O-linked attachment of the monosaccharide β-N-acetyl-glucosamine (O-GlcNAcylation) is a post-translational modification occurring on serine and threonine residues, which is evolving as an important ...mechanism for the regulation of various cellular processes. The present review will, first, provide a general background on the molecular regulation of protein O-GlcNAcylation and will summarize the role of this post-translational modification in various acute cardiac pathologies including ischemia-reperfusion. Then, we will focus on research studies examining protein O-GlcNAcylation in the context of cardiac hypertrophy. A particular emphasis will be laid on the convergent but also divergent actions of O-GlcNAcylation according to the type of hypertrophy investigated, including physiological, pressure overload-induced and diabetes-linked cardiac hypertrophy. In an attempt to distinguish whether O-GlcNAcylation is detrimental or beneficial, this review will present the different O-GlcNAcylated targets involved in hypertrophy development. We will finally argue on potential interest to target O-GlcNAc processes to treat cardiac hypertrophy. This article is part of a Special Issue entitled: The role of post-translational protein modifications on heart and vascular metabolism edited by Jason R.B. Dyck & Jan F.C. Glatz.
•O-GlcNAcylation process is extremely intricate to cardiac hypertrophy development.•O-GlcNAcylation is promoted during pathological cardiac hypertrophy.•O-GlcNAcylation action is multifaceted and varies upon types of cardiac hypertrophy.
We report a case of sudden unexplained death in a young asymptomatic woman in whom postmortem genetic testing after a negative autopsy identified a homozygous pathogenic mutation in SLC22A5 which ...leads clinically to primary carnitine deficiency (PCD). Her brother was subsequently diagnosed clinically with short QT syndrome, received an implantable defibrillator, and was then found to carry the same pathogenic homozygous mutation and critically low levels of carnitine. His QT interval improved with the use of carnitine supplementation, highlighting the close relationship between electrophysiology and biochemistry, and the importance of postmortem genetic testing in the clinical management of surviving relatives.
Nous décrivons le cas de mort subite inexpliquée d’une jeune femme asymptomatique chez qui les analyses génétiques post-mortem effectuées après une autopsie négative ont révélé une mutation homozygote pathogène du gène SLC22A5 entraînant sur le plan clinique un déficit primaire en carnitine (DPC). Son frère, chez qui un diagnostic clinique de syndrome du QT court a été posé ultérieurement, a reçu un défibrillateur implantable. On a alors constaté qu’il était porteur de la même mutation homozygote pathogène et qu’il présentait des concentrations de carnitine dangereusement faibles. La prise de supplément de carnitine a donné lieu à une amélioration de l’intervalle QT chez le patient, ce qui met en lumière la relation étroite qui existe entre l’électrophysiologie et la biochimie, de même que l’importance des analyses génétiques post-mortem dans la prise en charge clinique des proches survivants.
The AMP-activated protein kinase (AMPK) is an important cellular energy sensor. Its activation under energetic stress is known to activate energy-producing pathways and to inactivate energy-consuming ...pathways, promoting ATP preservation and cell survival. AMPK has been shown to play protective role in many pathophysiological processes including cardiovascular diseases, diabetes, and cancer. Its action is multi-faceted and comprises short-term regulation of enzymes by direct phosphorylation as well as long-term adaptation via control of transcription factors and cellular events such as autophagy. During the last decade, several studies underline the particular importance of the interaction between AMPK and the post-translational modification called O-GlcNAcylation. O-GlcNAcylation means the O-linked attachment of a single N-acetylglucosamine moiety on serine or threonine residues. O-GlcNAcylation plays a role in multiple physiological cellular processes but is also associated with the development of various diseases. The first goal of the present review is to present the tight molecular relationship between AMPK and enzymes regulating O-GlcNAcylation. We then draw the attention of the reader on the putative importance of this interaction in different pathophysiological events.
Genetic causes of many familial arrhythmia syndromes remain elusive. In this study, whole‐exome sequencing (WES) was carried out on patients from three different families that presented with ...life‐threatening arrhythmias and high risk of sudden cardiac death (SCD). Two French Canadian probands carried identical homozygous rare variant in TECRL gene (p.Arg196Gln), which encodes the trans‐2,3‐enoyl‐CoA reductase‐like protein. Both patients had cardiac arrest, stress‐induced atrial and ventricular tachycardia, and QT prolongation on adrenergic stimulation. A third patient from a consanguineous Sudanese family diagnosed with catecholaminergic polymorphic ventricular tachycardia (CPVT) had a homozygous splice site mutation (c.331+1G>A) in TECRL. Analysis of intracellular calcium (Ca2+i) dynamics in human induced pluripotent stem cell‐derived cardiomyocytes (hiPSC‐CMs) generated from this individual (TECRLHom‐hiPSCs), his heterozygous but clinically asymptomatic father (TECRLHet‐hiPSCs), and a healthy individual (CTRL‐hiPSCs) from the same Sudanese family, revealed smaller Ca2+i transient amplitudes as well as elevated diastolic Ca2+i in TECRLHom‐hiPSC‐CMs compared with CTRL‐hiPSC‐CMs. The Ca2+i transient also rose markedly slower and contained lower sarcoplasmic reticulum (SR) calcium stores, evidenced by the decreased magnitude of caffeine‐induced Ca2+i transients. In addition, the decay phase of the Ca2+i transient was slower in TECRLHom‐hiPSC‐CMs due to decreased SERCA and NCX activities. Furthermore, TECRLHom‐hiPSC‐CMs showed prolonged action potentials (APs) compared with CTRL‐hiPSC‐CMs. TECRL knockdown in control human embryonic stem cell‐derived CMs (hESC‐CMs) also resulted in significantly longer APs. Moreover, stimulation by noradrenaline (NA) significantly increased the propensity for triggered activity based on delayed afterdepolarizations (DADs) in TECRLHom‐hiPSC‐CMs and treatment with flecainide, a class Ic antiarrhythmic drug, significantly reduced the triggered activity in these cells. In summary, we report that mutations in TECRL are associated with inherited arrhythmias characterized by clinical features of both LQTS and CPVT. Patient‐specific hiPSC‐CMs recapitulated salient features of the clinical phenotype and provide a platform for drug screening evidenced by initial identification of flecainide as a potential therapeutic. These findings have implications for diagnosis and treatment of inherited cardiac arrhythmias.
Synopsis
Mutations in the novel TECRL gene were identified in patients with malignant exercise‐induced arrhythmias. Increased triggered electrical activity upon stimulation in patient‐specific hiPSC‐CMs was rescued by the antiarrhythmic drug flecainide.
Trans‐2,3‐enoyl‐CoA reductase‐like (TECRL) is preferentially expressed in the heart.
Mutations in TECRL cause lethal arrhythmias in humans.
Cardiac defects in TECRL patients are characterized by overlapping features of long QT syndrome (LQTS) and catecholaminergic polymorphic ventricular tachycardia (CPVT).
Cardiomyocytes differentiated from patient‐specific human induced pluripotent stem cells (hiPSCs) recapitulate the electrical abnormalities observed in TECRL patients.
Mutations in the novel TECRL gene were identified in patients with malignant exercise‐induced arrhythmias. Increased triggered electrical activity upon stimulation in patient‐specific hiPSC‐CMs was rescued by the antiarrhythmic drug flecainide.
Leigh syndrome French Canadian type (LSFC) is a recessive neurodegenerative disease characterized by tissue-specific deficiency in cytochrome c oxidase (COX), the fourth complex in the oxidative ...phosphorylation system. LSFC is caused by mutations in the leucine rich pentatricopeptide repeat containing gene (
). Most LSFC patients in Quebec are homozygous for an A354V substitution that causes a decrease in the expression of the LRPPRC protein. While LRPPRC is ubiquitously expressed and is involved in multiple cellular functions, tissue-specific expression of LRPPRC and COX activity is correlated with clinical features. In this proof-of-principle study, we developed human induced pluripotent stem cell (hiPSC)-based models from fibroblasts taken from a patient with LSFC, homozygous for the
*354V allele, and from a control, homozygous for the
*A354 allele. Specifically, for both of these fibroblast lines we generated hiPSC, hiPSC-derived cardiomyocytes (hiPSC-CMs) and hepatocyte-like cell (hiPSC-HLCs) lines, as well as the three germ layers. We observed that LRPPRC protein expression is reduced in all cell lines/layers derived from LSFC patient compared to control cells, with a reduction ranging from ∼70% in hiPSC-CMs to undetectable levels in hiPSC-HLC, reflecting tissue heterogeneity observed in patient tissues. We next performed exploratory analyses of these cell lines and observed that COX protein expression was reduced in all cell lines derived from LSFC patient compared to control cells. We also observed that mutant LRPPRC was associated with altered expression of key markers of endoplasmic reticulum stress response in hiPSC-HLCs but not in other cell types that were tested. While this demonstrates feasibility of the approach to experimentally study genotype-based differences that have tissue-specific impacts, this study will need to be extended to a larger number of patients and controls to not only validate the current observations but also to delve more deeply in the pathogenic mechanisms of LSFC.
There has been a resurgence of interest for the field of cardiac metabolism catalyzed by evidence demonstrating a role of metabolic dysregulation in the pathogenesis of heart disease as well as the ...increased need for new therapeutic targets for patients with these diseases. In this regard, measuring substrate fluxes is critical in providing insight into the dynamics of cellular metabolism and in delineating the regulation of metabolite production and utilization. This chapter provides a comprehensive description of concepts, guidelines, and tips to assess metabolic fluxes relevant to energy substrate metabolism using (13)C-labeled substrates and (13)C-isotopomer analysis by gas chromatography-mass spectrometry (GC-MS), and the ex vivo working heart as study model. The focus will be on the mouse and on flux parameters, which are commonly assessed in the field, namely, those relevant to substrate selection for energy metabolism, specifically the relative contribution of carbohydrate (glucose, lactate, and pyruvate) and fatty acid oxidation to acetyl-CoA formation for citrate synthesis, glycolysis, as well as anaplerosis. We provide detailed procedures for the heart isolation and perfusion in the working mode as well as for sample processing for metabolite extraction and analysis by GC-MS and subsequent data processing for calculation of metabolic flux parameters. Finally, we address practical considerations and discuss additional applications and future challenges.
High plasma leucine levels strongly correlate with type 2 diabetes. Studies of muscle cells have suggested that leucine alters the insulin response for glucose transport by activating an ...insulin-negative feedback loop driven by the mammalian target of rapamycin/p70 ribosomal S6 kinase (mTOR/p70S6K) pathway. Here, we examined the molecular mechanism involved in leucine's action on cardiac glucose uptake. Leucine was indeed able to curb glucose uptake after insulin stimulation in both cultured cardiomyocytes and perfused hearts. Although leucine activated mTOR/p70S6K, the mTOR inhibitor rapamycin did not prevent leucine's inhibitory action on glucose uptake, ruling out the contribution of the insulin-negative feedback loop. α-Ketoisocaproate, the first metabolite of leucine catabolism, mimicked leucine's effect on glucose uptake. Incubation of cardiomyocytes with
Cleucine ascertained its metabolism to ketone bodies (KBs), which had a similar negative impact on insulin-stimulated glucose transport. Both leucine and KBs reduced glucose uptake by affecting translocation of glucose transporter 4 (GLUT4) to the plasma membrane. Finally, we found that leucine elevated the global protein acetylation level. Pharmacological inhibition of lysine acetyltransferases counteracted this increase in protein acetylation and prevented leucine's inhibitory action on both glucose uptake and GLUT4 translocation. Taken together, these results indicate that leucine metabolism into KBs contributes to inhibition of cardiac glucose uptake by hampering the translocation of GLUT4-containing vesicles via acetylation. They offer new insights into the establishment of insulin resistance in the heart.
Catabolism of the branched-chain amino acid leucine into ketone bodies efficiently inhibits cardiac glucose uptake through decreased translocation of glucose transporter 4 to the plasma membrane. Leucine increases protein acetylation. Pharmacological inhibition of acetylation reverses leucine's action, suggesting acetylation involvement in this phenomenon.Listen to this article's corresponding podcast at http://ajpheart.podbean.com/e/leucine-metabolism-inhibits-cardiac-glucose-uptake/.
Very-long chain acyl-CoA dehydrogenase (VLCAD) catalyzes the initial step of mitochondrial long chain (LC) fatty acid β-oxidation (FAO). Inherited VLCAD deficiency (VLCADD) predisposes to neonatal ...arrhythmias whose pathophysiology is still not understood. We hypothesized that VLCADD results in global disruption of cardiac complex lipid homeostasis, which may set conditions predisposing to arrhythmia. To test this, we assessed the cardiac lipidome and related molecular markers in seven-month-old VLCAD
mice, which mimic to some extent the human cardiac phenotype. Mice were sacrificed in the fed or fasted state after receiving for two weeks a chow or a high-fat diet (HFD), the latter condition being known to worsen symptoms in human VLCADD. Compared to their littermate counterparts, HFD/fasted VLCAD
mouse hearts displayed the following lipid alterations: (1) Lower LC, but higher VLC-acylcarnitines accumulation, (2) higher levels of arachidonic acid (AA) and lower docosahexaenoic acid (DHA) contents in glycerophospholipids (GPLs), as well as (3) corresponding changes in pro-arrhythmogenic AA-derived isoprostanes and thromboxane B
(higher), and anti-arrythmogenic DHA-derived neuroprostanes (lower). These changes were associated with remodeling in the expression of gene or protein markers of (1) GPLs remodeling: higher calcium-dependent phospholipase A2 and lysophosphatidylcholine-acyltransferase 2, (2) calcium handling perturbations, and (3) endoplasmic reticulum stress. Altogether, these results highlight global lipid dyshomeostasis beyond FAO in VLCAD
mouse hearts, which may set conditions predisposing the hearts to calcium mishandling and endoplasmic reticulum stress and thereby may contribute to the pathogenesis of arrhythmias in VLCADD in mice as well as in humans.
The purpose of this study was to determine in vivo myocardial energy metabolism and function in a nutritional model of type 2 diabetes. Wistar rats rendered insulin-resistant and mildly ...hyperglycemic, hyperinsulinemic, and hypertriglyceridemic with a high-fructose/high-fat diet over a 6-wk period with injection of a small dose of streptozotocin (HFHFS) and control rats were studied using micro-PET (microPET) without or with a euglycemic hyperinsulinemic clamp. During glucose clamp, myocardial metabolic rate of glucose measured with (18)Ffluorodeoxyglucose ((18)FFDG) was reduced by approximately 81% (P < 0.05), whereas myocardial plasma nonesterified fatty acid (NEFA) uptake as determined by (18)Ffluorothia-6-heptadecanoic acid ((18)FFTHA) was not significantly changed in HFHFS vs. control rats. Myocardial oxidative metabolism as assessed by (11)Cacetate and myocardial perfusion index as assessed by (13)Nammonia were similar in both groups, whereas left ventricular ejection fraction as assessed by microPET was reduced by 26% in HFHFS rats (P < 0.05). Without glucose clamp, NEFA uptake was approximately 40% lower in HFHFS rats (P < 0.05). However, myocardial uptake of (18)FFTHA administered by gastric gavage was significantly higher in HFHFS rats (P < 0.05). These abnormalities were associated with reduced Glut4 mRNA expression and increased Cd36 mRNA expression and mitochondrial carnitine palmitoyltransferase 1 activity (P < 0.05). HFHFS rats display type 2 diabetes complicated by left ventricular contractile dysfunction with profound reduction in myocardial glucose utilization, activation of fatty acid metabolic pathways, and preserved myocardial oxidative metabolism, suggesting reduced myocardial metabolic efficiency. In this model, increased myocardial fatty acid exposure likely occurs from circulating triglyceride, but not from circulating plasma NEFA.
Ventricular arrhythmias and sudden cardiac deaths are among the leading causes of mortality in patients with heart failure, and the underlying mechanisms remain incompletely understood. Chronic ...elevation of angiotensin II (ANGII) is known to be one of the main contributors to heart failure.
We tested whether ANGII can alter ventricular conduction and Na(+) current using transgenic mice with cardiomyocyte-restricted overexpression of ANGII type 1 receptor (AT1R).
We used surface electrocardiograms along with current- and voltage-clamp techniques to characterize the electrophysiological properties of AT1R mice while the underlying regulatory mechanisms were explored using reverse transcription/quantitative polymerase chain reaction, Western blots, and immunofluorescence techniques.
Electrophysiological data indicated that chronic AT1R activation in ventricular myocytes caused a 60% reduction in Na(+) current density that slowed the maximal velocity of the action potential upstroke, leading to a prolongation of the QRS complex. These changes occur independently of cardiac hypertrophy, suggesting a direct role for ANGII/AT1R in slowing ventricular conduction. Western blots demonstrated a selective increase in sarcolemmal protein kinase Cα (PKCα) in AT1R mice, indicating PKCα activation. Furthermore, immunofluorescence analysis showed reorganization of PKCα expression to sarcolemma and colocalization with NaV1.5 in AT1R myocytes. The involvement of PKCα in regulating Na(+) current was subsequently demonstrated in human-induced pluripotent stem cell-derived cardiomyocytes where ANGII treatment reduced Na(+) current density. Concomitant treatment with αV5-3, a PKCα translocation inhibitor peptide, blocked the ANGII effect.
Overall, this study suggests that in mouse and human cardiomyocytes, PKCα is an important mediator of the ANGII-induced reduction in Na(+) current and may contribute to ventricular arrhythmias.
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