Mitochondrial Metabolic Pathways Research Articles

Pharmacologic Modulation of Mitochondrial Pathogenic Changes in Alzheimer’s Aβ‐Challenged Neurons

Mitochondria play essential roles in maintaining the high levels of brain energy demands required to maintain physiological ion gradients across membranes critical for the generation of action potentials and to sustain transport systems at endothelial barriers. About 90% of the energy requirement is produced through glucose oxidation, a tightly controlled mitochondrial process leading to ATP production. As the major consumers of oxygen, mitochondria are also the most important generators of reactive oxygen species (ROS) as by‐products of the electron transport chain. Thus, it is not surprising that mitochondrial dysfunction is emerging as a major contributor to the pathobiology of age‐associated neurodegenerative disorders and is being considered a crucial player in the pathophysiology of Alzheimer’s disease (AD). In spite of numerous studies, the mechanistic links among ROS homeostasis, cellular metabolic alterations, and changes in cell bioenergetics, particularly at the level of synapsis and in relation to oligomeric forms of amyloid‐β (oligAβ), still remain elusive. Through a combination of classical biochemical and immunocytochemical approaches together with the real‐time assessment of global energy metabolism our studies provide insights into the detrimental role of oligAβ in neuronal cell lines and murine primary cortical neurons. Our findings demonstrate that oligAβ induces detrimental changes in mitochondrial function with loss of mitochondrial membrane potential, release of cytochrome C, and enhanced ROS generation. Assessment of global energy metabolism in the Seahorse metabolic analyzer after sequential addition of different modulators of the mitochondrial metabolic pathways, demonstrated an oligAβ‐mediated reduction in oxygen consumption affecting basal and maximal respiration capacity and causing decreased ATP production. Pharmacologic targeting of Aβ‐challenged neurons with compounds of known antioxidant activity restored mitochondrial integrity/function, rescuing metabolic and bioenergetic changes induced by oligAβ. Overall, our studies provide insights into the complex molecular mechanisms triggered by oligAβ which profoundly affect mitochondrial performance and highlight the potential of antioxidant pharmacologic targeting to ameliorate the metabolic/bioenergetics alterations encompassing the complex and multifactorial pathways affected in AD.Support or Funding InformationNIH grants AG059595 and AG051266, the BrightFocus Foundation grant A2015275S, and the Alzheimer’s Association ZEN‐14‐283969

Characterization of Crohn's Disease Subtypes in Treatment-Naïve Paediatric Patients Based on Differential Intestinal Mucosal Transcriptomic Biomarkers

Background: Crohn’s disease (CD) is a chronic idiopathic mucosal disease of the gastrointestinal tract that exhibits different phenotypes according to its location. Efforts have been made to differentiate CD subtypes at the genetic level, but the diagnosis of CD subtypes using biomarkers still requires standardization. The aim of this study is to identify differentially expressed genes (DEGs) between the ileum and colon of CD patients using RNA-seq data. Methods: We analysed previously published RNA-seq data from samples collected at diagnostic endoscopy from terminal ileum and colon of 25 treatment-naive paediatric CD patients and 23 controls. We implemented functional profiling and proposed a statistic method for both gene selection and classification using a logistic regression (LR) model. Findings: We found that 550 genes with six different expression types were specifically expressed in CD patients compared to healthy controls (CI 95%, p<0·05): colonic CD genes (CCGs) represented 240 of these genes, and ileal CD genes (ICGs) composed 310. The DEGs were classified into categories such as mitochondrial dysfunction, oxidative phosphorylation, and metabolic pathways. In addition, mitochondrial dysfunction was only detected in the colon of CD patients. Fifty nine genes considered potentially useful for distinguishing Crohn’s disease subtypes were obtained, and average area under curve (AUC) and accuracy were 0·976 and 0·917, respectively. We also validated our method by applying it to an independent cohort of adult CD patients (accuracy = 0·886, AUC = 0·755). Interpretation: Our results show biomarkers for two CD subtypes, which will be useful for the personalized treatment of inflammatory bowel disease (IBD) patients. Funding Statement: This study was supported by the Basic Science Research Program of the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2019R1I1A2A01060140, 2014M3A9A5034349, 2018M3A9H3023077), and by the KRIBB Research Initiative Program. Declaration of Interests: The authors have no conflicts of interests to declare. Ethics Approval Statement: Mucosal biopsies were collected from colons and terminal ileums (TI) of children newly diagnosed with CD and children without IBD with full ethical approval in the University of Cambridge Department of Paediatrics, as reported in a previous study (EBI study ID: PRJEB24645).

Redox-mediated kick-start of mitochondrial energy metabolism drives resource-efficient seed germination

Seeds preserve a far developed plant embryo in a quiescent state. Seed metabolism relies on stored resources and is reactivated to drive germination when the external conditions are favorable. Since the switchover from quiescence to reactivation provides a remarkable case of a cell physiological transition we investigated the earliest events in energy and redox metabolism of Arabidopsis seeds at imbibition. By developing fluorescent protein biosensing in intact seeds, we observed ATP accumulation and oxygen uptake within minutes, indicating rapid activation of mitochondrial respiration, which coincided with a sharp transition from an oxidizing to a more reducing thiol redox environment in the mitochondrial matrix. To identify individual operational protein thiol switches, we captured the fast release of metabolic quiescence in organello and devised quantitative iodoacetyl tandem mass tag (iodoTMT)-based thiol redox proteomics. The redox state across all Cys peptides was shifted toward reduction from 27.1% down to 13.0% oxidized thiol. A large number of Cys peptides (412) were redox switched, representing central pathways of mitochondrial energy metabolism, including the respiratory chain and each enzymatic step of the tricarboxylic acid (TCA) cycle. Active site Cys peptides of glutathione reductase 2, NADPH-thioredoxin reductase a/b, and thioredoxin-o1 showed the strongest responses. Germination of seeds lacking those redox proteins was associated with markedly enhanced respiration and deregulated TCA cycle dynamics suggesting decreased resource efficiency of energy metabolism. Germination in aged seeds was strongly impaired. We identify a global operation of thiol redox switches that is required for optimal usage of energy stores by the mitochondria to drive efficient germination.

Mitochondrial metabolism: a common link between neuroinflammation and neurodegeneration.

Neurodegenerative disorders have been considered as a growing health concern for decades. Increasing risk of neurodegenerative disorders creates a socioeconomic burden to both patients and care givers. Mitochondria are organelle that are involved in both neuroinflammation and neurodegeneration. There are few reports on the effect of mitochondrial metabolism on the progress of neurodegeneration and neuroinflammation. Therefore, the present review summarizes the potential contribution of mitochondrial metabolic pathways in the pathogenesis of neuroinflammation and neurodegeneration. Mitochondrial pyruvate metabolism plays a critical role in the pathogenesis of neurodegenerative disorders such as Parkinson's disease and Alzheimer's disease. However, there its potential contribution in other neurodegenerative disorders is as yet unproven. The mitochondrial pyruvate carrier and pyruvate dehydrogenase can modulate mitochondrial pyruvate metabolism to attenuate neuroinflammation and neurodegeneration. Further, it has been observed that the mitochondrial citric acid cycle can regulate the pathogenesis of neuroinflammation and neurodegeneration. Additional research should be undertaken to target tricarboxylic acid cycle enzymes to minimize the progress of neuroinflammation and neurodegeneration. It has also been observed that the mitochondrial urea cycle can potentially contribute to the progression of neurodegenerative disorders. Therefore, targeting this pathway may control the mitochondrial dysfunction-induced neuroinflammation and neurodegeneration. Furthermore, the mitochondrial malate-aspartate shuttle could be another target to control mitochondrial dysfunction-induced neuroinflammation and neurodegeneration in neurodegenerative disorders.

Mitochondrial Dysfunction in Critical Illness: Implications for Nutritional Therapy.

This paper will review the evidence for mitochondrial dysfunction in critical illness, describe the mechanisms which lead to multiple organ failure, and detail the implications of this pathophysiologic process on nutritional therapy. Mitochondria are particularly sensitive to increased oxidative stress in critical illness. The functional and structural abnormalities which occur in this organelle contribute further to the excessive production of reactive oxygen species and the reduction in generation of adenosine triphosphate (ATP). To reduce metabolic demand, mitochondrial dysfunction develops (a process likened to hibernation), which helps sustain the life of the cell at a cost of organ system failure. Aggressive feeding in the early phases of critical illness might inappropriately increase demand at a time when ATP production is limited, further jeopardizing cell survival and potentiating the processes leading to multiple organ failure. Several potential therapies exist which would promote mitochondrial function in the intensive care setting through support of autophagy, antioxidant defense systems, and the biogenesis and recovery of the organelle itself. Nutritional therapy should supplement micronutrients required in the mitochondrial metabolic pathways and provide reduced delivery of macronutrients through slower advancement of feeding in the early phases of critical illness. A better understanding of mitochondrial dysfunction in the critically ill patient should lead to more innovative therapies in the future.

P.58No effect of resveratrol supplementation in patients with mitochondrial myopathy - a randomized, double-blind, placebo-controlled, cross-over study

Mitochondrial myopathies (MM) are caused by mutations that affect proteins involved in oxidative phosphorylation. Main symptoms are exercise intolerance and fatigue. Currently, there is no specific treatment for MM. Resveratrol (RSV) is a well-known nutritional supplement that may target several mitochondrial metabolic pathways. It has been shown to enhance oxidative capacity and have antioxidant and anti-inflammatory properties in cells and animal models. The study is the first to asses if RSV can improve mitochondrial function and thereby improve exercise capacity in patients with MM. The study design was randomized, double-blind, crossover and placebo-controlled. Ten patients with genetically verified MM were randomized to receive either 1000 mg/day RSV or placebo (P) for 8 weeks followed by 8 weeks with the opposite treatment separated by a 4-week washout. Primary outcomes were changes in heart rate (HR) during submaximal cycling exercise, and peak oxygen utilization (VO2max) during maximal exercise. Secondary outcomes included reduction in perceived exertion (Borg scale), changes in lactate production, and self-rated function (SF-36) and fatigue scores (FSS). Values are presented as mean change from baseline to end-treatments ±SD. The paired non-parametric Wilcoxon test was used to compare treatment effects with ≤0.05 as level of significance. The results show no changes in primary or secondary outcomes between treatments. Mean HR changes were -0.3±4.2 (RSV) vs 1.8±4.9 bpm (P), p= 0.241. Delta VO2max were 54±90 (RSV) vs -50±209 l/min (P), p=0.203. Delta lactate concentrations and perceived exertion during submaximal exercise, and SF-36 and FSS did not change (p>0.05). This study provides evidence that RSV 1000 mg daily is ineffective in improving exercise capacity in adults with MM. These findings indicate that previous in vitro studies suggesting a therapeutic potential for RSV in MM do not translate into clinically meaningful effects in vivo.

Proteolytic Control of Lipid Metabolism.

Synthesis and regulation of lipid levels and identities is critical for a wide variety of cellular functions, including structural and morphological properties of organelles, energy storage, signaling, and stability and function of membrane proteins. Proteolytic cleavage events regulate and/or influence some of these lipid metabolic processes and as a result help modulate their pleiotropic cellular functions. Proteins involved in lipid regulation are proteolytically cleaved for the purpose of their relocalization, processing, turnover, and quality control, among others. The scope of this review includes proteolytic events governing cellular lipid dynamics. After an initial discussion of the classic example of sterol regulatory element-binding proteins, our focus will shift to the mitochondrion, where a range of proteolytic events are critical for normal mitochondrial phospholipid metabolism and enforcing quality control therein. Recently, mitochondrial phospholipid metabolic pathways have been implicated as important for the proliferative capacity of cancers. Thus, the assorted proteases that regulate, monitor, or influence the activity of proteins that are important for phospholipid metabolism represent attractive targets to be manipulated for research purposes and clinical applications.

Loss of mitochondrial energetics is associated with poor recovery of muscle function but not mass following disuse atrophy.

Skeletal muscle atrophy is a clinically important outcome of disuse because of injury, immobilization, or bed rest. Disuse atrophy is accompanied by mitochondrial dysfunction, which likely contributes to activation of the muscle atrophy program. However, the linkage of muscle mass and mitochondrial energetics during disuse atrophy and its recovery is incompletely understood. Transcriptomic analysis of muscle biopsies from healthy older adults subject to complete bed rest revealed marked inhibition of mitochondrial energy metabolic pathways. To determine the temporal sequence of muscle atrophy and changes in intramyocellular lipid and mitochondrial energetics, we conducted a time course of hind limb unloading-induced atrophy in adult mice. Mitochondrial respiration and calcium retention capacity were diminished, whereas H2O2 emission was increased within 3 days of unloading before significant muscle atrophy. These changes were associated with a decrease in total cardiolipin and profound changes in remodeled cardiolipin species. Hind limb unloading performed in muscle-specific peroxisome proliferator-activated receptor-γ coactivator-1α/β knockout mice, a model of mitochondrial dysfunction, did not affect muscle atrophy but impacted muscle function. These data suggest early mitochondrial remodeling affects muscle function but not mass during disuse atrophy. Early alterations in mitochondrial energetics and lipid remodeling may represent novel targets to prevent muscle functional impairment caused by disuse and to enhance recovery from periods of muscle atrophy.

Nicotinamide nucleotide transhydrogenase expression analysis in multiple sclerosis patients

Background: Nicotinamide nucleotide transhydrogenase (NNT) is a mitochondrial redox-induced proton pump that links NADPH synthesis to the mitochondrial metabolic pathway. It also participates in the regulation of immune responses. A long non-coding RNA namely NNT-antisense 1 (NNT-AS1) has been shown to be transcribed from the same locus and exert anti-proliferative effects in some tissues.Methods: In the current study, we evaluated expression of NNT and NNT-AS1 in peripheral blood of 50 relapsing-remitting multiple sclerosis patients compared with healthy subjects. The difference in NNT expression was significant in only in male subjects aged over 50 when compared with the corresponding control subgroup.Results: For NNT-AS1, based on the results of Quantile regression and adjustment of the effects of age and sex as well as the interaction between sex and disease status, no significant difference was found between cases and controls. Moreover, NNT and NNT-AS1 expressions were correlated with age in controls and in female subjects respectively.Conclusion: Finally, we assessed correlations between expressions of these genes and detected significant pairwise correlations between transcript levels of NNT and NNT-AS1 genes in both cases and controls. The current study highlights a gender-specific role for NNT in the pathogenesis of MS.

Abstract 782: Human-induced Pluripotent Stem Cell-derived Cardiomyocytes as a Model for Trastuzumab-Induced Cardiac Dysfunction

Background and Objective: Trastuzumab (Herceptin)-based chemotherapy regimens play a prominent role in treatments of breast cancer, but a significant number of patients develop cardiac disfunction during the therapy. In this study, we examined whether human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) could serve as an effective platform to study this particular cardiotoxicity. Methods and Results: We assessed the effects of trastuzumab on structural and functional properties in three control iPSC-CMs from healthy individuals and found that clinically relevant doses of trastuzumab (1 μM) significantly reduced the contraction velocity (-7.4%, P &lt; 0.01) and calcium amplitude (-14.1%, P &lt; 0.01) in iPSC-CMs without inducing cardiomyocyte death or sarcomeric structural abnormalities. RNA-sequencing analysis revealed mitochondrial dysfunction and altered cardiac energy metabolism pathway as primary causes of this cardiotoxicity. Subsequently, we found that significant reductions in basal, maximal, and ATP-linked oxygen consumption (-19.8%, -10.7%, and -19.0%, respectively, P &lt; 0.01 for all) and cellular ATP content (-10.3%, P &lt; 0.01) in iPSC-CMs after trastuzumab treatment. Next, we recruited seven breast cancer patients with trastuzumab therapy, including five patients diagnosed with trastuzumab-induced cardiac dysfunction and generated iPSCs from these patients. We found that contraction velocity and maximal oxygen consumption were more severely reduced by trastuzumab in iPSC-CMs generated from patients who experienced severe cardiac dysfunction from trastuzumab therapy, compared to iPSC-CMs generated from patients who did not experience cardiac dysfunction (-16.6% vs. -1.4%, and -29.4% vs. -3.7%, respectively, P &lt; 0.01 for all). Lastly, we demonstrated that metabolic modulation with AMPK activators, such as metformin, could avert the detrimental effects of trastuzumab in iPSC-CMs. Conclusion: Our results indicate that human iPSC-CM model can recapitulate the clinical phenotype of trastuzumab-induced cardiac dysfunction, and therefore targeting altered metabolism may be a promising therapeutic approach for treating this cardiotoxicity.

Mitochondrial fatty acid oxidation and the electron transport chain comprise a multifunctional mitochondrial protein complex

Three mitochondrial metabolic pathways are required for efficient energy production in eukaryotic cells: the electron transfer chain (ETC), fatty acid β-oxidation (FAO), and the tricarboxylic acid cycle. The ETC is organized into inner mitochondrial membrane supercomplexes that promote substrate channeling and catalytic efficiency. Although previous studies have suggested functional interaction between FAO and the ETC, their physical interaction has never been demonstrated. In this study, using blue native gel and two-dimensional electrophoreses, nano-LC-MS/MS, immunogold EM, and stimulated emission depletion microscopy, we show that FAO enzymes physically interact with ETC supercomplexes at two points. We found that the FAO trifunctional protein (TFP) interacts with the NADH-binding domain of complex I of the ETC, whereas the electron transfer enzyme flavoprotein dehydrogenase interacts with ETC complex III. Moreover, the FAO enzyme very-long-chain acyl-CoA dehydrogenase physically interacted with TFP, thereby creating a multifunctional energy protein complex. These findings provide a first view of an integrated molecular architecture for the major energy-generating pathways in mitochondria that ensures the safe transfer of unstable reducing equivalents from FAO to the ETC. They also offer insight into clinical ramifications for individuals with genetic defects in these pathways.

Changes in blood-brain barrier permeability and ultrastructure, and protein expression in a rat model of cerebral hypoperfusion

Cerebral hypoperfusion involved a reduction in cerebral blood flow, leading to neuronal dysfunction, microglial activation and white matter degeneration. The effects on the blood-brain barrier (BBB) however, have not been well-documented. Here, two-vessel occlusion model was adopted to mimic the condition of cerebral hypoperfusion in Sprague-Dawley rats. The BBB permeability to high and low molecular weight exogenous tracers i.e. Evans blue dye and sodium fluorescein respectively, showed marked extravasation of the Evans blue dye in the frontal cortex, posterior cortex and thalamus-midbrain at day 1 following induction of cerebral hypoperfusion. Transmission electron microscopy revealed brain endothelial cell and astrocyte damages including increased pinocytotic vesicles and formation of membrane invaginations in the endothelial cells, and swelling of the astrocytes’ end-feet. Investigation on brain microvessel protein expressions using two-dimensional (2D) gel electrophoresis coupled with LC–MS/MS showed that proteins involved in mitochondrial energy metabolism, transcription regulation, cytoskeleton maintenance and signaling pathways were differently expressed. The expression of aconitate hydratase, heterogeneous nuclear ribonucleoprotein, enoyl Co-A hydratase and beta-synuclein were downregulated, while the opposite observed for calreticulin and enhancer of rudimentary homolog. These findings provide insights into the BBB molecular responses to cerebral hypoperfusion, which may assist development of future therapeutic strategies.

Effect of cigarette smoke extract on mitochondrial heme-metabolism: An in vitro model of oral cancer progression

Tobacco smoking is considered as one of the major risk factors for development of oral cancer. In vitro studies indicate that cigarette smoke initiates transformation of epithelial cells toward development of oral cancer through altering mitochondrial metabolic pathways. However the present in vitro models need to be improved to correlate these molecular changes with epithelial transformations. In present study, we investigated the association of mitochondrial metabolic events with oral cancer progression under cigarette smoke extract (CSE). In this regard, an in vitro model of oral keratinocyte cell line (MOE1A) was developed by exposing them with different concentrations of CSE. Alterations in cellular phenomena were confirmed by Fourier-transform infrared spectroscopy (FTIR) study, which indicated changes in important functional groups of CSE-induced oral cells. Enhanced reactive oxygen species (ROS) of exposed cells altered the mitochondrial metabolic activities in terms of increased mitochondrial mass and DNA content. Further, mitochondrial heme-metabolism was investigated and real-time PCR study showed altered expression of important genes like ALAS1, ABCB6, CPOX, FECH, HO-1. Both transcriptomic and proteomic studies showed up- and down-regulation of important biomarkers related to cellular cancer progression. Overall data suggest that CSE alters mitochondrial heme metabolic pathway and initiates cancer progression through modifying cellar biomarkers in oral epithelial cells.

Spermatozoa motility in bivalves: Signaling, flagellar beating behavior, and energetics

Though bivalve mollusks are keystone species and major species groups in aquaculture production worldwide, gamete biology is still largely unknown. This review aims to provide a synthesis of current knowledge in the field of sperm biology, including spermatozoa motility, flagellar beating, and energy metabolism; and to illustrate cellular signaling controlling spermatozoa motility initiation in bivalves. Serotonin (5-HT) induces hyper-motility in spermatozoa via a 5-HT receptor, suggesting a serotoninergic system in the male reproductive tract that might regulate sperm physiology. Acidic pH and high concentration of K+ are inhibitory factors of spermatozoa motility in the testis. Motility is initiated at spawning by a Na+-dependent alkalization of intracellular pH mediated by a Na+/H+ exchanger. Increase of 5-HT in the testis and decrease of extracellular K+ when sperm is released in seawater induce hyperpolarization of spermatozoa membrane potential mediated by K+ efflux and associated with an increase in intracellular Ca2+ via opening of voltage-dependent Ca2+ channels under alkaline conditions. These events activate dynein ATPases and Ca2+/calmodulin-dependent proteins resulting in flagellar beating. It may be possible that 5-HT is also involved in intracellular cAMP rise controlling cAMP-dependent protein kinase phosphorylation in the flagellum. Once motility is triggered, flagellum beats in asymmetric wave pattern leading to circular trajectories of spermatozoa. Three different flagellar wave characteristics are reported, including “full”, “twitching”, and “declining” propagation of wave, which are described and illustrated in the present review. Mitochondrial respiration, ATP content, and metabolic pathways producing ATP in bivalve spermatozoa are discussed. Energy metabolism of Pacific oyster spermatozoa differs from previously studied marine species since oxidative phosphorylation synthetizes a stable level of ATP throughout 24-h motility period and the end of movement is not explained by a low intracellular ATP content, revealing different strategy to improve oocyte fertilization success. Finally, our review highlights physiological mechanisms that require further researches and points out some advantages of bivalve spermatozoa to extend knowledge on mechanisms of motility.

Low-intensity exercise induces acute shifts in liver and skeletal muscle substrate metabolism but not chronic adaptations in tissue oxidative capacity.

Adaptations in hepatic and skeletal muscle substrate metabolism following acute and chronic (6 wk; 5 days/wk; 1 h/day) low-intensity treadmill exercise were tested in healthy male C57BL/6J mice. Low-intensity exercise maximizes lipid utilization; therefore, we hypothesized pathways involved in lipid metabolism would be most robustly affected. Acute exercise nearly depleted liver glycogen immediately postexercise (0 h), whereas hepatic triglyceride (TAG) stores increased in the early stages after exercise (0-3 h). Also, hepatic peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) gene expression and fat oxidation (mitochondrial and peroxisomal) increased immediately postexercise (0 h), whereas carbohydrate and amino acid oxidation in liver peaked 24-48 h later. Alternatively, skeletal muscle exhibited a less robust response to acute exercise as stored substrates (glycogen and TAG) remained unchanged, induction of PGC-1α gene expression was delayed (up at 3 h), and mitochondrial substrate oxidation pathways (carbohydrate, amino acid, and lipid) were largely unaltered. Peroxisomal lipid oxidation exhibited the most dynamic changes in skeletal muscle substrate metabolism after acute exercise; however, this response was also delayed (peaked 3-24 h postexercise), and expression of peroxisomal genes remained unaffected. Interestingly, 6 wk of training at a similar intensity limited weight gain, increased muscle glycogen, and reduced TAG accrual in liver and muscle; however, substrate oxidation pathways remained unaltered in both tissues. Collectively, these results suggest changes in substrate metabolism induced by an acute low-intensity exercise bout in healthy mice are more rapid and robust in liver than in skeletal muscle; however, training at a similar intensity for 6 wk is insufficient to induce remodeling of substrate metabolism pathways in either tissue. NEW & NOTEWORTHY Effects of low-intensity exercise on substrate metabolism pathways were tested in liver and skeletal muscle of healthy mice. This is the first study to describe exercise-induced adaptations in peroxisomal lipid metabolism and also reports comprehensive adaptations in mitochondrial substrate metabolism pathways (carbohydrate, lipid, and amino acid). Acute low-intensity exercise induced shifts in mitochondrial and peroxisomal metabolism in both tissues, but training at this intensity did not induce adaptive remodeling of metabolic pathways in healthy mice.

Cardioprotective effects of Aconiti Lateralis Radix Praeparata combined with Zingiberis Rhizoma on doxorubicin-induced chronic heart failure in rats and potential mechanisms

BackgroundThe combined use of Aconiti Lateralis Radix Praeparata (ALRP) and Zingiberis Rhizoma (ZR) are classic compatibilities in China for the treatment of cardiovascular diseases such as increasing myocardial contractility, anti-arrhythmia, reducing myocardial oxygen consumption, and dilating organ blood vessels, etc, thereby exerting anti-heart failure (HF) effects in traditional Chinese herbal medicine. However, comprehensive approaches for understanding the therapeutic effects and mechanisms underlying chronic heart failure (CHF) from the perspective of energy metabolism have not been pursued. AimThis research was aimed to investigate the effectiveness and potential mechanism of ALRP combined with ZR (1:1) on doxorubicin (DOX)-induced CHF in rats based on an integrated approach that combines network pharmacology analyses and molecular biology. Material and methodsCHF model was established by the intraperitoneal injection of DOX. ALRP and ZR were intragastrically administrated for three weeks. The detection indices including hemodynamic measurements, myocardial injury marker, and myocardial pathological changes were measured. Network pharmacology analysis was used to illustrate the pathways and network of ALRP and ZR against HF. Mitochondrial energy metabolism pathway associated gene and protein levels of PPARα, PGC-1α and Sirt3 in myocardial tissue were detected by real-time PCR and western blotting, respectively. ResultsThe results indicated that ALRP-ZR herbal couple significantly improved the left ventricular function and cardiac enzyme activities in comparison with their single use. Network pharmacology analysis results showed that the pharmacological mechanisms of ALRP-ZR may be related to PPAR energy metabolism pathway. Besides, the outcomes of western-blot and real-time PCR analysis showed that ALRP-ZR significantly upregulates the protein and gene level of PPARα, PGC-1α, and Sirt3. ConclusionsNetwork pharmacology analysis would be an effective network analyze workflow which was feasible for evaluating the pharmacological effect of a multi-drug complex system. The Chinese herbal couple ALRP-ZR had a better therapeutic effect than their single-use against DOX-induced CHF, which may be related to enhancing left ventricular function by activating the PPARα/PGC-1α/Sirt3 pathway.

Effects of AMPK on Apoptosis and Energy Metabolism of Gastric Smooth Muscle Cells in Rats with Diabetic Gastroparesis.

This study aimed to investigate the effect of AMPK on apoptosis and energy metabolism of gastric smooth muscle cells in diabetic rats and to explore the role of AMPK in the pathogenesis of diabetic gastroparesis (DGP). After establishment of a diabetic rat model, rats were divided into normal control (NC), 4-week (DM4W), 6-week (DM6W), and 8-week (DM8W) diabetic model groups. The gastric residual pigment ratio, intestinal transit rate, and intestinal propulsion rate in each group were detected to confirm the successful establishment of the DGP model. The spontaneous contraction in isolated gastric smooth muscle strips of the NC and DM8W groups was experimentally observed. The expression of phospho-AMPK, AMPK, phospho-LKB1, LKB1, phospho-TAK1, TAK1, and CaMMKβ in rat gastric smooth muscle tissues was detected by western blot analysis; ADP, AMP, ATP contents, and the energy charge were detected using Elisa; and apoptosis of gastric smooth muscle cells was detected by flow cytometry. The rat gastric smooth muscle cells were cultured in vitro, and treated with an AMPK inhibitor and an agonist. At 24 and 48 h, the effects of AMPK on apoptosis and energy metabolism of gastric smooth muscle cells were observed. Reduced spontaneous contractions, AMPK activation, cell apoptosis, and energy metabolism disorders were observed in gastric smooth muscle tissues of a diabetic rat, and AMPK activation was associated with an increased ratio of ADP/ATP, AMP/ATP, LKB1 activity, and CaMMKβ expression. From in vitro cell culture experiments, we found that AMPK activation of high-glucose conditions promoted cell apoptosis. Inhibition of AMPK had no obvious effect on apoptosis at the early stage with high glucose, but the inhibitory effect was significant at the late stage with high glucose. AMPK can regulate both mitochondrial metabolism and glycolysis pathways under high-glucose conditions. During the early stage with high glucose, AMPK was the main promotion factor of the mitochondrial metabolism pathway, but did not increase the ATP production, AMPK also promoted the glycolysis pathway. During the late stage with high glucose, AMPK was a major inhibitor of the mitochondrial pathway, and still played a role in promoting the glycolytic pathway, which acted as the main regulator. Apoptosis and energy metabolism disorders were present in gastric smooth muscle cells during the occurrence of DGP. Under high-glucose condition, AMPK was activated, which can promote apoptosis, change the energetic metabolism pathway of cells, inhibit mitochondrial energy metabolism, and promote glycolysis.

Quantitative Variation in m.3243A\u2009>\u2009G Mutation Produce Discrete Changes in Energy Metabolism

Mitochondrial DNA (mtDNA) 3243A > G tRNALeu(UUR) heteroplasmic mutation (m.3243A > G) exhibits clinically heterogeneous phenotypes. While the high mtDNA heteroplasmy exceeding a critical threshold causes mitochondrial encephalomyopathy, lactic acidosis with stroke-like episodes (MELAS) syndrome, the low mtDNA heteroplasmy causes maternally inherited diabetes with or without deafness (MIDD) syndrome. How quantitative differences in mtDNA heteroplasmy produces distinct pathological states has remained elusive. Here we show that despite striking similarities in the energy metabolic gene expression signature, the mitochondrial bioenergetics, biogenesis and fuel catabolic functions are distinct in cells harboring low or high levels of the m.3243 A > G mutation compared to wild type cells. We further demonstrate that the low heteroplasmic mutant cells exhibit a coordinate induction of transcriptional regulators of the mitochondrial biogenesis, glucose and fatty acid metabolism pathways that lack in near homoplasmic mutant cells compared to wild type cells. Altogether, these results shed new biological insights on the potential mechanisms by which low mtDNA heteroplasmy may progressively cause diabetes mellitus.

Exome sequencing and bioinformatic approaches reveals rare sequence variants involved in cell signalling and elastic fibre homeostasis: new evidence in the development of ectopic calcification

Elastic fibres undergo aberrant mineralization in genetic as well as in acquired pathologic conditions causing severe impairment of tissue mechanical properties. Despite the number of investigations performed so far, the pathogenesis of these alterations is still elusive, due to both the complexity of the elastin network and the involvement of many genes and/or pro-osteogenic signalling pathways.Whole Exome Sequencing (WES) was performed on DNA from three patients affected by beta-thalassemia exhibiting soft connective tissue calcification. WES data were analysed with a bioinformatic approach, allowing to screen and to select genes carrying rare sequence variants. These genes were matched with those present in Extracellular Matrix DB. This approach enables to shed light on the involvement of the extracellular matrix in the occurrence of ectopic calcification.Results revealed a number of rare sequence variants in genes related to elastic fibre assembly and integrity. For instance, the involvement of fibrillins and collagen type VI in the formation of a modified microfibrillar scaffold may lead to elastic fibres less resilient and more prone to hydroxyapatite deposition. Moreover, data reveal that changes in mitochondrial metabolic pathways are sustained by a genetic background and emphasize that a persistent chronic oxidative stress can further influence extracellular matrix homeostasis and cell signalling through the TGFβ-BMP axis. Eventually, the presence of multiple rare sequence variants in the Solute Carrier Family 25 Member 5 (SLC25A5) gene is suggestive of the role of this gene as a key factor linking mitochondria metabolism, ADP/ATP ratio and oxidative stress thus affecting extracellular matrix homeostasis and activation of pro-osteogenic factors.

Succinylation Links Metabolism to Protein Functions.

Post-translational modifications (PTMs) are important regulators of protein function, and integrate metabolism with physiological and pathological processes. Phosphorylation and acetylation are particularly well studied PTMs. A relatively recently discovered novel PTM is succinylation in which metabolically derived succinyl CoA modifies protein lysine groups. Succinylation causes a protein charge flip from positive to negative and a relatively large increase in mass compared to other PTMs. Hundreds of protein succinylation sites are present in proteins of multiple tissues and species, and the significance is being actively investigated. The few completed studies demonstrate that succinylation alters rates of enzymes and pathways, especially mitochondrial metabolic pathways. Thus, succinylation provides an elegant and efficient mechanism to coordinate metabolism and signaling by utilizing metabolic intermediates as sensors to regulate metabolism. Even though the brain is one of the most metabolically active organs, an understanding of the role succinylation in the nervous system is largely unknown. Data from other tissues and other PTMs suggest that succinylation provides a coupling between metabolism and protein function in the nervous system and in neurological diseases. This review provides a new insight into metabolism in neurological diseases and suggests that the drug development for these diseases requires a better understanding of succinylation and de-succinylation in the brain and other tissues.