Emmanuel E. Egom

Activation of Pak1/Akt/eNOS signaling following sphingosine-1-phosphate release as part of a mechanism protecting cardiomyocytes against ischemic cell injury.

We investigated whether plasma long-chain sphingoid base (LCSB) concentrations are altered by transient cardiac ischemia during percutaneous coronary intervention (PCI) in humans and examined the signaling through the sphingosine-1-phosphate (S1P) cascade as a mechanism underlying the S1P cardioprotective effect in cardiac myocytes. Venous samples were collected from either the coronary sinus (n = 7) or femoral vein (n = 24) of 31 patients at 1 and 5 min and 12 h, following induction of transient myocardial ischemia during elective PCI. Coronary sinus levels of LCSB were increased by 1,072% at 1 min and 941% at 5 min (n = 7), while peripheral blood levels of LCSB were increased by 579% at 1 min, 617% at 5 min, and 436% at 12 h (n = 24). In cultured cardiac myocytes, S1P, sphingosine (SPH), and FTY720, a sphingolipid drug candidate, showed protective effects against CoCl induced hypoxia/ischemic cell injury by reducing lactate dehydrogenase activity. Twenty-five nanomolars of FTY720 significantly increased phospho-Pak1 and phospho-Akt levels by 56 and 65.6% in cells treated with this drug for 15 min. Further experiments demonstrated that FTY720 triggered nitric oxide release from cardiac myocytes is through pertussis toxin-sensitive phosphatidylinositol 3-kinase/Akt/endothelial nitric oxide synthase signaling. In ex vivo hearts, ischemic preconditioning was cardioprotective in wild-type control mice (Pak1f/f), but this protection appeared to be ineffective in cardiomyocyte-specific Pak1 knockout (Pak1cko) hearts. The present study provides the first direct evidence of the behavior of plasma sphingolipids following transient cardiac ischemia with dramatic and early increases in LCSB in humans. We also demonstrated that S1P, SPH, and FTY720 have protective effects against hypoxic/ischemic cell injury, likely a Pak1/Akt1 signaling cascade and nitric oxide release. Further study on a mouse model of cardiac specific deletion of Pak1 demonstrates a crucial role of Pak1 in cardiac protection against ischemia/reperfusion injury.

Serum sphingolipids level as a novel potential marker for early detection of human myocardial ischaemic injury.

Background: Ventricular tachyarrhythmias are the most common and often the first manifestation of coronary heart disease and lead to sudden cardiac death (SCD). Early detection/identification of acute myocardial ischaemic injury at risk for malignant ventricular arrhythmias in patients remains an unmet medical need. In the present study, we examined the sphingolipids level after transient cardiac ischaemia following temporary coronary artery occlusion during percutaneous coronary intervention (PCI) in patients and determined the role of sphingolipids level as a novel marker for early detection of human myocardial ischaemic injury. Methods and Results: Venous samples were collected from either the coronary sinus (n = 7) or femoral vein (n = 24) from 31 patients aged 40–73 years-old at 1, 5 min, and 12 h, following elective PCI. Plasma sphingolipids levels were assessed by HPLC. At 1 min coronary sinus levels of sphingosine 1-phosphate (S1P), sphingosine (SPH), and sphinganine (SA) were increased by 314, 115, and 614%, respectively (n = 7), while peripheral blood levels increased by 79, 68, and 272% (n = 24). By 5 min, coronary sinus S1P and SPH levels increased further (720%, 117%), as did peripheral levels of S1P alone (792%). Where troponin T was detectable at 12 h (10 of 31), a strong correlation was found with peak S1P (R2 = 0.818; P < 0.0001). Conclusion: For the first time, we demonstrate the behavior of plasma sphingolipids following transient cardiac ischaemia in humans. The observation supports the important role of sphingolipids level as a potential novel marker of transient or prolonged myocardial ischaemia.

Impaired sinoatrial node function and increased susceptibility to atrial fibrillation in mice lacking natriuretic peptide receptor C

Natriuretic peptides (NPs) elicit their effects via multiple NP receptors (including NPR‐A, NPR‐B and NPR‐C, with NPR‐C being relatively poorly understood). We have studied the effects of NPR‐C ablation on cardiac structure, function and arrhythmogenesis using NPR‐C knockout (NPR‐C−/−) mice. NPR‐C−/− mice are characterized by sinoatrial node (SAN) dysfunction and a profound increase in susceptibility to atrial fibrillation. Increased susceptibility to arrhythmias in NPR‐C−/− mice was associated with slowed electrical conduction in the SAN as well as the right and left atria due to enhanced collagen expression and deposition in the atria (structural remodelling), but without changes in action potential morphology (electrical remodelling) in isolated SAN or atrial myocytes. This study demonstrates a critical protective role for NPR‐C in the heart.

HDL quality or cholesterol cargo: What really matters - Spotlight on sphingosine-1-phosphate-rich HDL

Purpose of review The absolute level of HDL cholesterol (HDL-C) may not be the only criterion contributing to their antiatherothrombotic effects. This review focuses on evidence in support of the concept that HDL-bound sphingosine-1-phosphate (S1P) plays a role in different HDL atheroprotective properties and may represent a potential target for therapeutic interventions. Recent findings Recent large randomized clinical trials testing the hypothesis of raising HDL-C with niacin and dalcetrapib in statin-treated patients failed to improve cardiovascular outcomes. Emerging evidence suggests that many of the cardioprotective functions of HDL, such as vasodilation, angiogenesis and endothelial barrier function, protection against ischemia/reperfusion injury, and inhibition of atherosclerosis, may be attributable to its S1P cargo. HDL-associated S1P may represent a future therapeutic target. Summary HDL functionality is affected by its composition and there is evidence to suggest S1P plays a role in some of HDLs functions and atheroprotective properties.

The potential role of sphingolipid-mediated cell signaling in the interaction between hyperglycemia, acute myocardial infarction and heart failure.

Introduction: Patients with acute myocardial infarction or heart failure frequently have abnormalities of glucose metabolism and insulin resistance, both of which are associated with a poor outcome. Sphingolipids are a class of lipids, which play important roles in cellular biological processes including insulin resistance and myocardial ischemia. Areas covered: This review examines the available evidence linking abnormalities in sphingolipids, glucose tolerance and insulin resistance to acute myocardial infarction and heart failure. Expert opinion: Pharmacological and genetic activation of enzymes controlling key sphingolipids synthesis, such as sphingosine-1-phosphate, increases insulin sensitivity in rodents and increases myocardial tolerance to ischemia. Major projects are being realized to develop clinical strategies to manipulate sphingolipid metabolism in this clinical settings, with the ultimate goal of increasing insulin sensitivity and augmenting myocardial tolerance to ischemia. Thus, a clear understanding of the sphingolipid-mediated signaling in ischemic heart disease is required to devise strategies to develop novel agents and technologies that directly target this signaling pathway.

Cardioprotection in ischemia/reperfusion injury: Spotlight on sphingosine-1-phosphate and bradykinin signalling

Complex signal-transduction cascades are known to be involved in regulating cardiomyocyte function, death and survival during acute cardiac ischemia-reperfusion process, but detailed survival signalling pathways are not clear. This review presents and discusses the recent findings bearing upon the evidence on the cardioprotective effect of sphingosine-1-phosphate (S1P) and bradykinin in acute cardiac ischemia-reperfusion and underlying signalling mechanisms, particularly, through activation of P21 activated kinase.

Novel bradykinin signaling in adult rat cardiac myocytes through activation of p21-activated kinase

Although bradykinin (BK) is known to exert effects on the myocardium, its intracellular signaling pathways remain poorly understood. Experiments in other cell types indicated that p21-activated kinase-1 (Pak1), a Ser/Thr kinase downstream of small monomeric G proteins, is activated by BK. We previously reported that the expression of active Pak1 in adult cardiac myocytes induced activation of protein phosphatase 2A and dephosphorylation of myofilament proteins (Ke et al. Circ Res 94: 194–200, 2004). In experiments reported here, we tested the hypothesis that BK signals altered protein phosphorylation in adult rat cardiac myocytes through the activation and translocation of Pak1. Treatment of myocytes with BK resulted in the activation of Pak1 as demonstrated by increased autophosphorylation at Thr423 and a diminished striated localization, which is present in the basal state. BK induced dephosphorylation of both cardiac troponin I and phospholamban. Treatment of isolated myocytes with BK also blunted the effect of isoproterenol to enhance peak Ca2+ and relaxation of Ca2+ transients. Protein phosphatase 2A was demonstrated to associate with both Pak 1 and phospholamban. Our studies indicate a novel signaling mechanism for BK in adult rat cardiac myocytes and support our hypothesis that Pak 1 is a significant regulator of phosphatase activity in the heart.

Activation of sphingosine-1-phosphate signalling as a potential underlying mechanism of the pleiotropic effects of statin therapy*

Abstract The mechanisms by which statins are beneficial are incompletely understood. While the lowering of low-density lipoprotein concentration is associated with regression of atherosclerosis, the observed benefit of statin therapy begins within months after its initiation, making regression an unlikely cause. Although LDL-C lowering is the main mechanism by which statin therapy reduces cardiovascular events, evidence suggests that at least some of the beneficial actions of statins may be mediated by their pleiotropic effects. Thus, statins may modulate the function of cardiovascular cells and key signalling proteins, including small G-proteins, to ultimately exert their pleiotropic effects. Sphingosine-1-phosphate (S1P) is a naturally occurring bioactive lysophospholipid that regulates diverse physiological functions in a variety of different organ systems. Within the cardiovascular system, S1P mediates cardioprotection following ischemia/reperfusion injury, anti-inflammatory response, improvement of endothelial function, increased mobilization and differentiation of endothelial progenitor cells, inhibition of oxidation, and anti-atherogenic and anti-thrombotic actions. Early evidence suggests that the pleiotropic effects of statins may be related to an increase in S1P signalling. This review focuses on S1P signalling as the potential mechanism underlying the pleiotropic effects of statins. An improved understanding of this mechanism may be vital for establishing the clinical relevance of statins and their importance in the treatment and prevention of coronary artery disease. Key points Several studies have demonstrated a benefit from lowering serum LDL-C with statins in patients with and without clinical evidence of CAD. These may be mediated by the pleiotropic effects of statins—the mechanisms of which are incompletely understood. Early evidence suggests that statins may increase S1P signalling pathways through upregulation of the expression of S1P receptors and an increase in plasma levels of S1P to ultimately exert their pleiotropic effects. Future clinical trials and basic science research aimed at the underlying mechanisms of the pleiotropic effects of statins should enlighten us to their relative clinical relevance and importance.

Altered parasympathetic nervous system regulation of the sinoatrial node in Akita diabetic mice

Cardiovascular autonomic neuropathy (CAN) is a serious complication of diabetes mellitus that impairs autonomic regulation of heart rate (HR). This has been attributed to damage to the nerves that modulate spontaneous pacemaker activity in the sinoatrial node (SAN). Our objective was to test the hypothesis that impaired parasympathetic regulation of HR in diabetes is due to reduced responsiveness of the SAN to parasympathetic agonists. We used the Akita mouse model of type 1 diabetes to study the effects of the parasympathetic agonist carbachol (CCh) on SAN function using intracardiac programmed stimulation, high resolution optical mapping and patch-clamping of SAN myocytes. CCh decreased HR by 30% and increased corrected SAN recovery time (cSNRT) by 123% in wildtype mice. In contrast, CCh only decreased HR by 12%, and only increased cSNRT by 37% in Akita mice. These alterations were due to smaller effects of CCh on SAN electrical conduction and spontaneous action potential firing in isolated SAN myocytes. Voltage clamp experiments demonstrate that the acetylcholine-activated K(+) current (IKACh) is reduced in Akita SAN myocytes due to enhanced desensitization and faster deactivation kinetics. These IKACh alterations were normalized by treating Akita SAN myocytes with PI(3,4,5)P3 or an inhibitor of regulator of G-protein signaling 4 (RGS4). There was no difference in the effects of CCh on the hyperpolarization-activated current (If) between wildtype and Akita mice. Our study demonstrates that Akita diabetic mice demonstrate impaired parasympathetic regulation of HR and SAN function due to reduced responses of the SAN to parasympathetic agonists. Our experiments demonstrate a key role for insulin-dependent phosphoinositide 3-kinase (PI3K) signaling in the parasympathetic dysfunction seen in the SAN in diabetes.

The effect of the sphingosine-1-phosphate analogue FTY720 on atrioventricular nodal tissue

The sphingosine‐1‐phosphate (S1P) receptor modulator, fingolimod (FTY720), has been used for the treatment of patients with relapsing forms of multiple sclerosis, but atrioventricular (AV) conduction block have been reported in some patients after the first dose. The underlying mechanism of this AV node conduction blockade is still not well‐understood. In this study, we hypothesize that expression of this particular arrhythmia might be related to a direct effect of FTY720 on AV node rather than a parasympathetic mimetic action. We, therefore, investigated the effect of FTY720 on AV nodal, using in vitro rat model preparation, under both basal as well as ischaemia/reperfusion conditions. We first look at the expression pattern of S1P receptors on the AV node using real‐time PCR. Although all three S1P receptor isoforms were expressed in AVN tissues, S1P1 receptor isoform expression level was higher than S1P2 and S1P3. The effect of 25 nM FTY720 on cycle length (CL) was subsequently studied via extracellular potentials recordings. FTY720 caused a mild to moderate prolongation in CL by an average 9% in AVN (n = 10, P < 0.05) preparations. We also show that FTY720 attenuated both ischaemia and reperfusion induced AVN rhythmic disturbance. To our knowledge, these remarkable findings have not been previously reported in the literature, and stress the importance for extensive monitoring period in certain cases, especially in patients taking concurrently AV node blocker agents.