprotection [53]. 19,20-EDP provided the greatest protection against myocardial infarction through reduction in the NLPR3 inflammasome to preserve mitochondrial function [53]. These findings demonstrate the potential for EPA, DHA, and their epoxylipid metabolites to protect cardiomyocytes via cell signaling mechanisms that impact mitochondrial function, oxidative stress, and apoptosis.

Experiments conducted in rodents with cardiac ischemia reperfusion injury provide additional evidence for EPA, DHA, and their epoxylipid metabolites to decrease apoptosis and decrease mitochondrial dysfunction [14,16]. Mice fed a diet enriched with EPA and DHA protects against thoracic aortic constriction induced left ventricular dysfunction and fibrosis [54]. Cardiomyocyte studies demonstrate that the anti-fibrotic actions for EPA and DHA are due to inhibiting transforming growth factor β1 (TGF-β1) [54]. Dietary EPA was also found to oppose pressure overload cardiac fibrosis through decreasing TGF-β1 signaling [55]. Anti-inflammatory actions for dietary EPA have been demonstrated in rodents that decrease cardiac remodeling following myocardial infarction [46,56]. Clinical studies have provided additional evidence for cardiac benefits for diets enriched with EPA and DHA [57,58,59]. Consumption of EPA/DHA supplements reduces cardiac sudden death and the severity of ventricular arrhythmias [57,58]. The findings that EPA, DHA, and their epoxylipid cardioprotective actions has led to therapeutically targeting these fatty acids for various heart diseases.

3. Pharmacological Manipulation of sEH and Epoxylipids in Heart Diseases

The development of therapeutics to manipulate sEH and epoxylipids has provided promising results in animal models of hypertension, vascular diseases, and heart diseases [13–17]. Inhibitors of sEH are orally active and have advanced to clinical trials for several diseases including cardiovascular diseases [15,18]. The primary action for sEH inhibitors under normal fatty acid diet conditions is to increase arachidonic acid epoxylipids, EETs [15,18]. Epoxylipid mimics (analogs) that are orally active are also under development by pharmaceutical companies and are in preclinical development and the early stages of clinical trials [13,37]. Epoxylipid mimics for arachidonic acid, EPA, and DHA epoxylipids have been developed and tested in cardiac diseases [13,14,16]. More recent animal studies are evaluating combinations of sEH inhibitors and epoxylipid mimics to treat cardiac diseases [13,37].

Cardioprotective action for sEH inhibitors have been well documented in animal models of left ventricular hypertrophy and ischemia reperfusion injury (Figure 3) [14,15]. The heart protective actions for sEH inhibition is due to effects on the coronary vasculature, inflammatory cells, and cardiomyocytes [14,15]. Left ventricular hypertrophy in mice subjected to pressure overload is prevented by sEH inhibitor actions to decrease cardiac myocyte inflammation [60]. Administration of sEH inhibitors to obese mice decreases inflammation to reduce cardiac fibrosis and hypertrophy [61]. These finding have provided strong evidence that sEH inhibitors have potential to combat the progression of cardiac fibrosis and left ventricular hypertrophy.

Figure 3 – Epoxylipid Drugs to Combat Heart Disease.

Soluble epoxide hydrolase inhibitors (sEH), epoxyeicosatrienoic acids (EET) analogs, epoxyeicosatetraenoic acids (EEQ) analogs, and epoxydocosapentaenoic acids (EDP) analogs act to combat cardiac inflammation and oxidative stress, decrease cardiac cell mitochondrial dysfunction and endoplasmic reticulum (ER) stress, and reduce cardiac hypertrophy and fibrosis. These positive cardiac actions of sEH inhibitors, EET analogs, EEQ analogs, and EDP analogs result in a decrease in arrythmias and heart failure

There is also ample evidence in animal models that sEH inhibitors improve cardiac function following an ischemic event [14,15]. Experimental evidence in rodents and canines using the Langendorff-perfused heart and in vivo studies have demonstrated that sEH inhibition decreased heart injury following ischemia [62,63,64]. Coronary artery administration of the sEH inhibitor, AUDA, reduced heart ischemia reperfusion injury in dogs [64]. Interestingly, AUDA also enhanced the cardioprotective actions of 14,15-EET by preventing conversion to the inactive diol [64]. In rodents, sEH inhibitors have been demonstrated to protect against cardiac ischemia reperfusion injury when administered prior to, during, or following reperfusion [62,63,65,66]. The recovery of postischemic left ventricular pressure has also been demonstrated to be improved by sEH inhibition [14,66]. Signaling mechanisms responsible for the sEH inhibitor cardiomyocyte actions include K⁺ channel activation, anti-apoptosis actions, and improved mitochondrial function [14,15]. Reduction in infarct size with sEH inhibitors is due to anti-apoptotic extracellular signal-regulated kinase (ERK) / mitogen-activated protein kinase (MAPK) and Janus kinase/signal transducer and activator of transcription (JAK/STAT) cell signaling pathways [67,68]. Cardiomyocyte activation of K_ATP channels to enhance PI3 kinase signaling and the subsequent protein kinase B (Akt) phosphorylation to increase phospho-glycogen kinase-3β (p-GSK-β) is critical for sEH to reduce myocardial ischemia reperfusion injury [65,66]. Activation of cardiac K_ATP channels also delays or inhibits mitochondrial permeability transition pore (mPTP) opening to prevent apoptosis and necrosis [69]. Taken together, sEH inhibitors have strong cardioprotective effects from ischemia reperfusion injury through activation of cardiomyocyte K_ATP channels that improves mitochondrial function and decreases apoptosis.

Cardiac protection from the long-term consequences of myocardial infarction have also been described for sEH inhibitors [14,15]. This is an important finding because following the acute phase recovery after a myocardial infarction there is extensive cardiac inflammation, apoptosis, and left ventricular remodeling [15,16,70]. The myocardial infarction induced cardiac hypertrophy and fibrosis can be diminished with sEH inhibition [16,70]. Long-term treatment with sEH inhibitors acts to reduce inflammation that occurs after ischemia reperfusion injury to improve cardiac outcomes [71]. Treatment of sEH inhibitors to rats subjected to permanent coronary artery occlusion reduced cardiac fibrosis [72]. Administration of sEH inhibitors days and weeks following coronary artery ligation also improved left ventricular function [72,73,74]. Mice subjected to acute left anterior descending artery and administered an sEH inhibitor for three weeks had reduced cardiac fibrosis and improved left ventricular function [72]. The improvement in cardiac function with sEH inhibition following myocardial infarction also reduces electrical conductance abnormalities and arrythmias [28,73,74]. Myocardial infarction studies in hypertensive rats have also demonstrated long-term cardioprotective actions for sEH inhibitors [75,76]. Chronic administration of the sEH inhibitor c-AUCB to TGR (Ren-2 renin transgenic rats) increased EET levels, reduced blood pressure, and significantly decreased infarct size following coronary artery occlusion [75]. Selective EET antagonism with 14,15-EEZE completely prevented c-AUCB cardioprotective actions confirming that cardiomyocyte EET actions are responsible for reducing cardiac injury [75]. These experimental findings demonstrate long-term protective sEH inhibitor actions following myocardial infarction that appear to be mediated by cardiomyocyte actions of the epoxylipid EETs.

The cardioprotective actions for sEH inhibitors extend to non-ischemic heart injury [15,16,70]. Atrial fibrillation, arrhythmias, and congestive heart failure are cardiac disorders where sEH inhibitors demonstrate potential as a therapeutic [15,16,70]. Treatment with sEH inhibitors reduced cardiac atrial and ventricular arrhythmias in cardiac hypertrophy rodent models [28,73,74]. Decreased inflammation in response to sEH inhibition is a key mechanism responsible for preventing atrial arrhythmias in cardiac hypertrophy [73,74]. Administration of sEH inhibitors to mice with thoracic aortic constriction resulted in inhibition of NF-κB activation and reduced inflammatory cytokine levels [74]. This reduction in inflammation led to decreased MAPK and endoplasmic reticulum stress to reduce adverse cardiac structural and electrical remodeling of the heart which reduced arrhythmias in mice [74]. Isoproterenol-induced cardiac hypertrophy is also decreased by sEH inhibition [77]. Likewise, inhibition of sEH in obese mice improves coronary artery endothelial function and prevents cardiac remodeling [61]. Mechanisms responsible for the decreased cardiac remodeling and reduced arrythmias include decreased inflammation, improved coronary artery endothelial function, and decreased cardiomyocyte AMP-activated protein kinase (AMPK) signaling resulting in decreased apoptosis and endoplasmic reticulum stress [61].

Less clear are the cardioprotective actions for sEH inhibitors in congestive heart failure. Evidence for a contribution for sEH to congestive heart failure comes from the spontaneously heart failure rat that has an alteration in the EPHX2 gene that facilitates progression of congestive heart failure [28]. Experimental studies that administered an sEH inhibitor to Fawn-hooded hypertensive rats with congestive heart failure and chronic kidney disease demonstrated beneficial actions [76]. Survival rates and urinary albumin levels were improved by sEH inhibition, and the effects were more pronounced in the hypertensive rats [76]. The protective actions of sEH inhibition in the hypertensive Fawn-hooded rats was a result of decreased cardiac hypertrophy and lung congestion [76]. On the other hand, sEH inhibition in aorto-caval fistula in Hannover Sprague-Dawley rats did not improve the course of congestive heart failure or chronic kidney disease [78]. The sEH inhibitor c-AUCB increased renal and cardiac EET availability in the aorto-caval fistula congestive heart failure rats; however, cardiac systolic and diastolic function failed to improve [78]. These findings in congestive heart failure rat models suggest that the experimental conditions and disease progression can influence the beneficial actions for sEH inhibitors. Additional studies are required in congestive heart failure to better understand the potential for sEH inhibitors to combat disease progression.

EET analogs represent a class of epoxylipid mimics that have been extensively evaluated in heart disease animal models [14,15,37]. These EET analogs mimic 11,12-EET and 14,15-EET and resist sEH metabolism [37]. Three generations of EET analogs have been developed with good water solubility, oral bioavailability, and favorable half-life to allow for chronic administration [37]. Initial studies in cardiac diseases with EET analogs have demonstrated excellent therapeutic potential [14,15,37].

The cardiac protective actions for EET analogs have been demonstrated in ischemic reperfusion injury studies (Figure 3) [14,37]. Administration of the 11,12-EET analog, NUDSA to mice following myocardial infarction resulted in improved left ventricular function [79]. Mice treated five days following left anterior descending coronary artery ligation resulted in improved left ventricular diastolic function and decreased cardiac fibrosis at one month [79]. Another 11,12-EET analog, UA-8 also improved cardiac ischemic tolerance in isolated mice hearts [80]. Like 11,12-EET analogs, the 14,15-EET analog, EET-A administered for two weeks can reduce cardiac hypertrophy [81]. Ventricular arrythmias following myocardial infarction in hypertensive rats are also reduced by EET-A treatment [81]. Cardiac hypertrophy in TGR rats was reduced by EET-A treatment which could be to a certain extent due to the anti-hypertensive actions [81]. More recently, TGR hypertensive rats with aorto-caval fistula induced congestive heart failure were treated with EET-A which improved survival [82]. The increased survival in this congestive heart failure was greatly improved by the combination of EET-A and angiotensin converting enzyme inhibition [82]. Another 14,15-EET analog, EET-B can reduce cardiac hypertrophy and fibrosis in the Dahl salt-sensitive hypertensive rat independent of blood pressure lowering [83]. EET-B also limited infarct size in rats subjected to regional ischemia in the heart in vivo [84]. The cardioprotective actions for EET-B extends to spontaneously hypertensive rats (SHR) subjected to 30 minutes of left coronary artery occlusion [84]. EET-B administered for several weeks did not lower blood pressure in SHR but improved cardiac fractional shortening, reduced cardiac inflammation, and decreased cardiac fibrosis [84]. These experimental findings demonstrate that 11,12-EET and 14,15-EET mimics can combat heart disease.

The mechanisms by which EET analogs decrease fibrosis and improve heart function has been investigated in cardiac ischemia reperfusion and other cardiac disease animal models [14,15,37]. Cardioprotective actions for EET analogs include improved mitochondrial function, anti-inflammatory, and anti-apoptotic actions [14,15]. EET analogs increase OPA1 oligomers and activate mitochondrial K_ATP channels in cardiac myocytes [39]. Anti-apoptotic actions for EET analogs are partially mediated by MAPK signaling activation to prevent ischemic cardiac cell stress [39]. Along these lines, EET-B was demonstrated to improve cardiac ischemic tolerance via hypoxia-inducible factor-1α (HIF-1α) [85]. These studies found that EET-B blunted the decrease in cadiomyocyte HIF-1α following reperfusion via downregulation of its degrading enzyme prolyl hydroxylase domain protein 3 (PHD3) [85]. This EET-B mediated change in HIF-α and PHD3 balance resulted in favoring the cardioprotective actions of HIF-1α [85]. Anti-inflammatory actions for EET-B are also responsible for improved cardiac function and attenuation of congestive heart failure in SHR following myocardial infarction [84]. Decreased cardiac CD68 positive macrophage/monocyte levels in SHR following myocardial infarction and treated with EET-B [84]. The EET-B anti-inflammatory actions could be partly attributed to the increase in cardiac HO-1 levels [84]. Future studies with EET analogs are needed to further examine cardiac cellular mechanisms responsible for improved cardiac function.

More recently combinations of sEH inhibitors, EET analogs, and renin-angiotensin system inhibitors have been evaluated for their ability to combat cardiac disease. TGR hypertensive rats were evaluated for cardiac hypertrophy and a subset subjected to acute ischemia reperfusion injury and treated with an sEH inhibitor, EET-A, or the combination [81]. Chronic administration of c-AUCB, EET-A or the combination decreased blood pressure and reduced ventricular fibrillation induced by myocardial infarction in the TGR hypertensive rats [81]. Importantly, the combination of c-AUCB and EET-A did not provide additive anti-hypertensive or cardioprotective actions [81]. Likewise, these treatments alone or in combination had similar actions to improve left ventricular function and slow the progression of heart failure in Hannover Sprague-Dawley but not in TGR hypertensive rats subjected to myocardial infarction [86]. Experimental findings in aorto-caval fistula induced congestive heart failure rats demonstrated differences between the combination of sEH inhibition and angiotensin converting enzyme inhibition and the combination of an EET analog and angiotensin converting enzyme inhibition [87]. The sEH inhibitor c-AUCB was given alone or in combination with the angiotensin converting enzyme inhibitor, trandolapril to TGR hypertensive rats with aorto-caval fistula induced congestive heart failure [87]. Intriguingly, c-AUCB in combination with trandolapril worsened the survival rate when compared to trandolapril alone in TGR hypertensive congestive heart failure rats [87]. In contrast, EET-A in combination with an angiotensin converting enzyme inhibitor improved survival to a greater extent than either treatment alone in TGR hypertensive congestive heart failure rats [87]. This combined EET-A and angiotensin converting enzyme inhibitor treatment had beneficial actions on cardiac morphology and left ventricular function [87]. Regrettably, these findings with combination treatments of sEH inhibitor, EET analog, and angiotensin converting enzyme inhibitor in hypertensive congestive heart failure rats do not provide strong evidence for additive actions to improve cardiac function.

Analogs mimicking EPA and DHA epoxy fatty acids have recently been synthesized and tested for their ability to decrease inflammation and combat cardiac diseases (Figure 3) [16,88,89]. The initial focus has been synthesizing EPA epoxylipid, 17,18-EEQ analogs [16,88]. Analogs of 17,18-EEQ were demonstrated to decrease ventricular tachyarrhythmias following coronary ligation in rats [90]. Additional studies demonstrated that 17,18-EEQ analogs can reduce atrial fibrillation in mice [91]. Further preclinical development of 17,18-EEQ analogs has led to clinical phase 1 trials in 2017 [16]. A third generation of 17,18-EEQ analogs have been synthesized and tested for cardioprotective actions [88]. This generation of 17,18-EEQ analogs demonstrated a similar action as previous generations to decrease ventricular tachyarrhythmias following coronary ligation [88]. Development of 16,17-EDP and 19,20-EDP analogs are in the initial stages and there has been initial testing for their ability to provide cardioprotection. Three novel 16,17-EDP and 19,20-EDP analogs were tested for their ability to improve cardiac function in isolated mice hearts exposed to ischemia reperfusion injury [53,89]. The 16,17-EDP analog, SA-26 improved postischemic cardiac function, reduced oxidative stress, preserved mitochondrial SIRT3 activity, and attenuated the NLPR3 inflammasome response [89]. Therefore, the cardiac therapeutic potential for EEQ and EDP analogs is promising; however, there remains much to be explored to fully understand the cardiac mechanisms of action to reach the ability to treat cardiac disease in humans.

4. Conclusions and Future Directions

The investigation of epoxylipids and their regulation by sEH in cardiac diseases has advanced from determining broad cardioprotective actions in myocardial infarction and progressive heart failure to determining cardiomyocyte cell signaling mechanisms that improve mitochondrial function, decrease oxidative stress, reduce inflammation, and oppose apoptosis. Subsequently, development of therapeutics to inhibit sEH and mimic epoxylipids have advanced to human clinical trials. Despite this outstanding progress, there is still much to be explored.

Novel pharmacological and genetic approaches to manipulate epoxylipids to combat heart disease are needed. Development of multi-target drugs that manipulate these epoxylipids could be beneficial for cardiac diseases. The concept for multi-target drugs to combat heart diseases is intriguing given the success of a combined neprilysin and angiotensin type 1 receptor blocker for heart failure [92]. Multi-target drugs that include sEH inhibitor activity have demonstrated to decrease inflammation and organ fibrosis in hypertension, kidney diseases, and metabolic diseases [91–96]. These multi-target drugs have sEH inhibitor activity combined with inhibition against other fatty acid metabolizing enzymes or transcription factor agonists [93,95,96]. Positive cardiovascular actions for these sEH inhibitor multi-target include lowering blood pressure and decreasing plasma lipids [95,96]. In addition, sEH inhibitors with EET mimic activity have been developed but these compounds have not been extensively tested in cardiac diseases [37,97]. Thus, multi-target drugs that act to manipulate epoxylipids combined with another activity could provide a therapeutic approach for heart diseases.

Likewise, although significant progress has been made with 17,18-EEQ mimics, 16,17-EDP mimics and 19,20-EDP mimics are in their infancy. The precise molecular and protein target for EETs, 17,18-EEQ, 16,17- EDP, and 19,20-EDP have yet to be identified. Attractively, a G-protein receptor for another CYP hydroxylase metabolite, 20-HETE has been identified as GPR75 [98,99]. This 20-HETE receptor has been implicated in vascular function and cardiovascular disease [98]. There has been extensive characterization for EETs to identify G-protein coupled EET receptors [37]. EETs have been demonstrated to act through G-proteins and bind to cells and cell membranes with low and high affinity binding [100,101,102]. However, a protein receptor for EETs has yet to be identified. Receptor or target protein identification for epoxylipids would allow for target-based screening drug design approaches.

Combining genetic and metabolomic data in humans is another approach that could guide treatment by sEH inhibitors and epoxylipid mimics. There are known genetic EPHX2 polymorphisms in humans that associate with endothelial dysfunction and cardiovascular diseases [19,21–24]. Testing for these genetic variants combined with measurements of epoxylipids could determine cardiac disease patients that would benefit from sEH inhibitors and epoxylipid mimics.

Ultimately, there has been significant progress with evaluating sEH and epoxylipids in heart diseases and the prospects that manipulation of epoxylipids can treat several heart diseases in humans is eminent.