Energetic Dysfunction Research Articles

Energetic Dysfunction Is Mediated by Mitochondrial Reactive Oxygen Species and Precedes Structural Remodeling in Metabolic Heart Disease.

Aims: Metabolic syndrome is associated with metabolic heart disease (MHD) that is characterized by left ventricular (LV) hypertrophy, interstitial fibrosis, contractile dysfunction, and mitochondrial dysfunction. Overexpression of catalase in mitochondria (transgenic expression of catalase targeted to the mitochondria [mCAT]) prevents the structural and functional features of MHD caused by a high-fat, high-sucrose (HFHS) diet for ≥4 months. However, it is unclear whether the effect of mCAT is due to prevention of reactive oxygen species (ROS)-mediated cardiac remodeling, a direct effect on mitochondrial function, or both. To address this question, we measured myocardial function and energetics in mice, with or without mCAT, after 1 month of HFHS, before the development of cardiac structural remodeling. Results: HFHS diet for 1 month had no effect on body weight, heart weight, LV structure, myocyte size, or interstitial fibrosis. Isolated cardiac mitochondria from HFHS-fed mice produced 2.2- to 3.8-fold more H2O2, and 16%-29% less adenosine triphosphate (ATP). In isolated beating hearts from HFHS-fed mice, [phosphocreatine (PCr)] and the free energy available for ATP hydrolysis (ΔG∼ATP) were decreased, and they failed to increase with work demands. Overexpression of mCAT normalized ROS and ATP production in isolated mitochondria, and it corrected myocardial [PCr] and ΔG∼ATP in the beating heart. Innovation: This is the first demonstration that in MHD, mitochondrial ROS mediate energetic dysfunction that is sufficient to impair contractile function. Conclusion: ROS produced and acting in the mitochondria impair myocardial energetics, leading to slowed relaxation and decreased contractile reserve. These effects precede structural remodeling and are corrected by mCAT, indicating that ROS-mediated energetic impairment, per se, is sufficient to cause contractile dysfunction in MHD.

Метаболічна підтримка хворих з інфарктом міокарда з порушеннями функціонального стану печінки

The aim – to improve the existing therapeutic programs for patients with myocardial infarction (MI) by differentiated approach to therapy depending on the presence or absence of liver dysfunction. Materials and methods. 149 patients with acute MI were included to the study – 114 (76.5 %) men and 35 (23.5 %) women of average age 59.9±1.0 years. A comparative clinical-laboratory and instrumental examination was performed in these groups: 107 patients with MI with functional liver disorders (main group) and 42 patients with MI without functional liver disorders (comparison group). For the study of intracardiac hemodynamics, echocardiography was performed in B-mode. 52 patients of main group and 22 patients of comparison group received standard treatment for MI (ACE-inhibitors, β-blockers, clopidogrel and acetylsalicylic acid, anticoagulant therapy, statins). The other 55 patients of main group and 20 patients of comparison group received modified treatment with the addition of the antioxidant bioflavonoid – quercetin to the standard scheme.Results and discussion. In the initial condition of patients of main group there are more pronounced violations of central and peripheral hemodynamics which cause the disorders of protein synthesis, detoxification, energetic dysfunction of liver and lipid metabolism. At the same time there is an excessive activation of lipid peroxidation, suppressed activity of enzymes of the antioxidant protection system, deepening of pathological processes in the heart and in the liver. Inclusion of bioflavonoid quercetin to the complex therapy of patients with myocardial infarction provided the improvement of hemodynamic parameters – increased EF by 22.89 % and elimination of diastolic dysfunction, which were accompanied by normalization of clinical and laboratory parameters of liver function disorders – normalization of levels of bilirubin and ALT, GGT, ALP on 14-th day and reduction of LDL-level by 25.61 % after 3 weeks was marked. Conclusions. Introduction of bioflavonoid quercetin to the complex therapy of patients with MI provided improvement of hemodynamic parameters and restoration of oxidation-reduction equilibrium, reduction of clinical and laboratory manifestations of liver dysfunction and contributed to a significant decrease in the frequency of MI complications development. Restoration of parameters of cardiac hemodynamics, metabolic processes and liver functions provides a significant reduction in the rate of development of acute heart failure by 63.9 %, early post-infarction angina by 22.2 %, rhythm and conduction disorders by 43.6 %.

Role of cerebral hypoperfusion in multiple sclerosis (ROCHIMS): study protocol for a proof-of-concept randomized controlled trial with bosentan

BackgroundAxonal degeneration is related to long-term disability in patients with multiple sclerosis (MS). The underlying mechanism remains ill understood but appears to involve axonal energetic dysfunction. A globally impaired cerebral blood flow (CBF) has been observed in the normal-appearing white matter (NAWM) of patients with MS, which is probably related to astrocytic overexpression of endothelin-1 (ET-1). Cerebral hypoperfusion has been associated with reduced mitochondrial activity and disabling symptoms (e.g. fatigue and cognitive decline) of MS. Countering this process could therefore be beneficial in the disease course. Short-term CBF restoration with a single 62.5-mg dose of the ET-1 receptor antagonist bosentan has already been demonstrated in patients with MS.MethodsThe ROCHIMS study is a proof-of-concept double-blind randomized clinical trial in which patients with relapsing-remitting MS will receive either 62.5 mg bosentan or matching placebo twice daily during 28 ± 2 days. Clinical evaluation and brain magnetic resonance imaging (MRI) will be performed at baseline and treatment termination. Based on previous work, we expect a global increase of CBF in the individuals treated with bosentan. The primary outcome measure is the change of N-acetyl aspartate in centrum semiovale NAWM, which is a marker of regional axonal mitochondrial activity. Other parameters of interest include changes in fatigue, cognition, motor function, depression, and brain volume.DiscussionWe hypothesize that restoring cerebral hypoperfusion in MS patients improves axonal metabolism.Early positive effects on fatigue and cognitive dysfunction related to MS might additionally be detected. There is a medical need for drugs that can slow down the progressive axonal degeneration in MS, making this an important topic of interest.Trial registrationClinical Trials Register, EudraCT 2017-001253-13. Registered on 15 February 2018.

A new approach to find biomarkers in chronic fatigue syndrome/myalgic encephalomyelitis (CFS/ME) by single-cell Raman micro-spectroscopy.

Chronic fatigue syndrome (CFS), also called myalgic encephalomyelitis (ME), is a debilitating disorder characterized by physical and mental exhaustion. Mitochondrial and energetic dysfunction has been investigated in CFS patients due to a hallmark relationship with fatigue; however, no consistent conclusion has yet been achieved. Single-cell Raman spectra (SCRS) are label-free biochemical profiles, indicating phenotypic fingerprints of single cells. In this study, we applied a new approach using single-cell Raman microspectroscopy (SCRM) to examine ρ0 cells that lack mitochondrial DNA (mtDNA), and peripheral blood mononuclear cells (PBMCs) from CFS patients and healthy controls. The experimental results show that Raman bands associated with phenylalanine in ρ0 cells and CFS patient PBMCs were significantly higher than those of the wild-type model and healthy controls. As similar changes were observed in the ρ0 cell model with a known deficiency in the mitochondrial respiratory chain as well as in CFS patients, our results suggest that the increase in cellular phenylalanine may be related to mitochondrial/energetic dysfunction in both systems. Interestingly, phenylalanine can be used as a potential biomarker for the diagnosis of CFS by SCRM. A machine learning classification model achieved an accuracy rate of 98% correctly assigning Raman spectra to either the CFS group or the control group. SCRM combined with a machine learning algorithm therefore has the potential to become a diagnostic tool for CFS.

Reduced cardiomyocyte Na+ current in the age-dependent murine Pgc-1β-/- model of ventricular arrhythmia.

Peroxisome proliferator‐activated receptor‐γ coactivator‐1 deficient (Pgc‐1β −/−) murine hearts model the increased, age‐dependent, ventricular arrhythmic risks attributed to clinical conditions associated with mitochondrial energetic dysfunction. These were accompanied by compromised action potential (AP) upstroke rates and impaired conduction velocities potentially producing arrhythmic substrate. We tested a hypothesis implicating compromised Na+ current in these electrophysiological phenotypes by applying loose patch‐clamp techniques in intact young and aged, wild‐type (WT) and Pgc‐1β −/−, ventricular cardiomyocyte preparations for the first time. This allowed conservation of their in vivo extracellular and intracellular conditions. Depolarising steps elicited typical voltage‐dependent activating and inactivating inward Na+ currents with peak amplitudes increasing or decreasing with their respective activating or preceding inactivating voltage steps. Two‐way analysis of variance associated Pgc‐1β −/− genotype with independent reductions in maximum peak ventricular Na+ currents from −36.63 ± 2.14 (n = 20) and −35.43 ± 1.96 (n = 18; young and aged WT, respectively), to −29.06 ± 1.65 (n = 23) and −27.93 ± 1.63 (n = 20; young and aged Pgc‐1β −/−, respectively) pA/μm2 (p < 0.0001), without independent effects of, or interactions with age. Voltages at half‐maximal current V*, and steepness factors k in plots of voltage dependences of both Na+ current activation and inactivation, and time constants for its postrepolarisation recovery from inactivation, remained indistinguishable through all experimental groups. So were the activation and rectification properties of delayed outward (K+) currents, demonstrated from tail currents reflecting current recoveries from respective varying or constant voltage steps. These current–voltage properties directly implicate decreases specifically in maximum available Na+ current with unchanged voltage dependences and unaltered K+ current properties, in proarrhythmic reductions in AP conduction velocity in Pgc‐1β−/− ventricles.

Target and untargeted GC–MS based metabolomic study of mouse optic nerve and its potential in the study of neurological visual diseases

The optic nerve is made of highly specialized neurons and the energetic supply to their axons is crucial due to their great demand. The energy comes basically through the oxidative phosphorylation in the mitochondria, supported by glial cells metabolism. Mitochondrial dysfunction is a shared feature encountered within the optic neuropathies, including Leber’s Hereditary Optic Neuropathy, Leigh’s Syndrome, or Kjer’s syndrome.In an effort to investigate the metabolic alterations produced within the optic nerve in a mutant mouse model of Neurological Visual Disease (NVD), a rapid, robust, and efficient one-single phase extraction methodology has been developed and validated for the GC–MS platform. Once the method was successfully validated for lactic acid and pyruvic acid as markers of an adequate optic nerve function, the protocol was applied to unveil the metabolomic signature of the wild-type mouse optic nerve. Along the chromatographic profile of the optic nerve, 94 peaks were identified and, to our knowledge, for the first time. Afterwards, a targeted metabolomics analysis was performed to quantify lactic acid and pyruvic acid in the NDV mice group (n = 8) and its corresponding wild-type (n = 8). Finally, an untargeted metabolomic study was carried out and univariate and multivariate data analyses showed 34 compounds modified in the optic nerve of the mouse with NVD mutation. Then, the metabolic reaction network of the identified metabolites highlighted alterations in the catabolism of proteins, TCA cycle, and urea cycle, reflecting a mitochondrial energetic dysfunction. Taken together, this metabolomic study has proven to be suited for the study of optic neuropathies.

Ex Vivo Cardiotoxicity of Antineoplastic Casiopeinas Is Mediated through Energetic Dysfunction and Triggered Mitochondrial-Dependent Apoptosis.

Casiopeinas are a group of copper-based antineoplastic molecules designed as a less toxic and more therapeutic alternative to cisplatin or Doxorubicin; however, there is scarce evidence about their toxic effects on the whole heart and cardiomyocytes. Given this, rat hearts were perfused with Casiopeinas or Doxorubicin and the effects on mechanical performance, energetics, and mitochondrial function were measured. As well, the effects of Casiopeinas-triggered cell death were explored in isolated cardiomyocytes. Casiopeinas III-Ea, II-gly, and III-ia induced a progressive and sustained inhibition of heart contractile function that was dose- and time-dependent with an IC50 of 1.3 ± 0.2, 5.5 ± 0.5, and 10 ± 0.7 μM, correspondingly. Myocardial oxygen consumption was not modified at their respective IC50, although ATP levels were significantly reduced, indicating energy impairment. Isolated mitochondria from Casiopeinas-treated hearts showed a significant loss of membrane potential and reduction of mitochondrial Ca2+ retention capacity. Interestingly, Cyclosporine A inhibited Casiopeinas-induced mitochondrial Ca2+ release, which suggests the involvement of the mitochondrial permeability transition pore opening. In addition, Casiopeinas reduced the viability of cardiomyocytes and stimulated the activation of caspases 3, 7, and 9, demonstrating a cell death mitochondrial-dependent mechanism. Finally, the early perfusion of Cyclosporine A in isolated hearts decreased Casiopeinas-induced dysfunction with reduction of their toxic effect. Our results suggest that heart cardiotoxicity of Casiopeinas is similar to that of Doxorubicin, involving heart mitochondrial dysfunction, loss of membrane potential, changes in energetic metabolites, and apoptosis triggered by mitochondrial permeability.

Age-dependent electrocardiographic changes in Pgc-1β deficient murine hearts.

SummaryIncreasing evidence implicates chronic energetic dysfunction in human cardiac arrhythmias. Mitochondrial impairment through Pgc‐1β knockout is known to produce a murine arrhythmic phenotype. However, the cumulative effect of this with advancing age and its electrocardiographic basis have not been previously studied. Young (12‐16 weeks) and aged (>52 weeks), wild type (WT) (n = 5 and 8) and Pgc‐1β−/− (n = 9 and 6), mice were anaesthetised and used for electrocardiographic (ECG) recordings. Time intervals separating successive ECG deflections were analysed for differences between groups before and after β1‐adrenergic (intraperitoneal dobutamine 3 mg/kg) challenge. Heart rates before dobutamine challenge were indistinguishable between groups. The Pgc‐1β−/− genotype however displayed compromised nodal function in response to adrenergic challenge. This manifested as an impaired heart rate response suggesting a functional defect at the level of the sino‐atrial node, and a negative dromotropic response suggesting an atrioventricular conduction defect. Incidences of the latter were most pronounced in the aged Pgc‐1β−/− mice. Moreover, Pgc‐1β−/− mice displayed electrocardiographic features consistent with the existence of a pro‐arrhythmic substrate. Firstly, ventricular activation was prolonged in these mice consistent with slowed action potential conduction and is reported here for the first time. Additionally, Pgc‐1β−/− mice had shorter repolarisation intervals. These were likely attributable to altered K+ conductance properties, ultimately resulting in a shortened QTc interval, which is also known to be associated with increased arrhythmic risk. ECG analysis thus yielded electrophysiological findings bearing on potential arrhythmogenicity in intact Pgc‐1β−/− systems in widespread cardiac regions.

Pro-arrhythmic atrial phenotypes in incrementally paced murine Pgc1β-/- hearts: effects of age.

New Findings What is the central question of this study? Can we experimentally replicate atrial pro‐arrhythmic phenotypes associated with important chronic clinical conditions, including physical inactivity, obesity, diabetes mellitus and metabolic syndrome, compromising mitochondrial function, and clarify their electrophysiological basis? What is the main finding and its importance? Electrocardiographic and intracellular cardiomyocyte recording at progressively incremented pacing rates demonstrated age‐dependent atrial arrhythmic phenotypes in Langendorff‐perfused murine Pgc1β −/− hearts for the first time. We attributed these to compromised action potential conduction and excitation wavefronts, whilst excluding alterations in recovery properties or temporal electrophysiological instabilities, clarifying these pro‐arrhythmic changes in chronic metabolic disease. Atrial arrhythmias, most commonly manifesting as atrial fibrillation, represent a major clinical problem. The incidence of atrial fibrillation increases with both age and conditions associated with energetic dysfunction. Atrial arrhythmic phenotypes were compared in young (12–16 week) and aged (>52 week) wild‐type (WT) and peroxisome proliferative activated receptor, gamma, coactivator 1 beta (Ppargc1b)‐deficient (Pgc1β −/−) Langendorff‐perfused hearts, previously used to model mitochondrial energetic disorder. Electrophysiological explorations were performed using simultaneous whole‐heart ECG and intracellular atrial action potential (AP) recordings. Two stimulation protocols were used: an S1S2 protocol, which imposed extrasystolic stimuli at successively decremented intervals following regular pulse trains; and a regular pacing protocol at successively incremented frequencies. Aged Pgc1β −/− hearts showed greater atrial arrhythmogenicity, presenting as atrial tachycardia and ectopic activity. Maximal rates of AP depolarization (dV/dt max) were reduced in Pgc1β −/− hearts. Action potential latencies were increased by the Pgc1β −/− genotype, with an added interactive effect of age. In contrast, AP durations to 90% recovery (APD90) were shorter in Pgc1β −/− hearts despite similar atrial effective recovery periods amongst the different groups. These findings accompanied paradoxical decreases in the incidence and duration of alternans in the aged and Pgc1β −/− hearts. Limiting slopes of restitution curves of APD90 against diastolic interval were correspondingly reduced interactively by Pgc1β −/− genotype and age. In contrast, reduced AP wavelengths were associated with Pgc1β −/− genotype, both independently and interacting with age, through the basic cycle lengths explored, with the aged Pgc1β −/− hearts showing the shortest wavelengths. These findings thus implicate AP wavelength in possible mechanisms for the atrial arrhythmic changes reported here.

Effects of ageing on pro-arrhythmic ventricular phenotypes in incrementally paced murine Pgc-1\u03b2\u2212/\u2212 hearts

A range of chronic clinical conditions accompany cardiomyocyte energetic dysfunction and constitute independent risk factors for cardiac arrhythmia. We investigated pro-arrhythmic and arrhythmic phenotypes in energetically deficient C57BL mice with genetic ablation of the mitochondrial promoter peroxisome proliferator-activated receptor-γ coactivator-1β (Pgc-1β), a known model of ventricular arrhythmia. Pro-arrhythmic and cellular action potential (AP) characteristics were compared in intact Langendorff-perfused hearts from young (12–16 week) and aged (> 52 week), wild-type (WT) and Pgc-1β−/− mice. Simultaneous electrocardiographic and intracellular microelectrode recordings were made through successive trains of 100 regular stimuli at progressively incremented heart rates. Aged Pgc-1β−/− hearts displayed an increased incidence of arrhythmia compared to other groups. Young and aged Pgc-1β−/− hearts showed higher incidences of alternans in both AP activation (maximum AP upshoot velocity (dV/dt)max and latency), recovery (action potential duration (APD90) and resting membrane potential (RMP) characteristics compared to WT hearts. This was particularly apparent at lower pacing frequencies. These findings accompanied reduced (dV/dt)max and increased AP latency values in the Pgc-1β−/− hearts. APs observed prior to termination of the protocol showed lower (dV/dt)max and longer AP latencies, but indistinguishable APD90 and RMPs in arrhythmic compared to those in non-arrhythmic hearts. APD restitution analysis showed that Pgc-1β−/− and WT hearts showed similar limiting gradients. However, Pgc-1β−/− hearts had shortened plateau AP wavelengths, particularly in aged Pgc-1β−/− hearts. Pgc-1β−/− hearts therefore show pro-arrhythmic instabilities attributable to altered AP conduction and activation rather than recovery characteristics.

Mesencephalic astrocyte-derived neurotrophic factor alleviated 6-OHDA-induced cell damage via ROS-AMPK/mTOR mediated autophagic inhibition

Autophagy and apoptosis are commonly involved in the dopaminergic neuron damage in the pathogenesis of Parkinson's disease. Recently, the autophagy pathway is thought to be critical to the process of PD. Therefore, the regulation of autophagy may be a potential strategy for PD treatment. Mesencephalic astrocyte-derived neurotrophic factor (MANF) has been reported to have neuroprotective effects through anti-apoptosis, anti-oxidative, and anti-inflammatory mechanisms in PD. In this study, we investigated the role of autophagy system in MANF-mediated neuroprotection against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity. Our results showed that MANF protected SH-SY5Y cells against 6-OHDA-induced cell viability decrease and apoptosis by inhibiting autophagy. Mitochondrion damage and energetic dysfunction triggered by reactive oxidative stress (ROS) accumulation were also alleviated by MANF treatment. Furthermore, MANF downregulated phosphorylation of AMP-activated protein kinase (AMPK), a cellular energy sensor and regulator, but upregulated phosphorylation of Mammalian target of rapamycin (mTOR) under energy depletion conditions, indicating AMPK/mTOR signaling pathway is involved in the autophagic inhibition of MANF. These results suggest that autophagic inhibition provides protective mechanism of MANF in 6-OHDA-induced SH-SY5Y cell death and this inhibition is associated with AMPK/mTOR pathway.

An impaired metabolism of nucleotides underpins a novel mechanism of cardiac remodeling leading to Huntington's disease related cardiomyopathy

Huntington's disease (HD) is mainly thought of as a neurological disease, but multiple epidemiological studies have demonstrated a number of cardiovascular events leading to heart failure in HD patients. Our recent studies showed an increased risk of heart contractile dysfunction and dilated cardiomyopathy in HD pre-clinical models. This could potentially involve metabolic remodeling, that is a typical feature of the failing heart, with reduced activities of high energy phosphate generating pathways. In this study, we sought to identify metabolic abnormalities leading to HD-related cardiomyopathy in pre-clinical and clinical settings. We found that HD mouse models developed a profound deterioration in cardiac energy equilibrium, despite AMP-activated protein kinase hyperphosphorylation. This was accompanied by a reduced glucose usage and a significant deregulation of genes involved in de novo purine biosynthesis, in conversion of adenine nucleotides, and in adenosine metabolism. Consequently, we observed increased levels of nucleotide catabolites such as inosine, hypoxanthine, xanthine and uric acid, in murine and human HD serum. These effects may be caused locally by mutant HTT, via gain or loss of function effects, or distally by a lack of trophic signals from central nerve stimulation. Either may lead to energy equilibrium imbalances in cardiac cells, with activation of nucleotide catabolism plus an inhibition of re-synthesis. Our study suggests that future therapies should target cardiac mitochondrial dysfunction to ameliorate energetic dysfunction. Importantly, we describe the first set of biomarkers related to heart and skeletal muscle dysfunction in both pre-clinical and clinical HD settings.

Influence of metabolic dysfunction on cardiac mechanics in decompensated hypertrophy and heart failure

Alterations in energetic state of the myocardium are associated with decompensated heart failure in humans and in animal models. However, the functional consequences of the observed changes in energetic state on mechanical function are not known. The primary aim of the study was to quantify mechanical/energetic coupling in the heart and to determine if energetic dysfunction can contribute to mechanical failure. A secondary aim was to apply a quantitative systems pharmacology analysis to investigate the effects of drugs that target cross-bridge cycling kinetics in heart failure-associated energetic dysfunction. Herein, a model of metabolite- and calcium-dependent myocardial mechanics was developed from calcium concentration and tension time courses in rat cardiac muscle obtained at different lengths and stimulation frequencies. The muscle dynamics model accounting for the effect of metabolites was integrated into a model of the cardiac ventricles to simulate pressure-volume dynamics in the heart. This cardiac model was integrated into a simple model of the circulation to investigate the effects of metabolic state on whole-body function. Simulations predict that reductions in metabolite pools observed in canine models of heart failure can cause systolic dysfunction, blood volume expansion, venous congestion, and ventricular dilation. Simulations also predict that myosin-activating drugs may partially counteract the effects of energetic state on cross-bridge mechanics in heart failure while increasing myocardial oxygen consumption. Our model analysis demonstrates how metabolic changes observed in heart failure are alone sufficient to cause systolic dysfunction and whole-body heart failure symptoms.

Sepsis induces long-term metabolic and mitochondrial muscle stem cell dysfunction amenable by mesenchymal stem cell therapy.

Sepsis, or systemic inflammatory response syndrome, is the major cause of critical illness resulting in admission to intensive care units. Sepsis is caused by severe infection and is associated with mortality in 60% of cases. Morbidity due to sepsis is complicated by neuromyopathy, and patients face long-term disability due to muscle weakness, energetic dysfunction, proteolysis and muscle wasting. These processes are triggered by pro-inflammatory cytokines and metabolic imbalances and are aggravated by malnutrition and drugs. Skeletal muscle regeneration depends on stem (satellite) cells. Herein we show that mitochondrial and metabolic alterations underlie the sepsis-induced long-term impairment of satellite cells and lead to inefficient muscle regeneration. Engrafting mesenchymal stem cells improves the septic status by decreasing cytokine levels, restoring mitochondrial and metabolic function in satellite cells, and improving muscle strength. These findings indicate that sepsis affects quiescent muscle stem cells and that mesenchymal stem cells might act as a preventive therapeutic approach for sepsis-related morbidity.

Obesity, diet and nutraceuticals.

Migraine is the most prevalent neurological disorder worldwide, affecting on average 12% of the general population. Recent data support migraine as part of the metabolic syndrome (MetSyn) spectrum. The evidence of an association between obesity, insulin resistance, and migraine, all having in common the ability to favor a general pro-inflammatory state, reinforces this view. Under a neuronal point of view, another possible link between migraine and MetSyn is, other than the inflammatory state, the cellular energetic deficit. Indeed, it is well known that a mitochondrial energetic dysfunction may play a role in migraine pathogenesis, that MetSyn could further impair mitochondrial metabolism increasing oxidative stress, and that nutraceuticals involved in mitochondrial oxidative phosphorylation (OXPHOS) metabolism (Riboflavin, CoQ10, and magnesium) can be used as migraine prophylactic treatments. Here we review the evidence of MetSyn and migraine association and possible therapeutic implications. There is a close relationship between migraine and MetSyn, especially insulin resistance and obesity. More in detail, obesity is regarded as an important factor of chronification for migraineur patients, while weight-loss seems to have a protective effect. The main player of this association seems to be the obesity-related inflammation and an adequate slimming diet can act to reduce this inflammatory mechanism. In fact, inflammatory cytokines are higher in obese subjects and normalized by weight reduction. Moreover, several foods could act as triggers for some patients by an inflammatory antibody mediated pathway, by a direct histamine or other vasoactive peptides effect, or by modifying the quality and quantity of fat intake. Also, altered insulin metabolism could be related to leptins and nitric oxide (NO) stress, both inducing inflammatory effects involved in migraine pathogenesis. Some prophylactic treatments induce increase of weight, insulin and leptins that can counteract the therapeutic effect; it is the “prophylactic paradox”: a long lasting migraine could be worsened by the drug-induced obesity. Therefore, there is the rationale for the adoption of specific dietetic patterns as complementary treatment in migraine management. In particular, the ketogenic diet is worthy of mention because of its multiple effects on the above-mentioned mechanisms, since it induces a significant weight-loss (when prescribed as very-low calorie diet), normalizes leptins and hyperinsulinemia, improves mitochondrial OXPHOS metabolism, and has a ketone bodies induced anti-inflammatory effect. In summary, more attention should be paid to metabolic implications of migraine and its treatment to further improve, or at least not worsen, patients’ conditions and reduce migraine-related comorbidities and complications.

Mitochondrial iron and energetic dysfunction distinguish fibroblasts and induced neurons from pantothenate kinase-associated neurodegeneration patients

Pantothenate kinase-associated neurodegeneration is an early onset autosomal recessive movement disorder caused by mutation of the pantothenate kinase-2 gene, which encodes a mitochondrial enzyme involved in coenzyme A synthesis. The disorder is characterised by high iron levels in the brain, although the pathological mechanism leading to this accumulation is unknown. To address this question, we tested primary skin fibroblasts from three patients and three healthy subjects, as well as neurons induced by direct fibroblast reprogramming, for oxidative status, mitochondrial functionality and iron parameters. The patients' fibroblasts showed altered oxidative status, reduced antioxidant defence, and impaired cytosolic and mitochondrial aconitase activities compared to control cells. Mitochondrial iron homeostasis and functionality analysis of patient fibroblasts indicated increased labile iron pool content and reactive oxygen species development, altered mitochondrial shape, decreased membrane potential and reduced ATP levels. Furthermore, analysis of induced neurons, performed at a single cell level, confirmed some of the results obtained in fibroblasts, indicating an altered oxidative status and signs of mitochondrial dysfunction, possibly due to iron mishandling. Thus, for the first time, altered biological processes have been identified in vitro in live diseased neurons. Moreover, the obtained induced neurons can be considered a suitable human neuronal model for the identification of candidate therapeutic compounds for this disease.

Neonatal iron supplementation potentiates oxidative stress, energetic dysfunction and neurodegeneration in the R6/2 mouse model of Huntington's disease

Huntington’s disease (HD) is a progressive neurodegenerative disorder caused by a CAG repeat expansion that encodes a polyglutamine tract in huntingtin (htt) protein. Dysregulation of brain iron homeostasis, oxidative stress and neurodegeneration are consistent features of the HD phenotype. Therefore, environmental factors that exacerbate oxidative stress and iron dysregulation may potentiate HD. Iron supplementation in the human population is common during infant and adult-life stages. In this study, iron supplementation in neonatal HD mice resulted in deterioration of spontaneous motor running activity, elevated levels of brain lactate and oxidized glutathione consistent with increased energetic dysfunction and oxidative stress, and increased striatal and motor cortical neuronal atrophy, collectively demonstrating potentiation of the disease phenotype. Oxidative stress, energetic, and anatomic markers of degeneration were not affected in wild-type littermate iron-supplemented mice. Further, there was no effect of elevated iron intake on disease outcomes in adult HD mice. We have demonstrated an interaction between the mutant huntingtin gene and iron supplementation in neonatal HD mice. Findings indicate that elevated neonatal iron intake potentiates mouse HD and promotes oxidative stress and energetic dysfunction in brain. Neonatal-infant dietary iron intake level may be an environmental modifier of human HD.

Abstract 1839: LKB1 deficiency enhances sensitivity to energetic stress induced by erlotinib treatment in non-small cell lung cancer (NSCLC) cells

Abstract The tumor suppressor LKB1 is inactivated in 20-30% of non-small cell lung cancer (NSCLC) patient tumors. Because LKB1 is the master kinase phosphorylating the energy-sensing kinase AMPK, dysregulation of the LKB1-AMPK signaling axis causes loss of cellular homeostasis in response to metabolic inhibition or external stress-induced mitochondrial insults. Erlotinib is used clinically in NSCLC patients to target mutationally activated EGFR and is known to induce mitochondrial-mediated apoptosis. We investigated the metabolic response and mitochondrial function of LKB1-deficient NSCLC cells treated with erlotinib. LKB1-deficient cells exhibited enhanced sensitivity to erlotinib treatment in vitro and in vivo despite having wild-type EGFR. We found that this enhanced response was due to altered energetic metabolism and mitochondrial dysfunction in LKB1-deficient cells, which prevents homestatic maintenance of ATP and reactive oxygen species (ROS) levels in response to erlotinib treatment. Despite negative regulation of mTOR activity by LKB1-AMPK signaling, erlotinib effectively blocked mTOR signaling in LKB1-deficient cells, resulting in inhibition of cell growth, as well as activation of apoptosis. These findings will be important for designing targeted treatments for LKB1-deficient NSCLC patient tumors harboring wild-type EGFR, which disrupt metabolic and signaling pathways known to regulate energy metabolism in these cells. Citation Format: Young Mi Whang, Serk In Park, Irina A. Trenary, Changki Lee, Jacob M. Kaufman, David P. Carbone, Jamey D. Young. LKB1 deficiency enhances sensitivity to energetic stress induced by erlotinib treatment in non-small cell lung cancer (NSCLC) cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1839. doi:10.1158/1538-7445.AM2014-1839

Impaired left ventricular mechanical and energetic function in mice after cardiomyocyte-specific excision of Serca2

Sarco(endo)plasmic reticulum Ca2+ -ATPase (SERCA)2 transports Ca2+ from the cytosol into the sarcoplasmic reticulum of cardiomyocytes and is essential for maintaining myocardial Ca2+ handling and thus the mechanical function of the heart. SERCA2 is a major ATP consumer in excitation-contraction coupling but is regarded to contribute to energetically efficient Ca2+ handling in the cardiomyocyte. Previous studies using cardiomyocyte-specific SERCA2 knockout (KO) mice have demonstrated that decreased SERCA2 activity reduces the Ca2+ transient amplitude and induces compensatory Ca2+ transport mechanisms that may lead to more inefficient Ca2+ transport. In this study, we examined the relationship between left ventricular (LV) function and myocardial O2 consumption (MVo2) in ex vivo hearts from SERCA2 KO mice to directly measure how SERCA2 elimination influences mechanical and energetic features of the heart. Ex vivo hearts from SERCA2 KO hearts developed mechanical dysfunction at 4 wk and demonstrated virtually no working capacity at 7 wk. In accordance with the reported reduction in Ca2+ transient amplitude in cardiomyocytes from SERCA2 KO mice, work-independent MVo2 was decreased due to a reduced energy cost of excitation-contraction coupling. As these hearts also showed a marked impairment in the efficiency of chemomechanical energy transduction (contractile efficiency, i.e, work-dependent MVo2), hearts from SERCA2 KO mice were found to be mechanically inefficient. This ex vivo evaluation of mechanical and energetic function in hearts from SERCA2 KO mice brings together findings from previous experimental and mathematical modeling-based studies and demonstrates that reduced SERCA2 activity not only leads to mechanical dysfunction but also to energetic dysfunction.

Metabolomic profiling of aging rat skeletal muscle

Skeletal muscles undergo progressive atrophy and pathologic remodeling with age. To better understand muscle senescence, we used metabolomic profiling to characterize ‘adult’ (15‐month‐old; n=8) and ‘old’ (32‐month‐old; n=8) FBN male rats. Gastrocnemius (gastroc) and soleus muscles were analyzed, as well as plasma. Compared to adult gastroc, old gastroc showed evidence of altered glucose metabolism, including accumulation of glycolytic, glycogenolytic, and pentose phosphate pathway intermediates. Pyruvate was elevated with age, yet tricarboxylic acid (TCA) cycle intermediates were reduced, and nicotinamide adenine dinucleotide was reduced 82% (p&lt;0.0001). Indicative of muscle atrophy, 3‐methylhistidine and free amino acids were elevated in old gastroc. The monounsaturated fatty acids oleate, cis‐vaccenate, and palmitoleate, also increased in old gastroc. Old soleus showed reductions in glycolytic and TCA cycle intermediates, and 39% (p&lt;0.0001) reduction in flavin adenine dinucleotide. Age‐related plasma biomarkers showing the largest % increases included glycocholate (218%), heme (122%), 1,5‐anhydroglucitol (119%), 1‐palmitoleoyl‐glycerophosphocholine (95%), palmitoleate (64%), and creatine (53%). These changes are consistent with reduced insulin sensitivity in aging. In sum, energetic dysfunction appears central to pathological metabolic processes in aging skeletal muscle.