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Sample records for mouse mitochondrial voltage

  1. Rosiglitazone induces mitochondrial biogenesis in mouse brain.

    PubMed

    Strum, Jay C; Shehee, Ron; Virley, David; Richardson, Jill; Mattie, Michael; Selley, Paula; Ghosh, Sujoy; Nock, Christina; Saunders, Ann; Roses, Allen

    2007-03-01

    Rosiglitazone was found to simulate mitochondrial biogenesis in mouse brain in an apolipoprotein (Apo) E isozyme-independent manner. Rosiglitazone induced both mitochondrial DNA (mtDNA) and estrogen-stimulated related receptor alpha (ESRRA) mRNA, a key regulator of mitochondrial biogenesis. Transcriptomics and proteomics analysis suggested the mitochondria produced in the presence of human ApoE3 and E4 were not as metabolically efficient as those in the wild type or ApoE knockout mice. Thus, we propose that PPARgamma agonism induces neuronal mitochondrial biogenesis and improves glucose utilization leading to improved cellular function and provides mechanistic support for the improvement in cognition observed in treatment of Alzheimer's patients with rosiglitazone.

  2. The pathophysiology of mitochondrial disease as modeled in the mouse

    PubMed Central

    Wallace, Douglas C.; Fan, WeiWei

    2009-01-01

    It is now clear that mitochondrial defects are associated with a plethora of clinical phenotypes in man and mouse. This is the result of the mitochondria's central role in energy production, reactive oxygen species (ROS) biology, and apoptosis, and because the mitochondrial genome consists of roughly 1500 genes distributed across the maternal mitochondrial DNA (mtDNA) and the Mendelian nuclear DNA (nDNA). While numerous pathogenic mutations in both mtDNA and nDNA mitochondrial genes have been identified in the past 21 years, the causal role of mitochondrial dysfunction in the common metabolic and degenerative diseases, cancer, and aging is still debated. However, the development of mice harboring mitochondrial gene mutations is permitting demonstration of the direct cause-and-effect relationship between mitochondrial dysfunction and disease. Mutations in nDNA-encoded mitochondrial genes involved in energy metabolism, antioxidant defenses, apoptosis via the mitochondrial permeability transition pore (mtPTP), mitochondrial fusion, and mtDNA biogenesis have already demonstrated the phenotypic importance of mitochondrial defects. These studies are being expanded by the recent development of procedures for introducing mtDNA mutations into the mouse. These studies are providing direct proof that mtDNA mutations are sufficient by themselves to generate major clinical phenotypes. As more different mtDNA types and mtDNA gene mutations are introduced into various mouse nDNA backgrounds, the potential functional role of mtDNA variation in permitting humans and mammals to adapt to different environments and in determining their predisposition to a wide array of diseases should be definitively demonstrated. PMID:19651984

  3. Mouse models of age-related mitochondrial neurosensory hearing loss.

    PubMed

    Han, Chul; Someya, Shinichi

    2013-07-01

    Hearing loss is the most common sensory disorder in the elderly population. Overall, 10% of the population has a hearing loss in the US, and this age-related hearing disorder is projected to afflict more than 28 million Americans by 2030. Age-related hearing loss is associated with loss of sensory hair cells (sensory hearing loss) and/or spiral ganglion neurons (neuronal hearing loss) in the cochlea of the inner ear. Many lines of evidence indicate that oxidative stress and associated mitochondrial dysfunction play a central role in age-related neurodegenerative diseases and are a cause of age-related neurosensory hearing loss. Yet, the molecular mechanisms of how oxidative stress and/or mitochondrial dysfunction lead to hearing loss during aging remain unclear, and currently there is no treatment for this age-dependent disorder. Several mouse models of aging and age-related diseases have been linked to age-related mitochondrial neurosensory hearing loss. Evaluation of these animal models has offered basic knowledge of the mechanism underlying hearing loss associated with oxidative stress, mitochondrial dysfunction, and aging. Here we review the evidence that specific mutations in the mitochondrial DNA or nuclear DNA that affect mitochondrial function result in increased oxidative damage and associated loss of sensory hair cells and/or spiral ganglion neurons in the cochlea during aging, thereby causing hearing loss in these mouse models. Future studies comparing these models will provide further insight into fundamental knowledge about the disordered process of hearing and treatments to improve the lives of individuals with communication disorders. This article is part of a Special Issue entitled 'Mitochondrial function and dysfunction in neurodegeneration'.

  4. Exercise increases mitochondrial glutamate oxidation in the mouse cerebral cortex.

    PubMed

    Herbst, Eric A F; Holloway, Graham P

    2016-07-01

    The present study investigated the impact of acute exercise on stimulating mitochondrial respiratory function in mouse cerebral cortex. Where pyruvate-stimulated respiration was not affected by acute exercise, glutamate respiration was enhanced following the exercise bout. Additional assessment revealed that this affect was dependent on the presence of malate and did not occur when substituting glutamine for glutamate. As such, our results suggest that glutamate oxidation is enhanced with acute exercise through activation of the malate-aspartate shuttle. PMID:27184881

  5. Purinergic signalling mobilizes mitochondrial Ca2+ in mouse Sertoli cells

    PubMed Central

    Veitinger, Sophie; Veitinger, Thomas; Cainarca, Silvia; Fluegge, Daniela; Engelhardt, Corinna H; Lohmer, Stefan; Hatt, Hanns; Corazza, Sabrina; Spehr, Jennifer; Neuhaus, Eva M; Spehr, Marc

    2011-01-01

    Abstract Intimate bidirectional communication between Sertoli cells and developing germ cells ensures the integrity and efficiency of spermatogenesis. Yet, a conceptual mechanistic understanding of the physiological principles that underlie Sertoli cell autocrine and paracrine signalling is lacking. Here, we characterize a purinergic Ca2+ signalling network in immature mouse Sertoli cells that consists of both P2X2 and P2Y2 purinoceptor subtypes, the endoplasmic reticulum and, notably, mitochondria. By combining a transgenic mouse model with a dedicated bioluminescence imaging device, we describe a novel method to monitor mitochondrial Ca2+ mobilization in Sertoli cells at subcellular spatial and millisecond temporal resolution. Our data identify mitochondria as essential components of the Sertoli cell signalling ‘toolkit’ that control the shape of purinergic Ca2+ responses, and probably several other paracrine Ca2+-dependent signals. PMID:21859825

  6. Auditory Pathology in a Transgenic mtTFB1 Mouse Model of Mitochondrial Deafness.

    PubMed

    McKay, Sharen E; Yan, Wayne; Nouws, Jessica; Thormann, Maximilian J; Raimundo, Nuno; Khan, Abdul; Santos-Sacchi, Joseph; Song, Lei; Shadel, Gerald S

    2015-12-01

    The A1555G mutation in the 12S rRNA gene of human mitochondrial DNA causes maternally inherited, nonsyndromic deafness, an extreme case of tissue-specific mitochondrial pathology. A transgenic mouse strain that robustly overexpresses the mitochondrial 12S ribosomal RNA methyltransferase TFB1M (Tg-mtTFB1 mice) exhibits progressive hearing loss that we proposed models aspects of A1555G-related pathology in humans. Although our previous studies of Tg-mtTFB1 mice implicated apoptosis in the spiral ganglion and stria vascularis because of mitochondrial reactive oxygen species-mediated activation of AMP kinase (AMPK) and the nuclear transcription factor E2F1, detailed auditory pathology was not delineated. Herein, we show that Tg-mtTFB1 mice have reduced endocochlear potential, indicative of significant stria vascularis dysfunction, but without obvious signs of strial atrophy. We also observed decreased auditory brainstem response peak 1 amplitude and prolonged wave I latency, consistent with apoptosis of spiral ganglion neurons. Although no major loss of hair cells was observed, there was a mild impairment of voltage-dependent electromotility of outer hair cells. On the basis of these results, we propose that these events conspire to produce the progressive hearing loss phenotype in Tg-mtTFB1 mice. Finally, genetically reducing AMPK α1 rescues hearing loss in Tg-mtTFB1 mice, confirming that aberrant up-regulation of AMPK signaling promotes the observed auditory pathology. The relevance of these findings to human A1555G patients and the potential therapeutic value of reducing AMPK activity are discussed.

  7. Regulation of Mitochondrial Function by Voltage Dependent Anion Channels in Ethanol Metabolism and the Warburg Effect

    PubMed Central

    Lemasters, John J.; Holmuhamedov, Ekhson L.; Czerny, Christoph; Zhong, Zhi; Maldonado, Eduardo N.

    2012-01-01

    Voltage dependent anion channels (VDAC) are highly conserved proteins that are responsible for permeability of the mitochondrial outer membrane to hydrophilic metabolites like ATP, ADP and respiratory substrates. Although previously assumed to remain open, VDAC closure is emerging as an important mechanism for regulation of global mitochondrial metabolism in apoptotic cells and also in cells that are not dying. During hepatic ethanol oxidation to acetaldehyde, VDAC closure suppresses exchange of mitochondrial metabolites, resulting in inhibition of ureagenesis. In vivo, VDAC closure after ethanol occurs coordinately with mitochondrial uncoupling. Since acetaldehyde passes through membranes independently of channels and transporters, VDAC closure and uncoupling together foster selective and more rapid oxidative metabolism of toxic acetaldehyde to nontoxic acetate by mitochondrial aldehyde dehydrogenase. In single reconstituted VDAC, tubulin decreases VDAC conductance, and in HepG2 hepatoma cells, free tubulin negatively modulates mitochondrial membrane potential, an effect enhanced by protein kinase A. Tubulin-dependent closure of VDAC in cancer cells contributes to suppression of mitochondrial metabolism and may underlie the Warburg phenomenon of aerobic glycolysis. PMID:22172804

  8. Alterations in voltage-sensing of the mitochondrial permeability transition pore in ANT1-deficient cells

    PubMed Central

    Doczi, Judit; Torocsik, Beata; Echaniz-Laguna, Andoni; Mousson de Camaret, Bénédicte; Starkov, Anatoly; Starkova, Natalia; Gál, Aniko; Molnár, Mária J; Kawamata, Hibiki; Manfredi, Giovanni; Adam-Vizi, Vera; Chinopoulos, Christos

    2016-01-01

    The probability of mitochondrial permeability transition (mPT) pore opening is inversely related to the magnitude of the proton electrochemical gradient. The module conferring sensitivity of the pore to this gradient has not been identified. We investigated mPT’s voltage-sensing properties elicited by calcimycin or H2O2 in human fibroblasts exhibiting partial or complete lack of ANT1 and in C2C12 myotubes with knocked-down ANT1 expression. mPT onset was assessed by measuring in situ mitochondrial volume using the ‘thinness ratio’ and the ‘cobalt-calcein’ technique. De-energization hastened calcimycin-induced swelling in control and partially-expressing ANT1 fibroblasts, but not in cells lacking ANT1, despite greater losses of mitochondrial membrane potential. Matrix Ca2+ levels measured by X-rhod-1 or mitochondrially-targeted ratiometric biosensor 4mtD3cpv, or ADP-ATP exchange rates did not differ among cell types. ANT1-null fibroblasts were also resistant to H2O2-induced mitochondrial swelling. Permeabilized C2C12 myotubes with knocked-down ANT1 exhibited higher calcium uptake capacity and voltage-thresholds of mPT opening inferred from cytochrome c release, but intact cells showed no differences in calcimycin-induced onset of mPT, irrespective of energization and ANT1 expression, albeit the number of cells undergoing mPT increased less significantly upon chemically-induced hypoxia than control cells. We conclude that ANT1 confers sensitivity of the pore to the electrochemical gradient. PMID:27221760

  9. Alterations in voltage-sensing of the mitochondrial permeability transition pore in ANT1-deficient cells.

    PubMed

    Doczi, Judit; Torocsik, Beata; Echaniz-Laguna, Andoni; Mousson de Camaret, Bénédicte; Starkov, Anatoly; Starkova, Natalia; Gál, Aniko; Molnár, Mária J; Kawamata, Hibiki; Manfredi, Giovanni; Adam-Vizi, Vera; Chinopoulos, Christos

    2016-01-01

    The probability of mitochondrial permeability transition (mPT) pore opening is inversely related to the magnitude of the proton electrochemical gradient. The module conferring sensitivity of the pore to this gradient has not been identified. We investigated mPT's voltage-sensing properties elicited by calcimycin or H2O2 in human fibroblasts exhibiting partial or complete lack of ANT1 and in C2C12 myotubes with knocked-down ANT1 expression. mPT onset was assessed by measuring in situ mitochondrial volume using the 'thinness ratio' and the 'cobalt-calcein' technique. De-energization hastened calcimycin-induced swelling in control and partially-expressing ANT1 fibroblasts, but not in cells lacking ANT1, despite greater losses of mitochondrial membrane potential. Matrix Ca(2+) levels measured by X-rhod-1 or mitochondrially-targeted ratiometric biosensor 4mtD3cpv, or ADP-ATP exchange rates did not differ among cell types. ANT1-null fibroblasts were also resistant to H2O2-induced mitochondrial swelling. Permeabilized C2C12 myotubes with knocked-down ANT1 exhibited higher calcium uptake capacity and voltage-thresholds of mPT opening inferred from cytochrome c release, but intact cells showed no differences in calcimycin-induced onset of mPT, irrespective of energization and ANT1 expression, albeit the number of cells undergoing mPT increased less significantly upon chemically-induced hypoxia than control cells. We conclude that ANT1 confers sensitivity of the pore to the electrochemical gradient. PMID:27221760

  10. Tools for assessing mitochondrial dynamics in mouse tissues and neurodegenerative models

    NASA Astrophysics Data System (ADS)

    Pham, Anh H.

    Mitochondria are dynamic organelles that undergo membrane fusion and fission and transport. The dynamic properties of mitochondria are important for regulating mitochondrial function. Defects in mitochondrial dynamics are linked neurodegenerative diseases and affect the development of many tissues. To investigate the role of mitochondrial dynamics in diseases, versatile tools are needed to explore the physiology of these dynamic organelles in multiple tissues. Current tools for monitoring mitochondrial dynamics have been limited to studies in cell culture, which may be inadequate model systems for exploring the network of tissues. Here, we have generated mouse models for monitoring mitochondrial dynamics in a broad spectrum of tissues and cell types. The Photo-Activatable Mitochondrial (PhAM floxed) line enables Cre-inducible expression of a mitochondrial targeted photoconvertible protein, Dendra2 (mito-Dendra2). In the PhAMexcised line, mito-Dendra2 is ubiquitously expressed to facilitate broad analysis of mitochondria at various developmental processes. We have utilized these models to study mitochondrial dynamics in the nigrostriatal circuit of Parkinson's disease (PD) and in the development of skeletal muscles. Increasing evidences implicate aberrant regulation of mitochondrial fusion and fission in models of PD. To assess the function of mitochondrial dynamics in the nigrostriatal circuit, we utilized transgenic techniques to abrogate mitochondrial fusion. We show that deletion of the Mfn2 leads to the degeneration of dopaminergic neurons and Parkinson's-like features in mice. To elucidate the dynamic properties of mitochondria during muscle development, we established a platform for examining mitochondrial compartmentalization in skeletal muscles. This model system may yield clues to the role of mitochondrial dynamics in mitochondrial myopathies.

  11. Complete mitochondrial genome of the gray mouse lemur, Microcebus murinus (Primates, Cheirogaleidae).

    PubMed

    Lecompte, Emilie; Crouau-Roy, Brigitte; Aujard, Fabienne; Holota, Hélène; Murienne, Jérôme

    2016-09-01

    We report the high-coverage complete mitochondrial genome sequence of the gray mouse lemur Microcebus murinus. The sequencing has been performed on an Illumina Hiseq 2500 platform, with a genome skimming strategy. The total length of this mitogenome is 16 963 bp, containing 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes and 1 non-coding region (D-loop region). The genome organization, nucleotide composition and codon usage are similar to those reported from other primate's mitochondrial genomes. The complete mitochondrial genome sequence reported here will be useful for comparative genomics studies in primates. PMID:27158869

  12. Complete mitochondrial genome of the gray mouse lemur, Microcebus murinus (Primates, Cheirogaleidae).

    PubMed

    Lecompte, Emilie; Crouau-Roy, Brigitte; Aujard, Fabienne; Holota, Hélène; Murienne, Jérôme

    2016-09-01

    We report the high-coverage complete mitochondrial genome sequence of the gray mouse lemur Microcebus murinus. The sequencing has been performed on an Illumina Hiseq 2500 platform, with a genome skimming strategy. The total length of this mitogenome is 16 963 bp, containing 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes and 1 non-coding region (D-loop region). The genome organization, nucleotide composition and codon usage are similar to those reported from other primate's mitochondrial genomes. The complete mitochondrial genome sequence reported here will be useful for comparative genomics studies in primates.

  13. Mitochondrial organization and motility probed by two-photon microscopy in cultured mouse brainstem neurons

    SciTech Connect

    Mueller, Michael . E-mail: mike@neuro-physiol.med.uni-goettingen.de; Mironov, Sergej L.; Ivannikov, Maxim V.; Schmidt, Joerg; Richter, Diethelm W.

    2005-02-01

    Two-photon microscopy of rhodamine 123-labeled mitochondria revealed that mitochondria of neurons cultured from mouse respiratory center form functionally coupled, dynamically organized aggregates such as chains and clusters, while single mitochondria were rarely seen. Mitochondrial chain structures predominate in dendrites, while irregularly shaped mitochondrial clusters are mostly found in the soma. Both types of mitochondrial structures showed chaotic Brownian motions and the mitochondrial chains also revealed well-directed movements. The latter dislocations were arrested upon mitochondrial depolarization or blockade of mitochondrial ATP synthesis. Depolymerization of microtubules by colchicine or nocodazole or inhibition of protein phosphatases by calyculin A disrupted mitochondrial chains and the mitochondria accumulated in the soma. Forskolin and IBMX reversibly blocked directed movements of mitochondria, but did not affect their overall spatial distribution. Thus, protein phosphorylation seems to control both mitochondrial transport and organization. Protein phosphorylation downstream of enhanced cytosolic cAMP levels apparently regulates the transition from motile to non-motile mitochondria, while phosphorylation resulting from inhibition of types 1 and 2A protein phosphatases massively disturbs mitochondrial organization. The complex phosphorylation processes seem to control the close interaction of mitochondria and cytoskeleton which may guarantee that mitochondria are immobilized at energetic hot spots and rearranged in response to changes in local energy demands.

  14. Tubulin binding blocks mitochondrial voltage-dependent anion channel and regulates respiration.

    PubMed

    Rostovtseva, Tatiana K; Sheldon, Kely L; Hassanzadeh, Elnaz; Monge, Claire; Saks, Valdur; Bezrukov, Sergey M; Sackett, Dan L

    2008-12-01

    Regulation of mitochondrial outer membrane (MOM) permeability has dual importance: in normal metabolite and energy exchange between mitochondria and cytoplasm and thus in control of respiration, and in apoptosis by release of apoptogenic factors into the cytosol. However, the mechanism of this regulation, dependent on the voltage-dependent anion channel (VDAC), the major channel of MOM, remains controversial. A long-standing puzzle is that in permeabilized cells, adenine nucleotide translocase (ANT) is less accessible to cytosolic ADP than in isolated mitochondria. We solve this puzzle by finding a missing player in the regulation of MOM permeability: the cytoskeletal protein tubulin. We show that nanomolar concentrations of dimeric tubulin induce voltage-sensitive reversible closure of VDAC reconstituted into planar phospholipid membranes. Tubulin strikingly increases VDAC voltage sensitivity and at physiological salt conditions could induce VDAC closure at <10 mV transmembrane potentials. Experiments with isolated mitochondria confirm these findings. Tubulin added to isolated mitochondria decreases ADP availability to ANT, partially restoring the low MOM permeability (high apparent K(m) for ADP) found in permeabilized cells. Our findings suggest a previously unknown mechanism of regulation of mitochondrial energetics, governed by VDAC and tubulin at the mitochondria-cytosol interface. This tubulin-VDAC interaction requires tubulin anionic C-terminal tail (CTT) peptides. The significance of this interaction may be reflected in the evolutionary conservation of length and anionic charge in CTT throughout eukaryotes, despite wide changes in the exact sequence. Additionally, tubulins that have lost significant length or anionic character are only found in cells that do not have mitochondria. PMID:19033201

  15. Translational Targeted Proteomics Profiling of Mitochondrial Energy Metabolic Pathways in Mouse and Human Samples.

    PubMed

    Wolters, Justina C; Ciapaite, Jolita; van Eunen, Karen; Niezen-Koning, Klary E; Matton, Alix; Porte, Robert J; Horvatovich, Peter; Bakker, Barbara M; Bischoff, Rainer; Permentier, Hjalmar P

    2016-09-01

    Absolute measurements of protein abundance are important in the understanding of biological processes and the precise computational modeling of biological pathways. We developed targeted LC-MS/MS assays in the selected reaction monitoring (SRM) mode to quantify over 50 mitochondrial proteins in a single run. The targeted proteins cover the tricarboxylic acid cycle, fatty acid β-oxidation, oxidative phosphorylation, and the detoxification of reactive oxygen species. Assays used isotopically labeled concatemers as internal standards designed to target murine mitochondrial proteins and their human orthologues. Most assays were also suitable to quantify the corresponding protein orthologues in rats. After exclusion of peptides that did not pass the selection criteria, we arrived at SRM assays for 55 mouse, 52 human, and 51 rat proteins. These assays were optimized in isolated mitochondrial fractions from mouse and rat liver and cultured human fibroblasts and in total liver extracts from mouse, rat, and human. The developed proteomics approach is suitable for the quantification of proteins in the mitochondrial energy metabolic pathways in mice, rats, and humans as a basis for translational research. Initial data show that the assays have great potential for elucidating the adaptive response of human patients to mutations in mitochondrial proteins in a clinical setting.

  16. Cyclophilin D deficiency improves mitochondrial function and learning/memory in aging Alzheimer disease mouse model.

    PubMed

    Du, Heng; Guo, Lan; Zhang, Wensheng; Rydzewska, Monika; Yan, Shidu

    2011-03-01

    Mitochondrial stress is one of the early features of Alzheimer disease (AD). Mitochondrial Aβ has been linked to mitochondrial toxicity. Our recent study demonstrated that cyclophilin D (CypD) mediated mitochondrial permeability transition pore (mPTP) is an important mechanism for neuronal and synaptic stress induced by both Aβ and oxidative stress. In transgenic AD-type mice overexpressing mutant amyloid precursor protein (APP) and Aβ (mAPP), CypD deficiency improves mitochondrial and synaptic function and learning/memory up to 12 months old. Here we provide evidence of the protective effects of CypD deficiency in aged AD mice (22-24 months). Cyp D deficient mAPP mice demonstrate less calcium-induced mitochondrial swelling, increased mitochondrial calcium uptake capacity, preserved mitochondrial respiratory function and improved spatial learning/memory even in old age (known to be the age for late stage AD pathology and synaptic dysfunction). These data demonstrate that abrogation of CypD results in persistent life-long protection against Aβ toxicity in an Alzheimer's disease mouse model, thereby suggesting that blockade of CypD may be of benefit for Alzheimer disease treatment.

  17. High-Resolution Respirometry for Mitochondrial Characterization of Ex Vivo Mouse Tissues.

    PubMed

    Cantó, Carles; Garcia-Roves, Pablo M

    2015-06-01

    This article describes methodologies to examine mitochondrial respiration in fresh preparations of mouse tissues, including skeletal muscle, heart, liver, white and brown adipose tissue, and brain. Reference values and tips to maximize experimental efficiencies are also provided. Finally, correction methods and complementary techniques to properly interpret the results are presented and contrasted.

  18. Increased mitochondrial biogenesis in muscle improves aging phenotypes in the mtDNA mutator mouse.

    PubMed

    Dillon, Lloye M; Williams, Siôn L; Hida, Aline; Peacock, Jacqueline D; Prolla, Tomas A; Lincoln, Joy; Moraes, Carlos T

    2012-05-15

    Aging is an intricate process that increases susceptibility to sarcopenia and cardiovascular diseases. The accumulation of mitochondrial DNA (mtDNA) mutations is believed to contribute to mitochondrial dysfunction, potentially shortening lifespan. The mtDNA mutator mouse, a mouse model with a proofreading-deficient mtDNA polymerase γ, was shown to develop a premature aging phenotype, including sarcopenia, cardiomyopathy and decreased lifespan. This phenotype was associated with an accumulation of mtDNA mutations and mitochondrial dysfunction. We found that increased expression of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), a crucial regulator of mitochondrial biogenesis and function, in the muscle of mutator mice increased mitochondrial biogenesis and function and also improved the skeletal muscle and heart phenotypes of the mice. Deep sequencing analysis of their mtDNA showed that the increased mitochondrial biogenesis did not reduce the accumulation of mtDNA mutations but rather caused a small increase. These results indicate that increased muscle PGC-1α expression is able to improve some premature aging phenotypes in the mutator mice without reverting the accumulation of mtDNA mutations.

  19. Turn up the power –pharmacological activation of mitochondrial biogenesis in mouse models

    PubMed Central

    Komen, J C; Thorburn, D R

    2014-01-01

    The oxidative phosphorylation (OXPHOS) system in mitochondria is responsible for the generation of the majority of cellular energy in the form of ATP. Patients with genetic OXPHOS disorders form the largest group of inborn errors of metabolism. Unfortunately, there is still a lack of efficient therapies for these disorders other than management of symptoms. Developing therapies has been complicated because, although the total group of OXPHOS patients is relatively large, there is enormous clinical and genetic heterogeneity within this patient population. Thus there has been a lot of interest in generating relevant mouse models for the different kinds of OXPHOS disorders. The most common treatment strategies tested in these mouse models have aimed to up-regulate mitochondrial biogenesis, in order to increase the residual OXPHOS activity present in affected animals and thereby to ameliorate the energy deficiency. Drugs such as bezafibrate, resveratrol and AICAR target the master regulator of mitochondrial biogenesis PGC-1α either directly or indirectly to manipulate mitochondrial metabolism. This review will summarize the outcome of preclinical treatment trials with these drugs in mouse models of OXPHOS disorders and discuss similar treatments in a number of mouse models of common diseases in which pathology is closely linked to mitochondrial dysfunction. In the majority of these studies the pharmacological activation of the PGC-1α axis shows true potential as therapy; however, other effects besides mitochondrial biogenesis may be contributing to this as well. Linked Articles This article is part of a themed issue on Mitochondrial Pharmacology: Energy, Injury & Beyond. To view the other articles in this issue visit http://dx.doi.org/10.1111/bph.2014.171.issue-8 PMID:24102298

  20. Mitochondrial bioenergetics and drug-induced toxicity in a panel of mouse embryonic fibroblasts with mitochondrial DNA single nucleotide polymorphisms

    SciTech Connect

    Pereira, Claudia V.; Oliveira, Paulo J.; Will, Yvonne; Nadanaciva, Sashi

    2012-10-15

    Mitochondrial DNA (mtDNA) variations including single nucleotide polymorphisms (SNPs) have been proposed to be involved in idiosyncratic drug reactions. However, current in vitro and in vivo models lack the genetic diversity seen in the human population. Our hypothesis is that different cell strains with distinct mtDNA SNPs may have different mitochondrial bioenergetic profiles and may therefore vary in their response to drug-induced toxicity. Therefore, we used an in vitro system composed of four strains of mouse embryonic fibroblasts (MEFs) with mtDNA polymorphisms. We sequenced mtDNA from embryonic fibroblasts isolated from four mouse strains, C57BL/6J, MOLF/EiJ, CZECHII/EiJ and PERA/EiJ, with the latter two being sequenced for the first time. The bioenergetic profile of the four strains of MEFs was investigated at both passages 3 and 10. Our results showed that there were clear differences among the four strains of MEFs at both passages, with CZECHII/EiJ having a lower mitochondrial robustness when compared to C57BL/6J, followed by MOLF/EiJ and PERA/EiJ. Seven drugs known to impair mitochondrial function were tested for their effect on the ATP content of the four strains of MEFs in both glucose- and galactose-containing media. Our results showed that there were strain-dependent differences in the response to some of the drugs. We propose that this model is a useful starting point to study compounds that may cause mitochondrial off-target toxicity in early stages of drug development, thus decreasing the number of experimental animals used. -- Highlights: ► mtDNA SNPs may be linked to individual predisposition to drug-induced toxicity. ► CZECHII/EiJ and PERA/EiJ mtDNA was sequenced for the first time in this study. ► Strain-dependent mitochondrial capacity differences were measured. ► Strain-dependent differences in response to mitochondrial toxicants were observed.

  1. Mitochondrial Diseases Part III: Therapeutic interventions in mouse models of OXPHOS deficiencies.

    PubMed

    Peralta, Susana; Torraco, Alessandra; Iommarini, Luisa; Diaz, Francisca

    2015-07-01

    Mitochondrial defects are the cause of numerous disorders affecting the oxidative phosphorylation system (OXPHOS) in humans leading predominantly to neurological and muscular degeneration. The molecular origin, manifestations, and progression of mitochondrial diseases have a broad spectrum, which makes very challenging to find a globally effective therapy. The study of the molecular mechanisms underlying the mitochondrial dysfunction indicates that there is a wide range of pathways, enzymes and molecules that can be potentially targeted for therapeutic purposes. Therefore, focusing on the pathology of the disease is essential to design new treatments. In this review, we will summarize and discuss the different therapeutic interventions tested in some mouse models of mitochondrial diseases emphasizing the molecular mechanisms of action and their potential applications.

  2. Permeabilization of brain tissue in situ enables multiregion analysis of mitochondrial function in a single mouse brain

    PubMed Central

    Herbst, Eric AF; Holloway, Graham P

    2015-01-01

    Abstract Mitochondria function as the core energy providers in the brain and symptoms of neurodegenerative diseases are often attributed to their dysregulation. Assessing mitochondrial function is classically performed in isolated mitochondria; however, this process requires significant isolation time, demand for abundant tissue and disruption of the cooperative mitochondrial reticulum, all of which reduce reliability when attempting to assess in vivo mitochondrial bioenergetics. Here we introduce a method that advances the assessment of mitochondrial respiration in the brain by permeabilizing existing brain tissue to grant direct access to the mitochondrial reticulum in situ. The permeabilized brain preparation allows for instant analysis of mitochondrial function with unaltered mitochondrial morphology using significantly small sample sizes (∼2 mg), which permits the analysis of mitochondrial function in multiple subregions within a single mouse brain. Here this technique was applied to assess regional variation in brain mitochondrial function with acute ischaemia–reperfusion injuries and to determine the role of reactive oxygen species in exacerbating dysfunction through the application of a transgenic mouse model overexpressing catalase within mitochondria. Through creating accessibility to small regions for the investigation of mitochondrial function, the permeabilized brain preparation enhances the capacity for examining regional differences in mitochondrial regulation within the brain, as the majority of genetic models used for unique approaches exist in the mouse model. PMID:25529987

  3. Preservation of mitochondrial functional integrity in mitochondria isolated from small cryopreserved mouse brain areas.

    PubMed

    Valenti, Daniela; de Bari, Lidia; De Filippis, Bianca; Ricceri, Laura; Vacca, Rosa Anna

    2014-01-01

    Studies of mitochondrial bioenergetics in brain pathophysiology are often precluded by the need to isolate mitochondria immediately after tissue dissection from a large number of brain biopsies for comparative studies. Here we present a procedure of cryopreservation of small brain areas from which mitochondrial enriched fractions (crude mitochondria) with high oxidative phosphorylation efficiency can be isolated. Small mouse brain areas were frozen and stored in a solution containing glycerol as cryoprotectant. Crude mitochondria were isolated by differential centrifugation from both cryopreserved and freshly explanted brain samples and were compared with respect to their ability to generate membrane potential and produce ATP. Intactness of outer and inner mitochondrial membranes was verified by polarographic ascorbate and cytochrome c tests and spectrophotometric assay of citrate synthase activity. Preservation of structural integrity and oxidative phosphorylation efficiency was successfully obtained in crude mitochondria isolated from different areas of cryopreserved mouse brain samples. Long-term cryopreservation of small brain areas from which intact and phosphorylating mitochondria can be isolated for the study of mitochondrial bioenergetics will significantly expand the study of mitochondrial defects in neurological pathologies, allowing large comparative studies and favoring interlaboratory and interdisciplinary analyses.

  4. MITOCHONDRIAL DISEASES PART II: MOUSE MODELS OF OXPHOS DEFICIENCIES CAUSED BY DEFECTS IN REGULATORY FACTORS AND OTHER COMPONENTS REQUIRED FOR MITOCHONDRIAL FUNCTION

    PubMed Central

    Iommarini, Luisa; Peralta, Susana; Torraco, Alessandra; Diaz, Francisca

    2015-01-01

    Mitochondrial disorders are defined as defects that affect the oxidative phosphorylation system (OXPHOS). They are characterized by a heterogeneous array of clinical presentations due in part to a wide variety of factors required for proper function of the components of the OXPHOS system. There is no cure for these disorders owing our poor knowledge of the pathogenic mechanisms of disease. To understand the mechanisms of human disease numerous mouse models have been developed in recent years. Here we summarize the features of several mouse models of mitochondrial diseases directly related to those factors affecting mtDNA maintenance, replication, transcription, translation as well to other proteins that are involved in mitochondrial dynamics and quality control which affect mitochondrial OXPHOS function without been intrinsic components of the system. We discuss how these models have contributed to our understanding of mitochondrial diseases and their pathogenic mechanisms. PMID:25640959

  5. Bioluminescence imaging of mitochondrial Ca2+ dynamics in soma and neurites of individual adult mouse sympathetic neurons

    PubMed Central

    Núñez, Lucía; Senovilla, Laura; Sanz-Blasco, Sara; Chamero, Pablo; Alonso, María T; Villalobos, Carlos; García-Sancho, Javier

    2007-01-01

    Changes in the cytosolic Ca2+ concentration ([Ca2+]c) are essential for triggering neurotransmitter release from presynaptic nerve terminals. Calcium-induced Ca2+ release (CICR) from the endoplasmic reticulum (ER) may amplify the [Ca2+]c signals and facilitate neurotransmitter release in sympathetic neurons. In adrenal chromaffin cells, functional triads are formed by voltage-operated Ca2+ channels (VOCCs), CICR sites and mitochondria. In fact, mitochondria take up most of the Ca2+ load entering the cells and are essential for shaping [Ca2+]c signals and exocytosis. Here we have investigated the existence of such functional triads in sympathetic neurons. The mitochondrial Ca2+ concentration ([Ca2+]m) in soma and neurites of individual mouse superior cervical ganglion (SCG) neurons was monitored by bioluminescence imaging of targeted aequorins. In soma, Ca2+ entry through VOCCs evoked rapid, near millimolar [Ca2+]m increases in a subpopulation of mitochondria containing about 40% of the aequorin. Caffeine evoked a similar [Ca2+]m increase in a mitochondrial pool containing about 30% of the aequorin and overlapping with the VOCC-sensitive pool. These observations suggest the existence of functional triads similar to the ones described in chromaffin cells. In neurites, mitochondria were able to buffer [Ca2+]c increases resulting from activation of VOCCs but not those mediated by caffeine-induced Ca2+ release from the ER. The weaker Ca2+ buffering by mitochondria in neurites could contribute to facilitate Ca2+-induced exocytosis at the presynaptic sites. PMID:17234693

  6. Mitochondrial pyruvate dehydrogenase phosphatase 1 regulates the early differentiation of cardiomyocytes from mouse embryonic stem cells.

    PubMed

    Heo, Hye Jin; Kim, Hyoung Kyu; Youm, Jae Boum; Cho, Sung Woo; Song, In-Sung; Lee, Sun Young; Ko, Tae Hee; Kim, Nari; Ko, Kyung Soo; Rhee, Byoung Doo; Han, Jin

    2016-01-01

    Mitochondria are crucial for maintaining the properties of embryonic stem cells (ESCs) and for regulating their subsequent differentiation into diverse cell lineages, including cardiomyocytes. However, mitochondrial regulators that manage the rate of differentiation or cell fate have been rarely identified. This study aimed to determine the potential mitochondrial factor that controls the differentiation of ESCs into cardiac myocytes. We induced cardiomyocyte differentiation from mouse ESCs (mESCs) and performed microarray assays to assess messenger RNA (mRNA) expression changes at differentiation day 8 (D8) compared with undifferentiated mESCs (D0). Among the differentially expressed genes, Pdp1 expression was significantly decreased (27-fold) on D8 compared to D0, which was accompanied by suppressed mitochondrial indices, including ATP levels, membrane potential, ROS and mitochondrial Ca(2+). Notably, Pdp1 overexpression significantly enhanced the mitochondrial indices and pyruvate dehydrogenase activity and reduced the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate compared to a mock control. In confirmation of this, a knockdown of the Pdp1 gene promoted the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate. In conclusion, our results suggest that mitochondrial PDP1 is a potential regulator that controls cardiac differentiation at an early differentiation stage in ESCs. PMID:27538372

  7. Mitochondrial pyruvate dehydrogenase phosphatase 1 regulates the early differentiation of cardiomyocytes from mouse embryonic stem cells

    PubMed Central

    Heo, Hye Jin; Kim, Hyoung Kyu; Youm, Jae Boum; Cho, Sung Woo; Song, In-Sung; Lee, Sun Young; Ko, Tae Hee; Kim, Nari; Ko, Kyung Soo; Rhee, Byoung Doo; Han, Jin

    2016-01-01

    Mitochondria are crucial for maintaining the properties of embryonic stem cells (ESCs) and for regulating their subsequent differentiation into diverse cell lineages, including cardiomyocytes. However, mitochondrial regulators that manage the rate of differentiation or cell fate have been rarely identified. This study aimed to determine the potential mitochondrial factor that controls the differentiation of ESCs into cardiac myocytes. We induced cardiomyocyte differentiation from mouse ESCs (mESCs) and performed microarray assays to assess messenger RNA (mRNA) expression changes at differentiation day 8 (D8) compared with undifferentiated mESCs (D0). Among the differentially expressed genes, Pdp1 expression was significantly decreased (27-fold) on D8 compared to D0, which was accompanied by suppressed mitochondrial indices, including ATP levels, membrane potential, ROS and mitochondrial Ca2+. Notably, Pdp1 overexpression significantly enhanced the mitochondrial indices and pyruvate dehydrogenase activity and reduced the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate compared to a mock control. In confirmation of this, a knockdown of the Pdp1 gene promoted the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate. In conclusion, our results suggest that mitochondrial PDP1 is a potential regulator that controls cardiac differentiation at an early differentiation stage in ESCs. PMID:27538372

  8. Glycolysis and mitochondrial respiration in mouse LDHC-null sperm.

    PubMed

    Odet, Fanny; Gabel, Scott; London, Robert E; Goldberg, Erwin; Eddy, Edward M

    2013-04-01

    We demonstrated previously that a knockout (KO) of the lactate dehydrogenase type C (Ldhc) gene disrupted male fertility and caused a considerable reduction in sperm glucose consumption, ATP production, and motility. While that study used mice with a mixed genetic background, the present study used C57BL/6 (B6) and 129S6 (129) Ldhc KO mice. We found that B6 KO males were subfertile and 129 KO males were infertile. Sperm from 129 wild-type (WT) mice have a lower glycolytic rate than sperm from B6 WT mice, resulting in a greater reduction in ATP production in 129 KO sperm than in B6 KO sperm. The lower glycolytic rate in 129 sperm offered a novel opportunity to examine the role of mitochondrial respiration in sperm ATP production and motility. We observed that in media containing a mitochondrial substrate (pyruvate or lactate) as the sole energy source, ATP levels and progressive motility in 129 KO sperm were similar to those in 129 WT sperm. However, when glucose was added, lactate was unable to maintain ATP levels or progressive motility in 129 KO sperm. The rate of respiration (ZO2) was high when 129 KO or WT sperm were incubated with lactate alone, but addition of glucose caused a reduction in ZO2. These results indicate that in the absence of glucose, 129 sperm can produce ATP via oxidative phosphorylation, but in the presence of glucose, oxidative phosphorylation is suppressed and the sperm utilize aerobic glycolysis, a phenomenon known as the Crabtree effect.

  9. Voltage-gated Ca2+ influx and mitochondrial Ca2+ initiate secretion from Aplysia neuroendocrine cells.

    PubMed

    Hickey, C M; Groten, C J; Sham, L; Carter, C J; Magoski, N S

    2013-10-10

    Neuroendocrine secretion often requires prolonged voltage-gated Ca(2+) entry; however, the ability of Ca(2+) from intracellular stores, such as endoplasmic reticulum or mitochondria, to elicit secretion is less clear. We examined this using the bag cell neurons, which trigger ovulation in Aplysia by releasing egg-laying hormone (ELH) peptide. Secretion from cultured bag cell neurons was observed as an increase in plasma membrane capacitance following Ca(2+) influx evoked by a 5-Hz, 1-min train of depolarizing steps under voltage-clamp. The response was similar for step durations of ≥ 50 ms, but fell off sharply with shorter stimuli. The capacitance change was attenuated by replacing external Ca(2+) with Ba(2+), blocking Ca(2+) channels, buffering intracellular Ca(2+) with EGTA, disrupting synaptic protein recycling, or genetic knock-down of ELH. Regarding intracellular stores, liberating mitochondrial Ca(2+) with the protonophore, carbonyl cyanide-p-trifluoromethoxyphenyl-hydrazone (FCCP), brought about an EGTA-sensitive elevation of capacitance. Conversely, no change was observed to Ca(2+) released from the endoplasmic reticulum or acidic stores. Prior exposure to FCCP lessened the train-induced capacitance increase, suggesting overlap in the pool of releasable vesicles. Employing GTP-γ-S to interfere with endocytosis delayed recovery (presumed membrane retrieval) of the capacitance change following FCCP, but not the train. Finally, secretion was correlated with reproductive behavior, in that neurons isolated from animals engaged in egg-laying presented a greater train-induced capacitance elevation vs quiescent animals. The bag cell neuron capacitance increase is consistent with peptide secretion requiring high Ca(2+), either from influx or stores, and may reflect the all-or-none nature of reproduction.

  10. Voltage-gated Ca2+ influx and mitochondrial Ca2+ initiate secretion from Aplysia neuroendocrine cells.

    PubMed

    Hickey, C M; Groten, C J; Sham, L; Carter, C J; Magoski, N S

    2013-10-10

    Neuroendocrine secretion often requires prolonged voltage-gated Ca(2+) entry; however, the ability of Ca(2+) from intracellular stores, such as endoplasmic reticulum or mitochondria, to elicit secretion is less clear. We examined this using the bag cell neurons, which trigger ovulation in Aplysia by releasing egg-laying hormone (ELH) peptide. Secretion from cultured bag cell neurons was observed as an increase in plasma membrane capacitance following Ca(2+) influx evoked by a 5-Hz, 1-min train of depolarizing steps under voltage-clamp. The response was similar for step durations of ≥ 50 ms, but fell off sharply with shorter stimuli. The capacitance change was attenuated by replacing external Ca(2+) with Ba(2+), blocking Ca(2+) channels, buffering intracellular Ca(2+) with EGTA, disrupting synaptic protein recycling, or genetic knock-down of ELH. Regarding intracellular stores, liberating mitochondrial Ca(2+) with the protonophore, carbonyl cyanide-p-trifluoromethoxyphenyl-hydrazone (FCCP), brought about an EGTA-sensitive elevation of capacitance. Conversely, no change was observed to Ca(2+) released from the endoplasmic reticulum or acidic stores. Prior exposure to FCCP lessened the train-induced capacitance increase, suggesting overlap in the pool of releasable vesicles. Employing GTP-γ-S to interfere with endocytosis delayed recovery (presumed membrane retrieval) of the capacitance change following FCCP, but not the train. Finally, secretion was correlated with reproductive behavior, in that neurons isolated from animals engaged in egg-laying presented a greater train-induced capacitance elevation vs quiescent animals. The bag cell neuron capacitance increase is consistent with peptide secretion requiring high Ca(2+), either from influx or stores, and may reflect the all-or-none nature of reproduction. PMID:23876326

  11. NAD(+)-dependent activation of Sirt1 corrects the phenotype in a mouse model of mitochondrial disease.

    PubMed

    Cerutti, Raffaele; Pirinen, Eija; Lamperti, Costanza; Marchet, Silvia; Sauve, Anthony A; Li, Wei; Leoni, Valerio; Schon, Eric A; Dantzer, Françoise; Auwerx, Johan; Viscomi, Carlo; Zeviani, Massimo

    2014-06-01

    Mitochondrial disorders are highly heterogeneous conditions characterized by defects of the mitochondrial respiratory chain. Pharmacological activation of mitochondrial biogenesis has been proposed as an effective means to correct the biochemical defects and ameliorate the clinical phenotype in these severely disabling, often fatal, disorders. Pathways related to mitochondrial biogenesis are targets of Sirtuin1, a NAD(+)-dependent protein deacetylase. As NAD(+) boosts the activity of Sirtuin1 and other sirtuins, intracellular levels of NAD(+) play a key role in the homeostatic control of mitochondrial function by the metabolic status of the cell. We show here that supplementation with nicotinamide riboside, a natural NAD(+) precursor, or reduction of NAD(+) consumption by inhibiting the poly(ADP-ribose) polymerases, leads to marked improvement of the respiratory chain defect and exercise intolerance of the Sco2 knockout/knockin mouse, a mitochondrial disease model characterized by impaired cytochrome c oxidase biogenesis. This strategy is potentially translatable into therapy of mitochondrial disorders in humans. PMID:24814483

  12. Mitochondrial Alterations and Oxidative Stress in an Acute Transient Mouse Model of Muscle Degeneration

    PubMed Central

    Ramadasan-Nair, Renjini; Gayathri, Narayanappa; Mishra, Sudha; Sunitha, Balaraju; Mythri, Rajeswara Babu; Nalini, Atchayaram; Subbannayya, Yashwanth; Harsha, Hindalahalli Chandregowda; Kolthur-Seetharam, Ullas; Bharath, Muchukunte Mukunda Srinivas

    2014-01-01

    Muscular dystrophies (MDs) and inflammatory myopathies (IMs) are debilitating skeletal muscle disorders characterized by common pathological events including myodegeneration and inflammation. However, an experimental model representing both muscle pathologies and displaying most of the distinctive markers has not been characterized. We investigated the cardiotoxin (CTX)-mediated transient acute mouse model of muscle degeneration and compared the cardinal features with human MDs and IMs. The CTX model displayed degeneration, apoptosis, inflammation, loss of sarcolemmal complexes, sarcolemmal disruption, and ultrastructural changes characteristic of human MDs and IMs. Cell death caused by CTX involved calcium influx and mitochondrial damage both in murine C2C12 muscle cells and in mice. Mitochondrial proteomic analysis at the initial phase of degeneration in the model detected lowered expression of 80 mitochondrial proteins including subunits of respiratory complexes, ATP machinery, fatty acid metabolism, and Krebs cycle, which further decreased in expression during the peak degenerative phase. The mass spectrometry (MS) data were supported by enzyme assays, Western blot, and histochemistry. The CTX model also displayed markers of oxidative stress and a lowered glutathione reduced/oxidized ratio (GSH/GSSG) similar to MDs, human myopathies, and neurogenic atrophies. MS analysis identified 6 unique oxidized proteins from Duchenne muscular dystrophy samples (n = 6) (versus controls; n = 6), including two mitochondrial proteins. Interestingly, these mitochondrial proteins were down-regulated in the CTX model thereby linking oxidative stress and mitochondrial dysfunction. We conclude that mitochondrial alterations and oxidative damage significantly contribute to CTX-mediated muscle pathology with implications for human muscle diseases. PMID:24220031

  13. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    SciTech Connect

    Han, Q.; Robinson, H.; Cai, T.; Tagle, D. A.; Li, J.

    2011-10-01

    Mammalian mAspAT (mitochondrial aspartate aminotransferase) is recently reported to have KAT (kynurenine aminotransferase) activity and plays a role in the biosynthesis of KYNA (kynurenic acid) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen oxo acids were tested for the co-substrate specificity of mouse mAspAT and 14 of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor-binding residues of mAspAT are similar to those of other KATs. The substrate-binding residues of mAspAT are slightly different from those of other KATs. Our results provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain.

  14. Biochemical and structural characterization of mouse mitochondrial aspartate aminotransferase, a newly identified kynurenine aminotransferase-IV

    PubMed Central

    Han, Qian; Robinson, Howard; Cai, Tao; Tagle, Danilo A.; Li, Jianyong

    2010-01-01

    Synopsis Mammalian mitochondrial aspartate aminotransferase (mAspAT) is recently reported to have kynurenine aminotransferase (KAT) activity and plays a role in the biosynthesis of kynurenic acid (KYNA) in rat, mouse and human brains. This study concerns the biochemical and structural characterization of mouse mAspAT. In this study, mouse mAspAT cDNA was amplified from mouse brain first stand cDNA and its recombinant protein was expressed in an Escherichia coli expression system. Sixteen keto acids were tested for the co-substrate specificity of mouse mAspAT and fourteen of them were shown to be capable of serving as co-substrates for the enzyme. Structural analysis of mAspAT by macromolecular crystallography revealed that the cofactor binding residues of mAspAT are similar to those of other KATs. The substrate binding residues of mAspAT are slightly different from those of other KATs. Our data provide a biochemical and structural basis towards understanding the overall physiological role of mAspAT in vivo and insight into controlling the levels of endogenous KYNA through modulation of the enzyme in the mouse brain. PMID:20977429

  15. Comparative studies of early liver dysfunction in senescence-accelerated mouse using mitochondrial proteomics approaches.

    PubMed

    Liu, Yashu; He, Jintang; Ji, Shaoyi; Wang, Qingsong; Pu, Hai; Jiang, Tingting; Meng, Lingyao; Yang, Xiuwei; Ji, Jianguo

    2008-09-01

    The liver is a complex and unique organ responsible for a breadth of functions crucial to sustaining life, especially for various metabolic processes in its mitochondria. Senescence-accelerated mouse prone/8 (SAMP8), a widely used aging model, exhibits an oxidative stress-induced aging phenotype and severe mitochondria-related liver pathology that are not seen in senescence-accelerated mouse resistant/1 (SAMR1). Here we used both two-dimensional electrophoresis- and ICAT-based mitochondrial proteomics analysis to view the liver mitochondrial protein alterations between SAMP8 and SAMR1. Compared with SAMR1, decreased expression and activity of mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase were detected in SAMP8 at 6 months old (SAMP8-6m). As the key enzyme of ketogenesis, 3-hydroxy-3-methylglutaryl-CoA synthase is well known to be transcriptionally regulated by peroxisome proliferator-activated receptor alpha, which was also expressed at lower levels in SAMP8-6m livers. In addition, down-regulation of two peroxisome proliferator-activated receptor alpha target gene products (acyl-CoA oxidase and enoyl-CoA hydratase), elevation of triglyceride, and reduction of acetyl-CoA were observed, indicating abnormal fatty acid metabolism in SAMP8-6m livers. In addition eight proteins (NDUAA, NDUBA, NDUB7, NDUS1, NDUS3, NDUV1, ETFA, and UCRI) of mitochondrial complexes were down-regulated in SAMP8-6m, resulting in mitochondria-related liver dysfunction characterized by enhanced oxidative stress-induced molecular damage (lipid peroxide and oxidized protein) and depressed energy production (ATP). Glutamine synthetase and ornithine aminotransferase involved in glutamine synthesis were up-regulated in SAMP8 livers at both 1 and 6 months old that may be related to the accumulation of glutamate and glutamine. Our work provided useful clues to understanding the molecular mechanism underlying liver dysfunction in senescence-accelerated mouse.

  16. Embryonic Lethality of Mitochondrial Pyruvate Carrier 1 Deficient Mouse Can Be Rescued by a Ketogenic Diet

    PubMed Central

    Krznar, Petra; Hörl, Manuel; Ammar, Zeinab; Montessuit, Sylvie; Pierredon, Sandra; Zamboni, Nicola; Martinou, Jean-Claude

    2016-01-01

    Mitochondrial import of pyruvate by the mitochondrial pyruvate carrier (MPC) is a central step which links cytosolic and mitochondrial intermediary metabolism. To investigate the role of the MPC in mammalian physiology and development, we generated a mouse strain with complete loss of MPC1 expression. This resulted in embryonic lethality at around E13.5. Mouse embryonic fibroblasts (MEFs) derived from mutant mice displayed defective pyruvate-driven respiration as well as perturbed metabolic profiles, and both defects could be restored by reexpression of MPC1. Labeling experiments using 13C-labeled glucose and glutamine demonstrated that MPC deficiency causes increased glutaminolysis and reduced contribution of glucose-derived pyruvate to the TCA cycle. Morphological defects were observed in mutant embryonic brains, together with major alterations of their metabolome including lactic acidosis, diminished TCA cycle intermediates, energy deficit and a perturbed balance of neurotransmitters. Strikingly, these changes were reversed when the pregnant dams were fed a ketogenic diet, which provides acetyl-CoA directly to the TCA cycle and bypasses the need for a functional MPC. This allowed the normal gestation and development of MPC deficient pups, even though they all died within a few minutes post-delivery. This study establishes the MPC as a key player in regulating the metabolic state necessary for embryonic development, neurotransmitter balance and post-natal survival. PMID:27176894

  17. Embryonic Lethality of Mitochondrial Pyruvate Carrier 1 Deficient Mouse Can Be Rescued by a Ketogenic Diet.

    PubMed

    Vanderperre, Benoît; Herzig, Sébastien; Krznar, Petra; Hörl, Manuel; Ammar, Zeinab; Montessuit, Sylvie; Pierredon, Sandra; Zamboni, Nicola; Martinou, Jean-Claude

    2016-05-01

    Mitochondrial import of pyruvate by the mitochondrial pyruvate carrier (MPC) is a central step which links cytosolic and mitochondrial intermediary metabolism. To investigate the role of the MPC in mammalian physiology and development, we generated a mouse strain with complete loss of MPC1 expression. This resulted in embryonic lethality at around E13.5. Mouse embryonic fibroblasts (MEFs) derived from mutant mice displayed defective pyruvate-driven respiration as well as perturbed metabolic profiles, and both defects could be restored by reexpression of MPC1. Labeling experiments using 13C-labeled glucose and glutamine demonstrated that MPC deficiency causes increased glutaminolysis and reduced contribution of glucose-derived pyruvate to the TCA cycle. Morphological defects were observed in mutant embryonic brains, together with major alterations of their metabolome including lactic acidosis, diminished TCA cycle intermediates, energy deficit and a perturbed balance of neurotransmitters. Strikingly, these changes were reversed when the pregnant dams were fed a ketogenic diet, which provides acetyl-CoA directly to the TCA cycle and bypasses the need for a functional MPC. This allowed the normal gestation and development of MPC deficient pups, even though they all died within a few minutes post-delivery. This study establishes the MPC as a key player in regulating the metabolic state necessary for embryonic development, neurotransmitter balance and post-natal survival. PMID:27176894

  18. A mouse model of mitochondrial complex III dysfunction induced by myxothiazol

    SciTech Connect

    Davoudi, Mina; Kallijärvi, Jukka; Marjavaara, Sanna; Kotarsky, Heike; Hansson, Eva; Levéen, Per; Fellman, Vineta

    2014-04-18

    Highlights: • Reversible chemical inhibition of complex III in wild type mouse. • Myxothiazol causes decreased complex III activity in mouse liver. • The model is useful for therapeutic trials to improve mitochondrial function. - Abstract: Myxothiazol is a respiratory chain complex III (CIII) inhibitor that binds to the ubiquinol oxidation site Qo of CIII. It blocks electron transfer from ubiquinol to cytochrome b and thus inhibits CIII activity. It has been utilized as a tool in studies of respiratory chain function in in vitro and cell culture models. We developed a mouse model of biochemically induced and reversible CIII inhibition using myxothiazol. We administered myxothiazol intraperitoneally at a dose of 0.56 mg/kg to C57Bl/J6 mice every 24 h and assessed CIII activity, histology, lipid content, supercomplex formation, and gene expression in the livers of the mice. A reversible CIII activity decrease to 50% of control value occurred at 2 h post-injection. At 74 h only minor histological changes in the liver were found, supercomplex formation was preserved and no significant changes in the expression of genes indicating hepatotoxicity or inflammation were found. Thus, myxothiazol-induced CIII inhibition can be induced in mice for four days in a row without overt hepatotoxicity or lethality. This model could be utilized in further studies of respiratory chain function and pharmacological approaches to mitochondrial hepatopathies.

  19. Maternal diet-induced obesity alters mitochondrial activity and redox status in mouse oocytes and zygotes.

    PubMed

    Igosheva, Natalia; Abramov, Andrey Y; Poston, Lucilla; Eckert, Judith J; Fleming, Tom P; Duchen, Michael R; McConnell, Josie

    2010-01-01

    The negative impact of obesity on reproductive success is well documented but the stages at which development of the conceptus is compromised and the mechanisms responsible for the developmental failure still remain unclear. Recent findings suggest that mitochondria may be a contributing factor. However to date no studies have directly addressed the consequences of maternal obesity on mitochondria in early embryogenesis.Using an established murine model of maternal diet induced obesity and a live cell dynamic fluorescence imaging techniques coupled with molecular biology we have investigated the underlying mechanisms of obesity-induced reduced fertility. Our study is the first to show that maternal obesity prior to conception is associated with altered mitochondria in mouse oocytes and zygotes. Specifically, maternal diet-induced obesity in mice led to an increase in mitochondrial potential, mitochondrial DNA content and biogenesis. Generation of reactive oxygen species (ROS) was raised while glutathione was depleted and the redox state became more oxidised, suggestive of oxidative stress. These altered mitochondrial properties were associated with significant developmental impairment as shown by the increased number of obese mothers who failed to support blastocyst formation compared to lean dams. We propose that compromised oocyte and early embryo mitochondrial metabolism, resulting from excessive nutrient exposure prior to and during conception, may underlie poor reproductive outcomes frequently reported in obese women.

  20. A mitochondrial therapeutic reverses visual decline in mouse models of diabetes.

    PubMed

    Alam, Nazia M; Mills, William C; Wong, Aimee A; Douglas, Robert M; Szeto, Hazel H; Prusky, Glen T

    2015-07-01

    Diabetic retinopathy is characterized by progressive vision loss and the advancement of retinal micoraneurysms, edema and angiogenesis. Unfortunately, managing glycemia or targeting vascular complications with anti-vascular endothelial growth factor agents has shown only limited efficacy in treating the deterioration of vision in diabetic retinopathy. In light of growing evidence that mitochondrial dysfunction is an independent pathophysiology of diabetes and diabetic retinopathy, we investigated whether selectively targeting and improving mitochondrial dysfunction is a viable treatment for visual decline in diabetes. Measures of spatial visual behavior, blood glucose, bodyweight and optical clarity were made in mouse models of diabetes. Treatment groups were administered MTP-131, a water-soluble tetrapeptide that selectively targets mitochondrial cardiolipin and promotes efficient electron transfer, either systemically or in eye drops. Progressive visual decline emerged in untreated animals before the overt symptoms of metabolic and ophthalmic abnormalities were manifest, but with time, visual dysfunction was accompanied by compromised glucose clearance, and elevated blood glucose and bodyweight. MTP-131 treatment reversed the visual decline without improving glycemic control or reducing bodyweight. These data provide evidence that visuomotor decline is an early complication of diabetes. They also indicate that selectively treating mitochondrial dysfunction with MTP-131 has the potential to remediate the visual dysfunction and to complement existing treatments for diabetic retinopathy. PMID:26035391

  1. Reducing Mitochondrial ROS Improves Disease-related Pathology in a Mouse Model of Ataxia-telangiectasia

    PubMed Central

    D'Souza, Anthony D; Parish, Ian A; Krause, Diane S; Kaech, Susan M; Shadel, Gerald S

    2013-01-01

    The disease ataxia-telangiectasia (A-T) has no cure and few treatment options. It is caused by mutations in the ATM kinase, which functions in the DNA-damage response and redox sensing. In addition to severe cerebellar degeneration, A-T pathology includes cancer predisposition, sterility, immune system dysfunction, and bone marrow abnormalities. These latter phenotypes are recapitulated in the ATM null (ATM−/−) mouse model of the disease. Since oxidative stress and mitochondrial dysfunction are implicated in A-T, we determined whether reducing mitochondrial reactive oxygen species (ROS) via overexpression of catalase targeted to mitochondria (mCAT) alleviates A-T–related pathology in ATM−/− mice. We found that mCAT has many beneficial effects in this context, including reduced propensity to develop thymic lymphoma, improved bone marrow hematopoiesis and macrophage differentiation in vitro, and partial rescue of memory T-cell developmental defects. Our results suggest that positive effects observed on cancer development may be linked to mCAT reducing mitochondrial ROS, lactate production, and TORC1 signaling in transforming double-positive cells, whereas beneficial effects in memory T cells appear to be TORC1-independent. Altogether, this study provides proof-of-principle that reducing mitochondrial ROS production per se may be therapeutic for the disease, which may have advantages compared with more general antioxidant strategies. PMID:23011031

  2. Identification of nonferritin mitochondrial iron deposits in a mouse model of Friedreich ataxia

    PubMed Central

    Whitnall, Megan; Rahmanto, Yohan Suryo; Huang, Michael L.-H.; Saletta, Federica; Lok, Hiu Chuen; Gutiérrez, Lucía; Lázaro, Francisco J.; Fleming, Adam J.; St. Pierre, Tim G.; Mikhael, Marc R.; Ponka, Prem; Richardson, Des R.

    2012-01-01

    There is no effective treatment for the cardiomyopathy of the most common autosomal recessive ataxia, Friedreich ataxia (FA). This disease is due to decreased expression of the mitochondrial protein, frataxin, which leads to alterations in mitochondrial iron (Fe) metabolism. The identification of potentially toxic mitochondrial Fe deposits in FA suggests Fe plays a role in its pathogenesis. Studies using the muscle creatine kinase (MCK) conditional frataxin knockout mouse that mirrors the disease have demonstrated frataxin deletion alters cardiac Fe metabolism. Indeed, there are pronounced changes in Fe trafficking away from the cytosol to the mitochondrion, leading to a cytosolic Fe deficiency. Considering Fe deficiency can induce apoptosis and cell death, we examined the effect of dietary Fe supplementation, which led to body Fe loading and limited the cardiac hypertrophy in MCK mutants. Furthermore, this study indicates a unique effect of heart and skeletal muscle-specific frataxin deletion on systemic Fe metabolism. Namely, frataxin deletion induces a signaling mechanism to increase systemic Fe levels and Fe loading in tissues where frataxin expression is intact (i.e., liver, kidney, and spleen). Examining the mutant heart, native size-exclusion chromatography, transmission electron microscopy, Mössbauer spectroscopy, and magnetic susceptibility measurements demonstrated that in the absence of frataxin, mitochondria contained biomineral Fe aggregates, which were distinctly different from isolated mammalian ferritin molecules. These mitochondrial aggregates of Fe, phosphorus, and sulfur, probably contribute to the oxidative stress and pathology observed in the absence of frataxin. PMID:23169664

  3. Methoxychlor causes mitochondrial dysfunction and oxidative damage in the mouse ovary.

    PubMed

    Gupta, R K; Schuh, R A; Fiskum, G; Flaws, J A

    2006-11-01

    Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by causing ovarian atrophy, persistent estrous cyclicity, and antral follicle atresia (apoptotic cell death). Oxidative damage resulting from reactive oxygen species (ROS) generation has been demonstrated to lead to toxicant-induced cell death. Thus, this work tested the hypothesis that MXC causes oxidative damage to the mouse ovary and affects mitochondrial respiration in a manner that stimulates ROS production. For the in vitro experiments, mitochondria were collected from adult cycling mouse ovaries, treated with vehicle (dimethyl sulfoxide; DMSO) or MXC, and subjected to polarographic measurements of respiration. For the in vivo experiments, adult cycling CD-1 mice were dosed with either vehicle (sesame oil) or MXC for 20 days. After treatment, ovarian mitochondria were isolated and subjected to measurements of respiration and fluorimetric measurements of H2O2 production. Some ovaries were also fixed and processed for immunohistochemistry using antibodies for ROS production markers: nitrotyrosine and 8-hydroxy-2'-deoxyguanosine (8-OHG). Ovaries from in vivo experiments were also used to measure the mRNA expression and activity of antioxidants such as Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX), and catalase (CAT). The results indicate that MXC significantly impairs mitochondrial respiration, increases production of H2O2, causes more staining for nitrotyrosine and 8-OHG in antral follicles, and decreases the expression and activity of SOD1, GPX, and CAT as compared to controls. Collectively, these data indicate that MXC inhibits mitochondrial respiration, causes ROS production, and decreases antioxidant expression and activity in the ovary, specifically in the antral follicles. Therefore, it is possible that MXC causes atresia of ovarian antral follicles by inducing oxidative stress through mitochondrial production of ROS.

  4. Methoxychlor causes mitochondrial dysfunction and oxidative damage in the mouse ovary

    SciTech Connect

    Gupta, R.K.; Schuh, R.A.; Fiskum, G.; Flaws, J.A. . E-mail: jflaws@epi.umaryland.edu

    2006-11-01

    Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by causing ovarian atrophy, persistent estrous cyclicity, and antral follicle atresia (apoptotic cell death). Oxidative damage resulting from reactive oxygen species (ROS) generation has been demonstrated to lead to toxicant-induced cell death. Thus, this work tested the hypothesis that MXC causes oxidative damage to the mouse ovary and affects mitochondrial respiration in a manner that stimulates ROS production. For the in vitro experiments, mitochondria were collected from adult cycling mouse ovaries, treated with vehicle (dimethyl sulfoxide; DMSO) or MXC, and subjected to polarographic measurements of respiration. For the in vivo experiments, adult cycling CD-1 mice were dosed with either vehicle (sesame oil) or MXC for 20 days. After treatment, ovarian mitochondria were isolated and subjected to measurements of respiration and fluorimetric measurements of H{sub 2}O{sub 2} production. Some ovaries were also fixed and processed for immunohistochemistry using antibodies for ROS production markers: nitrotyrosine and 8-hydroxy-2'-deoxyguanosine (8-OHG). Ovaries from in vivo experiments were also used to measure the mRNA expression and activity of antioxidants such as Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX), and catalase (CAT). The results indicate that MXC significantly impairs mitochondrial respiration, increases production of H{sub 2}O{sub 2}, causes more staining for nitrotyrosine and 8-OHG in antral follicles, and decreases the expression and activity of SOD1, GPX, and CAT as compared to controls. Collectively, these data indicate that MXC inhibits mitochondrial respiration, causes ROS production, and decreases antioxidant expression and activity in the ovary, specifically in the antral follicles. Therefore, it is possible that MXC causes atresia of ovarian antral follicles by inducing oxidative stress through mitochondrial production of ROS.

  5. Purinergic signalling mobilizes mitochondrial Ca²⁺ in mouse Sertoli cells.

    PubMed

    Veitinger, Sophie; Veitinger, Thomas; Cainarca, Silvia; Fluegge, Daniela; Engelhardt, Corinna H; Lohmer, Stefan; Hatt, Hanns; Corazza, Sabrina; Spehr, Jennifer; Neuhaus, Eva M; Spehr, Marc

    2011-11-01

    Intimate bidirectional communication between Sertoli cells and developing germ cells ensures the integrity and efficiency of spermatogenesis. Yet, a conceptual mechanistic understanding of the physiological principles that underlie Sertoli cell autocrine and paracrine signalling is lacking. Here, we characterize a purinergic Ca(2+) signalling network in immature mouse Sertoli cells that consists of both P2X2 and P2Y2 purinoceptor subtypes, the endoplasmic reticulum and, notably, mitochondria. By combining a transgenic mouse model with a dedicated bioluminescence imaging device, we describe a novel method to monitor mitochondrial Ca(2+) mobilization in Sertoli cells at subcellular spatial and millisecond temporal resolution. Our data identify mitochondria as essential components of the Sertoli cell signalling 'toolkit' that control the shape of purinergic Ca(2+) responses, and probably several other paracrine Ca(2+)-dependent signals. PMID:21859825

  6. Mitochondrial APE1/Ref-1 suppressed protein kinase C-induced mitochondrial dysfunction in mouse endothelial cells.

    PubMed

    Joo, Hee Kyoung; Lee, Yu Ran; Park, Myoung Soo; Choi, Sunga; Park, Kyoungsook; Lee, Sang Ki; Kim, Cuk-Seong; Park, Jin Bong; Jeon, Byeong Hwa

    2014-07-01

    Protein kinase C (PKC) induces mitochondrial dysfunction, which is an important pathological factor in cardiovascular diseases. The role of apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE1/Ref-1) on PKC-induced mitochondrial dysfunction has not been variously investigated. In this study, phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, induced mitochondrial hyperpolarization and reactive oxygen species generation and also increased mitochondrial translocation of APE1/Ref-1. APE1/Ref-1 overexpression suppressed PMA-induced mitochondrial dysfunction. In contrast, gene silencing of APE1/Ref-1 increased the sensitivity of mitochondrial dysfunction. Moreover, mitochondrial targeting sequence (MTS)-fused APE1/Ref-1 more effectively suppressed PMA-induced mitochondrial dysfunctions. These results suggest that mitochondrial APE1/Ref-1 is contributed to the protective role to protein kinase C-induced mitochondrial dysfunction in endothelial cells.

  7. Palmitate induces ER calcium depletion and apoptosis in mouse podocytes subsequent to mitochondrial oxidative stress

    PubMed Central

    Xu, S; Nam, S M; Kim, J-H; Das, R; Choi, S-K; Nguyen, T T; Quan, X; Choi, S J; Chung, C H; Lee, E Y; Lee, I-K; Wiederkehr, A; Wollheim, C B; Cha, S-K; Park, K-S

    2015-01-01

    Pathologic alterations in podocytes lead to failure of an essential component of the glomerular filtration barrier and proteinuria in chronic kidney diseases. Elevated levels of saturated free fatty acid (FFA) are harmful to various tissues, implemented in the progression of diabetes and its complications such as proteinuria in diabetic nephropathy. Here, we investigated the molecular mechanism of palmitate cytotoxicity in cultured mouse podocytes. Incubation with palmitate dose-dependently increased cytosolic and mitochondrial reactive oxygen species, depolarized the mitochondrial membrane potential, impaired ATP synthesis and elicited apoptotic cell death. Palmitate not only evoked mitochondrial fragmentation but also caused marked dilation of the endoplasmic reticulum (ER). Consistently, palmitate upregulated ER stress proteins, oligomerized stromal interaction molecule 1 (STIM1) in the subplasmalemmal ER membrane, abolished the cyclopiazonic acid-induced cytosolic Ca2+ increase due to depletion of luminal ER Ca2+. Palmitate-induced ER Ca2+ depletion and cytotoxicity were blocked by a selective inhibitor of the fatty-acid transporter FAT/CD36. Loss of the ER Ca2+ pool induced by palmitate was reverted by the phospholipase C (PLC) inhibitor edelfosine. Palmitate-dependent activation of PLC was further demonstrated by following cytosolic translocation of the pleckstrin homology domain of PLC in palmitate-treated podocytes. An inhibitor of diacylglycerol (DAG) kinase, which elevates cytosolic DAG, strongly promoted ER Ca2+ depletion by low-dose palmitate. GF109203X, a PKC inhibitor, partially prevented palmitate-induced ER Ca2+ loss. Remarkably, the mitochondrial antioxidant mitoTEMPO inhibited palmitate-induced PLC activation, ER Ca2+ depletion and cytotoxicity. Palmitate elicited cytoskeletal changes in podocytes and increased albumin permeability, which was also blocked by mitoTEMPO. These data suggest that oxidative stress caused by saturated FFA leads to

  8. Expression of deoxynucleoside kinases and 5'-nucleotidases in mouse tissues: implications for mitochondrial toxicity.

    PubMed

    Rylova, Svetlana N; Mirzaee, Saeedeh; Albertioni, Freidoun; Eriksson, Staffan

    2007-06-30

    Anti-HIV nucleoside therapy can result in mitochondrial toxicity affecting muscles, peripheral nerves, pancreas and adipose tissue. The cytosolic deoxycytidine kinase (dCK; EC 2.7.1.74) and thymidine kinase (TK1; EC 2.7.1.21), the mitochondrial thymidine kinase (TK2) and deoxyguanosine kinase (dGK; EC 2.7.1.113) as well as 5'-deoxynucleotidases (5'-dNT; EC 3.1.3.5) are enzymes that control rate-limiting steps in formation of intracellular and intra-mitochondrial nucleotides. The mRNA levels and activities of these enzymes were determined in mouse tissues, using real-time PCR and selective enzyme assays. The expression of mRNA for all these enzymes and the mitochondrial deoxynucleotide carrier was detected in all tissues with a 5-10-fold variation. TK1 activities were only clearly detected in spleen and testis, while TK2, dGK and dCK activities were found in all tissues. dGK activities were higher than any other dNK in all tissues, except spleen and testis. In skeletal muscle dGK activity was 5-fold lower, TK2 and dCK levels were 10-fold lower as compared with other tissues. The variation in 5'-dNT activities was about eight-fold with the highest levels in brain and lowest in brown fat. Thus, the salvage of deoxynucleosides in muscles is 5-10-fold lower as compared to other non-proliferating tissues and 100-fold lower compared to spleen. These results may help to explain tissue specific toxicity observed with nucleoside analogs used in HIV treatment as well as symptoms in inherited mitochondrial TK2 deficiencies.

  9. Defects in Mitochondrial Dynamics and Metabolomic Signatures of Evolving Energetic Stress in Mouse Models of Familial Alzheimer's Disease

    PubMed Central

    Trushina, Eugenia; Nemutlu, Emirhan; Zhang, Song; Christensen, Trace; Camp, Jon; Mesa, Janny; Siddiqui, Ammar; Tamura, Yasushi; Sesaki, Hiromi; Wengenack, Thomas M.; Dzeja, Petras P.; Poduslo, Joseph F.

    2012-01-01

    Background The identification of early mechanisms underlying Alzheimer's Disease (AD) and associated biomarkers could advance development of new therapies and improve monitoring and predicting of AD progression. Mitochondrial dysfunction has been suggested to underlie AD pathophysiology, however, no comprehensive study exists that evaluates the effect of different familial AD (FAD) mutations on mitochondrial function, dynamics, and brain energetics. Methods and Findings We characterized early mitochondrial dysfunction and metabolomic signatures of energetic stress in three commonly used transgenic mouse models of FAD. Assessment of mitochondrial motility, distribution, dynamics, morphology, and metabolomic profiling revealed the specific effect of each FAD mutation on the development of mitochondrial stress and dysfunction. Inhibition of mitochondrial trafficking was characteristic for embryonic neurons from mice expressing mutant human presenilin 1, PS1(M146L) and the double mutation of human amyloid precursor protein APP(Tg2576) and PS1(M146L) contributing to the increased susceptibility of neurons to excitotoxic cell death. Significant changes in mitochondrial morphology were detected in APP and APP/PS1 mice. All three FAD models demonstrated a loss of the integrity of synaptic mitochondria and energy production. Metabolomic profiling revealed mutation-specific changes in the levels of metabolites reflecting altered energy metabolism and mitochondrial dysfunction in brains of FAD mice. Metabolic biomarkers adequately reflected gender differences similar to that reported for AD patients and correlated well with the biomarkers currently used for diagnosis in humans. Conclusions Mutation-specific alterations in mitochondrial dynamics, morphology and function in FAD mice occurred prior to the onset of memory and neurological phenotype and before the formation of amyloid deposits. Metabolomic signatures of mitochondrial stress and altered energy metabolism indicated

  10. Neural stem cell transplantation enhances mitochondrial biogenesis in a transgenic mouse model of Alzheimer's disease-like pathology.

    PubMed

    Zhang, Wei; Gu, Guo-Jun; Shen, Xing; Zhang, Qi; Wang, Gang-Min; Wang, Pei-Jun

    2015-03-01

    Mitochondrial dysfunction, especially a defect in mitochondrial biogenesis, is an early and prominent feature of Alzheimer's disease (AD). Previous studies demonstrated that the number of mitochondria is significantly reduced in susceptible hippocampal neurons from AD patients. Neural stem cell (NSC) transplantation in AD-like mice can compensate for the neuronal loss resulting from amyloid-beta protein deposition. The effects of NSC transplantation on mitochondrial biogenesis and cognitive function in AD-like mice, however, are poorly understood. In this study, we injected NSCs or vehicle into 12-month-old amyloid precursor protein (APP)/PS1 transgenic mice, a mouse model of AD-like pathology. The effects of NSC transplantation on cognitive function, the amount of mitochondrial DNA, the expression of mitochondrial biogenesis factors and mitochondria-related proteins, and mitochondrial morphology were investigated. Our results show that in NSC-injected APP/PS1 (Tg-NSC) mice, the cognitive function, number of mitochondria, and expression of mitochondria-related proteins, specifically the mitochondrial fission factors (dynamin-related protein 1 [Drp1] and fission 1 [Fis1]) and the mitochondrial fusion factor optic atrophy 1 (OPA1), were significantly increased compared with those in age-matched vehicle-injected APP/PS1 (Tg-Veh) mice, whereas the expression of mitochondrial fusion factors mitofusion 1 (Mfn1) and Mfn2 was significantly decreased. These data indicate that NSC transplantation may enhance mitochondria biogenesis and further rescue cognitive deficits in AD-like mice. PMID:25582749

  11. Mitoguardin-1 and -2 promote maturation and the developmental potential of mouse oocytes by maintaining mitochondrial dynamics and functions.

    PubMed

    Liu, Xiao-Man; Zhang, Yong-Ping; Ji, Shu-Yan; Li, Bo-Tai; Tian, Xuejun; Li, Dali; Tong, Chao; Fan, Heng-Yu

    2016-01-12

    Mitochondrial dynamics change mitochondrial morphological features and numbers as a part of adaptive cellular metabolism, which is vital for most eukaryotic cells and organisms. A disease or even death of an animal can occur if these dynamics are disrupted. Using large-scale genetic screening in fruit flies, we previously found the gene mitoguardin (Miga), which encodes a mitochondrial outer-membrane protein and promotes mitochondrial fusion. Knockout mouse strains were generated for the mammalian Miga homologs Miga1 and Miga2. Miga1/2-/- females show greatly reduced quality of oocytes and early embryos and are subfertile. Mitochondria became clustered in the cytoplasm of oocytes from the germinal-vesicle stage to meiosis II; production of reactive oxygen species increased in mitochondria and caused damage to mitochondrial ultrastructures. Additionally, reduced ATP production, a decreased mitochondrial-DNA copy number, and lower mitochondrial membrane potential were detected in Miga1/2-/- oocytes during meiotic maturation. These changes resulted in low rates of polar-body extrusion during oocyte maturation, reduced developmental potential of the resulting early embryos, and consequently female subfertility. We provide direct evidence that MIGA1/2-regulated mitochondrial dynamics is crucial for mitochondrial functions, ensure oocyte maturation, and maintain the developmental potential. PMID:26716412

  12. Mitochondrial dysfunction in a transgenic mouse model expressing human amyloid precursor protein (APP) with the Arctic mutation.

    PubMed

    Rönnbäck, Annica; Pavlov, Pavel F; Mansory, Mansorah; Gonze, Prisca; Marlière, Nicolas; Winblad, Bengt; Graff, Caroline; Behbahani, Homira

    2016-02-01

    Accumulation of amyloid β-peptide (Aβ) in the brain is an important event in the pathogenesis of Alzheimer disease. We have used a transgenic mouse model expressing human amyloid precursor protein (APP) with the Arctic mutation to investigate whether Aβ deposition is correlated with mitochondrial functions in these animals. We found evidence of mitochondrial dysfunction (i.e., decreased mitochondrial membrane potential, increased production of reactive oxygen species and oxidative DNA damage) at 6 months of age, when the mice showed very mild Aβ deposition. More pronounced mitochondrial abnormalities were present in 24-month-old TgAPParc mice with more extensive Aβ pathology. This study demonstrates for the first time mitochondrial dysfunction in transgenic mice with a mutation within the Aβ peptide (the Arctic APP mutation), and confirms previous studies suggesting that mitochondrial dysfunction and oxidative stress is an early event in the pathogenesis of Alzheimer disease. This study demonstrates mitochondrial dysfunction in transgenic mice with a mutation within the amyloid beta (Aβ) peptide (the Arctic amyloid precursor protein (APP) mutation). We found evidence of mitochondrial dysfunction (i.e. decreased mitochondrial membrane potential (MMP), increased production of reactive oxygen species (ROS) and oxidative DNA damage) at 6 months of age, when very mild Aβ deposition is present in the mice. Also, the cytochrome c (COX) activity was significantly decreased in mitochondria from transgenic mice at 24 months of age.

  13. Mitoguardin-1 and -2 promote maturation and the developmental potential of mouse oocytes by maintaining mitochondrial dynamics and functions

    PubMed Central

    Liu, Xiao-Man; Zhang, Yong-Ping; Ji, Shu-Yan; Li, Bo-Tai; Tian, Xuejun; Li, Dali; Tong, Chao; Fan, Heng-Yu

    2016-01-01

    Mitochondrial dynamics change mitochondrial morphological features and numbers as a part of adaptive cellular metabolism, which is vital for most eukaryotic cells and organisms. A disease or even death of an animal can occur if these dynamics are disrupted. Using large-scale genetic screening in fruit flies, we previously found the gene mitoguardin (Miga), which encodes a mitochondrial outer-membrane protein and promotes mitochondrial fusion. Knockout mouse strains were generated for the mammalian Miga homologs Miga1 and Miga2. Miga1/2−/− females show greatly reduced quality of oocytes and early embryos and are subfertile. Mitochondria became clustered in the cytoplasm of oocytes from the germinal-vesicle stage to meiosis II; production of reactive oxygen species increased in mitochondria and caused damage to mitochondrial ultrastructures. Additionally, reduced ATP production, a decreased mitochondrial-DNA copy number, and lower mitochondrial membrane potential were detected in Miga1/2−/− oocytes during meiotic maturation. These changes resulted in low rates of polar-body extrusion during oocyte maturation, reduced developmental potential of the resulting early embryos, and consequently female subfertility. We provide direct evidence that MIGA1/2-regulated mitochondrial dynamics is crucial for mitochondrial functions, ensure oocyte maturation, and maintain the developmental potential. PMID:26716412

  14. Neural stem cell transplantation enhances mitochondrial biogenesis in a transgenic mouse model of Alzheimer's disease-like pathology.

    PubMed

    Zhang, Wei; Gu, Guo-Jun; Shen, Xing; Zhang, Qi; Wang, Gang-Min; Wang, Pei-Jun

    2015-03-01

    Mitochondrial dysfunction, especially a defect in mitochondrial biogenesis, is an early and prominent feature of Alzheimer's disease (AD). Previous studies demonstrated that the number of mitochondria is significantly reduced in susceptible hippocampal neurons from AD patients. Neural stem cell (NSC) transplantation in AD-like mice can compensate for the neuronal loss resulting from amyloid-beta protein deposition. The effects of NSC transplantation on mitochondrial biogenesis and cognitive function in AD-like mice, however, are poorly understood. In this study, we injected NSCs or vehicle into 12-month-old amyloid precursor protein (APP)/PS1 transgenic mice, a mouse model of AD-like pathology. The effects of NSC transplantation on cognitive function, the amount of mitochondrial DNA, the expression of mitochondrial biogenesis factors and mitochondria-related proteins, and mitochondrial morphology were investigated. Our results show that in NSC-injected APP/PS1 (Tg-NSC) mice, the cognitive function, number of mitochondria, and expression of mitochondria-related proteins, specifically the mitochondrial fission factors (dynamin-related protein 1 [Drp1] and fission 1 [Fis1]) and the mitochondrial fusion factor optic atrophy 1 (OPA1), were significantly increased compared with those in age-matched vehicle-injected APP/PS1 (Tg-Veh) mice, whereas the expression of mitochondrial fusion factors mitofusion 1 (Mfn1) and Mfn2 was significantly decreased. These data indicate that NSC transplantation may enhance mitochondria biogenesis and further rescue cognitive deficits in AD-like mice.

  15. Trihalomethanes in liver pathology: Mitochondrial dysfunction and oxidative stress in the mouse.

    PubMed

    Faustino-Rocha, Ana I; Rodrigues, D; da Costa, R Gil; Diniz, C; Aragão, S; Talhada, D; Botelho, M; Colaço, A; Pires, M J; Peixoto, F; Oliveira, P A

    2016-08-01

    Trihalomethanes (THMs) are disinfection byproducts found in chlorinated water, and are associated with several different kinds of cancer in human populations and experimental animal models. Metabolism of THMs proceeds through enzymes such as GSTT1 and CYP2E1 and gives rise to reactive intermediates, which form the basis for their toxic activities. The aim of this study was to assess the mitochondrial dysfunction caused by THMs at low levels, and the resulting hepatic histological and biochemical changes in the mouse. Male ICR mice were administered with two THMs: dibromochloromethane (DBCM) and bromodichloromethane (BDCM); once daily, by gavage, to a total of four administrations. Animals were sacrificed four weeks after DBCM and BDCM administrations. Blood biochemistry was performed for alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin (TB), albumin (Alb), total protein (TP), creatinine, and urea. Animals exposed to DBCM and BDCM showed elevated ALT and TB levels (p < 0.05) as compared with controls. Histological analysis confirmed the presence of vacuolar degenerescence and a multifocal necrotizing hepatitis in 33% of animals (n = 2). Mitochondrial analysis showed that THMs reduced mitochondrial bioenergetic activity (succinate dehydrogenase (SQR), cytochrome c oxidase (COX), and ATP synthase) and increased oxidative stress (glutathione S-transferase (GST)) in hepatic tissues (p < 0.05). These results add detail to the current understanding of the mechanisms underlying THM-induced toxicity, supporting the role of mitochondrial dysfunction and oxidative stress in liver toxicity caused by DBCM and BDCM. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1009-1016, 2016. PMID:25640707

  16. Trihalomethanes in liver pathology: Mitochondrial dysfunction and oxidative stress in the mouse.

    PubMed

    Faustino-Rocha, Ana I; Rodrigues, D; da Costa, R Gil; Diniz, C; Aragão, S; Talhada, D; Botelho, M; Colaço, A; Pires, M J; Peixoto, F; Oliveira, P A

    2016-08-01

    Trihalomethanes (THMs) are disinfection byproducts found in chlorinated water, and are associated with several different kinds of cancer in human populations and experimental animal models. Metabolism of THMs proceeds through enzymes such as GSTT1 and CYP2E1 and gives rise to reactive intermediates, which form the basis for their toxic activities. The aim of this study was to assess the mitochondrial dysfunction caused by THMs at low levels, and the resulting hepatic histological and biochemical changes in the mouse. Male ICR mice were administered with two THMs: dibromochloromethane (DBCM) and bromodichloromethane (BDCM); once daily, by gavage, to a total of four administrations. Animals were sacrificed four weeks after DBCM and BDCM administrations. Blood biochemistry was performed for alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin (TB), albumin (Alb), total protein (TP), creatinine, and urea. Animals exposed to DBCM and BDCM showed elevated ALT and TB levels (p < 0.05) as compared with controls. Histological analysis confirmed the presence of vacuolar degenerescence and a multifocal necrotizing hepatitis in 33% of animals (n = 2). Mitochondrial analysis showed that THMs reduced mitochondrial bioenergetic activity (succinate dehydrogenase (SQR), cytochrome c oxidase (COX), and ATP synthase) and increased oxidative stress (glutathione S-transferase (GST)) in hepatic tissues (p < 0.05). These results add detail to the current understanding of the mechanisms underlying THM-induced toxicity, supporting the role of mitochondrial dysfunction and oxidative stress in liver toxicity caused by DBCM and BDCM. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1009-1016, 2016.

  17. Methylene Blue Improves Brain Mitochondrial ABAD Functions and Decreases Aβ in a Neuroinflammatory Alzheimer's Disease Mouse Model.

    PubMed

    Zakaria, Aya; Hamdi, Nabila; Abdel-Kader, Reham Mahmoud

    2016-03-01

    Methylene blue (MB) phase II clinical trials reported improvements in cognitive functions of Alzheimer's disease (AD) patients. Regarding MB mechanism of action, its antioxidant and mitochondrial protective effects have been previously described. In relation to AD, it has been recently reported that MB reduced amyloid beta (Aβ) levels in AD models. The mitochondrial enzyme amyloid-binding alcohol dehydrogenase (ABAD) has been shown to bind Aβ inducing mitochondrial dysfunction, providing a direct relation between Aβ toxicity and mitochondrial dysfunction occurring in AD. Since it has been reported that inhibiting ABAD protects mitochondrial functions and prevents Aβ-induced toxicity, the aim of the current study was to investigate if the protective effects of MB could be associated with an effect on ABAD levels and functions. The current study shows that MB is able to enhance cell viability, reduce both reactive oxygen species levels and importantly Aβ oligomers in a lipopolysaccharide (LPS) mouse model. Interestingly, ABAD levels were increased in the brains of the LPS mouse model and MB treatment was able to reduce its levels. Given that regulation of the estradiol level is a well-established function of ABAD, brain estradiol level was compared in LPS mouse model and in MB-treated mice. The results of the current study show that MB treatment is able to improve significantly the LPS-induced decrease of estradiol levels in mice brains, indicating its ability to modulate both levels and function of ABAD. These results give a new insight to possible mechanisms of MB in AD.

  18. Validation of optical voltage reporting by the genetically encoded voltage indicator VSFP-Butterfly from cortical layer 2/3 pyramidal neurons in mouse brain slices

    PubMed Central

    Empson, Ruth M; Goulton, Chelsea; Scholtz, David; Gallero-Salas, Yasir; Zeng, Hongkui; Knöpfel, Thomas

    2015-01-01

    Understanding how behavior emerges from brain electrical activity is one of the ultimate goals of neuroscience. To achieve this goal we require methods for large-scale recording of the electrical activity of specific neuronal circuits. A very promising approach is to use optical reporting of membrane voltage transients, particularly if the voltage reporter is genetically targeted to specific neuronal populations. Targeting in this way allows population signals to be recorded and interpreted without blindness to neuronal diversity. Here, we evaluated the voltage-sensitive fluorescent protein, VSFP Butterfly 2.1, a genetically encoded voltage indicator (GEVI), for monitoring electrical activity of layer 2/3 cortical pyramidal neurons in mouse brain slices. Standard widefield fluorescence and two-photon imaging revealed robust, high signal-to-noise ratio read-outs of membrane voltage transients that are predominantly synaptic in nature and can be resolved as discrete areas of synaptically connected layer 2/3 neurons. We find that targeted expression of this GEVI in the cortex provides a flexible and promising tool for the analysis of L2/3 cortical network function. PMID:26229003

  19. Multilayered Genetic and Omics Dissection of Mitochondrial Activity in a Mouse Reference Population

    PubMed Central

    Wu, Yibo; Williams, Evan G.; Dubuis, Sébastien; Mottis, Adrienne; Jovaisaite, Virginija; Houten, Sander M.; Argmann, Carmen A.; Faridi, Pouya; Wolski, Witold; Kutalik, Zoltán; Zamboni, Nicola; Auwerx, Johan; Aebersold, Ruedi

    2014-01-01

    SUMMARY The manner by which genotype and environment affect complex phenotypes is one of the fundamental questions in biology. In this study, we quantified the transcriptome—a subset of the metabolome—and, using targeted proteomics, quantified a subset of the liver proteome from 40 strains of the BXD mouse genetic reference population on two diverse diets. We discovered dozens of transcript, protein, and metabolite QTLs, several of which linked to metabolic phenotypes. Most prominently, Dhtkd1 was identified as a primary regulator of 2-aminoadipate, explaining variance in fasted glucose and diabetes status in both mice and humans. These integrated molecular profiles also allowed further characterization of complex pathways, particularly the mitochondrial unfolded protein response (UPRmt). UPRmt shows strikingly variant responses at the transcript and protein level that are remarkably conserved among C. elegans, mice, and humans. Overall, these examples demonstrate the value of an integrated multilayered omics approach to characterize complex metabolic phenotypes. PMID:25215496

  20. Neuroinflammation, mitochondrial defects and neurodegeneration in mucopolysaccharidosis III type C mouse model

    PubMed Central

    Martins, Carla; Hůlková, Helena; Dridi, Larbi; Dormoy-Raclet, Virginie; Grigoryeva, Lubov; Choi, Yoo; Langford-Smith, Alexander; Wilkinson, Fiona L.; Ohmi, Kazuhiro; DiCristo, Graziella; Hamel, Edith; Ausseil, Jerôme; Cheillan, David; Moreau, Alain; Svobodová, Eva; Hájková, Zuzana; Tesařová, Markéta; Hansíková, Hana; Bigger, Brian W.; Hrebícek, Martin

    2015-01-01

    Severe progressive neurological paediatric disease mucopolysaccharidosis III type C is caused by mutations in the HGSNAT gene leading to deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase involved in the lysosomal catabolism of heparan sulphate. To understand the pathophysiology of the disease we generated a mouse model of mucopolysaccharidosis III type C by germline inactivation of the Hgsnat gene. At 6–8 months mice showed hyperactivity, and reduced anxiety. Cognitive memory decline was detected at 10 months and at 12–13 months mice showed signs of unbalanced hesitant walk and urinary retention. Lysosomal accumulation of heparan sulphate was observed in hepatocytes, splenic sinus endothelium, cerebral microglia, liver Kupffer cells, fibroblasts and pericytes. Starting from 5 months, brain neurons showed enlarged, structurally abnormal mitochondria, impaired mitochondrial energy metabolism, and storage of densely packed autofluorescent material, gangliosides, lysozyme, phosphorylated tau, and amyloid-β. Taken together, our data demonstrate for the first time that deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase causes lysosomal accumulation of heparan sulphate in microglial cells followed by their activation and cytokine release. They also show mitochondrial dysfunction in the neurons and neuronal loss explaining why mucopolysaccharidosis III type C manifests primarily as a neurodegenerative disease. PMID:25567323

  1. Myotubularin controls desmin intermediate filament architecture and mitochondrial dynamics in human and mouse skeletal muscle

    PubMed Central

    Hnia, Karim; Tronchère, Helene; Tomczak, Kinga K.; Amoasii, Leonela; Schultz, Patrick; Beggs, Alan H.; Payrastre, Bernard; Mandel, Jean Louis; Laporte, Jocelyn

    2010-01-01

    Muscle contraction relies on a highly organized intracellular network of membrane organelles and cytoskeleton proteins. Among the latter are the intermediate filaments (IFs), a large family of proteins mutated in more than 30 human diseases. For example, mutations in the DES gene, which encodes the IF desmin, lead to desmin-related myopathy and cardiomyopathy. Here, we demonstrate that myotubularin (MTM1), which is mutated in individuals with X-linked centronuclear myopathy (XLCNM; also known as myotubular myopathy), is a desmin-binding protein and provide evidence for direct regulation of desmin by MTM1 in vitro and in vivo. XLCNM-causing mutations in MTM1 disrupted the MTM1-desmin complex, resulting in abnormal IF assembly and architecture in muscle cells and both mouse and human skeletal muscles. Adeno-associated virus–mediated ectopic expression of WT MTM1 in Mtm1-KO muscle reestablished normal desmin expression and localization. In addition, decreased MTM1 expression and XLCNM-causing mutations induced abnormal mitochondrial positioning, shape, dynamics, and function. We therefore conclude that MTM1 is a major regulator of both the desmin cytoskeleton and mitochondria homeostasis, specifically in skeletal muscle. Defects in IF stabilization and mitochondrial dynamics appear as common physiopathological features of centronuclear myopathies and desmin-related myopathies. PMID:21135508

  2. Decline of cell viability and mitochondrial activity in mouse skeletal muscle cell in a hypomagnetic field.

    PubMed

    Fu, Jing-Peng; Mo, Wei-Chuan; Liu, Ying; He, Rong-Qiao

    2016-05-01

    Hypomagnetic field (HMF), one of the key environmental risk factors for astronauts traveling in outer space, has previously been shown to repress locomotion of mammalians. However, underlying mechanisms of how HMF affects the motor system remains poorly understood. In this study, we created an HMF (<3 μT) by eliminating geomagnetic field (GMF, ∼50 μT) and exposed primary mouse skeletal muscle cells to this low magnetic field condition for a period of three days. HMF-exposed cells showed a decline in cell viability relative to GMF control, even though cells appeared normal in terms of morphology and survival rate. After a 3-day HMF-exposure, glucose consumption of skeletal muscle cells was significantly lower than GMF control, accompanied by less adenosine triphosphate (ATP) and adenosine diphosphate (ADP) content and higher ADP/ATP ratio. In agreement with these findings, mitochondrial membrane potential of HMF-exposed cells was also lower, whereas levels of cellular Reactive Oxygen Species were higher. Moreover, viability and membrane potential of isolated mitochondria were reduced after 1 h HMF-exposure in vitro. Our results indicate that mitochondria can directly respond to HMF at functional level, and suggest that HMF-induced decline in cell functionality results from a reduction in energy production and mitochondrial activity. PMID:27003876

  3. Neuroinflammation, mitochondrial defects and neurodegeneration in mucopolysaccharidosis III type C mouse model.

    PubMed

    Martins, Carla; Hůlková, Helena; Dridi, Larbi; Dormoy-Raclet, Virginie; Grigoryeva, Lubov; Choi, Yoo; Langford-Smith, Alexander; Wilkinson, Fiona L; Ohmi, Kazuhiro; DiCristo, Graziella; Hamel, Edith; Ausseil, Jerôme; Cheillan, David; Moreau, Alain; Svobodová, Eva; Hájková, Zuzana; Tesařová, Markéta; Hansíková, Hana; Bigger, Brian W; Hrebícek, Martin; Pshezhetsky, Alexey V

    2015-02-01

    Severe progressive neurological paediatric disease mucopolysaccharidosis III type C is caused by mutations in the HGSNAT gene leading to deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase involved in the lysosomal catabolism of heparan sulphate. To understand the pathophysiology of the disease we generated a mouse model of mucopolysaccharidosis III type C by germline inactivation of the Hgsnat gene. At 6-8 months mice showed hyperactivity, and reduced anxiety. Cognitive memory decline was detected at 10 months and at 12-13 months mice showed signs of unbalanced hesitant walk and urinary retention. Lysosomal accumulation of heparan sulphate was observed in hepatocytes, splenic sinus endothelium, cerebral microglia, liver Kupffer cells, fibroblasts and pericytes. Starting from 5 months, brain neurons showed enlarged, structurally abnormal mitochondria, impaired mitochondrial energy metabolism, and storage of densely packed autofluorescent material, gangliosides, lysozyme, phosphorylated tau, and amyloid-β. Taken together, our data demonstrate for the first time that deficiency of acetyl-CoA: α-glucosaminide N-acetyltransferase causes lysosomal accumulation of heparan sulphate in microglial cells followed by their activation and cytokine release. They also show mitochondrial dysfunction in the neurons and neuronal loss explaining why mucopolysaccharidosis III type C manifests primarily as a neurodegenerative disease. PMID:25567323

  4. Metabolomic analysis of exercise effects in the POLG mitochondrial DNA mutator mouse brain.

    PubMed

    Clark-Matott, Joanne; Saleem, Ayesha; Dai, Ying; Shurubor, Yevgeniya; Ma, Xiaoxing; Safdar, Adeel; Beal, Myron Flint; Tarnopolsky, Mark; Simon, David K

    2015-11-01

    Mitochondrial DNA (mtDNA) mutator mice express a mutated form of mtDNA polymerase gamma that results an accelerated accumulation of somatic mtDNA mutations in association with a premature aging phenotype. An exploratory metabolomic analysis of cortical metabolites in sedentary and exercised mtDNA mutator mice and wild-type littermate controls at 9-10 months of age was performed. Pathway analysis revealed deficits in the neurotransmitters acetylcholine, glutamate, and aspartate that were ameliorated by exercise. Nicotinamide adenine dinucleotide (NAD) depletion and evidence of increased poly(adenosine diphosphate-ribose) polymerase 1 (PARP1)activity were apparent in sedentary mtDNA mutator mouse cortex, along with deficits in carnitine metabolites and an upregulated antioxidant response that largely normalized with exercise. These data highlight specific pathways that are altered in the brain in association with an accelerated age-related accumulation of somatic mtDNA mutations. These results may have relevance to age-related neurodegenerative diseases associated with mitochondrial dysfunction, such as Alzheimer's disease and Parkinson's disease and provide insights into potential mechanisms of beneficial effects of exercise on brain function.

  5. Isoniazid-induced cell death is precipitated by underlying mitochondrial complex I dysfunction in mouse hepatocytes.

    PubMed

    Lee, Kang Kwang; Fujimoto, Kazunori; Zhang, Carmen; Schwall, Christine T; Alder, Nathan N; Pinkert, Carl A; Krueger, Winfried; Rasmussen, Theodore; Boelsterli, Urs A

    2013-12-01

    Isoniazid (INH) is an antituberculosis drug that has been associated with idiosyncratic liver injury in susceptible patients. The underlying mechanisms are still unclear, but there is growing evidence that INH and/or its major metabolite, hydrazine, may interfere with mitochondrial function. However, hepatic mitochondria have a large reserve capacity, and minor disruption of energy homeostasis does not necessarily induce cell death. We explored whether pharmacologic or genetic impairment of mitochondrial complex I may amplify mitochondrial dysfunction and precipitate INH-induced hepatocellular injury. We found that INH (≤ 3000 μM) did not induce cell injury in cultured mouse hepatocytes, although it decreased hepatocellular respiration and ATP levels in a concentration-dependent fashion. However, coexposure of hepatocytes to INH and nontoxic concentrations of the complex I inhibitors rotenone (3 μM) or piericidin A (30 nM) resulted in massive ATP depletion and cell death. Although both rotenone and piericidin A increased MitoSox-reactive fluorescence, Mito-TEMPO or N-acetylcysteine did not attenuate the extent of cytotoxicity. However, preincubation of cells with the acylamidase inhibitor bis-p-nitrophenol phosphate provided protection from hepatocyte injury induced by rotenone/INH (but not rotenone/hydrazine), suggesting that hydrazine was the cell-damaging species. Indeed, we found that hydrazine directly inhibited the activity of solubilized complex II. Hepatocytes isolated from mutant Ndufs4(+/-) mice, although featuring moderately lower protein expression levels of this complex I subunit in liver mitochondria, exhibited unchanged hepatic complex I activity and were therefore not sensitized to INH. These data indicate that underlying inhibition of complex I, which alone is not acutely toxic, can trigger INH-induced hepatocellular injury.

  6. Mouse Cytotoxic T Cell-derived Granzyme B Activates the Mitochondrial Cell Death Pathway in a Bim-dependent Fashion*

    PubMed Central

    Catalán, Elena; Jaime-Sánchez, Paula; Aguiló, Nacho; Simon, Markus M.; Froelich, Christopher J.; Pardo, Julián

    2015-01-01

    Cytotoxic T cells (Tc) use perforin and granzyme B (gzmB) to kill virus-infected cells and cancer cells. Recent evidence suggests that human gzmB primarily induces apoptosis via the intrinsic mitochondrial pathway by either cleaving Bid or activating Bim leading to the activation of Bak/Bax and subsequent generation of active caspase-3. In contrast, mouse gzmB is thought to predominantly induce apoptosis by directly processing pro-caspase-3. However, in certain mouse cell types gzmB-mediated apoptosis mainly occurs via the mitochondrial pathway. To investigate whether Bim is involved under the latter conditions, we have now employed ex vivo virus-immune mouse Tc that selectively kill by using perforin and gzmB (gzmB+Tc) as effector cells and wild type as well as Bim- or Bak/Bax-deficient spontaneously (3T9) or virus-(SV40) transformed mouse embryonic fibroblast cells as targets. We show that gzmB+Tc-mediated apoptosis (phosphatidylserine translocation, mitochondrial depolarization, cytochrome c release, and caspase-3 activation) was severely reduced in 3T9 cells lacking either Bim or both Bak and Bax. This outcome was related to the ability of Tc cells to induce the degradation of Mcl-1 and Bcl-XL, the anti-apoptotic counterparts of Bim. In contrast, gzmB+Tc-mediated apoptosis was not affected in SV40-transformed mouse embryonic fibroblast cells lacking Bak/Bax. The data provide evidence that Bim participates in mouse gzmB+Tc-mediated apoptosis of certain targets by activating the mitochondrial pathway and suggest that the mode of cell death depends on the target cell. Our results suggest that the various molecular events leading to transformation and/or immortalization of cells have an impact on their relative resistance to the multiple gzmB+Tc-induced death pathways. PMID:25605735

  7. The role of voltage-gated potassium channels in the regulation of mouse uterine contractility

    PubMed Central

    Smith, Ryan C; McClure, Marisa C; Smith, Margaret A; Abel, Peter W; Bradley, Michael E

    2007-01-01

    Background Uterine smooth muscle cells exhibit ionic currents that appear to be important in the control of uterine contractility, but how these currents might produce the changes in contractile activity seen in pregnant myometrium has not been established. There are conflicting reports concerning the role of voltage-gated potassium (Kv) channels and large-conductance, calcium-activated potassium (BK) channels in the regulation of uterine contractility. In this study we provide molecular and functional evidence for a role for Kv channels in the regulation of spontaneous contractile activity in mouse myometrium, and also demonstrate a change in Kv channel regulation of contractility in pregnant mouse myometrium. Methods Functional assays which evaluated the effects of channel blockers and various contractile agonists were accomplished by quantifying contractility of isolated uterine smooth muscle obtained from nonpregnant mice as well as mice at various stages of pregnancy. Expression of Kv channel proteins in isolated uterine smooth muscle was evaluated by Western blots. Results The Kv channel blocker 4-aminopyridine (4-AP) caused contractions in nonpregnant mouse myometrium (EC50 = 54 micromolar, maximal effect at 300 micromolar) but this effect disappeared in pregnant mice; similarly, the Kv4.2/Kv4.3 blocker phrixotoxin-2 caused contractions in nonpregnant, but not pregnant, myometrium. Contractile responses to 4-AP were not dependent upon nerves, as neither tetrodotoxin nor storage of tissues at room temperature significantly altered these responses, nor were responses dependent upon the presence of the endometrium. Spontaneous contractions and contractions in response to 4-AP did not appear to be mediated by BK, as the BK channel-selective blockers iberiotoxin, verruculogen, or tetraethylammonium failed to affect either spontaneous contractions or 4-AP-elicited responses. A number of different Kv channel alpha subunit proteins were found in isolated myometrium

  8. Voltage-Dependent Regulation of Complex II Energized Mitochondrial Oxygen Flux

    PubMed Central

    Bai, Fan; Fink, Brian D.; Yu, Liping; Sivitz, William I.

    2016-01-01

    Oxygen consumption by isolated mitochondria is generally measured during state 4 respiration (no ATP production) or state 3 (maximal ATP production at high ADP availability). However, mitochondria in vivo do not function at either extreme. Here we used ADP recycling methodology to assess muscle mitochondrial function over intermediate clamped ADP concentrations. In so doing, we uncovered a previously unrecognized biphasic respiratory pattern wherein O2 flux on the complex II substrate, succinate, initially increased and peaked over low clamped ADP concentrations then decreased markedly at higher clamped concentrations. Mechanistic studies revealed no evidence that the observed changes in O2 flux were due to altered opening or function of the mitochondrial permeability transition pore or to changes in reactive oxygen. Based on metabolite and functional metabolic data, we propose a multifactorial mechanism that consists of coordinate changes that follow from reduced membrane potential (as the ADP concentration in increased). These changes include altered directional electron flow, altered NADH/NAD+ redox cycling, metabolite exit, and OAA inhibition of succinate dehydrogenase. In summary, we report a previously unrecognized pattern for complex II energized O2 flux. Moreover, our findings suggest that the ADP recycling approach might be more widely adapted for mitochondrial studies. PMID:27153112

  9. Radiolabeled Phosphonium Salts as Mitochondrial Voltage Sensors for Positron Emission Tomography Myocardial Imaging Agents.

    PubMed

    Kim, Dong-Yeon; Min, Jung-Joon

    2016-09-01

    Despite substantial advances in the diagnosis of cardiovascular disease, (18)F-labeled positron emission tomography (PET) radiopharmaceuticals remain necessary to diagnose heart disease because clinical use of current PET tracers is limited by their short half-life. Lipophilic cations such as phosphonium salts penetrate the mitochondrial membranes and accumulate in mitochondria of cardiomyocytes in response to negative inner-transmembrane potentials. Radiolabeled tetraphenylphosphonium cation derivatives have been developed as myocardial imaging agents for PET. In this review, a general overview of these radiotracers, including their radiosynthesis, in vivo characterization, and evaluation is provided and clinical perspectives are discussed. PMID:27540422

  10. Respiratory complex I dysfunction due to mitochondrial DNA mutations shifts the voltage threshold for opening of the permeability transition pore toward resting levels.

    PubMed

    Porcelli, Anna Maria; Angelin, Alessia; Ghelli, Anna; Mariani, Elisa; Martinuzzi, Andrea; Carelli, Valerio; Petronilli, Valeria; Bernardi, Paolo; Rugolo, Michela

    2009-01-23

    We have studied mitochondrial bioenergetics in HL180 cells (a cybrid line harboring the T14484C/ND6 and G14279A/ND6 mtDNA mutations of Leber hereditary optic neuropathy, leading to an approximately 50% decrease of ATP synthesis) and XTC.UC1 cells (derived from a thyroid oncocytoma bearing a disruptive frameshift mutation in MT-ND1, which impairs complex I assembly). The addition of rotenone to HL180 cells and of antimycin A to XTC.UC1 cells caused fast mitochondrial membrane depolarization that was prevented by treatment with cyclosporin A, intracellular Ca2+ chelators, and antioxidant. Both cell lines also displayed an anomalous response to oligomycin, with rapid onset of depolarization that was prevented by cyclosporin A and by overexpression of Bcl-2. These findings indicate that depolarization by respiratory chain inhibitors and oligomycin was due to opening of the mitochondrial permeability transition pore (PTP). A shift of the threshold voltage for PTP opening close to the resting potential may therefore be the underlying cause facilitating cell death in diseases affecting complex I activity. This study provides a unifying reading frame for previous observations on mitochondrial dysfunction, bioenergetic defects, and Ca2+ deregulation in mitochondrial diseases. Therapeutic strategies aimed at normalizing the PTP voltage threshold may be instrumental in ameliorating the course of complex I-dependent mitochondrial diseases.

  11. Mitochondrial phenotype of marsupial torpor: Fuel metabolic switch in the Chilean mouse-opossum Thylamys elegans.

    PubMed

    Cortés, Pablo Andres; Bacigalupe, Leonardo Daniel; Mondaca, Fredy; Desrosiers, Véronique; Blier, Pierre U

    2016-01-01

    Torpor is a phenotype characterized by a controlled decline of metabolic rate and body temperature. During arousal from torpor, organs undergo rapid metabolic reactivation and rewarming to near normal levels. As torpor progress, animals show a preference for fatty acids over glucose as primary source of energy. Here, we analyzed for first time the changes in the maximal activity of key enzymes related to fatty acid (Carnitine palmitoyltransferase and β-Hydroxyacyl CoA dehydrogenase) and carbohydrate (Pyruvate kinase, Phosphofructokinase and Lactate dehydrogenase) catabolism, as well as mitochondrial oxidative capacity (Citrate synthase), in six organs of torpid, arousing and euthermic Chilean mouse-opossums (Thylamys elegans). Our results showed that activity of enzymes related to fatty acid and carbohydrate catabolism were different among torpor phases and the pattern of variation differs among tissues. In terms of lipid utilization, maximal enzymatic activities differ in tissues with high oxidative capacity such as heart, kidney, and liver. In terms of carbohydrate use, lower enzymatic activities were observed during torpor in brain and liver. Interestingly, citrate synthase activity did not differ thought torpor-arousal cycle in any tissues analyzed, suggesting no modulation of mitochondrial content in T. elegans. Overall results provide an indication that modulation of enzymes associated with carbohydrate and fatty-acid pathways is mainly oriented to limit energy expensive processes and sustain energy metabolism during transition from torpor to euthermy. Future studies are required to elucidate if physiological events observed for T. elegans are unique from other marsupials, or represents a general response in marsupials. J. Exp. Zool. 325A:41-51, 2016. © 2015 Wiley Periodicals, Inc. PMID:26553608

  12. Reactive nitrogen species in acetaminophen-induced mitochondrial damage and toxicity in mouse hepatocytes.

    PubMed

    Burke, Angela S; MacMillan-Crow, Lee Ann; Hinson, Jack A

    2010-07-19

    Acetaminophen (APAP) toxicity in primary mouse hepatocytes occurs in two phases. The initial phase (0-2 h) occurs with metabolism to N-acetyl-p-benzoquinoneimine which depletes glutathione, and covalently binds to proteins, but little toxicity is observed. Subsequent washing of hepatocytes to remove APAP and reincubating in media alone (2-5 h) results in toxicity. We previously reported that the reincubation phase occurs with mitochondrial permeability transition (MPT) and increased oxidative stress (dichlorodihydrofluorescein fluorescence) (DCFH(2)). Since DCFH(2) may be oxidized by multiple oxidative mechanisms, we investigated the role of reactive nitrogen species (RNS) leading to 3-nitrotyrosine in proteins by ELISA and by immunoblots. Incubation of APAP with hepatocytes for 2 h did not result in toxicity or protein nitration; however, washing hepatocytes and reincubating in media alone (2-5 h) resulted in protein nitration which correlated with toxicity. Inclusion of the MPT inhibitor, cyclosporine A, in the reincubation media eliminated toxicity and protein nitration. The general nitric oxide synthase (NOS) inhibitor L-NMMA and the neuronal NOS (NOS1) inhibitor, 7-nitroindazole, added in the reincubation media decreased toxicity and protein nitration; however, neither the inducible NOS (NOS2) inhibitors L-NIL (N6-(1-iminoethyl)-L-lysine) nor SAIT (S-(2-aminoethyl)isothiourea) decreased protein nitration or toxicity. The RNS scavengers, N-acetylcysteine, and high concentrations of APAP, added in the reincubation phase decreased toxicity and protein nitration. 7-Nitroindazole and cyclosporine A inhibited the APAP-induced loss of mitochondrial membrane potential when added in the reincubation phase. The data indicate a role for RNS in APAP induced toxicity.

  13. Mitochondrial phenotype of marsupial torpor: Fuel metabolic switch in the Chilean mouse-opossum Thylamys elegans.

    PubMed

    Cortés, Pablo Andres; Bacigalupe, Leonardo Daniel; Mondaca, Fredy; Desrosiers, Véronique; Blier, Pierre U

    2016-01-01

    Torpor is a phenotype characterized by a controlled decline of metabolic rate and body temperature. During arousal from torpor, organs undergo rapid metabolic reactivation and rewarming to near normal levels. As torpor progress, animals show a preference for fatty acids over glucose as primary source of energy. Here, we analyzed for first time the changes in the maximal activity of key enzymes related to fatty acid (Carnitine palmitoyltransferase and β-Hydroxyacyl CoA dehydrogenase) and carbohydrate (Pyruvate kinase, Phosphofructokinase and Lactate dehydrogenase) catabolism, as well as mitochondrial oxidative capacity (Citrate synthase), in six organs of torpid, arousing and euthermic Chilean mouse-opossums (Thylamys elegans). Our results showed that activity of enzymes related to fatty acid and carbohydrate catabolism were different among torpor phases and the pattern of variation differs among tissues. In terms of lipid utilization, maximal enzymatic activities differ in tissues with high oxidative capacity such as heart, kidney, and liver. In terms of carbohydrate use, lower enzymatic activities were observed during torpor in brain and liver. Interestingly, citrate synthase activity did not differ thought torpor-arousal cycle in any tissues analyzed, suggesting no modulation of mitochondrial content in T. elegans. Overall results provide an indication that modulation of enzymes associated with carbohydrate and fatty-acid pathways is mainly oriented to limit energy expensive processes and sustain energy metabolism during transition from torpor to euthermy. Future studies are required to elucidate if physiological events observed for T. elegans are unique from other marsupials, or represents a general response in marsupials. J. Exp. Zool. 325A:41-51, 2016. © 2015 Wiley Periodicals, Inc.

  14. Mitochondrial Ca2+ uptake by the voltage-dependent anion channel 2 regulates cardiac rhythmicity

    PubMed Central

    Shimizu, Hirohito; Schredelseker, Johann; Huang, Jie; Lu, Kui; Naghdi, Shamim; Lu, Fei; Franklin, Sarah; Fiji, Hannah DG; Wang, Kevin; Zhu, Huanqi; Tian, Cheng; Lin, Billy; Nakano, Haruko; Ehrlich, Amy; Nakai, Junichi; Stieg, Adam Z; Gimzewski, James K; Nakano, Atsushi; Goldhaber, Joshua I; Vondriska, Thomas M; Hajnóczky, György; Kwon, Ohyun; Chen, Jau-Nian

    2015-01-01

    Tightly regulated Ca2+ homeostasis is a prerequisite for proper cardiac function. To dissect the regulatory network of cardiac Ca2+ handling, we performed a chemical suppressor screen on zebrafish tremblor embryos, which suffer from Ca2+ extrusion defects. Efsevin was identified based on its potent activity to restore coordinated contractions in tremblor. We show that efsevin binds to VDAC2, potentiates mitochondrial Ca2+ uptake and accelerates the transfer of Ca2+ from intracellular stores into mitochondria. In cardiomyocytes, efsevin restricts the temporal and spatial boundaries of Ca2+ sparks and thereby inhibits Ca2+ overload-induced erratic Ca2+ waves and irregular contractions. We further show that overexpression of VDAC2 recapitulates the suppressive effect of efsevin on tremblor embryos whereas VDAC2 deficiency attenuates efsevin's rescue effect and that VDAC2 functions synergistically with MCU to suppress cardiac fibrillation in tremblor. Together, these findings demonstrate a critical modulatory role for VDAC2-dependent mitochondrial Ca2+ uptake in the regulation of cardiac rhythmicity. DOI: http://dx.doi.org/10.7554/eLife.04801.001 PMID:25588501

  15. CGP37157, an inhibitor of the mitochondrial Na+/Ca2+ exchanger, protects neurons from excitotoxicity by blocking voltage-gated Ca2+ channels.

    PubMed

    Ruiz, A; Alberdi, E; Matute, C

    2014-04-10

    Inhibition of the mitochondrial Na(+)/Ca(2+) exchanger (NCLX) by CGP37157 is protective in models of neuronal injury that involve disruption of intracellular Ca(2+) homeostasis. However, the Ca(2+) signaling pathways and stores underlying neuroprotection by that inhibitor are not well defined. In the present study, we analyzed how intracellular Ca(2+) levels are modulated by CGP37157 (10 μM) during NMDA insults in primary cultures of rat cortical neurons. We initially assessed the presence of NCLX in mitochondria of cultured neurons by immunolabeling, and subsequently, we analyzed the effects of CGP37157 on neuronal Ca(2+) homeostasis using cameleon-based mitochondrial Ca(2+) and cytosolic Ca(2+) ([Ca(2+)]i) live imaging. We observed that NCLX-driven mitochondrial Ca(2+) exchange occurs in cortical neurons under basal conditions as CGP37157 induced a decrease in [Ca(2)]i concomitant with a Ca(2+) accumulation inside the mitochondria. In turn, CGP37157 also inhibited mitochondrial Ca(2+) efflux after the stimulation of acetylcholine receptors. In contrast, CGP37157 strongly prevented depolarization-induced [Ca(2+)]i increase by blocking voltage-gated Ca(2+) channels (VGCCs), whereas it did not induce depletion of ER Ca(2+) stores. Moreover, mitochondrial Ca(2+) overload was reduced as a consequence of diminished Ca(2+) entry through VGCCs. The decrease in cytosolic and mitochondrial Ca(2+) overload by CGP37157 resulted in a reduction of excitotoxic mitochondrial damage, characterized here by a reduction in mitochondrial membrane depolarization, oxidative stress and calpain activation. In summary, our results provide evidence that during excitotoxicity CGP37157 modulates cytosolic and mitochondrial Ca(2+) dynamics that leads to attenuation of NMDA-induced mitochondrial dysfunction and neuronal cell death by blocking VGCCs.

  16. Reg2 protects mouse insulinoma cells from streptozotocin-induced mitochondrial disruption and apoptosis.

    PubMed

    Liu, Lu; Liu, Jun-Li; Srikant, Coimbatore B

    2010-10-01

    We reported previously that pancreas-specific ablation of IGF-I in mice induced an increased expression of regenerating family proteins Reg2 and Reg3β in the pancreas and protected them from streptozotocin (Stz)-induced β-cell damage. We, therefore, assessed the effect of ectopically introduced Reg2 on Stz-induced apoptosis in MIN6 mouse insulinoma cells and report here that Reg2 protects MIN6 cells from Stz-induced apoptosis by attenuating its ability to disrupt mitochondrial membrane integrity, activate caspase-3 and promote poly-ADP ribose polymerase cleavage, and induce apoptosis. These changes correlated with suppression of c-jun N-terminal kinase (JNK) phosphorylation by Stz. Reg2 inhibited Stz-induced proapoptotic events as well as the inactivation of JNK. Inclusion of chemical inhibitor of JNK to Reg2 expressing cells rendered them sensitive to Stz. These data demonstrate that Reg2 protects insulin-producing cells against Stz-induced apoptosis by interfering with its cytotoxic signaling upstream of the intrinsic proapoptotic events by preventing its ability to inactivate JNK.

  17. Pathological consequences of long-term mitochondrial oxidative stress in the mouse retinal pigment epithelium.

    PubMed

    Seo, Soo-jung; Krebs, Mark P; Mao, Haoyu; Jones, Kyle; Conners, Mandy; Lewin, Alfred S

    2012-08-01

    Oxidative stress in the retinal pigment epithelium (RPE) is hypothesized to be a major contributor to the development of age-related macular degeneration (AMD). Mitochondrial manganese superoxide dismutase (MnSOD) is a critical antioxidant protein that scavenges the highly reactive superoxide radical. We speculated that specific reduction of MnSOD in the RPE will increase the level of reactive oxygen species in the retina/RPE/choroid complex leading to pathogenesis similar to geographic atrophy. To test this hypothesis, an Sod2-specific hammerhead ribozyme (Rz), delivered by AAV2/1 and driven by the human VMD2 promoter was injected subretinally into C57BL/6J mice. Dark-adapted full field electroretinogram (ERG) detected a decrease in the response to light. We investigated the age-dependent phenotypic and morphological changes of the outer retina using digital fundus imaging and SD-OCT measurement of ONL thickness. Fundus microscopy revealed pigmentary abnormalities in the retina and these corresponded to sub-retinal and sub-RPE deposits seen in SD-OCT B-scans. Light and electron microscopy documented the localization of apical deposits and thickening of the RPE. In RPE flat-mounts we observed abnormally displaced nuclei and regions of apparent fibrosis in the central retina of the oldest mice. This region was surrounded by enlarged and irregular RPE cells that have been observed in eyes donated by AMD patients and in other mouse models of AMD.

  18. 2,2',4,4'-Tetrabromodiphenyl ether injures cell viability and mitochondrial function of mouse spermatocytes by decreasing mitochondrial proteins Atp5b and Uqcrc1.

    PubMed

    Huang, Shaoping; Wang, Jing; Cui, Yiqiang

    2016-09-01

    Our object was to explore direct effects and mechanism of BDE47 on GC2 (immortalized mouse spermatocyte). GC2 were exposed to DMSO, 0.1, 1, 10, 100μM BDE47 for 48h. Cell viability was detected by trypan-blue exclusion; ultrastructure by electron-microscopy; cell cycle, mitochondrial membrane motential (MMP), reactive oxygen species (ROS) by flow-cytometry; ATP production by luminometer; Atp5b, Uqcrc1, Bcl-2 level by WB. To explore whether the decreased mitochondrial proteins play an important role in apoptosis, MMP and apoptosis were detected after Atp5b or Uqcrc1 knockdown in GC2. Results showed BDE47 reduced cell viability, caused condensation of nuclear and vacuolated mitochondria, decreased MMP and ATP, induced ROS, cell cycle arrest at S and G2/M phase, reduced Atp5b, Uqcrc1, Bcl-2 in GC2. Knockdown of Atp5b or Uqcrc1 decreased MMP, induced apoptosis in GC2. Results suggested that BDE47 reduced cell viability, injured mitochondria in spermatocytes probably by decreasing mitochondrial protein Atp5b and Uqcrc1. PMID:27525561

  19. Retinal ganglion cell responses to voltage and current stimulation in wild-type and rd1 mouse retinas

    NASA Astrophysics Data System (ADS)

    Goo, Yong Sook; Ye, Jang Hee; Lee, Seokyoung; Nam, Yoonkey; Ryu, Sang Baek; Kim, Kyung Hwan

    2011-06-01

    Retinal prostheses are being developed to restore vision for those with retinal diseases such as retinitis pigmentosa or age-related macular degeneration. Since neural prostheses depend upon electrical stimulation to control neural activity, optimal stimulation parameters for successful encoding of visual information are one of the most important requirements to enable visual perception. In this paper, we focused on retinal ganglion cell (RGC) responses to different stimulation parameters and compared threshold charge densities in wild-type and rd1 mice. For this purpose, we used in vitro retinal preparations of wild-type and rd1 mice. When the neural network was stimulated with voltage- and current-controlled pulses, RGCs from both wild-type and rd1 mice responded; however the temporal pattern of RGC response is very different. In wild-type RGCs, a single peak within 100 ms appears, while multiple peaks (approximately four peaks) with ~10 Hz rhythm within 400 ms appear in RGCs in the degenerated retina of rd1 mice. We find that an anodic phase-first biphasic voltage-controlled pulse is more efficient for stimulation than a biphasic current-controlled pulse based on lower threshold charge density. The threshold charge densities for activation of RGCs both with voltage- and current-controlled pulses are overall more elevated for the rd1 mouse than the wild-type mouse. Here, we propose the stimulus range for wild-type and rd1 retinas when the optimal modulation of a RGC response is possible.

  20. Overexpression of the mitochondrial methyltransferase TFB1M in the mouse does not impact mitoribosomal methylation status or hearing.

    PubMed

    Lee, Seungmin; Rose, Simon; Metodiev, Metodi D; Becker, Lore; Vernaleken, Alexandra; Klopstock, Thomas; Gailus-Durner, Valerie; Fuchs, Helmut; Hrabě De Angelis, Martin; Douthwaite, Stephen; Larsson, Nils-Göran

    2015-12-20

    Mitochondrial dysfunction is a well-established cause of sensorineural deafness, but the pathophysiological events are poorly understood. Non-syndromic deafness and predisposition to aminoglycoside-induced deafness can be caused by specific mutations in the 12S rRNA gene of mtDNA and are thus maternally inherited traits. The pathophysiology induced by mtDNA mutations has traditionally been attributed to deficient oxidative phosphorylation, which causes energy crisis with functional impairment of multiple cellular processes. In contrast, it was recently reported that signaling induced by 'hypermethylation' of two conserved adenosines of 12S rRNA in the mitoribosome is of key pathophysiological importance in sensorineural deafness. In support for this concept, it was reported that overexpression of the essential mitochondrial methyltransferase TFB1M in the mouse was sufficient to induce mitoribosomal hypermethylation and deafness. At variance with this model, we show here that 12S rRNA is near fully methylated in vivo in the mouse and thus cannot be further methylated to any significant extent. Furthermore, bacterial artificial chromosome transgenic mice overexpressing TFB1M have no increase of 12S rRNA methylation levels and hear normally. We thus conclude that therapies directed against mitoribosomal methylation are unlikely to be beneficial to patients with sensorineural hearing loss or other types of mitochondrial disease.

  1. Mitochondrial base excision repair in mouse synaptosomes during normal aging and in a model of Alzheimer's disease.

    PubMed

    Gredilla, Ricardo; Weissman, Lior; Yang, Jenq-Lin; Bohr, Vilhelm A; Stevnsner, Tinna

    2012-04-01

    Brain aging is associated with synaptic decline and synaptic function is highly dependent on mitochondria. Increased levels of oxidative DNA base damage and accumulation of mitochondrial DNA (mtDNA) mutations or deletions lead to mitochondrial dysfunction, playing an important role in the aging process and the pathogenesis of several neurodegenerative diseases. Here we have investigated the repair of oxidative base damage, in synaptosomes of mouse brain during normal aging and in an AD model. During normal aging, a reduction in the base excision repair (BER) capacity was observed in the synaptosomal fraction, which was associated with a decrease in the level of BER proteins. However, we did not observe changes between the synaptosomal BER activities of presymptomatic and symptomatic AD mice harboring mutated amyolid precursor protein (APP), Tau, and presinilin-1 (PS1) (3xTgAD). Our findings suggest that the age-related reduction in BER capacity in the synaptosomal fraction might contribute to mitochondrial and synaptic dysfunction during aging. The development of AD-like pathology in the 3xTgAD mouse model was, however, not associated with deficiencies of the BER mechanisms in the synaptosomal fraction when the whole brain was analyzed.

  2. Impaired mitochondrial functions contribute to 3-bromopyruvate toxicity in primary rat and mouse hepatocytes.

    PubMed

    Sobotka, Ondřej; Endlicher, René; Drahota, Zdeněk; Kučera, Otto; Rychtrmoc, David; Raad, Marjan; Hakeem, Khurum; Červinková, Zuzana

    2016-08-01

    A compound with promising anticancer properties, 3-bromopyruvate (3-BP) is a synthetic derivative of a pyruvate molecule; however, its toxicity in non-malignant cells has not yet been fully elucidated. Therefore, we elected to study the effects of 3-BP on primary hepatocytes in monolayer cultures, permeabilized hepatocytes and isolated mitochondria. After a 1-h treatment with 100 μM 3-BP cell viability of rat hepatocytes was decreased by 30 % as measured by the WST-1 test (p < 0.001); after 3-h exposure to ≥200 μM 3-BP lactate dehydrogenase leakage was increased (p < 0.001). Reactive oxygen species production was increased in the cell cultures after a 1-h treatment at concentrations ≥100 μmol/l (p < 0.01), and caspase 3 activity was increased after a 20-h incubation with 150 μM and 200 μM 3-BP (p < 0.001). This toxic effect of 3-BP was also proved using primary mouse hepatocytes. In isolated mitochondria, 3-BP induced a dose- and time-dependent decrease of mitochondrial membrane potential during a 10-min incubation both with Complex I substrates glutamate + malate or Complex II substrate succinate, although this decrease was more pronounced with the latter. We also measured the effect of 3-BP on respiration of isolated mitochondria. ADP-activated respiration was inhibited by 20 μM 3-BP within 10 min. Similar effects were also found in permeabilized hepatocytes of both species. PMID:27530389

  3. α-MHC MitoTimer mouse: In vivo mitochondrial turnover model reveals remarkable mitochondrial heterogeneity in the heart.

    PubMed

    Stotland, Aleksandr; Gottlieb, Roberta A

    2016-01-01

    In order to maintain an efficient, energy-producing network in the heart, dysfunctional mitochondria are cleared through the mechanism of autophagy, which is closely linked with mitochondrial biogenesis; these, together with fusion and fission comprise a crucial process known as mitochondrial turnover. Until recently, the lack of molecular tools and methods available to researchers has impeded in vivo investigations of turnover. To investigate the process at the level of a single mitochondrion, our laboratory has developed the MitoTimer protein. Timer is a mutant of DsRed fluorescent protein characterized by transition from green fluorescence to a more stable red conformation over 48 h, and its rate of maturation is stable under physiological conditions. We fused the Timer cDNA with the inner mitochondrial membrane signal sequence and placed it under the control of a cardiac-restricted promoter. This construct was used to create the alpha-MHC-MitoTimer mice. Surprisingly, initial analysis of the hearts from these mice demonstrated a high degree of heterogeneity in the ratio of red-to-green fluorescence of MitoTimer in cardiac tissue. Further, scattered solitary mitochondria within cardiomyocytes display a much higher red-to-green fluorescence (red-shifted) relative to other mitochondria in the cell, implying a block in import of newly synthesized MitoTimer likely due to lower membrane potential. These red-shifted mitochondria may represent older, senescent mitochondria. Concurrently, the cardiomyocytes also contain a subpopulation of mitochondria that display a lower red-to-green fluorescence (green-shifted) relative to other mitochondria, indicative of germinal mitochondria that are actively engaged in import of newly-synthesized mito-targeted proteins. These mitochondria can be isolated and sorted from the heart by flow cytometry for further analysis. Initial studies suggest that these mice represent an elegant tool for the investigation of mitochondrial turnover

  4. Methionine sulfoxide reductase A affects β-amyloid solubility and mitochondrial function in a mouse model of Alzheimer's disease.

    PubMed

    Moskovitz, Jackob; Du, Fang; Bowman, Connor F; Yan, Shirley S

    2016-03-15

    Accumulation of oxidized proteins, and especially β-amyloid (Aβ), is thought to be one of the common causes of Alzheimer's disease (AD). The current studies determine the effect of an in vivo methionine sulfoxidation of Aβ through ablation of the methionine sulfoxide reductase A (MsrA) in a mouse model of AD, a mouse that overexpresses amyloid precursor protein (APP) and Aβ in neurons. Lack of MsrA fosters the formation of methionine sulfoxide in proteins, and thus its ablation in the AD-mouse model will increase the formation of methionine sulfoxide in Aβ. Indeed, the novel MsrA-deficient APP mice (APP(+)/MsrAKO) exhibited higher levels of soluble Aβ in brain compared with APP(+) mice. Furthermore, mitochondrial respiration and the activity of cytochrome c oxidase were compromised in the APP(+)/MsrAKO compared with control mice. These results suggest that lower MsrA activity modifies Aβ solubility properties and causes mitochondrial dysfunction, and augmenting its activity may be beneficial in delaying AD progression.

  5. Prevention and reversal of severe mitochondrial cardiomyopathy by gene therapy in a mouse model of Friedreich's ataxia.

    PubMed

    Perdomini, Morgane; Belbellaa, Brahim; Monassier, Laurent; Reutenauer, Laurence; Messaddeq, Nadia; Cartier, Nathalie; Crystal, Ronald G; Aubourg, Patrick; Puccio, Hélène

    2014-05-01

    Cardiac failure is the most common cause of mortality in Friedreich's ataxia (FRDA), a mitochondrial disease characterized by neurodegeneration, hypertrophic cardiomyopathy and diabetes. FRDA is caused by reduced levels of frataxin (FXN), an essential mitochondrial protein involved in the biosynthesis of iron-sulfur (Fe-S) clusters. Impaired mitochondrial oxidative phosphorylation, bioenergetics imbalance, deficit of Fe-S cluster enzymes and mitochondrial iron overload occur in the myocardium of individuals with FRDA. No treatment exists as yet for FRDA cardiomyopathy. A conditional mouse model with complete frataxin deletion in cardiac and skeletal muscle (Mck-Cre-Fxn(L3/L-) mice) recapitulates most features of FRDA cardiomyopathy, albeit with a more rapid and severe course. Here we show that adeno-associated virus rh10 vector expressing human FXN injected intravenously in these mice fully prevented the onset of cardiac disease. Moreover, later administration of the frataxin-expressing vector, after the onset of heart failure, was able to completely reverse the cardiomyopathy of these mice at the functional, cellular and molecular levels within a few days. Our results demonstrate that cardiomyocytes with severe energy failure and ultrastructure disorganization can be rapidly rescued and remodeled by gene therapy and establish the preclinical proof of concept for the potential of gene therapy in treating FRDA cardiomyopathy.

  6. Mitochondrial dysfunction in an Opa1Q285STOP mouse model of dominant optic atrophy results from Opa1 haploinsufficiency

    PubMed Central

    Kushnareva, Y; Seong, Y; Andreyev, A Y; Kuwana, T; Kiosses, W B; Votruba, M; Newmeyer, D D

    2016-01-01

    Mutations in the opa1 (optic atrophy 1) gene lead to autosomal dominant optic atrophy (ADOA), a hereditary eye disease. This gene encodes the Opa1 protein, a mitochondrial dynamin-related GTPase required for mitochondrial fusion and the maintenance of normal crista structure. The majority of opa1 mutations encode truncated forms of the protein, lacking a complete GTPase domain. It is unclear whether the phenotype results from haploinsufficiency or rather a deleterious effect of truncated Opa1 protein. We studied a heterozygous Opa1 mutant mouse carrying a defective allele with a stop codon in the beginning of the GTPase domain at residue 285, a mutation that mimics human pathological mutations. Using an antibody raised against an N-terminal portion of Opa1, we found that the level of wild-type protein was decreased in the mutant mice, as predicted. However, no truncated Opa1 protein was expressed. In embryonic fibroblasts isolated from the mutant mice, this partial loss of Opa1 caused mitochondrial respiratory deficiency and a selective loss of respiratory Complex IV subunits. Furthermore, partial Opa1 deficiency resulted in a substantial resistance to endoplasmic reticulum stress-induced death. On the other hand, the enforced expression of truncated Opa1 protein in cells containing normal levels of wild-type protein did not cause mitochondrial defects. Moreover, cells expressing the truncated Opa1 protein showed reduced Bax activation in response to apoptotic stimuli. Taken together, our results exclude deleterious dominant-negative or gain-of-function mechanisms for this type of Opa1 mutation and affirm haploinsufficiency as the mechanism underlying mitochondrial dysfunction in ADOA. PMID:27468686

  7. Mitochondrial Bioenergetic Alterations in Mouse Neuroblastoma Cells Infected with Sindbis Virus: Implications to Viral Replication and Neuronal Death

    PubMed Central

    Silva da Costa, Leandro; Pereira da Silva, Ana Paula; Da Poian, Andrea T.; El-Bacha, Tatiana

    2012-01-01

    The metabolic resources crucial for viral replication are provided by the host. Details of the mechanisms by which viruses interact with host metabolism, altering and recruiting high free-energy molecules for their own replication, remain unknown. Sindbis virus, the prototype of and most widespread alphavirus, causes outbreaks of arthritis in humans and serves as a model for the study of the pathogenesis of neurological diseases induced by alphaviruses in mice. In this work, respirometric analysis was used to evaluate the effects of Sindbis virus infection on mitochondrial bioenergetics of a mouse neuroblastoma cell lineage, Neuro 2a. The modulation of mitochondrial functions affected cellular ATP content and this was synchronous with Sindbis virus replication cycle and cell death. At 15 h, irrespective of effects on cell viability, viral replication induced a decrease in oxygen consumption uncoupled to ATP synthesis and a 36% decrease in maximum uncoupled respiration, which led to an increase of 30% in the fraction of oxygen consumption used for ATP synthesis. Decreased proton leak associated to complex I respiration contributed to the apparent improvement of mitochondrial function. Cellular ATP content was not affected by infection. After 24 h, mitochondria dysfunction was clearly observed as maximum uncoupled respiration reduced 65%, along with a decrease in the fraction of oxygen consumption used for ATP synthesis. Suppressed respiration driven by complexes I- and II-related substrates seemed to play a role in mitochondrial dysfunction. Despite the increase in glucose uptake and glycolytic flux, these changes were followed by a 30% decrease in ATP content and neuronal death. Taken together, mitochondrial bioenergetics is modulated during Sindbis virus infection in such a way as to favor ATP synthesis required to support active viral replication. These early changes in metabolism of Neuro 2a cells may form the molecular basis of neuronal dysfunction and Sindbis

  8. Tubulin tail sequences and post-translational modifications regulate closure of mitochondrial voltage-dependent anion channel (VDAC).

    PubMed

    Sheldon, Kely L; Gurnev, Philip A; Bezrukov, Sergey M; Sackett, Dan L

    2015-10-30

    It was previously shown that tubulin dimer interaction with the mitochondrial outer membrane protein voltage-dependent anion channel (VDAC) blocks traffic through the channel and reduces oxidative metabolism and that this requires the unstructured anionic C-terminal tail peptides found on both α- and β-tubulin subunits. It was unclear whether the α- and β-tubulin tails contribute equally to VDAC blockade and what effects might be due to sequence variations in these tail peptides or to tubulin post-translational modifications, which mostly occur on the tails. The nature of the contribution of the tubulin body beyond acting as an anchor for the tails had not been clarified either. Here we present peptide-protein chimeras to address these questions. These constructs allow us to easily combine a tail peptide with different proteins or combine different tail peptides with a particular protein. The results show that a single tail grafted to an inert protein is sufficient to produce channel closure similar to that observed with tubulin. We show that the β-tail is more than an order of magnitude more potent than the α-tail and that the lower α-tail activity is largely due to the presence of a terminal tyrosine. Detyrosination activates the α-tail, and activation is reversed by the removal of the glutamic acid penultimate to the tyrosine. Nitration of tyrosine reverses the tyrosine inhibition of binding and even induces prolonged VDAC closures. Our results demonstrate that small changes in sequence or post-translational modification of the unstructured tails of tubulin result in substantial changes in VDAC closure.

  9. Tubulin tail sequences and post-translational modifications regulate closure of mitochondrial voltage-dependent anion channel (VDAC).

    PubMed

    Sheldon, Kely L; Gurnev, Philip A; Bezrukov, Sergey M; Sackett, Dan L

    2015-10-30

    It was previously shown that tubulin dimer interaction with the mitochondrial outer membrane protein voltage-dependent anion channel (VDAC) blocks traffic through the channel and reduces oxidative metabolism and that this requires the unstructured anionic C-terminal tail peptides found on both α- and β-tubulin subunits. It was unclear whether the α- and β-tubulin tails contribute equally to VDAC blockade and what effects might be due to sequence variations in these tail peptides or to tubulin post-translational modifications, which mostly occur on the tails. The nature of the contribution of the tubulin body beyond acting as an anchor for the tails had not been clarified either. Here we present peptide-protein chimeras to address these questions. These constructs allow us to easily combine a tail peptide with different proteins or combine different tail peptides with a particular protein. The results show that a single tail grafted to an inert protein is sufficient to produce channel closure similar to that observed with tubulin. We show that the β-tail is more than an order of magnitude more potent than the α-tail and that the lower α-tail activity is largely due to the presence of a terminal tyrosine. Detyrosination activates the α-tail, and activation is reversed by the removal of the glutamic acid penultimate to the tyrosine. Nitration of tyrosine reverses the tyrosine inhibition of binding and even induces prolonged VDAC closures. Our results demonstrate that small changes in sequence or post-translational modification of the unstructured tails of tubulin result in substantial changes in VDAC closure. PMID:26306046

  10. Mitochondrial dysfunction and decrease in body weight of a transgenic knock-in mouse model for TDP-43.

    PubMed

    Stribl, Carola; Samara, Aladin; Trümbach, Dietrich; Peis, Regina; Neumann, Manuela; Fuchs, Helmut; Gailus-Durner, Valerie; Hrabě de Angelis, Martin; Rathkolb, Birgit; Wolf, Eckhard; Beckers, Johannes; Horsch, Marion; Neff, Frauke; Kremmer, Elisabeth; Koob, Sebastian; Reichert, Andreas S; Hans, Wolfgang; Rozman, Jan; Klingenspor, Martin; Aichler, Michaela; Walch, Axel Karl; Becker, Lore; Klopstock, Thomas; Glasl, Lisa; Hölter, Sabine M; Wurst, Wolfgang; Floss, Thomas

    2014-04-11

    The majority of amyotrophic lateral sclerosis (ALS) cases as well as many patients suffering from frontotemporal lobar dementia (FTLD) with ubiquitinated inclusion bodies show TDP-43 pathology, the protein encoded by the TAR DNA-binding protein (Tardbp) gene. We used recombinase-mediated cassette exchange to introduce an ALS patient cDNA into the mouse Tdp-43 locus. Expression levels of human A315T TDP-43 protein were 300% elevated in heterozygotes, whereas the endogenous mouse Tdp-43 was decreased to 20% of wild type levels as a result of disturbed feedback regulation. Heterozygous TDP-43(A315TKi) mutants lost 10% of their body weight and developed insoluble TDP-43 protein starting as early as 3 months after birth, a pathology that was exacerbated with age. We analyzed the splicing patterns of known Tdp-43 target genes as well as genome-wide gene expression levels in different tissues that indicated mitochondrial dysfunction. In heterozygous mutant animals, we observed a relative decrease in expression of Parkin (Park2) and the fatty acid transporter CD36 along with an increase in fatty acids, HDL cholesterol, and glucose in the blood. As seen in transmission electron microscopy, neuronal cells in motor cortices of TDP-43(A315TKi) animals had abnormal neuronal mitochondrial cristae formation. Motor neurons were reduced to 90%, but only slight motoric impairment was detected. The observed phenotype was interpreted as a predisease model, which might be valuable for the identification of further environmental or genetic triggers of neurodegeneration.

  11. Mitochondrial Dysfunction and Decrease in Body Weight of a Transgenic Knock-in Mouse Model for TDP-43*

    PubMed Central

    Stribl, Carola; Samara, Aladin; Trümbach, Dietrich; Peis, Regina; Neumann, Manuela; Fuchs, Helmut; Gailus-Durner, Valerie; Hrabě de Angelis, Martin; Rathkolb, Birgit; Wolf, Eckhard; Beckers, Johannes; Horsch, Marion; Neff, Frauke; Kremmer, Elisabeth; Koob, Sebastian; Reichert, Andreas S.; Hans, Wolfgang; Rozman, Jan; Klingenspor, Martin; Aichler, Michaela; Walch, Axel Karl; Becker, Lore; Klopstock, Thomas; Glasl, Lisa; Hölter, Sabine M.; Wurst, Wolfgang; Floss, Thomas

    2014-01-01

    The majority of amyotrophic lateral sclerosis (ALS) cases as well as many patients suffering from frontotemporal lobar dementia (FTLD) with ubiquitinated inclusion bodies show TDP-43 pathology, the protein encoded by the TAR DNA-binding protein (Tardbp) gene. We used recombinase-mediated cassette exchange to introduce an ALS patient cDNA into the mouse Tdp-43 locus. Expression levels of human A315T TDP-43 protein were 300% elevated in heterozygotes, whereas the endogenous mouse Tdp-43 was decreased to 20% of wild type levels as a result of disturbed feedback regulation. Heterozygous TDP-43A315TKi mutants lost 10% of their body weight and developed insoluble TDP-43 protein starting as early as 3 months after birth, a pathology that was exacerbated with age. We analyzed the splicing patterns of known Tdp-43 target genes as well as genome-wide gene expression levels in different tissues that indicated mitochondrial dysfunction. In heterozygous mutant animals, we observed a relative decrease in expression of Parkin (Park2) and the fatty acid transporter CD36 along with an increase in fatty acids, HDL cholesterol, and glucose in the blood. As seen in transmission electron microscopy, neuronal cells in motor cortices of TDP-43A315TKi animals had abnormal neuronal mitochondrial cristae formation. Motor neurons were reduced to 90%, but only slight motoric impairment was detected. The observed phenotype was interpreted as a predisease model, which might be valuable for the identification of further environmental or genetic triggers of neurodegeneration. PMID:24515116

  12. Increased neuronal PreP activity reduces Aβ accumulation, attenuates neuroinflammation and improves mitochondrial and synaptic function in Alzheimer disease's mouse model.

    PubMed

    Fang, Du; Wang, Yongfu; Zhang, Zhihua; Du, Heng; Yan, Shiqiang; Sun, Qinru; Zhong, Changjia; Wu, Long; Vangavaragu, Jhansi Rani; Yan, Shijun; Hu, Gang; Guo, Lan; Rabinowitz, Molly; Glaser, Elzbieta; Arancio, Ottavio; Sosunov, Alexander A; McKhann, Guy M; Chen, John Xi; Yan, Shirley ShiDu

    2015-09-15

    Accumulation of amyloid-β (Aβ) in synaptic mitochondria is associated with mitochondrial and synaptic injury. The underlying mechanisms and strategies to eliminate Aβ and rescue mitochondrial and synaptic defects remain elusive. Presequence protease (PreP), a mitochondrial peptidasome, is a novel mitochondrial Aβ degrading enzyme. Here, we demonstrate for the first time that increased expression of active human PreP in cortical neurons attenuates Alzheimer disease's (AD)-like mitochondrial amyloid pathology and synaptic mitochondrial dysfunction, and suppresses mitochondrial oxidative stress. Notably, PreP-overexpressed AD mice show significant reduction in the production of proinflammatory mediators. Accordingly, increased neuronal PreP expression improves learning and memory and synaptic function in vivo AD mice, and alleviates Aβ-mediated reduction of long-term potentiation (LTP). Our results provide in vivo evidence that PreP may play an important role in maintaining mitochondrial integrity and function by clearance and degradation of mitochondrial Aβ along with the improvement in synaptic and behavioral function in AD mouse model. Thus, enhancing PreP activity/expression may be a new therapeutic avenue for treatment of AD.

  13. Mitochondrial free radical overproduction due to respiratory chain impairment in the brain of a mouse model of Rett syndrome: protective effect of CNF1.

    PubMed

    De Filippis, Bianca; Valenti, Daniela; de Bari, Lidia; De Rasmo, Domenico; Musto, Mattia; Fabbri, Alessia; Ricceri, Laura; Fiorentini, Carla; Laviola, Giovanni; Vacca, Rosa Anna

    2015-06-01

    Rett syndrome (RTT) is a pervasive neurodevelopmental disorder mainly caused by mutations in the X-linked MECP2 gene associated with severe intellectual disability, movement disorders, and autistic-like behaviors. Its pathogenesis remains mostly not understood and no effective therapy is available. High circulating levels of oxidative stress markers in patients and the occurrence of oxidative brain damage in MeCP2-deficient mouse models suggest the involvement of oxidative stress in RTT pathogenesis. However, the molecular mechanism and the origin of the oxidative stress have not been elucidated. Here we demonstrate that a redox imbalance arises from aberrant mitochondrial functionality in the brain of MeCP2-308 heterozygous female mice, a condition that more closely recapitulates that of RTT patients. The marked increase in the rate of hydrogen peroxide generation in the brain of RTT mice seems mainly produced by the dysfunctional complex II of the mitochondrial respiratory chain. In addition, both membrane potential generation and mitochondrial ATP synthesis are decreased in RTT mouse brains when succinate, the complex II respiratory substrate, is used as an energy source. Respiratory chain impairment is brain area specific, owing to a decrease in either cAMP-dependent phosphorylation or protein levels of specific complex subunits. Further, we investigated whether the treatment of RTT mice with the bacterial protein CNF1, previously reported to ameliorate the neurobehavioral phenotype and brain bioenergetic markers in an RTT mouse model, exerts specific effects on brain mitochondrial function and consequently on hydrogen peroxide production. In RTT brains treated with CNF1, we observed the reactivation of respiratory chain complexes, the rescue of mitochondrial functionality, and the prevention of brain hydrogen peroxide overproduction. These results provide definitive evidence of mitochondrial reactive oxygen species overproduction in RTT mouse brain and

  14. A Phenotype-Driven Approach to Generate Mouse Models with Pathogenic mtDNA Mutations Causing Mitochondrial Disease.

    PubMed

    Kauppila, Johanna H K; Baines, Holly L; Bratic, Ana; Simard, Marie-Lune; Freyer, Christoph; Mourier, Arnaud; Stamp, Craig; Filograna, Roberta; Larsson, Nils-Göran; Greaves, Laura C; Stewart, James B

    2016-09-13

    Mutations of mtDNA are an important cause of human disease, but few animal models exist. Because mammalian mitochondria cannot be transfected, the development of mice with pathogenic mtDNA mutations has been challenging, and the main strategy has therefore been to introduce mutations found in cell lines into mouse embryos. Here, we describe a phenotype-driven strategy that is based on detecting clonal expansion of pathogenic mtDNA mutations in colonic crypts of founder mice derived from heterozygous mtDNA mutator mice. As proof of concept, we report the generation of a mouse line transmitting a heteroplasmic pathogenic mutation in the alanine tRNA gene of mtDNA displaying typical characteristics of classic mitochondrial disease. In summary, we describe a straightforward and technically simple strategy based on mouse breeding and histology to generate animal models of mtDNA-mutation disease, which will be of great importance for studies of disease pathophysiology and preclinical treatment trials. PMID:27626666

  15. Aging Exacerbates Pressure-Induced Mitochondrial Oxidative Stress in Mouse Cerebral Arteries.

    PubMed

    Springo, Zsolt; Tarantini, Stefano; Toth, Peter; Tucsek, Zsuzsanna; Koller, Akos; Sonntag, William E; Csiszar, Anna; Ungvari, Zoltan

    2015-11-01

    Epidemiological studies demonstrate that in addition to the increased prevalence of hypertension in old patients, the deleterious cerebrovascular effects of hypertension (including atherosclerosis, stroke, and vascular cognitive impairment) are also exacerbated in elderly individuals. The cellular mechanisms by which aging and hypertension interact to promote cerebrovascular pathologies are not well understood. To test the hypothesis that aging exacerbates high pressure-induced mitochondrial oxidative stress, we exposed isolated segments of the middle cerebral arteries of young (3 months) and aged (24 months) C57BL/6 mice to 60 or 140 mmHg intraluminal pressure and assessed changes in mitochondrial reactive oxygen species production using a mitochondria-targeted redox-sensitive fluorescent indicator dye (MitoSox) by confocal microscopy. Perinuclear MitoSox fluorescence was significantly stronger in high pressure-exposed middle cerebral arteries compared with middle cerebral arteries of the same animals exposed to 60 mmHg, indicating that high pressure increases mitochondrial reactive oxygen species production in the smooth muscle cells of cerebral arteries. Comparison of young and aged middle cerebral arteries showed that aging exacerbates high pressure-induced mitochondrial reactive oxygen species production in cerebral arteries. We propose that increased mechanosensitive mitochondrial oxidative stress may potentially exacerbate cerebrovascular injury and vascular inflammation in aging.

  16. Differential age-related changes in mitochondrial DNA repair activities in mouse brain regions

    PubMed Central

    Gredilla, Ricardo; Garm, Christian; Holm, Rikke; Bohr, Vilhelm A.; Stevnsner, Tinna

    2008-01-01

    Aging in the brain is characterized by increased susceptibility to neuronal loss and functional decline, and mitochondrial DNA (mtDNA) mutations are thought to play an important role in these processes. Due to the proximity of mtDNA to the main sites of mitochondrial free radical generation, oxidative stress is a major source of DNA mutations in mitochondria. The base excision repair (BER) pathway removes oxidative lesions from mtDNA, thereby constituting an important mechanism to avoid accumulation of mtDNA mutations. The complexity of the brain implies that exposure and defence against oxidative stress varies among brain regions and hence some regions may be particularly prone to accumulation of mtDNA damages. In the current study we investigated the efficiency of the BER pathway throughout the murine lifespan in mitochondria from cortex and hippocampus, regions that are central in mammalian cognition, and which are severely affected during aging and in neurodegenerative diseases. A regional specific regulation of mitochondrial DNA repair activities was observed with aging. In cortical mitochondria, DNA glycosylase activities peaked at middle-age followed by a significant drop at old age. However, only minor changes were observed in hippocampal mitochondria during the whole lifespan of the animals. Furthermore, DNA glycosylase activities were lower in hippocampal than in cortical mitochondria. Mitochondrial AP endonuclease activity increased in old animals in both brain regions. Our data suggest an important regional specific regulation of mitochondrial BER during aging. PMID:18701195

  17. Development of synaptic networks in the mouse vagal pathway revealed by optical mapping with a voltage-sensitive dye.

    PubMed

    Momose-Sato, Yoko; Sato, Katsushige

    2016-07-01

    The central issue in developmental neuroscience is when and how neural synaptic networks are established and become functional within the central nervous system (CNS). Investigations of the neural network organization have been hampered because conventional electrophysiological means have some technical limitations. In this study, the multiple-site optical recording technique with a voltage-sensitive dye was employed to survey the developmental organization of the vagal system in the mouse embryo. Stimulation of the vagus nerve in E11-E14 mouse embryos elicited optical responses in areas corresponding to the vagal sensory and motor nuclei. Postsynaptic responses in the first-order sensory nucleus, the nucleus of the tractus solitarius (NTS), were identified from E11, suggesting that sensory information becomes transferred to the brain at this stage. In addition to the NTS, optical responses were identified in the rostral and contralateral brainstem regions, which corresponded to second/higher order nuclei of the vagus nerve including the parabrachial nucleus (PBN). Postsynaptic responses in the second/higher-order nuclei were detected from E12, suggesting that polysynaptic networks were functional at this stage. We discuss the results of our optical mapping, comparing them with previous findings obtained in the chick and rat embryos, and suggest some fundamental principles in the functional organization of synaptic networks in the embryonic brain. PMID:27207499

  18. Mechanisms of stress resistance in Snell dwarf mouse fibroblasts: enhanced antioxidant and DNA base excision repair capacity, but no differences in mitochondrial metabolism.

    PubMed

    Page, Melissa M; Salmon, Adam B; Leiser, Scott F; Robb, Ellen L; Brown, Melanie F; Miller, Richard A; Stuart, Jeffrey A

    2009-04-15

    Dermal fibroblasts from long-lived Snell dwarf mice can withstand a variety of oxidative and non-oxidative stressors compared to normal littermate controls. Here, we report differences in the levels and activities of intracellular antioxidant and DNA repair enzymes between normal and Snell dwarf mice fibroblasts cultured under a variety of conditions, including: 3% and 20% ambient O(2); the presence and absence of serum; and the addition of an exogenous oxidative stress. The only significant difference between normal and dwarf cells cultured in complete medium, at 20% O(2), was an approximately 40% elevation of glutathione peroxidase (GPx) activity in the mutant cells. Serum deprivation elicited increases in GPx in both genotypes, but these activities remained higher in dwarf mouse cells. Dwarf mouse cells deprived of serum and challenged with exposure to paraquat or hydrogen peroxide showed a generally greater upregulation of catalase and DNA base excision repair enzymes. As these toxins can interact with mitochondria to increase mitochondrial ROS production, we explored whether there were differences in mitochondrial metabolism between normal and dwarf mouse cells. However, neither mitochondrial content nor the apparent mitochondrial membrane potential differed between genotypes. Overall, the results suggest that superior hydrogen peroxide metabolism and a marginally greater DNA base excision repair capacity contribute to the stress resistance phenotype of Snell dwarf mouse fibroblasts.

  19. Mechanisms of stress resistance in Snell dwarf mouse fibroblasts: enhanced antioxidant and DNA base excision repair capacity, but no differences in mitochondrial metabolism.

    PubMed

    Page, Melissa M; Salmon, Adam B; Leiser, Scott F; Robb, Ellen L; Brown, Melanie F; Miller, Richard A; Stuart, Jeffrey A

    2009-04-15

    Dermal fibroblasts from long-lived Snell dwarf mice can withstand a variety of oxidative and non-oxidative stressors compared to normal littermate controls. Here, we report differences in the levels and activities of intracellular antioxidant and DNA repair enzymes between normal and Snell dwarf mice fibroblasts cultured under a variety of conditions, including: 3% and 20% ambient O(2); the presence and absence of serum; and the addition of an exogenous oxidative stress. The only significant difference between normal and dwarf cells cultured in complete medium, at 20% O(2), was an approximately 40% elevation of glutathione peroxidase (GPx) activity in the mutant cells. Serum deprivation elicited increases in GPx in both genotypes, but these activities remained higher in dwarf mouse cells. Dwarf mouse cells deprived of serum and challenged with exposure to paraquat or hydrogen peroxide showed a generally greater upregulation of catalase and DNA base excision repair enzymes. As these toxins can interact with mitochondria to increase mitochondrial ROS production, we explored whether there were differences in mitochondrial metabolism between normal and dwarf mouse cells. However, neither mitochondrial content nor the apparent mitochondrial membrane potential differed between genotypes. Overall, the results suggest that superior hydrogen peroxide metabolism and a marginally greater DNA base excision repair capacity contribute to the stress resistance phenotype of Snell dwarf mouse fibroblasts. PMID:19439226

  20. Two deeply divergent mitochondrial clades in the wild mouse Mus macedonicus reveal multiple glacial refuges south of Caucasus.

    PubMed

    Orth, A; Auffray, J-C; Bonhomme, F

    2002-11-01

    A survey of 77 individuals covering the range of Mus macedonicus from Georgia in the East to Greece and Bulgaria in the West and Israel in the South has shown the existence of two deeply divergent mitochondrial clades. The southern clade was until now undetected and characterises mice from Israel. Nuclear genes also show some amount of regional differentiation tending to separate the southern M. macedonicus from the northern ones. These results point towards the fact that the eastern Mediterranean short-tailed mouse, which was seen as a fairly homogeneous monotypic species, has in fact a more complex phylogeographic history than has been suspected, and that it warrants the existence of two subspecies. The reasons for this non-uniformity probably ought to be looked for in the history of faunal movements linked to glacial periods, underlining the possible existence of at least two refugia south of the Caucasus.

  1. Alternative Oxidase Expression in the Mouse Enables Bypassing Cytochrome c Oxidase Blockade and Limits Mitochondrial ROS Overproduction

    PubMed Central

    El-Khoury, Riyad; Dufour, Eric; Rak, Malgorzata; Ramanantsoa, Nelina; Grandchamp, Nicolas; Csaba, Zsolt; Duvillié, Bertrand; Bénit, Paule; Gallego, Jorge; Gressens, Pierre; Sarkis, Chamsy; Jacobs, Howard T.; Rustin, Pierre

    2013-01-01

    Cyanide-resistant non-phosphorylating respiration is known in mitochondria from plants, fungi, and microorganisms but is absent in mammals. It results from the activity of an alternative oxidase (AOX) that conveys electrons directly from the respiratory chain (RC) ubiquinol pool to oxygen. AOX thus provides a bypath that releases constraints on the cytochrome pathway and prevents the over-reduction of the ubiquinone pool, a major source of superoxide. RC dysfunctions and deleterious superoxide overproduction are recurrent themes in human pathologies, ranging from neurodegenerative diseases to cancer, and may be instrumental in ageing. Thus, preventing RC blockade and excess superoxide production by means of AOX should be of considerable interest. However, because of its energy-dissipating properties, AOX might produce deleterious effects of its own in mammals. Here we show that AOX can be safely expressed in the mouse (MitAOX), with major physiological parameters being unaffected. It neither disrupted the activity of other RC components nor decreased oxidative phosphorylation in isolated mitochondria. It conferred cyanide-resistance to mitochondrial substrate oxidation and decreased reactive oxygen species (ROS) production upon RC blockade. Accordingly, AOX expression was able to support cyanide-resistant respiration by intact organs and to afford prolonged protection against a lethal concentration of gaseous cyanide in whole animals. Taken together, these results indicate that AOX expression in the mouse is innocuous and permits to overcome a RC blockade, while reducing associated oxidative insult. Therefore, the MitAOX mice represent a valuable tool in order to investigate the ability of AOX to counteract the panoply of mitochondrial-inherited diseases originating from oxidative phosphorylation defects. PMID:23300486

  2. Glucose sensitivity of mouse olfactory bulb neurons is conveyed by a voltage-gated potassium channel

    PubMed Central

    Tucker, Kristal; Cho, Sukhee; Thiebaud, Nicolas; Henderson, Michael X; Fadool, Debra Ann

    2013-01-01

    The olfactory bulb has recently been proposed to serve as a metabolic sensor of internal chemistry, particularly that modified by metabolism. Because the voltage-dependent potassium channel Kv1.3 regulates a large proportion of the outward current in olfactory bulb neurons and gene-targeted deletion of the protein produces a phenotype of resistance to diet-induced obesity in mice, we hypothesized that this channel may play a role in translating energy availability into a metabolic signal. Here we explored the ability of extracellular glucose concentration to modify evoked excitability of the mitral neurons that principally regulate olfactory coding and processing of olfactory information. Using voltage-clamp electrophysiology of heterologously expressed Kv1.3 channels in HEK 293 cells, we found that Kv1.3 macroscopic currents responded to metabolically active (d-) rather than inactive (l-) glucose with a response profile that followed a bell-shaped curve. Olfactory bulb slices stimulated with varying glucose concentrations showed glucose-dependent mitral cell excitability as evaluated by current-clamp electrophysiology. While glucose could be either excitatory or inhibitory, the majority of the sampled neurons displayed a decreased firing frequency in response to elevated glucose concentration that was linked to increased latency to first spike and decreased action potential cluster length. Unlike modulation attributed to phosphorylation, glucose modulation of mitral cells was rapid, less than one minute, and was reversible within the time course of a patch recording. Moreover, we report that modulation targets properties of spike firing rather than action potential shape, involves synaptic activity of glutamate or GABA signalling circuits, and is dependent upon Kv1.3 expression. Given the rising incidence of metabolic disorders attributed to weight gain, changes in neuronal excitability in brain regions regulating sensory perception of food are of consequence

  3. Transcranial laser therapy alters amyloid precursor protein processing and improves mitochondrial function in a mouse model of Alzheimer's disease

    NASA Astrophysics Data System (ADS)

    McCarthy, Thomas; Yu, Jin; El-Amouri, Salim; Gattoni-Celli, Sebastiano; Richieri, Steve; De Taboada, Luis; Streeter, Jackson; Kindy, Mark S.

    2011-03-01

    Transcranial laser therapy (TLT) using a near-infrared energy laser system was tested in the 2x Tg amyloid precursor protein (APP) mouse model of Alzheimer's Disease (AD). TLT was administered 3 times/week at escalating doses, starting at 3 months of age, and was compared to a control group which received no laser treatment. Treatment sessions were continued for a total of six months. The brains were examined for amyloid plaque burden, Aβ peptides (Aβ1-40 and Aβ1-42 ), APP cleavage products (sAPPα, CTFβ) and mitochondrial activity. Administration of TLT was associated with a significant, dose-dependent reduction in amyloid load as indicated by the numbers of Aβ plaques. Levels of Aβ1-40 and Aβ1-42 levels were likewise reduced in a dose-dependent fashion. All TLT doses produced an increase in brain sAPPα and a decrease in CTFβ levels consistent with an increase in α-secretase activity and a decrease in β-secretase activity. In addition, TLT increased ATP levels and oxygen utilization in treated animals suggesting improved mitochondrial function. These studies suggest that TLT is a potential candidate for treatment of AD.

  4. Lysosomal iron mobilization and induction of the mitochondrial permeability transition in acetaminophen-induced toxicity to mouse hepatocytes.

    PubMed

    Kon, Kazuyoshi; Kim, Jae-Sung; Uchiyama, Akira; Jaeschke, Hartmut; Lemasters, John J

    2010-09-01

    Acetaminophen induces the mitochondrial permeability transition (MPT) in hepatocytes. Reactive oxygen species (ROS) trigger the MPT and play an important role in AAP-induced hepatocellular injury. Because iron is a catalyst for ROS formation, our aim was to investigate the role of chelatable iron in MPT-dependent acetaminophen toxicity to mouse hepatocytes. Hepatocytes were isolated from fasted male C3Heb/FeJ mice. Necrotic cell killing was determined by propidium iodide fluorometry. Mitochondrial membrane potential was visualized by confocal microscopy of tetramethylrhodamine methylester. Chelatable ferrous ion was monitored by calcein quenching, and 70 kDa rhodamine-dextran was used to visualize lysosomes. Cell killing after acetaminophen (10mM) was delayed and decreased by more than half after 6 h by 1mM desferal or 1mM starch-desferal. In a cell-free system, ferrous but not ferric iron quenched calcein fluorescence, an effect reversed by dipyridyl, a membrane-permeable iron chelator. In hepatocytes loaded with calcein, intracellular calcein fluorescence decreased progressively beginning about 4 h after acetaminophen. Mitochondria then depolarized after about 6 h. Dipyridyl (20mM) dequenched calcein fluorescence. Desferal and starch-desferal conjugate prevented acetaminophen-induced calcein quenching and mitochondrial depolarization. As calcein fluorescence became quenched, lysosomes disappeared, consistent with release of iron from ruptured lysosomes. In conclusion, an increase of cytosolic chelatable ferrous iron occurs during acetaminophen hepatotoxicity, which triggers the MPT and cell killing. Disrupted lysosomes are the likely source of iron, and chelation of this iron decreases acetaminophen toxicity to hepatocytes.

  5. L-Carnitine Prevents Progression of Non-Alcoholic Steatohepatitis in a Mouse Model with Upregulation of Mitochondrial Pathway

    PubMed Central

    Ishikawa, Hisashi; Takaki, Akinobu; Tsuzaki, Ryuichiro; Yasunaka, Tetsuya; Koike, Kazuko; Shimomura, Yasuyuki; Seki, Hiroyuki; Matsushita, Hiroshi; Miyake, Yasuhiro; Ikeda, Fusao; Shiraha, Hidenori; Nouso, Kazuhiro; Yamamoto, Kazuhide

    2014-01-01

    Non-alcoholic steatohepatitis (NASH) is a severe form of non-alcoholic fatty liver disease characterized by lobular inflammation, hepatocellular ballooning, and fibrosis with an inherent risk for progression to cirrhosis and hepatocellular carcinoma (HCC). Mitochondrial dysfunction appears to play a role in the progression from simple steatosis to NASH. L-carnitine (L-b-hydroxy-g-N-trimethylaminobutyric acid), an essential nutrient that converts fat into energy in mitochondria, has been shown to ameliorate liver damage. The aim of the present study was to explore the preventive and therapeutic effect of L-carnitine in NASH model mice. Eight-week-old male STAM mice, a NASH-cirrhosis-hepatocarcinogenic model, were divided into 3 experimental groups and fed as follows: 1) high-fat diet (HFD) (control group); 2) HFD mixed with 0.28% L-carnitine (L-carnitine group); and 3) HFD mixed with 0.01% α-tocopherol (α-tocopherol group). After 4 or 8 weeks, mice were sacrificed. Blood samples and livers were collected, and hepatic tumors were counted and measured. Livers were subjected to histological study, immunohistochemical staining of 4-hydroxynonenal and ferritin, determination of 8-OHdG levels, mRNA and protein expressions for multiple genes, and metabolomic analysis. The intestinal microbiome was also analyzed. L-carnitine increased hepatic expression of genes related to long-chain fatty acid transport, mitochondrial β-oxidation, and antioxidant enzymes following suppression of hepatic oxidative stress markers and inflammatory cytokines in NASH, and mice treated with L-carnitine developed fewer liver tumors. Although α-tocopherol resulted in NASH improvement in the same manner as L-carnitine, it increased periodontitis-related microbiotic changes and hepatic iron transport-related gene expression and led to less effective for anti-hepatocarcinogenesis. Conclusion L-carnitine prevents progression of non-alcoholic steatohepatitis in a mouse model by upregulating the

  6. Mitochondrial complex I deficiency leads to inflammation and retinal ganglion cell death in the Ndufs4 mouse

    PubMed Central

    Yu, Alfred K.; Song, Lanying; Murray, Karl D.; van der List, Deborah; Sun, Chao; Shen, Yan; Xia, Zhengui; Cortopassi, Gino A.

    2015-01-01

    Mitochondrial complex I (NADH dehydrogenase) is a major contributor to neuronal energetics, and mutations in complex I lead to vision loss. Functional, neuroanatomical and transcriptional consequences of complex I deficiency were investigated in retinas of the Ndufs4 knockout mouse. Whole-eye ERGs and multielectrode arrays confirmed a major retinal ganglion cell functional loss at P32, and retinal ganglion cell loss at P42. RNAseq demonstrated a mild and then sharp increase in innate immune and inflammatory retinal transcripts at P22 and P33, respectively, which were confirmed with QRT-PCR. Intraperitoneal injection of the inflammogen lipopolysaccharide further reduced retinal ganglion cell function in Ndufs4 KO, supporting the connection between inflammatory activation and functional loss. Complex I deficiency in the retina clearly caused innate immune and inflammatory markers to increase coincident with loss of vision, and RGC functional loss. How complex I incites inflammation and functional loss is not clear, but could be the result of misfolded complex I generating a ‘non-self’ response, and induction of innate immune response transcripts was observed before functional loss at P22, including β-2 microglobulin and Cx3cr1, and during vision loss at P31 (B2m, Tlr 2, 3, 4, C1qa, Cx3cr1 and Fas). These data support the hypothesis that mitochondrial complex I dysfunction in the retina triggers an innate immune and inflammatory response that results in loss of retinal ganglion cell function and death, as in Leber's hereditary Optic Neuropathy and suggests novel therapeutic routes to counter mitochondrial defects that contribute to vision loss. PMID:25652399

  7. Effects of Oxidative Alcohol Metabolism on the Mitochondrial Permeability Transition Pore and Necrosis in a Mouse Model of Alcoholic Pancreatitis

    PubMed Central

    SHALBUEVA, NATALIA; MARENINOVA, OLGA A.; GERLOFF, ANDREAS; YUAN, JINGZHEN; WALDRON, RICHARD T.; PANDOL, STEPHEN J.; GUKOVSKAYA, ANNA S.

    2013-01-01

    BACKGROUND & AIMS Opening of the mitochondrial permeability transition pore (MPTP) causes loss of the mitochondrial membrane potential (ΔΨm) and, ultimately, adenosine triphosphate depletion and necrosis. Cells deficient in cyclophilin D (CypD), a component of the MPTP, are resistant to MPTP opening, loss of ΔΨm, and necrosis. Alcohol abuse is a major risk factor for pancreatitis and is believed to sensitize the pancreas to stressors, by poorly understood mechanisms. We investigated the effects of ethanol on the pancreatic MPTP, the mechanisms of these effects, and their role in pancreatitis. METHODS We measured ΔΨm in mouse pancreatic acinar cells incubated with ethanol alone and in combination with physiologic and pathologic concentrations of cholecystokinin-8 (CCK). To examine the role of MPTP, we used ex vivo and in vivo models of pancreatitis, induced in wild-type and CypD−/− mice by a combination of ethanol and CCK. RESULTS Ethanol reduced basal ΔΨm and converted a transient depolarization, induced by physiologic concentrations of CCK, into a sustained decrease in ΔΨm, resulting in reduced cellular adenosine triphosphate and increased necrosis. The effects of ethanol and CCK were mediated by MPTP because they were not observed in CypD−/− acinar cells. Ethanol and CCK activated MPTP through different mechanisms— ethanol by reducing the ratio of oxidized nicotinamide adenine dinucleotide to reduced nicotinamide adenine dinucleotide, as a result of oxidative metabolism, and CCK by increasing cytosolic Ca2+. CypD−/− mice developed a less-severe form of pancreatitis after administration of ethanol and CCK. CONCLUSIONS Oxidative metabolism of ethanol sensitizes pancreatic mitochondria to activate MPTP, leading to mitochondrial failure; this makes the pancreas susceptible to necrotizing pancreatitis. PMID:23103769

  8. Cell-type-dependent action potentials and voltage-gated currents in mouse fungiform taste buds.

    PubMed

    Kimura, Kenji; Ohtubo, Yoshitaka; Tateno, Katsumi; Takeuchi, Keita; Kumazawa, Takashi; Yoshii, Kiyonori

    2014-01-01

    Taste receptor cells fire action potentials in response to taste substances to trigger non-exocytotic neurotransmitter release in type II cells and exocytotic release in type III cells. We investigated possible differences between these action potentials fired by mouse taste receptor cells using in situ whole-cell recordings, and subsequently we identified their cell types immunologically with cell-type markers, an IP3 receptor (IP3 R3) for type II cells and a SNARE protein (SNAP-25) for type III cells. Cells not immunoreactive to these antibodies were examined as non-IRCs. Here, we show that type II cells and type III cells fire action potentials using different ionic mechanisms, and that non-IRCs also fire action potentials with either of the ionic mechanisms. The width of action potentials was significantly narrower and their afterhyperpolarization was deeper in type III cells than in type II cells. Na(+) current density was similar in type II cells and type III cells, but it was significantly smaller in non-IRCs than in the others. Although outwardly rectifying current density was similar between type II cells and type III cells, tetraethylammonium (TEA) preferentially suppressed the density in type III cells and the majority of non-IRCs. Our mathematical model revealed that the shape of action potentials depended on the ratio of TEA-sensitive current density and TEA-insensitive current one. The action potentials of type II cells and type III cells under physiological conditions are discussed.

  9. Quantitative Proteomics of Sleep-Deprived Mouse Brains Reveals Global Changes in Mitochondrial Proteins

    PubMed Central

    Li, Tie-Mei; Zhang, Ju-en; Lin, Rui; Chen, She; Luo, Minmin; Dong, Meng-Qiu

    2016-01-01

    Sleep is a ubiquitous, tightly regulated, and evolutionarily conserved behavior observed in almost all animals. Prolonged sleep deprivation can be fatal, indicating that sleep is a physiological necessity. However, little is known about its core function. To gain insight into this mystery, we used advanced quantitative proteomics technology to survey the global changes in brain protein abundance. Aiming to gain a comprehensive profile, our proteomics workflow included filter-aided sample preparation (FASP), which increased the coverage of membrane proteins; tandem mass tag (TMT) labeling, for relative quantitation; and high resolution, high mass accuracy, high throughput mass spectrometry (MS). In total, we obtained the relative abundance ratios of 9888 proteins encoded by 6070 genes. Interestingly, we observed significant enrichment for mitochondrial proteins among the differentially expressed proteins. This finding suggests that sleep deprivation strongly affects signaling pathways that govern either energy metabolism or responses to mitochondrial stress. Additionally, the differentially-expressed proteins are enriched in pathways implicated in age-dependent neurodegenerative diseases, including Parkinson’s, Huntington’s, and Alzheimer’s, hinting at possible connections between sleep loss, mitochondrial stress, and neurodegeneration. PMID:27684481

  10. Reduced low-voltage activated K+ conductances and enhanced central excitability in a congenitally deaf (dn/dn) mouse

    PubMed Central

    Leao, Richardson N; Berntson, Amy; Forsythe, Ian D; Walmsley, Bruce

    2004-01-01

    We have investigated changes in the neuronal excitability of the auditory brainstem in a congenitally deaf mouse (deafness dn/dn). Whole cell patch recordings from principal neurones of the medial nucleus of the trapezoid body (MNTB) showed strikingly enhanced excitability in the deaf mice when compared to control CBA mice at 12–14 days postnatal. MNTB neurones in normal CBA mice showed the phenotypic single action potential response on depolarization in current clamp; however, recordings from CBA mice carrying the homozygous deafness mutation fired trains of action potentials on depolarization. We show here that these changes are associated with reduced functional expression of dendrotoxin-sensitive Kv1 potassium channels. In contrast, no differences were found in voltage-gated calcium currents between control and deaf mice. These results reveal that loss of hair cell function in the cochlea leads to changes in ion channel expression in the central nervous system and suggests that this deafness model will be an important tool in understanding central changes occurring in human congenital deafness and in exploring activity-dependent regulation of ion channel expression. PMID:15235085

  11. [Distribution of foreign mitochondrial DNA during the first splittings of the transmitochondrial mouse embryos].

    PubMed

    Kustova, M E; Sokolova, V A; Bass, M G; Zakharova, F M; Sorokin, A V; Vasil'ev, V B

    2008-01-01

    Distribution of human mitochondrial DNA (mtDNA) among separate murine blastomeres was analyzed during the splitting of embryos in which the suspension of human mitochondria had been injected at the one- or two-cell stage. Human mtDNA was detected by PCR with species specific primers. The total amount of the two- and four-cell murine embryos analyzed in the study was 339. In all embryos examined the copies of human mitochondrial genome were revealed along with murine mtDNA, which indicated the phenomenon of an artificially modeled heteroplasmy. The foreign mtDNA was not ubiquitous among the blastomeres of transmitochondrial embryos. Mathematical analysis of the results showed that in the period between the injection of human mitochondria and the subsequent splitting no equal distribution of the human mtDNA occurred in the cytoplasm. These results also point at the presence of more than 2-3 segregation units of mtDNA in the entire pool of mitochondria (about 5 x 10(2)) introduced into an embryo by microinjection.

  12. An integrase of endogenous retrovirus is involved in maternal mitochondrial DNA inheritance of the mouse.

    PubMed

    Hayashida, Kenji; Omagari, Katsuhisa; Masuda, Jun-Ichi; Kohno, Shigeru

    2008-02-01

    The mechanism of maternal mitochondrial DNA (mtDNA) inheritance in animals can be said to be the selective elimination of sperm mtDNA via the elimination factor of the egg and a sperm mitochondria-specific factor. In 2005, we clarified that t-tpis (Spag1 isoform 1) is a mitochondria-specific translocator and the sperm factor, and furthermore estimated that the elimination factors of the egg are the divalent cation-dependent endonuclease and s-tpis (Spag1 isoform 2 and isoform 3) as the elimination system-specific chaperone [K. Hayashida, K. Omagari, J. Masuda, H. Hazama, Y. Kadokawa, K. Ohba, S. Kohno, The sperm mitochondria-specific translocator has a key role in maternal mitochondrial inheritance, Cell Biol. Int. 29 (2005) 472-481]. This time, using a recombinant Spag1 isoform 1 protein, a pull-down assay of ovary cytosol was performed and the elimination factors searched for. Surprisingly, an endogenous retroviral integrase fragment (Eri15) was identified using mass spectrometry of the electrophoresis band of the pull-down protein. Eri15 was detected as a complex of approximately 500kDa with Spag1 isoform 2 or isoform 3 in native PAGE of the ovary cytosol. This strongly suggested that Eri15 is selectively transported into the sperm mitochondria matrix by Spag1 isoform 2 and 3 via Spag1 isoform 1 and that sperm mtDNA is destroyed, thus causing the establishment of maternal mtDNA inheritance.

  13. Autophagy and mitochondrial remodelling in mouse mesenchymal stromal cells challenged with Staphylococcus epidermidis.

    PubMed

    Gorbunov, Nikolai V; McDaniel, Dennis P; Zhai, Min; Liao, Pei-Jyun; Garrison, Bradley R; Kiang, Juliann G

    2015-05-01

    The bone marrow stroma constitutes the marrow-blood barrier, which sustains immunochemical homoeostasis and protection of the haematopoietic tissue in sequelae of systemic bacterial infections. Under these conditions, the bone marrow stromal cells affected by circulating bacterial pathogens shall elicit the adaptive stress-response mechanisms to maintain integrity of the barrier. The objective of this communication was to demonstrate (i) that in vitro challenge of mesenchymal stromal cells, i.e. colony-forming unit fibroblasts (CFU-F), with Staphylococcus epidermidis can activate the autophagy pathway to execute antibacterial defence response, and (ii) that homoeostatic shift because of the bacteria-induced stress includes the mitochondrial remodelling and sequestration of compromised organelles via mitophagy. Implication of Drp1 and PINK1-PARK2-dependent mechanisms in the mitophagy turnover of the aberrant mitochondria in mesenchymal stromal cells is investigated and discussed.

  14. Autophagy and mitochondrial remodelling in mouse mesenchymal stromal cells challenged with Staphylococcus epidermidis.

    PubMed

    Gorbunov, Nikolai V; McDaniel, Dennis P; Zhai, Min; Liao, Pei-Jyun; Garrison, Bradley R; Kiang, Juliann G

    2015-05-01

    The bone marrow stroma constitutes the marrow-blood barrier, which sustains immunochemical homoeostasis and protection of the haematopoietic tissue in sequelae of systemic bacterial infections. Under these conditions, the bone marrow stromal cells affected by circulating bacterial pathogens shall elicit the adaptive stress-response mechanisms to maintain integrity of the barrier. The objective of this communication was to demonstrate (i) that in vitro challenge of mesenchymal stromal cells, i.e. colony-forming unit fibroblasts (CFU-F), with Staphylococcus epidermidis can activate the autophagy pathway to execute antibacterial defence response, and (ii) that homoeostatic shift because of the bacteria-induced stress includes the mitochondrial remodelling and sequestration of compromised organelles via mitophagy. Implication of Drp1 and PINK1-PARK2-dependent mechanisms in the mitophagy turnover of the aberrant mitochondria in mesenchymal stromal cells is investigated and discussed. PMID:25721260

  15. Omega-3 fatty acid deficiency disrupts endocytosis, neuritogenesis, and mitochondrial protein pathways in the mouse hippocampus

    PubMed Central

    English, Jane A.; Harauma, Akiko; Föcking, Melanie; Wynne, Kieran; Scaife, Caitriona; Cagney, Gerard; Moriguchi, Toru; Cotter, David R.

    2013-01-01

    Omega-3 fatty acid (n-3 FA) deficiency is an environmental risk factor for schizophrenia, yet characterization of the consequences of deficiency at the protein level in the brain is limited. We aimed to identify the protein pathways disrupted as a consequence of chronic n-3 deficiency in the hippocampus of mice. Fatty acid analysis of the hippocampus following chronic dietary deficiency revealed a 3-fold decrease (p < 0.001) in n-3 FA levels. Label free LC-MS/MS analysis identified and profiled 1008 proteins, of which 114 were observed to be differentially expressed between n-3 deficient and control groups (n = 8 per group). The cellular processes that were most implicated were neuritogenesis, endocytosis, and exocytosis, while specific protein pathways that were most significantly dysregulated were mitochondrial dysfunction and clathrin mediated endocytosis (CME). In order to characterize whether these processes and pathways are ones influenced by antipsychotic medication, we used LC-MS/MS to test the differential expression of these 114 proteins in the hippocampus of mice chronically treated with the antipsychotic agent haloperidol. We observed 23 of the 114 proteins to be differentially expressed, 17 of which were altered in the opposite direction to that observed following n-3 deficiency. Overall, our findings point to disturbed synaptic function, neuritogenesis, and mitochondrial function as a consequence of dietary deficiency in n-3 FA. This study greatly aids our understanding of the molecular mechanism by which n-3 deficiency impairs normal brain function, and provides clues as to how n-3 FA exert their therapeutic effect in early psychosis. PMID:24194745

  16. Membrane voltage, resistance, and channel switching in isolated mouse fibroblasts (L cells): a patch-electrode analysis.

    PubMed Central

    Hosoi, S; Slayman, C L

    1985-01-01

    The whole-cell patch-electrode technique of Fenwick, Marty & Neher (1982) has been applied to single suspension-cultured mouse fibroblasts. Seals in the range of 10-50 G omega were obtained without special cleaning of the cell membranes. Rupture of the membrane patch inside the electrode was accompanied by a shift of measured potential into the range -10 to -25 mV, but in most cases with little change in the recorded resistance. The latter fact implied that the absolute resistance of the cell membrane must be in the same range as the seal resistance and the recorded potential is a poor measure of actual cell membrane potential. Steady-state current-voltage curves (range -160 mV to +80 mV) were generated before and after rupture of the membrane patch, and the difference between these gave (zero-current) membrane potentials of -50 to -75 mV, which represents a leak-corrected estimate of the true cell-membrane potential. The associated slope conductivity of the cell membrane was 5-15 microS/cm2 (assumed smooth-sphere geometry, cells 13-15 microns in diameter) and was K+-dominated. With 0.1 mM (or more) free Ca2+ filling the patch electrode, membrane potentials in the range -60 to -85 mV were observed following patch rupture, with associated slope conductivities of 200-400 microS/cm2, also K+-dominated. Similar voltages and conductivities were observed at the peak of pulse-induced 'hyperpolarizing activation' (Nelson, Peacock, & Minna, 1972), and the two phenomena probably reflect the behaviour of Ca2+-activated K+ channels. Both the pulse-induced conductance and the Ca2+-activated conductance spontaneously decayed, the latter over periods of 5-15 min following patch rupture. Sr2+, Ba2+, and Co2+ could also activate the putative K+ channels, but only Sr2+ really mimicked Ca2+. Co2+ and Ba2+ activated with a delay of several minutes following patch rupture, and deactivated quickly with a small decrease of conductance and a large decrease of membrane potential. Evidently

  17. The human and mouse SLC25A29 mitochondrial transporters rescue the deficient ornithine metabolism in fibroblasts of patients with the hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome.

    PubMed

    Camacho, José A; Rioseco-Camacho, Natalia

    2009-07-01

    The hyperornithinemia-hyperammonemia-homocitrullinuria (HHH) syndrome is a disorder of the urea cycle (UCD) and ornithine degradation pathway caused by mutations in the mitochondrial ornithine transporter (ORNT1). Unlike other UCDs, HHH syndrome is characterized by a less severe and variable phenotype that we believe may, in part, be due to genes with redundant function to ORNT1, such as the previously characterized ORNT2 gene. We reasoned that SLC25A29, a member of the same subfamily of mitochondrial carrier proteins as ORNT1 and ORNT2, might also have overlapping function with ORNT1. Here, we report that both the human and mouse SLC25A29, previously identified as mitochondrial carnitine/acyl-carnitine transporter-like, when overexpressed transiently also rescues the impaired ornithine transport in cultured HHH fibroblasts. Moreover, we observed that, in the mouse, the Slc25a29 message is more significantly expressed in the CNS and cultured astrocytes when compared with the liver and kidney. These results suggest a potential physiologic role for the SLC25A29 transporter in the oxidation of fatty acids, ornithine degradation pathway, and possibly the urea cycle. Our results show that SLC25A29 is the third human mitochondrial ornithine transporter, designated as ORNT3, which may contribute to the milder and variable phenotype seen in patients with HHH syndrome. PMID:19287344

  18. Restricted ADP movement in cardiomyocytes: Cytosolic diffusion obstacles are complemented with a small number of open mitochondrial voltage-dependent anion channels.

    PubMed

    Simson, Päivo; Jepihhina, Natalja; Laasmaa, Martin; Peterson, Pearu; Birkedal, Rikke; Vendelin, Marko

    2016-08-01

    Adequate intracellular energy transfer is crucial for proper cardiac function. In energy starved failing hearts, partial restoration of energy transfer can rescue mechanical performance. There are two types of diffusion obstacles that interfere with energy transfer from mitochondria to ATPases: mitochondrial outer membrane (MOM) with voltage-dependent anion channel (VDAC) permeable to small hydrophilic molecules and cytoplasmatic diffusion barriers grouping ATP-producers and -consumers. So far, there is no method developed to clearly distinguish the contributions of cytoplasmatic barriers and MOM to the overall diffusion restriction. Furthermore, the number of open VDACs in vivo remains unknown. The aim of this work was to establish the partitioning of intracellular diffusion obstacles in cardiomyocytes. We studied the response of mitochondrial oxidative phosphorylation of permeabilized rat cardiomyocytes to changes in extracellular ADP by recording 3D image stacks of NADH autofluorescence. Using cell-specific mathematical models, we determined the permeability of MOM and cytoplasmatic barriers. We found that only ~2% of VDACs are accessible to cytosolic ADP and cytoplasmatic diffusion barriers reduce the apparent diffusion coefficient by 6-10×. In cardiomyocytes, diffusion barriers in the cytoplasm and by the MOM restrict ADP/ATP diffusion to similar extents suggesting a major role of both barriers in energy transfer and other intracellular processes. PMID:27261153

  19. Mitochondria-targeted antioxidant mitotempo protects mitochondrial function against amyloid beta toxicity in primary cultured mouse neurons.

    PubMed

    Hu, Hongtao; Li, Mo

    2016-09-01

    Mitochondrial defects including excess reactive oxygen species (ROS) production and compromised ATP generation are featured pathology in Alzheimer's disease (AD). Amyloid beta (Aβ)-mediated mitochondrial ROS overproduction disrupts intra-neuronal Redox balance, in turn exacerbating mitochondrial dysfunction leading to neuronal injury. Previous studies have found the beneficial effects of mitochondria-targeted antioxidants in preventing mitochondrial dysfunction and neuronal injury in AD animal and cell models, suggesting that mitochondrial ROS scavengers hold promise for the treatment of this neurological disorder. In this study, we have determined that mitotempo, a novel mitochondria-targeted antioxidant protects mitochondrial function from the toxicity of Aβ in primary cultured neurons. Our results showed that Aβ-promoted mitochondrial superoxide production and neuronal lipid oxidation were significantly suppressed by the application of mitotempo. Moreover, mitotempo also demonstrated protective effects on mitochondrial bioenergetics evidenced by preserved mitochondrial membrane potential, cytochrome c oxidase activity as well as ATP production. In addition, the Aβ-induced mitochondrial DNA (mtDNA) depletion and decreased expression levels of mtDNA replication-related DNA polymerase gamma (DNA pol γ) and Twinkle were substantially mitigated by mitotempo. Therefore, our study suggests that elimination of excess mitochondrial ROS rescues mitochondrial function in Aβ-insulted neruons; and mitotempo has the potential to be a promising therapeutic agent to protect mitochondrial and neuronal function in AD.

  20. Mitochondria-targeted antioxidant mitotempo protects mitochondrial function against amyloid beta toxicity in primary cultured mouse neurons.

    PubMed

    Hu, Hongtao; Li, Mo

    2016-09-01

    Mitochondrial defects including excess reactive oxygen species (ROS) production and compromised ATP generation are featured pathology in Alzheimer's disease (AD). Amyloid beta (Aβ)-mediated mitochondrial ROS overproduction disrupts intra-neuronal Redox balance, in turn exacerbating mitochondrial dysfunction leading to neuronal injury. Previous studies have found the beneficial effects of mitochondria-targeted antioxidants in preventing mitochondrial dysfunction and neuronal injury in AD animal and cell models, suggesting that mitochondrial ROS scavengers hold promise for the treatment of this neurological disorder. In this study, we have determined that mitotempo, a novel mitochondria-targeted antioxidant protects mitochondrial function from the toxicity of Aβ in primary cultured neurons. Our results showed that Aβ-promoted mitochondrial superoxide production and neuronal lipid oxidation were significantly suppressed by the application of mitotempo. Moreover, mitotempo also demonstrated protective effects on mitochondrial bioenergetics evidenced by preserved mitochondrial membrane potential, cytochrome c oxidase activity as well as ATP production. In addition, the Aβ-induced mitochondrial DNA (mtDNA) depletion and decreased expression levels of mtDNA replication-related DNA polymerase gamma (DNA pol γ) and Twinkle were substantially mitigated by mitotempo. Therefore, our study suggests that elimination of excess mitochondrial ROS rescues mitochondrial function in Aβ-insulted neruons; and mitotempo has the potential to be a promising therapeutic agent to protect mitochondrial and neuronal function in AD. PMID:27444386

  1. A disulfide bond in the TIM23 complex is crucial for voltage gating and mitochondrial protein import.

    PubMed

    Ramesh, Ajay; Peleh, Valentina; Martinez-Caballero, Sonia; Wollweber, Florian; Sommer, Frederik; van der Laan, Martin; Schroda, Michael; Alexander, R Todd; Campo, María Luisa; Herrmann, Johannes M

    2016-08-15

    Tim17 is a central, membrane-embedded subunit of the mitochondrial protein import machinery. In this study, we show that Tim17 contains a pair of highly conserved cysteine residues that form a structural disulfide bond exposed to the intermembrane space (IMS). This disulfide bond is critical for efficient protein translocation through the TIM23 complex and for dynamic gating of its preprotein-conducting channel. The disulfide bond in Tim17 is formed during insertion of the protein into the inner membrane. Whereas the import of Tim17 depends on the binding to the IMS protein Mia40, the oxidoreductase activity of Mia40 is surprisingly dispensable for Tim17 oxidation. Our observations suggest that Tim17 can be directly oxidized by the sulfhydryl oxidase Erv1. Thus, import and oxidation of Tim17 are mediated by the mitochondrial disulfide relay, though the mechanism by which the disulfide bond in Tim17 is formed differs considerably from that of soluble IMS proteins. PMID:27502485

  2. In vivo evidence of mitochondrial dysfunction and altered redox homeostasis in a genetic mouse model of propionic acidemia: Implications for the pathophysiology of this disorder.

    PubMed

    Gallego-Villar, L; Rivera-Barahona, A; Cuevas-Martín, C; Guenzel, A; Pérez, B; Barry, M A; Murphy, M P; Logan, A; Gonzalez-Quintana, A; Martín, M A; Medina, S; Gil-Izquierdo, A; Cuezva, J M; Richard, E; Desviat, L R

    2016-07-01

    Accumulation of toxic metabolites has been described to inhibit mitochondrial enzymes, thereby inducing oxidative stress in propionic acidemia (PA), an autosomal recessive metabolic disorder caused by the deficiency of mitochondrial propionyl-CoA carboxylase. PA patients exhibit neurological deficits and multiorgan complications including cardiomyopathy. To investigate the role of mitochondrial dysfunction in the development of these alterations we have used a hypomorphic mouse model of PA that mimics the biochemical and clinical hallmarks of the disease. We have studied the tissue-specific bioenergetic signature by Reverse Phase Protein Microarrays and analysed OXPHOS complex activities, mtDNA copy number, oxidative damage, superoxide anion and hydrogen peroxide levels. The results show decreased levels and/or activity of several OXPHOS complexes in different tissues of PA mice. An increase in mitochondrial mass and OXPHOS complexes was observed in brain, possibly reflecting a compensatory mechanism including metabolic reprogramming. mtDNA depletion was present in most tissues analysed. Antioxidant enzymes were also found altered. Lipid peroxidation was present along with an increase in hydrogen peroxide and superoxide anion production. These data support the hypothesis that oxidative damage may contribute to the pathophysiology of PA, opening new avenues in the identification of therapeutic targets and paving the way for in vivo evaluation of compounds targeting mitochondrial biogenesis or reactive oxygen species production. PMID:27083476

  3. Paternal Diet-Induced Obesity Retards Early Mouse Embryo Development, Mitochondrial Activity and Pregnancy Health

    PubMed Central

    Binder, Natalie K.; Hannan, Natalie J.; Gardner, David K.

    2012-01-01

    Worldwide, 48% of adult males are overweight or obese. An association between infertility and excessive body weight is now accepted, although focus remains primarily on females. It has been shown that parental obesity results in compromised embryo development, disproportionate changes in embryo metabolism and reduced blastocyst cell number. The aim of this study was to determine whether paternal obesity has negative effects on the resultant embryo. Specifically, using in vitro fertilisation (IVF), we wanted to isolate the functional effects of obesity on sperm by examining the subsequent embryo both pre- and post-implantation. Epididymal sperm was collected from age matched normal and obese C57BL/6 mice and cryopreserved for subsequent IVF with oocytes collected from Swiss females (normal diet/weight). Obesity was induced in male mice by feeding a high fat diet of 22% fat for 10 weeks. Resultant embryos were cultured individually and development monitored using time-lapse microscopy. Paternal obesity resulted in a significant delay in preimplantation embryo development as early as syngamy (P<0.05). Metabolic parameters were measured across key developmental stages, demonstrating significant reduction in mitochondrial membrane potential (P<0.01). Blastocysts were stained to determine trophectoderm (TE) and inner cell mass (ICM) cell numbers, revealing significant differences in the ratio of cell allocation to TE and ICM lineages (P<0.01). Functional studies examining blastocyst attachment, growth and implantation demonstrated that blastocysts derived from sperm of obese males displayed significantly reduced outgrowth on fibronectin in vitro (P<0.05) and retarded fetal development in vivo following embryo transfer (P<0.05). Taken together, these data clearly demonstrate that paternal obesity has significant negative effects on the embryo at a variety of key early developmental stages, resulting in delayed development, reduced placental size and smaller offspring

  4. Estradiol and tamoxifen regulate NRF-1 and mitochondrial function in mouse mammary gland and uterus.

    PubMed

    Ivanova, Margarita M; Radde, Brandie N; Son, Jieun; Mehta, Fabiola F; Chung, Sang-Hyuk; Klinge, Carolyn M

    2013-10-01

    Nuclear respiratory factor-1 (NRF-1) stimulates the transcription of nuclear-encoded genes that regulate mitochondrial (mt) genome transcription and biogenesis. We reported that estradiol (E2) and 4-hydroxytamoxifen (4-OHT) stimulate NRF-1 transcription in an estrogen receptor α (ERα)- and ERβ-dependent manner in human breast cancer cells. The aim of this study was to determine whether E2 and 4-OHT increase NRF-1 in vivo. Here, we report that E2 and 4-OHT increase NRF-1 expression in mammary gland (MG) and uterus of ovariectomized C57BL/6 mice in a time-dependent manner. E2 increased NRF-1 protein in the uterus and MG; however, in MG, 4-OHT increased Nrf1 mRNA but not protein. Chromatin immunoprecipitation assays revealed increased in vivo recruitment of ERα to the Nrf1 promoter and intron 3 in MG and uterus 6 h after E2 and 4-OHT treatment, commensurate with increased NRF-1 expression. E2- and 4-OHT-induced increases in NRF-1 and its target genes Tfam, Tfb1m, and Tfb2m were coordinated in MG but not in uterus due to uterine-selective inhibition of the expression of the NRF-1 coactivators Ppargc1a and Ppargc1b by E2 and 4-OHT. E2 transiently increased NRF-1 and PGC-1α nuclear staining while reducing PGC-1α in uterus. E2, not 4-OHT, activates mt biogenesis in MG and uterus in a time-dependent manner. E2 increased mt outer membrane Tomm40 protein levels in MG and uterus whereas 4-OHT increased Tomm40 only in uterus. These data support the hypothesis of tissue-selective regulation of NRF-1 and its downstream targets by E2 and 4-OHT in vivo.

  5. Real-Time PCR Quantification of Heteroplasmy in a Mouse Model with Mitochondrial DNA of C57BL/6 and NZB/BINJ Strains

    PubMed Central

    Sangalli, Juliano Rodrigues; Rodrigues, Thiago Bittencourt; Smith, Lawrence Charles; Meirelles, Flávio Vieira; Chiaratti, Marcos Roberto

    2015-01-01

    Mouse models are widely employed to study mitochondrial inheritance, which have implications to several human diseases caused by mutations in the mitochondrial genome (mtDNA). These mouse models take advantage of polymorphisms between the mtDNA of the NZB/BINJ and the mtDNA of common inbred laboratory (i.e., C57BL/6) strains to generate mice with two mtDNA haplotypes (heteroplasmy). Based on PCR followed by restriction fragment length polymorphism (PCR-RFLP), these studies determine the level of heteroplasmy across generations and in different cell types aiming to understand the mechanisms underlying mitochondrial inheritance. However, PCR-RFLP is a time-consuming method of low sensitivity and accuracy that dependents on the use of restriction enzyme digestions. A more robust method to measure heteroplasmy has been provided by the use of real-time quantitative PCR (qPCR) based on allelic refractory mutation detection system (ARMS-qPCR). Herein, we report an ARMS-qPCR assay for quantification of heteroplasmy using heteroplasmic mice with mtDNA of NZB/BINJ and C57BL/6 origin. Heteroplasmy and mtDNA copy number were estimated in germline and somatic tissues, providing evidence of the reliability of the approach. Furthermore, it enabled single-step quantification of heteroplasmy, with sensitivity to detect as low as 0.1% of either NZB/BINJ or C57BL/6 mtDNA. These findings are relevant as the ARMS-qPCR assay reported here is fully compatible with similar heteroplasmic mouse models used to study mitochondrial inheritance in mammals. PMID:26274500

  6. High Glucose-Induced Mitochondrial Respiration and Reactive Oxygen Species in Mouse Cerebral Pericytes is Reversed by Pharmacological Inhibition of Mitochondrial Carbonic Anhydrases: Implications for Cerebral Microvascular Disease in Diabetes

    PubMed Central

    Shah, Gul N.; Morofuji, Yoichi; Banks, William A.; Price, Tulin O.

    2013-01-01

    Hyperglycemia-induced oxidative stress leads to diabetes-associated damage to the microvasculature of the brain. Pericytes in close proximity to endothelial cells in the brain microvessels are vital to the integrity of the blood-brain barrier and are especially susceptible to oxidative stress. According to our recently published results, streptozotocin-diabetic mouse brain exhibits oxidative stress and loose pericytes by twelve weeks of diabetes, and cerebral pericytes cultured in high glucose media suffer intracellular oxidative stress and apoptosis. Oxidative stress in diabetes is hypothesized to be caused by reactive oxygen species (ROS) produced during hyperglycemia-induced enhanced oxidative metabolism of glucose (respiration). To test this hypothesis, we investigated the effect of high glucose on respiration rate and ROS production in mouse cerebral pericytes. Previously, we showed that pharmacological inhibition of mitochondrial carbonic anhydrases protects the brain from oxidative stress and pericyte loss. The high glucose-induced intracellular oxidative stress and apoptosis of pericytes in culture were also reversed by inhibition of mitochondrial carbonic anhydrases. Therefore, we extended our current study to determine the effect of these inhibitors on high glucose-induced increases in pericyte respiration and ROS. We now report that both the respiration and ROS are significantly increased in pericytes challenged with high glucose. Furthermore, inhibition of mitochondrial carbonic anhydrases significantly slowed down both the rate of respiration and ROS production. These data provide new evidence that pharmacological inhibitors of mitochondrial carbonic anhydrases, already in clinical use, may prove beneficial in protecting the brain from oxidative stress caused by ROS produced as a consequence of hyperglycemia-induced enhanced respiration. PMID:24076121

  7. High glucose-induced mitochondrial respiration and reactive oxygen species in mouse cerebral pericytes is reversed by pharmacological inhibition of mitochondrial carbonic anhydrases: Implications for cerebral microvascular disease in diabetes.

    PubMed

    Shah, Gul N; Morofuji, Yoichi; Banks, William A; Price, Tulin O

    2013-10-18

    Hyperglycemia-induced oxidative stress leads to diabetes-associated damage to the microvasculature of the brain. Pericytes in close proximity to endothelial cells in the brain microvessels are vital to the integrity of the blood-brain barrier and are especially susceptible to oxidative stress. According to our recently published results, streptozotocin-diabetic mouse brain exhibits oxidative stress and loose pericytes by twelve weeks of diabetes, and cerebral pericytes cultured in high glucose media suffer intracellular oxidative stress and apoptosis. Oxidative stress in diabetes is hypothesized to be caused by reactive oxygen species (ROS) produced during hyperglycemia-induced enhanced oxidative metabolism of glucose (respiration). To test this hypothesis, we investigated the effect of high glucose on respiration rate and ROS production in mouse cerebral pericytes. Previously, we showed that pharmacological inhibition of mitochondrial carbonic anhydrases protects the brain from oxidative stress and pericyte loss. The high glucose-induced intracellular oxidative stress and apoptosis of pericytes in culture were also reversed by inhibition of mitochondrial carbonic anhydrases. Therefore, we extended our current study to determine the effect of these inhibitors on high glucose-induced increases in pericyte respiration and ROS. We now report that both the respiration and ROS are significantly increased in pericytes challenged with high glucose. Furthermore, inhibition of mitochondrial carbonic anhydrases significantly slowed down both the rate of respiration and ROS production. These data provide new evidence that pharmacological inhibitors of mitochondrial carbonic anhydrases, already in clinical use, may prove beneficial in protecting the brain from oxidative stress caused by ROS produced as a consequence of hyperglycemia-induced enhanced respiration.

  8. Arsenic Induces Insulin Resistance in Mouse Adipocytes and Myotubes Via Oxidative Stress-Regulated Mitochondrial Sirt3-FOXO3a Signaling Pathway.

    PubMed

    Padmaja Divya, Sasidharan; Pratheeshkumar, Poyil; Son, Young-Ok; Vinod Roy, Ram; Andrew Hitron, John; Kim, Donghern; Dai, Jin; Wang, Lei; Asha, Padmaja; Huang, Bin; Xu, Mei; Luo, Jia; Zhang, Zhuo

    2015-08-01

    Chronic exposure to arsenic via drinking water is associated with an increased risk for development of type 2 diabetes mellitus (T2DM). This study investigates the role of mitochondrial oxidative stress protein Sirtuin 3 (Sirt3) and its targeting proteins in chronic arsenic-induced T2DM in mouse adipocytes and myotubes. The results show that chronic arsenic exposure significantly decreased insulin-stimulated glucose uptake (ISGU) in correlation with reduced expression of insulin-regulated glucose transporter type 4 (Glut4). Expression of Sirt3, a mitochondrial deacetylase, was dramatically decreased along with its associated transcription factor, forkhead box O3 (FOXO3a) upon arsenic exposure. A decrease in mitochondrial membrane potential (Δψm) was observed in both 3T3L1 adipocytes and C2C12 myotubes treated by arsenic. Reduced FOXO3a activity by arsenic exhibited a decreased binding affinity to the promoters of both manganese superoxide dismutase (MnSOD) and peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1α, a broad and powerful regulator of reactive oxygen species (ROS) metabolism. Forced expression of Sirt3 or MnSOD in mouse myotubes elevated Δψm and restored ISGU inhibited by arsenic exposure. Our results suggest that Sirt3/FOXO3a/MnSOD signaling plays a significant role in the inhibition of ISGU induced by chronic arsenic exposure. PMID:25979314

  9. Diphenyl diselenide administration enhances cortical mitochondrial number and activity by increasing hemeoxygenase type 1 content in a methylmercury-induced neurotoxicity mouse model.

    PubMed

    Glaser, Viviane; Martins, Roberta de Paula; Vieira, Ana Julia Hoffmann; Oliveira, Eliana de Medeiros; Straliotto, Marcos Raniel; Mukdsi, Jorge Humberto; Torres, Alicia Inés; de Bem, Andreza Fabro; Farina, Marcelo; da Rocha, João Batista Teixeira; De Paul, Ana Lucia; Latini, Alexandra

    2014-05-01

    Interest in biochemistry of organoselenium compound has increased in the last decades, mainly due to their chemical and biological activities. Here, we investigated the protective effect of diphenyl diselenide (PhSe)2 (5 μmol/kg), in a mouse model of methylmercury (MeHg)-induced brain toxicity. Swiss male mice were divided into four experimental groups: control, (PhSe)2 (5 μmol/kg, subcutaneous administration), MeHg (40 mg/L, in tap water), and MeHg + (PhSe)2. After the treatment (21 days), the animals were killed and the cerebral cortex was analyzed. Electron microscopy indicated an enlarged and fused mitochondria leading to a reduced number of organelles, in the MeHg-exposed mice. Furthermore, cortical creatine kinase activity, a sensitive mitochondrial oxidative stress sensor, was almost abolished by MeHg. Subcutaneous (PhSe)2 co-treatment rescued from MeHg-induced mitochondrial alterations. (PhSe)2 also behaved as an enhancer of mitochondrial biogenesis, by increasing cortical mitochondria content in mouse-receiving (PhSe)2 alone. Mechanistically, (PhSe)2 (1 μM; 24 h) would trigger the cytoprotective Nrf-2 pathway for activating target genes, since astroglial cells exposed to the chalcogen showed increased content of hemeoxygenase type 1, a sensitive marker of the activation of this via. Thus, it is proposed that the (PhSe)2-neuroprotective effect might be linked to its mitoprotective activity. PMID:24623265

  10. Arsenic Induces Insulin Resistance in Mouse Adipocytes and Myotubes Via Oxidative Stress-Regulated Mitochondrial Sirt3-FOXO3a Signaling Pathway.

    PubMed

    Padmaja Divya, Sasidharan; Pratheeshkumar, Poyil; Son, Young-Ok; Vinod Roy, Ram; Andrew Hitron, John; Kim, Donghern; Dai, Jin; Wang, Lei; Asha, Padmaja; Huang, Bin; Xu, Mei; Luo, Jia; Zhang, Zhuo

    2015-08-01

    Chronic exposure to arsenic via drinking water is associated with an increased risk for development of type 2 diabetes mellitus (T2DM). This study investigates the role of mitochondrial oxidative stress protein Sirtuin 3 (Sirt3) and its targeting proteins in chronic arsenic-induced T2DM in mouse adipocytes and myotubes. The results show that chronic arsenic exposure significantly decreased insulin-stimulated glucose uptake (ISGU) in correlation with reduced expression of insulin-regulated glucose transporter type 4 (Glut4). Expression of Sirt3, a mitochondrial deacetylase, was dramatically decreased along with its associated transcription factor, forkhead box O3 (FOXO3a) upon arsenic exposure. A decrease in mitochondrial membrane potential (Δψm) was observed in both 3T3L1 adipocytes and C2C12 myotubes treated by arsenic. Reduced FOXO3a activity by arsenic exhibited a decreased binding affinity to the promoters of both manganese superoxide dismutase (MnSOD) and peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1α, a broad and powerful regulator of reactive oxygen species (ROS) metabolism. Forced expression of Sirt3 or MnSOD in mouse myotubes elevated Δψm and restored ISGU inhibited by arsenic exposure. Our results suggest that Sirt3/FOXO3a/MnSOD signaling plays a significant role in the inhibition of ISGU induced by chronic arsenic exposure.

  11. DRP1 inhibition rescues retinal ganglion cells and their axons by preserving mitochondrial integrity in a mouse model of glaucoma.

    PubMed

    Kim, K-Y; Perkins, G A; Shim, M S; Bushong, E; Alcasid, N; Ju, S; Ellisman, M H; Weinreb, R N; Ju, W-K

    2015-01-01

    Glaucoma is the leading cause of irreversible blindness and is characterized by slow and progressive degeneration of the optic nerve head axons and retinal ganglion cell (RGC), leading to loss of visual function. Although oxidative stress and/or alteration of mitochondrial (mt) dynamics induced by elevated intraocular pressure (IOP) are associated with this neurodegenerative disease, the mechanisms that regulate mt dysfunction-mediated glaucomatous neurodegeneration are poorly understood. Using a mouse model of glaucoma, DBA/2J (D2), which spontaneously develops elevated IOP, as well as an in vitro RGC culture system, we show here that oxidative stress, as evidenced by increasing superoxide dismutase 2 (SOD2) and mt transcription factor A (Tfam) protein expression, triggers mt fission and loss by increasing dynamin-related protein 1 (DRP1) in the retina of glaucomatous D2 mice as well as in cultured RGCs exposed to elevated hydrostatic pressure in vitro. DRP1 inhibition by overexpressing DRP1 K38A mutant blocks mt fission and triggers a subsequent reduction of oxidative stress, as evidenced by decreasing SOD2 and Tfam protein expression. DRP1 inhibition promotes RGC survival by increasing phosphorylation of Bad at serine 112 in the retina and preserves RGC axons by maintaining mt integrity in the glial lamina of glaucomatous D2 mice. These findings demonstrate an important vicious cycle involved in glaucomatous neurodegeneration that starts with elevated IOP producing oxidative stress; the oxidative stress then leads to mt fission and a specific form of mt dysfunction that generates further oxidative stress, thus perpetuating the cycle. Our findings suggest that DRP1 is a potential therapeutic target for ameliorating oxidative stress-mediated mt fission and dysfunction in RGC and its axons during glaucomatous neurodegeneration. Thus, DRP1 inhibition may provide a new therapeutic strategy for protecting both RGCs and their axons in glaucoma and other optic

  12. Antioxidant treatment normalizes mitochondrial energetics and myocardial insulin sensitivity independently of changes in systemic metabolic homeostasis in a mouse model of the metabolic syndrome.

    PubMed

    Ilkun, Olesya; Wilde, Nicole; Tuinei, Joseph; Pires, Karla M P; Zhu, Yi; Bugger, Heiko; Soto, Jamie; Wayment, Benjamin; Olsen, Curtis; Litwin, Sheldon E; Abel, E Dale

    2015-08-01

    Cardiac dysfunction in obesity is associated with mitochondrial dysfunction, oxidative stress and altered insulin sensitivity. Whether oxidative stress directly contributes to myocardial insulin resistance remains to be determined. This study tested the hypothesis that ROS scavenging will improve mitochondrial function and insulin sensitivity in the hearts of rodent models with varying degrees of insulin resistance and hyperglycemia. The catalytic antioxidant MnTBAP was administered to the uncoupling protein-diphtheria toxin A (UCP-DTA) mouse model of insulin resistance (IR) and obesity, at early and late time points in the evolution of IR, and to db/db mice with severe obesity and type-two diabetes. Mitochondrial function was measured in saponin-permeabilized cardiac fibers. Aconitase activity and hydrogen peroxide emission were measured in isolated mitochondria. Insulin-stimulated glucose oxidation, glycolysis and fatty acid oxidation rates were measured in isolated working hearts, and 2-deoxyglucose uptake was measured in isolated cardiomyocytes. Four weeks of MnTBAP attenuated glucose intolerance in 13-week-old UCP-DTA mice but was without effect in 24-week-old UCP-DTA mice and in db/db mice. Despite the absence of improvement in the systemic metabolic milieu, MnTBAP reversed cardiac mitochondrial oxidative stress and improved mitochondrial bioenergetics by increasing ATP generation and reducing mitochondrial uncoupling in all models. MnTBAP also improved myocardial insulin mediated glucose metabolism in 13 and 24-week-old UCP-DTA mice. Pharmacological ROS scavenging improves myocardial energy metabolism and insulin responsiveness in obesity and type 2 diabetes via direct effects that might be independent of changes in systemic metabolism. PMID:26004364

  13. Preservation of Cognitive Function by Lepidium meyenii (Maca) Is Associated with Improvement of Mitochondrial Activity and Upregulation of Autophagy-Related Proteins in Middle-Aged Mouse Cortex

    PubMed Central

    Guo, Shan-Shan; Gao, Xiao-Fang; Gu, Yan-Rong

    2016-01-01

    Maca has been used as a foodstuff and a traditional medicine in the Andean region for over 2,000 years. Recently the neuroprotective effects of maca also arouse interest of researchers. Decrease in mitochondrial function and decline in autophagy signaling may participate in the process of age-related cognitive decline. This study aimed to investigate if maca could improve cognitive function of middle-aged mice and if this effect was associated with improvement of mitochondrial activity and modulation of autophagy signaling in mouse cortex. Fourteen-month-old male ICR mice received maca powder administered by gavage for five weeks. Maca improved cognitive function, motor coordination, and endurance capacity in middle-aged mice, accompanied by increased mitochondrial respiratory function and upregulation of autophagy-related proteins in cortex. Our findings suggest that maca is a newly defined nutritional plant which can improve mitochondrial function and upregulate autophagy-related proteins and may be an effective functional food for slowing down age-related cognitive decline. PMID:27648102

  14. PGC-1alpha plays a functional role in exercise-induced mitochondrial biogenesis and angiogenesis but not fiber-type transformation in mouse skeletal muscle.

    PubMed

    Geng, Tuoyu; Li, Ping; Okutsu, Mitsuharu; Yin, Xinhe; Kwek, Jyeyi; Zhang, Mei; Yan, Zhen

    2010-03-01

    Endurance exercise stimulates peroxisome proliferator-activated receptor gamma coactivator-1alpha (PGC-1alpha) expression in skeletal muscle, and forced expression of PGC-1alpha changes muscle metabolism and exercise capacity in mice. However, it is unclear if PGC-1alpha is indispensible for endurance exercise-induced metabolic and contractile adaptations in skeletal muscle. In this study, we showed that endurance exercise-induced expression of mitochondrial enzymes (cytochrome oxidase IV and cytochrome c) and increases of platelet endothelial cell adhesion molecule-1 (PECAM-1, CD31)-positive endothelial cells in skeletal muscle, but not IIb-to-IIa fiber-type transformation, were significantly attenuated in muscle-specific Pgc-1alpha knockout mice. Interestingly, voluntary running effectively restored the compromised mitochondrial integrity and superoxide dismutase 2 (SOD2) protein expression in skeletal muscle in Pgc-1alpha knockout mice. Thus, PGC-1alpha plays a functional role in endurance exercise-induced mitochondrial biogenesis and angiogenesis, but not IIb-to-IIa fiber-type transformation in mouse skeletal muscle, and the improvement of mitochondrial morphology and antioxidant defense in response to endurance exercise may occur independently of PGC-1alpha function. We conclude that PGC-1alpha is required for complete skeletal muscle adaptations induced by endurance exercise in mice. PMID:20032509

  15. Preservation of Cognitive Function by Lepidium meyenii (Maca) Is Associated with Improvement of Mitochondrial Activity and Upregulation of Autophagy-Related Proteins in Middle-Aged Mouse Cortex

    PubMed Central

    Guo, Shan-Shan; Gao, Xiao-Fang; Gu, Yan-Rong

    2016-01-01

    Maca has been used as a foodstuff and a traditional medicine in the Andean region for over 2,000 years. Recently the neuroprotective effects of maca also arouse interest of researchers. Decrease in mitochondrial function and decline in autophagy signaling may participate in the process of age-related cognitive decline. This study aimed to investigate if maca could improve cognitive function of middle-aged mice and if this effect was associated with improvement of mitochondrial activity and modulation of autophagy signaling in mouse cortex. Fourteen-month-old male ICR mice received maca powder administered by gavage for five weeks. Maca improved cognitive function, motor coordination, and endurance capacity in middle-aged mice, accompanied by increased mitochondrial respiratory function and upregulation of autophagy-related proteins in cortex. Our findings suggest that maca is a newly defined nutritional plant which can improve mitochondrial function and upregulate autophagy-related proteins and may be an effective functional food for slowing down age-related cognitive decline.

  16. Behavioral stress causes mitochondrial dysfunction via ABAD up-regulation and aggravates plaque pathology in the brain of a mouse model of Alzheimer disease.

    PubMed

    Seo, Ji-Seon; Lee, Kang-Woo; Kim, Tae-Kyung; Baek, In-Sun; Im, Joo-Young; Han, Pyung-Lim

    2011-06-01

    Basic and clinical studies have reported that behavioral stress worsens the pathology of Alzheimer disease (AD), but the underlying mechanism has not been clearly understood. In this study, we determined the mechanism by which behavioral stress affects the pathogenesis of AD using Tg-APPswe/PS1dE9 mice, a murine model of AD. Tg-APPswe/PS1dE9 mice that were restrained for 2h daily for 16 consecutive days (2-h/16-day stress) from 6.5months of age had significantly increased Aβ(1-42) levels and plaque deposition in the brain. The 2-h/16-day stress increased oxidative stress and induced mitochondrial dysfunction in the brain. Treatment with glucocorticoid (corticosterone) and Aβ in SH-SY5Y cells increased the expression of 17β-hydroxysteroid dehydrogenase (ABAD), mitochondrial dysfunction, and levels of ROS, whereas blockade of ABAD expression by siRNA-ABAD in SH-SY5Y cells suppressed glucocorticoid-enhanced mitochondrial dysfunction and ROS accumulation. The 2-h/16-day stress up-regulated ABAD expression in mitochondria in the brain of Tg-APPswe/PS1dE9 mice. Moreover, all visible Aβ plaques were costained with anti-ABAD in the brains of Tg-APPswe/PS1dE9 mice. Together, these results suggest that behavioral stress aggravates plaque pathology and mitochondrial dysfunction via up-regulation of ABAD in the brain of a mouse model of AD.

  17. Preservation of Cognitive Function by Lepidium meyenii (Maca) Is Associated with Improvement of Mitochondrial Activity and Upregulation of Autophagy-Related Proteins in Middle-Aged Mouse Cortex.

    PubMed

    Guo, Shan-Shan; Gao, Xiao-Fang; Gu, Yan-Rong; Wan, Zhong-Xiao; Lu, A-Ming; Qin, Zheng-Hong; Luo, Li

    2016-01-01

    Maca has been used as a foodstuff and a traditional medicine in the Andean region for over 2,000 years. Recently the neuroprotective effects of maca also arouse interest of researchers. Decrease in mitochondrial function and decline in autophagy signaling may participate in the process of age-related cognitive decline. This study aimed to investigate if maca could improve cognitive function of middle-aged mice and if this effect was associated with improvement of mitochondrial activity and modulation of autophagy signaling in mouse cortex. Fourteen-month-old male ICR mice received maca powder administered by gavage for five weeks. Maca improved cognitive function, motor coordination, and endurance capacity in middle-aged mice, accompanied by increased mitochondrial respiratory function and upregulation of autophagy-related proteins in cortex. Our findings suggest that maca is a newly defined nutritional plant which can improve mitochondrial function and upregulate autophagy-related proteins and may be an effective functional food for slowing down age-related cognitive decline. PMID:27648102

  18. The N-Terminal Peptides of the Three Human Isoforms of the Mitochondrial Voltage-Dependent Anion Channel Have Different Helical Propensities.

    PubMed

    Guardiani, Carlo; Scorciapino, Mariano Andrea; Amodeo, Giuseppe Federico; Grdadolnik, Joze; Pappalardo, Giuseppe; De Pinto, Vito; Ceccarelli, Matteo; Casu, Mariano

    2015-09-15

    The voltage-dependent anion channel (VDAC) is the main mitochondrial porin allowing the exchange of ions and metabolites between the cytosol and the mitochondrion. In addition, VDAC was found to actively interact with proteins playing a fundamental role in the regulation of apoptosis and being of central interest in cancer research. VDAC is a large transmembrane β-barrel channel, whose N-terminal helical fragment adheres to the channel interior, partially closing the pore. This fragment is considered to play a key role in protein stability and function as well as in the interaction with apoptosis-related proteins. Three VDAC isoforms are differently expressed in higher eukaryotes, for which distinct and complementary roles are proposed. In this work, the folding propensity of their N-terminal fragments has been compared. By using multiple spectroscopic techniques, and complementing the experimental results with theoretical computer-assisted approaches, we have characterized their conformational equilibrium. Significant differences were found in the intrinsic helical propensity of the three peptides, decreasing in the following order: hVDAC2 > hVDAC3 > hVDAC1. In light of the models proposed in the literature to explain voltage gating, selectivity, and permeability, as well as interactions with functionally related proteins, our results suggest that the different chemicophysical properties of the N-terminal domain are possibly correlated to different functions for the three isoforms. The overall emerging picture is that a similar transmembrane water accessible conduit has been equipped with not identical domains, whose differences can modulate the functional roles of the three VDAC isoforms.

  19. Developmental regulation of mitochondrial biogenesis and function in the mouse mammary gland during a prolonged lactation cycle

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The regulation of mitochondrial biogenesis and function in the lactating mammary cell is poorly understood. The goal of this study was to use proteomics to relate temporal changes in mammary cell mitochondrial function during lactation to changes in the proteins that make up this organelle. The hypo...

  20. Lack of GDAP1 Induces Neuronal Calcium and Mitochondrial Defects in a Knockout Mouse Model of Charcot-Marie-Tooth Neuropathy

    PubMed Central

    Civera-Tregón, Azahara; Yndriago, Laura; Pla-Martin, David; Zenker, Jennifer; Cuevas-Martín, Carmen; Estela, Anna; Sánchez-Aragó, María; Forteza-Vila, Jerónimo; Cuezva, José M.; Chrast, Roman; Palau, Francesc

    2015-01-01

    Mutations in GDAP1, which encodes protein located in the mitochondrial outer membrane, cause axonal recessive (AR-CMT2), axonal dominant (CMT2K) and demyelinating recessive (CMT4A) forms of Charcot-Marie-Tooth (CMT) neuropathy. Loss of function recessive mutations in GDAP1 are associated with decreased mitochondrial fission activity, while dominant mutations result in impairment of mitochondrial fusion with increased production of reactive oxygen species and susceptibility to apoptotic stimuli. GDAP1 silencing in vitro reduces Ca2+ inflow through store-operated Ca2+ entry (SOCE) upon mobilization of endoplasmic reticulum (ER) Ca2+, likely in association with an abnormal distribution of the mitochondrial network. To investigate the functional consequences of lack of GDAP1 in vivo, we generated a Gdap1 knockout mouse. The affected animals presented abnormal motor behavior starting at the age of 3 months. Electrophysiological and biochemical studies confirmed the axonal nature of the neuropathy whereas histopathological studies over time showed progressive loss of motor neurons (MNs) in the anterior horn of the spinal cord and defects in neuromuscular junctions. Analyses of cultured embryonic MNs and adult dorsal root ganglia neurons from affected animals demonstrated large and defective mitochondria, changes in the ER cisternae, reduced acetylation of cytoskeletal α-tubulin and increased autophagy vesicles. Importantly, MNs showed reduced cytosolic calcium and SOCE response. The development and characterization of the GDAP1 neuropathy mice model thus revealed that some of the pathophysiological changes present in axonal recessive form of the GDAP1-related CMT might be the consequence of changes in the mitochondrial network biology and mitochondria–endoplasmic reticulum interaction leading to abnormalities in calcium homeostasis. PMID:25860513

  1. Decreased ovarian reserve, dysregulation of mitochondrial biogenesis, and increased lipid peroxidation in female mouse offspring exposed to an obesogenic maternal diet

    PubMed Central

    Aiken, Catherine E.; Tarry-Adkins, Jane L.; Penfold, Naomi C.; Dearden, Laura; Ozanne, Susan E.

    2016-01-01

    Maternal diet during pregnancy influences the later life reproductive potential of female offspring. We investigate the molecular mechanisms underlying the depletion of ovarian follicular reserve in young adult females following exposure to obesogenic diet in early life. Furthermore, we explore the interaction between adverse maternal diet and postweaning diet in generating reduced ovarian reserve. Female mice were exposed to either maternal obesogenic (high fat/high sugar) or maternal control diet in utero and during lactation, then weaned onto either obesogenic or control diet. At 12 wk of age, the offspring ovarian reserve was depleted following exposure to maternal obesogenic diet (P < 0.05), but not postweaning obesogenic diet. Maternal obesogenic diet was associated with increased mitochondrial DNA biogenesis (copy number P < 0.05; transcription factor A, mitochondrial expression P < 0.05), increased mitochondrial antioxidant defenses [manganese superoxide dismutase (MnSOD) P < 0.05; copper/zinc superoxide dismutase P < 0.05; glutathione peroxidase 4 P < 0.01] and increased lipoxygenase expression (arachidonate 12-lipoxygenase P < 0.05; arachidonate 15-lipoxygenase P < 0.05) in the ovary. There was also significantly increased expression of the transcriptional regulator NF-κB (P < 0.05). There was no effect of postweaning diet on any measured ovarian parameters. Maternal diet thus plays a central role in determining follicular reserve in adult female offspring. Our observations suggest that lipid peroxidation and mitochondrial biogenesis are the key intracellular pathways involved in programming of ovarian reserve.—Aiken, C. E., Tarry-Adkins, J. L., Penfold, N. C., Dearden, L., Ozanne, S. E. Decreased ovarian reserve, dysregulation of mitochondrial biogenesis, and increased lipid peroxidation in female mouse offspring exposed to an obesogenic maternal diet. PMID:26700734

  2. Hydrogen Sulfide Epigenetically Attenuates Homocysteine-Induced Mitochondrial Toxicity Mediated Through NMDA Receptor in Mouse Brain Endothelial (bEnd3) Cells.

    PubMed

    Kamat, Pradip K; Kalani, Anuradha; Tyagi, Suresh C; Tyagi, Neetu

    2015-02-01

    Previously we have shown that homocysteine (Hcy) caused oxidative stress and altered mitochondrial function. Hydrogen sulfide (H2S) has potent anti-inflammatory, anti-oxidative, and anti-apoptotic effects. Therefore, in the present study we examined whether H2S ameliorates Hcy-induced mitochondrial toxicity which led to endothelial dysfunction in part, by epigenetic alterations in mouse brain endothelial cells (bEnd3). The bEnd3 cells were exposed to 100 μM Hcy treatment in the presence or absence of 30 μM NaHS (donor of H2S) for 24 h. Hcy-activate NMDA receptor and induced mitochondrial toxicity by increased levels of Ca(2+), NADPH-oxidase-4 (NOX-4) expression, mitochondrial dehydrogenase activity and decreased the level of nitrate, superoxide dismutase (SOD-2) expression, mitochondria membrane potentials, ATP production. To confirm the role of epigenetic, 5'-azacitidine (an epigenetic modulator) treatment was given to the cells. Pretreatment with NaHS (30 μM) attenuated the Hcy-induced increased expression of DNMT1, DNMT3a, Ca(2+), and decreased expression of DNMT3b in bEND3 cells. Furthermore, NaHS treatment also mitigated mitochondrial oxidative stress (NOX4, ROS, and NO) and restored ATP that indicates its protective effects against mitochondrial toxicity. Additional, NaHS significantly alleviated Hcy-induced LC3-I/II, CSE, Atg3/7, and low p62 expression which confirm its effect on mitophagy. Likewise, NaHS also restored level of eNOS, CD31, VE-cadherin and ET-1 and maintains endothelial function in Hcy treated cells. Molecular inhibition of NMDA receptor by using small interfering RNA showed protective effect whereas inhibition of H2S production by propargylglycine (PG) (inhibitor of enzyme CSE) showed mitotoxic effect. Taken together, results demonstrate that, administration of H2S protected the cells from HHcy-induced mitochondrial toxicity and endothelial dysfunction.

  3. Hydrogen sulfide epigenetically attenuates homocysteine-induced mitochondrial toxicity mediated through NMDA receptor in mouse brain endothelial (bEnd3) cells†

    PubMed Central

    Kamat, Pradip K.; Kalani, Anuradha; Tyagi, Suresh C.; Tyagi, Neetu

    2014-01-01

    Previously we have showed that homocysteine (Hcy) caused oxidative stress and altered mitochondrial function. Hydrogen sulphide (H2S) has potent anti-inflammatory, anti-oxidative and anti-apoptotic effects. Therefore, in the present study we examined whether H2S ameliorates Hcy-induced mitochondrial toxicity which led to endothelial dysfunction in part, by epigenetic alterations in mouse brain endothelial cells (bEnd3). The bEnd3 cells were exposed to 100μM Hcy treatment in the presence or absence of 30μM NaHS (donor of H2S) for 24hrs. Hcy-activate NMDA receptor and induced mitochondrial toxicity by increased levels of Ca2+, NADPH-oxidase-4 (NOX-4) expression, mitochondrial dehydrogenase activity and decreased the level of nitrate, superoxide dismutase (SOD-2) expression, mitochondria membrane potentials, ATP production. To confirm the role of epigenetic, 5′-azacitidine (an epigenetic modulator) treatment was given to the cells. Pretreatment with NaHS (30μM) attenuated the Hcy-induced increased expression of DNMT1, DNMT3a, Ca2+ and decreased expression of DNMT3b in bEND3 cells. Furthermore, NaHS treatment also enhanced mitochondrial oxidative stress (NOX4, ROS, and NO) and restored ATP that indicates its protective effects against mitochondrial toxicity. Additional, NaHS significantly alleviated Hcy-induced LC3-I/II, CSE, Atg3/7 and low p62 expression which confirm its effect on mitophagy. Likewise, NaHS also restored level of eNOS, CD31, VE-Cadherin and ET-1 and maintains endothelial function in Hcy treated cells. Molecular inhibition of NMDA receptor by using small interfering RNA showed protective effect whereas inhibition of H2S production by propargylglycine (PG) (inhibitor of enzyme CSE) showed mitotoxic effect. Taken together, results demonstrate that, administration of H2S protected the cells from HHcy-induced mitochondrial toxicity and endothelial dysfunction. PMID:25056869

  4. Hydrogen Sulfide Epigenetically Attenuates Homocysteine-Induced Mitochondrial Toxicity Mediated Through NMDA Receptor in Mouse Brain Endothelial (bEnd3) Cells.

    PubMed

    Kamat, Pradip K; Kalani, Anuradha; Tyagi, Suresh C; Tyagi, Neetu

    2015-02-01

    Previously we have shown that homocysteine (Hcy) caused oxidative stress and altered mitochondrial function. Hydrogen sulfide (H2S) has potent anti-inflammatory, anti-oxidative, and anti-apoptotic effects. Therefore, in the present study we examined whether H2S ameliorates Hcy-induced mitochondrial toxicity which led to endothelial dysfunction in part, by epigenetic alterations in mouse brain endothelial cells (bEnd3). The bEnd3 cells were exposed to 100 μM Hcy treatment in the presence or absence of 30 μM NaHS (donor of H2S) for 24 h. Hcy-activate NMDA receptor and induced mitochondrial toxicity by increased levels of Ca(2+), NADPH-oxidase-4 (NOX-4) expression, mitochondrial dehydrogenase activity and decreased the level of nitrate, superoxide dismutase (SOD-2) expression, mitochondria membrane potentials, ATP production. To confirm the role of epigenetic, 5'-azacitidine (an epigenetic modulator) treatment was given to the cells. Pretreatment with NaHS (30 μM) attenuated the Hcy-induced increased expression of DNMT1, DNMT3a, Ca(2+), and decreased expression of DNMT3b in bEND3 cells. Furthermore, NaHS treatment also mitigated mitochondrial oxidative stress (NOX4, ROS, and NO) and restored ATP that indicates its protective effects against mitochondrial toxicity. Additional, NaHS significantly alleviated Hcy-induced LC3-I/II, CSE, Atg3/7, and low p62 expression which confirm its effect on mitophagy. Likewise, NaHS also restored level of eNOS, CD31, VE-cadherin and ET-1 and maintains endothelial function in Hcy treated cells. Molecular inhibition of NMDA receptor by using small interfering RNA showed protective effect whereas inhibition of H2S production by propargylglycine (PG) (inhibitor of enzyme CSE) showed mitotoxic effect. Taken together, results demonstrate that, administration of H2S protected the cells from HHcy-induced mitochondrial toxicity and endothelial dysfunction. PMID:25056869

  5. Diallyl disulfide attenuated carbon ion irradiation-induced apoptosis in mouse testis through changing the ratio of Tap73/ΔNp73 via mitochondrial pathway.

    PubMed

    Di, Cui-xia; Han, Lu; Zhang, Hong; Xu, Shuai; Mao, Ai-hong; Sun, Chao; Liu, Yang; Si, Jing; Li, Hong-yan; Zhou, Xin; Liu, Bing; Miao, Guo-ying

    2015-01-01

    Diallyl disulfide (DADS), a major organosulfur compound derived from garlic, has various biological properties, including anti-cancer effects. However, the protective mechanism of DADS against radiation-induced mouse testis cell apoptosis has not been elucidated. In this study, the magnitude of radiation effects evoked by carbon ion irradiation was marked by morphology changes, significant rise in apoptotic cells, activation expression of p53, up regulation the ratio of pro-apoptotic Tap73/anti-apoptotic ΔNp73, as well as alterations of crucial mediator of the mitochondrial pathway. Interestingly, pretreatment with DADS attenuated carbon ion irradiation-induced morphology damages and apoptotic cells. Additionally, DADS elevated radiation-induced p53 and p21 expression, suggesting that p53 might be involved in the inhibition of cell cycle progression through up regulation of p21. Furthermore, administration with DADS prevented radiation-induced Tap73/ΔNp73 expression and consequently down regulated Bax/Bcl-2 ratio, cytochrome c release and caspase-3 expression, indicating that the balance between Tap73 and ΔNp73 had potential to activate p53 responsive genes. Thus, our results showed that radio protection effect of DADS on mouse testis is mediated by blocking apoptosis through changing the ratio of Tap73/ΔNp73 via mitochondrial pathway, suggesting that DADS could be used as a potential radio protection agent for the testis against heavy-ion radiation.

  6. Computer prediction of peptide maps: assignment of polypeptides to human and mouse mitochondrial DNA genes by analysis of two-dimensional-proteolytic digest gels.

    PubMed

    Wallace, D C; Yang, J H; Ye, J H; Lott, M T; Oliver, N A; McCarthy, J

    1986-04-01

    We have prepared a computer program that predicts complete and partial peptide maps from amino acid sequences. The program fragments amino acid sequences at designated cleavage sites and calculates the molecular weight and relative labeling of each peptide. These data are graphed as log molecular weight of the original protein (X-axis) vs. log molecular weight of the component peptides (Y-axis). The program is interactive, permitting adjustment of a number of graphic parameters and alteration of the position of proteins in the first dimension to accommodate aberrations in protein mobility. The program has been used to predict the V8 protease peptide maps of the 13 open reading frames (ORFs) identified in the human and the mouse mitochondrial DNA (mtDNA) sequences. The results were compared to the V8 protease peptide maps obtained for mouse and human mitochondrially synthesized proteins by two-dimensional proteolytic digest gels. A high correlation was observed between the predicted and observed peptide maps. These results suggest the assignment of several proteins to mtDNA genes.

  7. Differential Gene Expression Reveals Mitochondrial Dysfunction in an Imprinting Center Deletion Mouse Model of Prader-Willi Syndrome

    PubMed Central

    Fan, Weiwei; Coskun, Pinar E.; Nalbandian, Angèle; Knoblach, Susan; Resnick, James L.; Hoffman, Eric; Wallace, Douglas C.; Kimonis, Virginia E.

    2013-01-01

    Prader-Willi syndrome (PWS) is a genetic disorder caused by deficiency of imprinted gene expression from the paternal chromosome 15q11-15q13 and clinically characterized by neonatal hypotonia, short stature, cognitive impairment, hypogonadism, hyperphagia, morbid obesity and diabetes. Previous clinical studies suggest that a defect in energy metabolism may be involved in the pathogenesis of PWS. We focused our attention on the genes associated with energy metabolism and found that there were 95 and 66 mitochondrial genes differentially expressed in PWS muscle and brain, respectively. Assessment of enzyme activities of mitochondrial oxidative phosphorylation (OXPHOS) complexes in the brain, heart, liver and muscle were assessed. We found the enzyme activities of the cardiac mitochondrial complexes II+III were upregulated in the imprinting center deletion (PWS-IC) mice compared to the wild type littermates. These studies suggest that differential gene expression, especially of the mitochondrial genes may contribute to the pathophysiology of PWS. PMID:24127921

  8. Analysis of the mouse mutant Cloth-ears shows a role for the voltage-gated sodium channel Scn8a in peripheral neural hearing loss.

    PubMed

    Mackenzie, F E; Parker, A; Parkinson, N J; Oliver, P L; Brooker, D; Underhill, P; Lukashkina, V A; Lukashkin, A N; Holmes, C; Brown, S D M

    2009-10-01

    Deafness is the most common sensory disorder in humans and the aetiology of genetic deafness is complex. Mouse mutants have been crucial in identifying genes involved in hearing. However, many deafness genes remain unidentified. Using N-ethyl N-nitrosourea (ENU) mutagenesis to generate new mouse models of deafness, we identified a novel semi-dominant mouse mutant, Cloth-ears (Clth). Cloth-ears mice show reduced acoustic startle response and mild hearing loss from approximately 30 days old. Auditory-evoked brainstem response (ABR) and distortion product otoacoustic emission (DPOAE) analyses indicate that the peripheral neural auditory pathway is impaired in Cloth-ears mice, but that cochlear function is normal. In addition, both Clth/Clth and Clth/+ mice display paroxysmal tremor episodes with behavioural arrest. Clth/Clth mice also show a milder continuous tremor during movement and rest. Longitudinal phenotypic analysis showed that Clth/+ and Clth/Clth mice also have complex defects in behaviour, growth, neurological and motor function. Positional cloning of Cloth-ears identified a point mutation in the neuronal voltage-gated sodium channel alpha-subunit gene, Scn8a, causing an aspartic acid to valine (D981V) change six amino acids downstream of the sixth transmembrane segment of the second domain (D2S6). Complementation testing with a known Scn8a mouse mutant confirmed that this mutation is responsible for the Cloth-ears phenotype. Our findings suggest a novel role for Scn8a in peripheral neural hearing loss and paroxysmal motor dysfunction. PMID:19737145

  9. Of mice and the 'Age of Discovery': the complex history of colonization of the Azorean archipelago by the house mouse (Mus musculus) as revealed by mitochondrial DNA variation.

    PubMed

    Gabriel, S I; Mathias, M L; Searle, J B

    2015-01-01

    Humans have introduced many species onto remote oceanic islands. The house mouse (Mus musculus) is a human commensal and has consequently been transported to oceanic islands around the globe as an accidental stowaway. The history of these introductions can tell us not only about the mice themselves but also about the people that transported them. Following a phylogeographic approach, we used mitochondrial D-loop sequence variation (within an 849- to 864-bp fragment) to study house mouse colonization of the Azores. A total of 239 sequences were obtained from all nine islands, and interpretation was helped by previously published Iberian sequences and 66 newly generated Spanish sequences. A Bayesian analysis revealed presence in the Azores of most of the D-loop clades previously described in the domesticus subspecies of the house mouse, suggesting a complex colonization history of the archipelago as a whole from multiple geographical origins, but much less heterogeneity (often single colonization?) within islands. The expected historical link with mainland Portugal was reflected in the pattern of D-loop variation of some of the islands but not all. A more unexpected association with a distant North European source area was also detected in three islands, possibly reflecting human contact with the Azores prior to the 15th century discovery by Portuguese mariners. Widening the scope to colonization of the Macaronesian islands as a whole, human linkages between the Azores, Madeira, the Canaries, Portugal and Spain were revealed through the sharing of mouse sequences between these areas. From these and other data, we suggest mouse studies may help resolve historical uncertainties relating to the 'Age of Discovery'.

  10. Administration of CoQ10 analogue ameliorates dysfunction of the mitochondrial respiratory chain in a mouse model of Angelman syndrome.

    PubMed

    Llewellyn, Katrina J; Nalbandian, Angèle; Gomez, Arianna; Wei, Don; Walker, Naomi; Kimonis, Virginia E

    2015-04-01

    Genetic defects in the UBE3A gene, which encodes for the imprinted E6-AP ubiquitin E3 ligase (UBE3A), is responsible for the occurrence of Angelman syndrome (AS), a neurodegenerative disorder which arises in 1 out of every 12,000-20,000 births. Classical symptoms of AS include delayed development, impaired speech, and epileptic seizures with characteristic electroencephalography (EEG) readings. We have previously reported impaired mitochondrial structure and reduced complex III in the hippocampus and cerebellum in the Ube3a(m-/p+) mice. CoQ10 supplementation restores the electron flow to the mitochondrial respiratory chain (MRC) to ultimately increase mitochondrial antioxidant capacity. A number of recent studies with CoQ10 analogues seem promising in providing therapeutic benefit to patients with a variety of disorders. CoQ10 therapy has been reported to be safe and relatively well-tolerated at doses as high as 3000mg/day in patients with disorders of CoQ10 biosynthesis and MRC disorders. Herein, we report administration of idebenone, a potent CoQ10 analogue, to the Ube3a(m-/p+) mouse model corrects motor coordination and anxiety levels, and also improves the expression of complexes III and IV in hippocampus CA1 and CA2 neurons and cerebellum in these Ube3a(m-/p+) mice. However, treatment with idebenone illustrated no beneficial effects in the reduction of oxidative stress. To our knowledge, this is the first study to suggest an improvement in mitochondrial respiratory chain dysfunction via bioenergetics modulation with a CoQ10 analogue. These findings may further elucidate possible cellular and molecular mechanism(s) and ultimately a clinical therapeutic approach/benefit for patients with Angelman syndrome. PMID:25684537

  11. Underlying mitochondrial dysfunction triggers flutamide-induced oxidative liver injury in a mouse model of idiosyncratic drug toxicity

    SciTech Connect

    Kashimshetty, Rohini; Desai, Varsha G.; Kale, Vijay M.; Lee, Taewon; Moland, Carrie L.; Branham, William S.; New, Lee S.; Chan, Eric C.Y.; Younis, Husam; Boelsterli, Urs A.

    2009-07-15

    Flutamide, a widely used nonsteroidal anti-androgen, but not its bioisostere bicalutamide, has been associated with idiosyncratic drug-induced liver injury. Although the susceptibility factors are unknown, mitochondrial injury has emerged as a putative hazard of flutamide. To explore the role of mitochondrial sensitization in flutamide hepatotoxicity, we determined the effects of superimposed drug stress in a murine model of underlying mitochondrial abnormalities. Male wild-type or heterozygous Sod2{sup +/-} mice were injected intraperitoneously with flutamide (0, 30 or 100 mg/kg/day) for 28 days. A kinetic pilot study revealed that flutamide (100 mg/kg/day) caused approximately 10-fold greater exposure than the reported therapeutic mean plasma levels. Mutant (5/10), but not wild-type, mice in the high-dose group exhibited small foci of hepatocellular necrosis and an increased number of apoptotic hepatocytes. Hepatic GSSG/GSH, protein carbonyl levels, and serum lactate levels were significantly increased, suggesting oxidant stress and mitochondrial dysfunction. Measurement of mitochondrial superoxide in cultured hepatocytes demonstrated that mitochondria were a significant source of flutamide-enhanced oxidant stress. Indeed, mitochondria isolated from flutamide-treated Sod2{sup +/-} mice exhibited decreased aconitase activity as compared to vehicle controls. A transcriptomics analysis using MitoChips revealed that flutamide-treated Sod2{sup +/-} mice exhibited a selective decrease in the expression of all complexes I and III subunits encoded by mitochondrial DNA. In contrast, Sod2{sup +/-} mice receiving bicalutamide (50 mg/kg/day) did not reveal any hepatic changes. These results are compatible with our concept that flutamide targets hepatic mitochondria and exerts oxidant stress that can lead to overt hepatic injury in the presence of an underlying mitochondrial abnormality.

  12. Unchanged mitochondrial organization and compartmentation of high-energy phosphates in creatine-deficient GAMT−/− mouse hearts

    PubMed Central

    Branovets, Jelena; Sepp, Mervi; Kotlyarova, Svetlana; Jepihhina, Natalja; Sokolova, Niina; Aksentijevic, Dunja; Lygate, Craig A.; Neubauer, Stefan; Birkedal, Rikke

    2013-01-01

    Disruption of the creatine kinase (CK) system in hearts of CK-deficient mice leads to changes in the ultrastructure and regulation of mitochondrial respiration. We expected to see similar changes in creatine-deficient mice, which lack the enzyme guanidinoacetate methyltransferase (GAMT) to produce creatine. The aim of this study was to characterize the changes in cardiomyocyte mitochondrial organization, regulation of respiration, and intracellular compartmentation associated with GAMT deficiency. Three-dimensional mitochondrial organization was assessed by confocal microscopy. On populations of permeabilized cardiomyocytes, we recorded ADP and ATP kinetics of respiration, competition between mitochondria and pyruvate kinase for ADP produced by ATPases, ADP kinetics of endogenous pyruvate kinase, and ATP kinetics of ATPases. These data were analyzed by mathematical models to estimate intracellular compartmentation. Quantitative analysis of morphological and kinetic data as well as derived model fits showed no difference between GAMT-deficient and wild-type mice. We conclude that inactivation of the CK system by GAMT deficiency does not alter mitochondrial organization and intracellular compartmentation in relaxed cardiomyocytes. Thus, our results suggest that the healthy heart is able to preserve cardiac function at a basal level in the absence of CK-facilitated energy transfer without compromising intracellular organization and the regulation of mitochondrial energy homeostasis. This raises questions on the importance of the CK system as a spatial energy buffer in unstressed cardiomyocytes. PMID:23792673

  13. Unchanged mitochondrial organization and compartmentation of high-energy phosphates in creatine-deficient GAMT-/- mouse hearts.

    PubMed

    Branovets, Jelena; Sepp, Mervi; Kotlyarova, Svetlana; Jepihhina, Natalja; Sokolova, Niina; Aksentijevic, Dunja; Lygate, Craig A; Neubauer, Stefan; Vendelin, Marko; Birkedal, Rikke

    2013-08-15

    Disruption of the creatine kinase (CK) system in hearts of CK-deficient mice leads to changes in the ultrastructure and regulation of mitochondrial respiration. We expected to see similar changes in creatine-deficient mice, which lack the enzyme guanidinoacetate methyltransferase (GAMT) to produce creatine. The aim of this study was to characterize the changes in cardiomyocyte mitochondrial organization, regulation of respiration, and intracellular compartmentation associated with GAMT deficiency. Three-dimensional mitochondrial organization was assessed by confocal microscopy. On populations of permeabilized cardiomyocytes, we recorded ADP and ATP kinetics of respiration, competition between mitochondria and pyruvate kinase for ADP produced by ATPases, ADP kinetics of endogenous pyruvate kinase, and ATP kinetics of ATPases. These data were analyzed by mathematical models to estimate intracellular compartmentation. Quantitative analysis of morphological and kinetic data as well as derived model fits showed no difference between GAMT-deficient and wild-type mice. We conclude that inactivation of the CK system by GAMT deficiency does not alter mitochondrial organization and intracellular compartmentation in relaxed cardiomyocytes. Thus, our results suggest that the healthy heart is able to preserve cardiac function at a basal level in the absence of CK-facilitated energy transfer without compromising intracellular organization and the regulation of mitochondrial energy homeostasis. This raises questions on the importance of the CK system as a spatial energy buffer in unstressed cardiomyocytes. PMID:23792673

  14. A subtle alternative splicing event of the Na(V)1.8 voltage-gated sodium channel is conserved in human, rat, and mouse.

    PubMed

    Schirmeyer, Jana; Szafranski, Karol; Leipold, Enrico; Mawrin, Christian; Platzer, Matthias; Heinemann, Stefan H

    2010-06-01

    The voltage-gated sodium channel subtype Na(V)1.8 (SCN10A) is exclusively expressed in dorsal root ganglia (DRG) and plays a critical role in pain perception. We isolated mRNA from human, rat, and mouse DRGs and screened for alternatively spliced isoforms of the SCN10A mRNA using 454 sequencing. In all three species, we found an event of subtle alternative splicing at a NAGNAG tandem acceptor that results in isoforms including or lacking glutamine 1030 (Na(V)1.8+Q and Na(V)1.8-Q, respectively) within the cytoplasmic loop between domains II and III. The relative amount of Na(V)1.8-Q mRNA in adult DRG was measured with 14.1 +/- 0.1% in humans and 11.2 +/- 0.2% in rats. This is in contrast to an abundance of 64.3 +/- 0.3% in mouse DRG. Thus, the NAGNAG tandem acceptor in SCN10A is conserved among rodents and humans but its alternative usage apparently occurs with species-specific abundance. Analysis of human Na(V)1.8+Q and -Q isoforms in whole-cell patch-clamp experiments after heterologous expression in the neuroblastoma cell line Neuro-2A revealed no obvious impact of the splicing event on channel function. PMID:19953341

  15. Effect of angiotensin II-induced arterial hypertension on the voltage-dependent contractions of mouse arteries.

    PubMed

    Fransen, Paul; Van Hove, Cor E; Leloup, Arthur J A; Schrijvers, Dorien M; De Meyer, Guido R Y; De Keulenaer, Gilles W

    2016-02-01

    Arterial hypertension (AHT) affects the voltage dependency of L-type Ca(2+) channels in cardiomyocytes. We analyzed the effect of angiotensin II (AngII)-induced AHT on L-type Ca(2+) channel-mediated isometric contractions in conduit arteries. AHT was induced in C57Bl6 mice with AngII-filled osmotic mini-pumps (4 weeks). Normotensive mice treated with saline-filled osmotic mini-pumps were used for comparison. Voltage-dependent contractions mediated by L-type Ca(2+) channels were studied in vaso-reactive studies in vitro in isolated aortic and femoral arteries by using extracellular K(+) concentration-response (KDR) experiments. In aortic segments, AngII-induced AHT significantly sensitized isometric contractions induced by elevated extracellular K(+) and depolarization. This sensitization was partly prevented by normalizing blood pressure with hydralazine, suggesting that it was caused by AHT rather than by direct AngII effects on aortic smooth muscle cells. The EC50 for extracellular K(+) obtained in vitro correlated significantly with the rise in arterial blood pressure induced by AngII in vivo. The AHT-induced sensitization persisted when aortic segments were exposed to levcromakalim or to inhibitors of basal nitric oxide release. Consistent with these observations, AngII-treatment also sensitized the vaso-relaxing effects of the L-type Ca(2+) channel blocker diltiazem during K(+)-induced contractions. Unlike aorta, AngII-treatment desensitized the isometric contractions to depolarization in femoral arteries pointing to vascular bed specific responses of arteries to hypertension. AHT affects the voltage-dependent L-type Ca(2+) channel-mediated contraction of conduit arteries. This effect may contribute to the decreased vascular compliance in AHT and explain the efficacy of Ca(2+) channel blockers to reduce vascular stiffness and central blood pressure in AHT.

  16. N-Acetylcysteine improves mitochondrial function and ameliorates behavioral deficits in the R6/1 mouse model of Huntington's disease

    PubMed Central

    Wright, D J; Renoir, T; Smith, Z M; Frazier, A E; Francis, P S; Thorburn, D R; McGee, S L; Hannan, A J; Gray, L J

    2015-01-01

    Huntington's disease (HD) is a neurodegenerative disorder, involving psychiatric, cognitive and motor symptoms, caused by a CAG-repeat expansion encoding an extended polyglutamine tract in the huntingtin protein. Oxidative stress and excitotoxicity have previously been implicated in the pathogenesis of HD. We hypothesized that N-acetylcysteine (NAC) may reduce both excitotoxicity and oxidative stress through its actions on glutamate reuptake and antioxidant capacity. The R6/1 transgenic mouse model of HD was used to investigate the effects of NAC on HD pathology. It was found that chronic NAC administration delayed the onset and progression of motor deficits in R6/1 mice, while having an antidepressant-like effect on both R6/1 and wild-type mice. A deficit in the astrocytic glutamate transporter protein, GLT-1, was found in R6/1 mice. However, this deficit was not ameliorated by NAC, implying that the therapeutic effect of NAC is not due to rescue of the GLT-1 deficit and associated glutamate-induced excitotoxicity. Assessment of mitochondrial function in the striatum and cortex revealed that R6/1 mice show reduced mitochondrial respiratory capacity specific to the striatum. This deficit was rescued by chronic treatment with NAC. There was a selective increase in markers of oxidative damage in mitochondria, which was rescued by NAC. In conclusion, NAC is able to delay the onset of motor deficits in the R6/1 model of Huntington's disease and it may do so by ameliorating mitochondrial dysfunction. Thus, NAC shows promise as a potential therapeutic agent in HD. Furthermore, our data suggest that NAC may also have broader antidepressant efficacy. PMID:25562842

  17. N-Acetylcysteine improves mitochondrial function and ameliorates behavioral deficits in the R6/1 mouse model of Huntington's disease.

    PubMed

    Wright, D J; Renoir, T; Smith, Z M; Frazier, A E; Francis, P S; Thorburn, D R; McGee, S L; Hannan, A J; Gray, L J

    2015-01-01

    Huntington's disease (HD) is a neurodegenerative disorder, involving psychiatric, cognitive and motor symptoms, caused by a CAG-repeat expansion encoding an extended polyglutamine tract in the huntingtin protein. Oxidative stress and excitotoxicity have previously been implicated in the pathogenesis of HD. We hypothesized that N-acetylcysteine (NAC) may reduce both excitotoxicity and oxidative stress through its actions on glutamate reuptake and antioxidant capacity. The R6/1 transgenic mouse model of HD was used to investigate the effects of NAC on HD pathology. It was found that chronic NAC administration delayed the onset and progression of motor deficits in R6/1 mice, while having an antidepressant-like effect on both R6/1 and wild-type mice. A deficit in the astrocytic glutamate transporter protein, GLT-1, was found in R6/1 mice. However, this deficit was not ameliorated by NAC, implying that the therapeutic effect of NAC is not due to rescue of the GLT-1 deficit and associated glutamate-induced excitotoxicity. Assessment of mitochondrial function in the striatum and cortex revealed that R6/1 mice show reduced mitochondrial respiratory capacity specific to the striatum. This deficit was rescued by chronic treatment with NAC. There was a selective increase in markers of oxidative damage in mitochondria, which was rescued by NAC. In conclusion, NAC is able to delay the onset of motor deficits in the R6/1 model of Huntington's disease and it may do so by ameliorating mitochondrial dysfunction. Thus, NAC shows promise as a potential therapeutic agent in HD. Furthermore, our data suggest that NAC may also have broader antidepressant efficacy. PMID:25562842

  18. Apicidin induces endoplasmic reticulum stress- and mitochondrial dysfunction-associated apoptosis via phospholipase Cγ1- and Ca(2+)-dependent pathway in mouse Neuro-2a neuroblastoma cells.

    PubMed

    Choi, Ji Hyun; Lee, Jung Yeon; Choi, A-Young; Hwang, Keun-Young; Choe, Wonchae; Yoon, Kyung-Sik; Ha, Joohun; Yeo, Eui-Ju; Kang, Insug

    2012-12-01

    Apicidin, a fungal metabolite that functions as a histone deacetylase inhibitor, induces apoptosis in cancer cells. We investigated the molecular mechanisms of the anti-cancer effects of apicidin in mouse Neuro-2a neuroblastoma cells. Apicidin induced apoptotic cell death and activation of caspase-12, -9, and -3. Apicidin induced expression of endoplasmic reticulum (ER) stress-associated proteins, including CCAAT/enhancer binding protein homologous protein (CHOP), cleavage of activating transcription factor 6α, and phosphorylation of eukaryotic initiation factor 2α. Inhibition of ER stress by CHOP knockdown or using the ER stress inhibitors, salubrinal and 4-phenylbutyric acid, reduced apicidin-induced cell death. Apicidin induced reactive oxygen species accumulation and mitochondrial membrane potential loss. An antioxidant, N-acetyl cysteine, reduced apicidin-induced cell death, CHOP expression, and mitochondrial dysfunction. In addition, apicidin increased cytosolic Ca(2+), which was blocked by 2-aminoethoxydiphenyl borate, an antagonist of inositol 1,4,5-trisphosphate receptor, and BAPTA-AM, an intracellular Ca(2+) chelator. 2-Aminoethoxydiphenyl borate and BAPTA-AM inhibited apicidin-induced cell death and ER stress. Interestingly, apicidin induced phosphorylation of phospholipase Cγ1 (PLCγ1) and epidermal growth factor receptor (EGFR), and inhibition of PLCγ1 and EGFR reduced cell death and ER stress. Finally, apicidin-induced histone H3 hyperacetylation and reduction of histone deacetylase 2 mRNA expression were not affected by either a PLCγ1 inhibitor, U73122, or the antioxidant, N-acetyl cysteine. Taken together, the results suggest that apicidin induces apoptosis by ER stress and mitochondrial dysfunction via PLCγ1 activation, Ca(2+) release, and reactive oxygen species accumulation in Neuro-2a neuroblastoma cells.

  19. N-Acetylcysteine improves mitochondrial function and ameliorates behavioral deficits in the R6/1 mouse model of Huntington's disease.

    PubMed

    Wright, D J; Renoir, T; Smith, Z M; Frazier, A E; Francis, P S; Thorburn, D R; McGee, S L; Hannan, A J; Gray, L J

    2015-01-01

    Huntington's disease (HD) is a neurodegenerative disorder, involving psychiatric, cognitive and motor symptoms, caused by a CAG-repeat expansion encoding an extended polyglutamine tract in the huntingtin protein. Oxidative stress and excitotoxicity have previously been implicated in the pathogenesis of HD. We hypothesized that N-acetylcysteine (NAC) may reduce both excitotoxicity and oxidative stress through its actions on glutamate reuptake and antioxidant capacity. The R6/1 transgenic mouse model of HD was used to investigate the effects of NAC on HD pathology. It was found that chronic NAC administration delayed the onset and progression of motor deficits in R6/1 mice, while having an antidepressant-like effect on both R6/1 and wild-type mice. A deficit in the astrocytic glutamate transporter protein, GLT-1, was found in R6/1 mice. However, this deficit was not ameliorated by NAC, implying that the therapeutic effect of NAC is not due to rescue of the GLT-1 deficit and associated glutamate-induced excitotoxicity. Assessment of mitochondrial function in the striatum and cortex revealed that R6/1 mice show reduced mitochondrial respiratory capacity specific to the striatum. This deficit was rescued by chronic treatment with NAC. There was a selective increase in markers of oxidative damage in mitochondria, which was rescued by NAC. In conclusion, NAC is able to delay the onset of motor deficits in the R6/1 model of Huntington's disease and it may do so by ameliorating mitochondrial dysfunction. Thus, NAC shows promise as a potential therapeutic agent in HD. Furthermore, our data suggest that NAC may also have broader antidepressant efficacy.

  20. Reduction of apoptosis through the mitochondrial pathway by the administration of acetyl-L-carnitine to mouse fibroblasts in culture

    SciTech Connect

    Pillich, Rudolf Tito; Scarsella, Gianfranco; Risuleo, Gianfranco . E-mail: gianfranco.risuleo@uniroma1.it

    2005-05-15

    It is shown in literature that stress, such as deprivation of trophic factors and hypoxia, induces apoptosis in cultured cells and in tissues. In light of these results, we explored the possibility of protecting cells from programmed death by improving the metabolism of the mitochondrion. To this end, acetyl-L-carnitine was administered at various concentrations under conditions of serum deprivation. The choice of this drug was based on the accepted notion that acetyl-L-carnitine is able to stabilize mitochondrial membranes and to increase the supply of energy to the organelle. The results presented here indicate that the drug protects cells from apoptotic death: this is demonstrated by a lower positivity to the TUNEL reaction and by a strong reduction of the apoptotic DNA ladder in serum-deprived cells. The involvement of the mitochondrial apoptotic pathway was assessed by cytochrome C release and immunoreactivity to caspase 3. Moreover, acetyl-L-carnitine stimulates cell proliferation.

  1. Fragmentation of mitochondrial cardiolipin by copper ions in the Atp7b-/- mouse model of Wilson's disease.

    PubMed

    Yurkova, Irina L; Arnhold, Juergen; Fitzl, Guenther; Huster, Dominik

    2011-07-01

    Cellular copper overload as found in Wilson's disease may disturb mitochondrial function and integrity. Atp7b(-/-) mice accumulate copper in the liver and serve as an animal model for this inherited disease. The molecular mechanism of copper toxicity in hepatocytes is poorly understood. Total mitochondrial lipids from liver of wild-type mice were subjected to oxidative stress by the Cu(2+)/H(2)O(2)/ascorbate system. Phosphatidic acid (PA) and phosphatidylhydroxyacetone (PHA) were detected as cardiolipin fragmentation products by thin-layer chromatography combined with MALDI-TOF mass spectrometry in oxidized samples, but not in unperturbed ones. The formation of PA and PHA in copper-treated model membrane correlated well with the decrease of cardiolipin. Mitochondrial lipids from Atp7b(-/-) mice of different age were analyzed for the presence of PA. While 32-weeks old wild-type (control) and Atp7b(-/-) mice did not show any PA, there was a steady increase in the amount of this lipid in Atp7b(-/-) mice in contrast to control with increasing age. Hepatocytes from elder Atp7b(-/-)mice contained morphologically changed mitochondria unlike cells from wild-type animals of the same age. We concluded that free-radical fragmentation of cardiolipin with the formation of PA is a likely mechanism that damages mitochondria under conditions of oxidative stress due to copper overload. Our findings are relevant for better understanding of molecular mechanisms for liver damage found in Wilson's disease.

  2. Galangin prevents aminoglycoside-induced ototoxicity by decreasing mitochondrial production of reactive oxygen species in mouse cochlear cultures.

    PubMed

    Kim, Ye-Ri; Kim, Min-A; Cho, Hyun-Ju; Oh, Se-Kyung; Lee, In-Kyu; Kim, Un-Kyung; Lee, Kyu-Yup

    2016-03-14

    Amikacin is a semi-synthetic aminoglycoside widely used to treat infections caused by gentamicin-resistant gram-negative organisms and nontuberculous mycobacteria. However, the use of this agent often results in ototoxicity due to the overproduction of reactive oxygen species (ROS). Galangin, a natural flavonoid, has been shown to play a protective role against mitochondrial dysfunction by reducing mitochondrial ROS production. In this study, the effect of galangin on amikacin-induced ototoxicity was examined using cultures of cochlear explants. Immunofluorescent staining showed that treatment of inner hair cells (IHCs) and outer hair cells (OHCs) with galangin significantly decreased damage induced by amikacin. Moreover, pretreatment with galangin resulted in decreased amikacin-provoked increase in ROS production in both types of hair cells by MitoSOX-red staining. Attenuation of apoptotic cell death was assessed immunohistochemically using active caspase-3 antibody and with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, compared to explants exposed to amikacin alone (P<0.05). These results indicate that galangin protects hair cells in the organ of Corti from amikacin-induced toxicity by reducing the production of mitochondrial ROS. The results of this study suggest that galangin can potentially be used as an antioxidant and antiapoptotic agent to prevent hearing loss caused by aminoglycoside induced-oxidative stress. PMID:26778349

  3. Ablation of Ca(V)2.1 voltage-gated Ca²⁺ channels in mouse forebrain generates multiple cognitive impairments.

    PubMed

    Mallmann, Robert Theodor; Elgueta, Claudio; Sleman, Faten; Castonguay, Jan; Wilmes, Thomas; van den Maagdenberg, Arn; Klugbauer, Norbert

    2013-01-01

    Voltage-gated Ca(V)2.1 (P/Q-type) Ca²⁺ channels located at the presynaptic membrane are known to control a multitude of Ca²⁺-dependent cellular processes such as neurotransmitter release and synaptic plasticity. Our knowledge about their contributions to complex cognitive functions, however, is restricted by the limited adequacy of existing transgenic Ca(V)2.1 mouse models. Global Ca(V)2.1 knock-out mice lacking the α1 subunit Cacna1a gene product exhibit early postnatal lethality which makes them unsuitable to analyse the relevance of Ca(V)2.1 Ca²⁺ channels for complex behaviour in adult mice. Consequently we established a forebrain specific Ca(V)2.1 knock-out model by crossing mice with a floxed Cacna1a gene with mice expressing Cre-recombinase under the control of the NEX promoter. This novel mouse model enabled us to investigate the contribution of Ca(V)2.1 to complex cognitive functions, particularly learning and memory. Electrophysiological analysis allowed us to test the specificity of our conditional knock-out model and revealed an impaired synaptic transmission at hippocampal glutamatergic synapses. At the behavioural level, the forebrain-specific Ca(V)2.1 knock-out resulted in deficits in spatial learning and reference memory, reduced recognition memory, increased exploratory behaviour and a strong attenuation of circadian rhythmicity. In summary, we present a novel conditional Ca(V)2.1 knock-out model that is most suitable for analysing the in vivo functions of Ca(V)2.1 in the adult murine forebrain.

  4. How mitochondrial dynamism orchestrates mitophagy

    PubMed Central

    Shirihai, Orian; Song, Moshi; Dorn, Gerald W

    2015-01-01

    Mitochondria are highly dynamic, except in adult cardiomyocytes. Yet, the fission and fusion-promoting proteins that mediate mitochondrial dynamism are highly expressed in, and essential to the normal functioning of, hearts. Here, we review accumulating evidence supporting important roles for mitochondrial fission and fusion in cardiac mitochondrial quality control, focusing on the PINK1-Parkin mitophagy pathway.Based in part on recent findings from in vivo mouse models in which mitofusin-mediated mitochondrial fusion or Drp1-mediated mitochondrial fission were conditionally interrupted in cardiac myocytes, we propose several new concepts that may provide insight into the cardiac mitochondrial dynamism-mitophagy interactome. PMID:25999423

  5. (−)-EPICATECHIN IMPROVES MITOCHONDRIAL RELATED PROTEIN LEVELS AND AMELIORATES OXIDATIVE STRESS IN DYSTROPHIC DELTA SARCOGLYCAN NULL MOUSE STRIATED MUSCLE

    PubMed Central

    Ramirez-Sanchez, Israel; De los Santos, Sergio; Gonzalez-Basurto, Silvia; Canto, Patricia; Mendoza-Lorenzo, Patricia; Palma-Flores, Carlos; Ceballos-Reyes, Guillermo; Villarreal, Francisco; Zentella-Dehesa, Alejandro; Coral-Vazquez, Ramon

    2014-01-01

    Muscular dystrophies (MD) are a group of heterogeneous genetic disorders characterized by progressive striated muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism for disease pathogenesis remains unclear. The presence of oxidative stress (OS) is known to contribute to the pathophysiology and severity of the MD. Mitochondrial dysfunction is observed in MD and likely represents an important determinant of increased OS. Experimental antioxidant therapies have been implemented with the aim of protecting against disease progression, but results from clinical trials have been disappointing. In this study, we explored the capacity of the cacao flavonoid (−)-epicatechin (Epi) to mitigate OS by acting as a positive regulator of mitochondrial structure/function endpoints and redox balance control systems in skeletal and cardiac muscles of dystrophic, δ-sarcoglycan (δ-SG) null mice. Wild type or δ-SG null 2.5 month old male mice were treated via oral gavage with either water (control animals) or Epi (1 mg/kg, twice/day) for 2 weeks. Results evidence a significant normalization of total protein carbonylation, recovery of reduced/oxidized glutathione (GSH/GSSG ratio) and enhanced superoxide dismutase 2, catalase and citrate synthase activities with Epi treatment. These effects were accompanied by increases in protein levels for thiolredoxin, glutathione peroxidase, superoxide dismutase 2, catalase and mitochondrial endpoints. Furthermore, we evidence decreases in heart and skeletal muscle fibrosis, accompanied with an improvement in skeletal muscle function with treatment. These results warrant the further investigation of Epi as a potential therapeutic agent to mitigate MD associated muscle degeneration. PMID:25284161

  6. (-)-Epicatechin improves mitochondrial-related protein levels and ameliorates oxidative stress in dystrophic δ-sarcoglycan null mouse striated muscle.

    PubMed

    Ramirez-Sanchez, Israel; De los Santos, Sergio; Gonzalez-Basurto, Silvia; Canto, Patricia; Mendoza-Lorenzo, Patricia; Palma-Flores, Carlos; Ceballos-Reyes, Guillermo; Villarreal, Francisco; Zentella-Dehesa, Alejandro; Coral-Vazquez, Ramon

    2014-12-01

    Muscular dystrophies (MDs) are a group of heterogeneous genetic disorders characterized by progressive striated muscle wasting and degeneration. Although the genetic basis for many of these disorders has been identified, the exact mechanism of disease pathogenesis remains unclear. The presence of oxidative stress (OS) is known to contribute to the pathophysiology and severity of the MD. Mitochondrial dysfunction is observed in MD, and probably represents an important determinant of increased OS. Experimental antioxidant therapies have been implemented with the aim of protecting against disease progression, but results from clinical trials have been disappointing. In this study, we explored the capacity of the cacao flavonoid (-)-epicatechin (Epi) to mitigate OS by acting as a positive regulator of mitochondrial structure/function endpoints and redox balance control systems in skeletal and cardiac muscles of dystrophic, δ-sarcoglycan (δ-SG) null mice. Wild-type or δ-SG null 2.5-month-old male mice were treated via oral gavage with either water (controls) or Epi (1 mg·kg(-1) , twice daily) for 2 weeks. The results showed significant normalization of total protein carbonylation, recovery of the glutathione/oxidized glutathione ratio and enhanced superoxide dismutase 2, catalase and citrate synthase activities with Epi treatment. These effects were accompanied by increases in the protein levels of thioredoxin, glutathione peroxidase, superoxide dismutase 2, catalase, and mitochondrial endpoints. Furthermore, we found decreases in heart and skeletal muscle fibrosis, accompanied by an improvement in skeletal muscle function, with treatment. These results warrant further investigation of Epi as a potential therapeutic agent to mitigate MD-associated muscle degeneration. PMID:25284161

  7. Apolipoprotein A1 regulates coenzyme Q10 absorption, mitochondrial function, and infarct size in a mouse model of myocardial infarction.

    PubMed

    Dadabayev, Alisher R; Yin, Guotian; Latchoumycandane, Calivarathan; McIntyre, Thomas M; Lesnefsky, Edward J; Penn, Marc S

    2014-07-01

    HDL and apolipoprotein A1 (apoA1) concentrations inversely correlate with risk of death from ischemic heart disease; however, the role of apoA1 in the myocardial response to ischemia has not been well defined. To test whether apoA1, the primary HDL apolipoprotein, has an acute anti-inflammatory role in ischemic heart disease, we induced myocardial infarction via direct left anterior descending coronary artery ligation in apoA1 null (apoA1(-/-)) and apoA1 heterozygous (apoA1(+/-)) mice. We observed that apoA1(+/-) and apoA1(-/-) mice had a 52% and 125% increase in infarct size as a percentage of area at risk, respectively, compared with wild-type (WT) C57BL/6 mice. Mitochondrial oxidation contributes to tissue damage in ischemia-reperfusion injury. A substantial defect was present at baseline in the electron transport chain of cardiac myocytes from apoA1(-/-) mice localized to the coenzyme Q (CoQ) pool with impaired electron transfer (67% decrease) from complex II to complex III. Administration of coenzyme Q10 (CoQ10) to apoA1 null mice normalized the cardiac mitochondrial CoQ pool and reduced infarct size to that observed in WT mice. CoQ10 administration did not significantly alter infarct size in WT mice. These data identify CoQ pool content leading to impaired mitochondrial function as major contributors to infarct size in the setting of low HDL/apoA1. These data suggest a previously unappreciated mechanism for myocardial stunning, cardiac dysfunction, and muscle pain associated with low HDL and low apoA1 concentrations that can be corrected by CoQ10 supplementation and suggest populations of patients that may benefit particularly from CoQ10 supplementation.

  8. Identification of the Sensory Neuron Specific Regulatory Region for the Mouse Gene Encoding the Voltage Gated Sodium Channel Nav1.8

    PubMed Central

    Puhl, Henry L.; Ikeda, Stephen R.

    2008-01-01

    Voltage-gated sodium channels (VGSC) are critical membrane components that participate in the electrical activity of excitable cells. The type one VGSC family includes the tetrodotoxin insensitive sodium channel, Nav1.8, encoded by the Scn10a gene. Nav1.8 expression is restricted to small and medium diameter nociceptive sensory neurons of the dorsal root (DRG) and cranial sensory ganglia. In order to understand the stringent transcriptional regulation of the Scn10a gene, the sensory neuron specific promoter was functionally identified. While identifying the mRNA 5’ end, alternative splicing within the 5’ UTR was observed to create heterogeneity in the RNA transcript. Four kilobases of upstream genomic DNA was cloned and the presence of tissue specific promoter activity was tested by microinjection and adenoviral infection of fluorescent protein reporter constructs into primary mouse and rat neurons, and cell lines. The region contained many putative transcription factor binding sites and strong homology with the predicted rat ortholog. Homology to the predicted human ortholog was limited to the proximal end and several conserved cis elements were noted. Two regulatory modules were identified by microinjection of reporter constructs into DRG and superior cervical ganglia neurons: a neuron specific proximal promoter region between −1.6 and −0.2kb of the transcription start site cluster, and a distal sensory neuron switch region beyond −1.6kb that restricted fluorescent protein expression to a subset of primary sensory neurons. PMID:18466327

  9. Chronic alcohol exposure affects pancreatic acinar mitochondrial thiamin pyrophosphate uptake: studies with mouse 266-6 cell line and primary cells.

    PubMed

    Srinivasan, Padmanabhan; Nabokina, Svetlana; Said, Hamid M

    2015-11-01

    Thiamin is essential for normal metabolic activity of all mammalian cells, including those of the pancreas. Cells obtain thiamin from their surroundings and enzymatically convert it into thiamin pyrophosphate (TPP) in the cytoplasm; TPP is then taken up by mitochondria via a specific carrier the mitochondrial TPP transporter (MTPPT; product of the SLC25A19 gene). Chronic alcohol exposure negatively impacts the health of pancreatic acinar cells (PAC), but its effect on physiological/molecular parameters of MTPPT is not known. We addressed this issue using mouse pancreatic acinar tumor cell line 266-6 and primary PAC of wild-type and transgenic mice carrying the SLC25A19 promoter that were fed alcohol chronically. Chronic alcohol exposure of 266-6 cells (but not to its nonoxidative metabolites ethyl palmitate and ethyl oleate) led to a significant inhibition in mitochondrial TPP uptake, which was associated with a decreased expression of MTPPT protein, mRNA, and activity of the SLC25A19 promoter. Similarly, chronic alcohol feeding of mice led to a significant inhibition in expression of MTPPT protein, mRNA, heterogeneous nuclear RNA, as well as in activity of SLC25A19 promoter in PAC. While chronic alcohol exposure did not affect DNA methylation of the Slc25a19 promoter, a significant decrease in histone H3 euchromatin markers and an increase in H3 heterochromatin marker were observed. These findings show, for the first time, that chronic alcohol exposure negatively impacts pancreatic MTPPT, and that this effect is exerted, at least in part, at the level of Slc25a19 transcription and appears to involve epigenetic mechanism(s).

  10. Mitochondrial dysfunction, oxidative stress, and apoptosis revealed by proteomic and transcriptomic analyses of the striata in two mouse models of Parkinson's disease.

    PubMed

    Chin, Mark H; Qian, Wei-Jun; Wang, Haixing; Petyuk, Vladislav A; Bloom, Joshua S; Sforza, Daniel M; Laćan, Goran; Liu, Dahai; Khan, Arshad H; Cantor, Rita M; Bigelow, Diana J; Melega, William P; Camp, David G; Smith, Richard D; Smith, Desmond J

    2008-02-01

    The molecular mechanisms underlying the changes in the nigrostriatal pathway in Parkinson's disease (PD) are not completely understood. Here, we use mass spectrometry and microarrays to study the proteomic and transcriptomic changes in the striatum of two mouse models of PD, induced by the distinct neurotoxins 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and methamphetamine (METH). Proteomic analyses resulted in the identification and relative quantification of 912 proteins with two or more unique peptides and 86 proteins with significant abundance changes following neurotoxin treatment. Similarly, microarray analyses revealed 181 genes with significant changes in mRNA, following neurotoxin treatment. The combined protein and gene list provides a clearer picture of the potential mechanisms underlying neurodegeneration observed in PD. Functional analysis of this combined list revealed a number of significant categories, including mitochondrial dysfunction, oxidative stress response, and apoptosis. These results constitute one of the largest descriptive data sets integrating protein and transcript changes for these neurotoxin models with many similar end point phenotypes but distinct mechanisms.

  11. Mitochondrial Dysfunction, Oxidative Stress, and Apoptosis Revealed by Proteomic and Transcriptomic Analyses of the Striata in Two Mouse Models of Parkinson’s Disease

    PubMed Central

    Chin, Mark H.; Qian, Wei-Jun; Wang, Haixing; Petyuk, Vladislav A.; Bloom, Joshua S.; Sforza, Daniel M.; Laćan, Goran; Liu, Dahai; Khan, Arshad H.; Cantor, Rita M.; Bigelow, Diana J.; Melega, William P.; Camp, David G.; Smith, Richard D.; Smith, Desmond J.

    2012-01-01

    The molecular mechanisms underlying the changes in the nigrostriatal pathway in Parkinson’s disease (PD) are not completely understood. Here, we use mass spectrometry and microarrays to study the proteomic and transcriptomic changes in the striatum of two mouse models of PD, induced by the distinct neurotoxins 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and methamphetamine (METH). Proteomic analyses resulted in the identification and relative quantification of 912 proteins with two or more unique peptides and 86 proteins with significant abundance changes following neurotoxin treatment. Similarly, microarray analyses revealed 181 genes with significant changes in mRNA, following neurotoxin treatment. The combined protein and gene list provides a clearer picture of the potential mechanisms underlying neurodegeneration observed in PD. Functional analysis of this combined list revealed a number of significant categories, including mitochondrial dysfunction, oxidative stress response, and apoptosis. These results constitute one of the largest descriptive data sets integrating protein and transcript changes for these neurotoxin models with many similar end point phenotypes but distinct mechanisms. PMID:18173235

  12. Mitochondrial dysfunction, oxidative stress and apoptosis revealed by proteomic and transcriptomic analyses of the striata in two mouse models of Parkinson’s disease

    SciTech Connect

    Chin, Mark H.; Qian, Weijun; Wang, Haixing; Petyuk, Vladislav A.; Bloom, Joshua S.; Sforza, Daniel M.; Lacan, Goran; Liu, Dahai; Khan, Arshad H.; Cantor, Rita M.; Bigelow, Diana J.; Melega, William P.; Camp, David G.; Smith, Richard D.; Smith, Desmond J.

    2008-02-10

    The molecular mechanisms underlying the changes in the nigrostriatal pathway in Parkinson disease (PD) are not completely understood. Here we use mass spectrometry and microarrays to study the proteomic and transcriptomic changes in the striatum of two mouse models of PD, induced by the distinct neurotoxins 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and methamphetamine (METH). Proteomic analyses resulted in the identification and relative quantification of 912 proteins with two or more unique peptides and 85 proteins with significant abundance changes following neurotoxin treatment. Similarly, microarray analyses revealed 181 genes with significant changes in mRNA following neurotoxin treatment. The combined protein and gene list provides a clearer picture of the potential mechanisms underlying neurodegeneration observed in PD. Functional analysis of this combined list revealed a number of significant categories, including mitochondrial dysfunction, oxidative stress response and apoptosis. Additionally, codon usage and miRNAs may play an important role in translational control in the striatum. These results constitute one of the largest datasets integrating protein and transcript changes for these neurotoxin models with many similar endpoint phenotypes but distinct mechanisms.

  13. A trend of central versus peripheral structuring in mitochondrial and nuclear gene sequences of the Japanese wood mouse, Apodemus speciosus.

    PubMed

    Tomozawa, Morihiko; Suzuki, Hitoshi

    2008-03-01

    A phylogeographic analysis was performed on Japanese endemic wood mice (Apodemus speciosus) using nuclear interphotoreceptor retinol binding protein (IRBP) gene sequences (1,152 bp), together with previously published mitochondrial cytochrome b (cyt b) data. In the IRBP analysis, 40 haplotypes were recovered from 84 individuals by statistical and subcloning methods. Substantial sequence variation was determined from the IRBP data (pi=0.0047), and no significant evidence of recombination was detected. From the phylogenetic analysis, the 40 haplotypes fell into two major groups with geographic associations, irrespective of the karyotype groups (2n=46 and 2n=48), yielding a trend of central (Hokkaido, Honshu, Shikoku, Kyushu, and Sado) and peripheral (Izu, Oki, Tsushima, and Satsunan Is.) groupings. This geographic pattern is similar to that observed in the cyt b data, with a different insular grouping of Sado, Hokkaido, Izu, and Satsunan islands, and also to that of morphological features. In both gene data sets, nested clade analyses revealed allopatric fragmentation in the "peripheral island clades" and range expansion in the "central island clades." A mismatch analysis using cyt b data also suggested expansion of the central islands clade. Thus, the trend of central vs. peripheral structuring may be attributable to past demographic dynamics in the two distinct haplotype clades, such as range expansion of one clade in the central area of the Japanese Islands, leaving the other clade in the periphery.

  14. Demethyleneberberine, a natural mitochondria-targeted antioxidant, inhibits mitochondrial dysfunction, oxidative stress, and steatosis in alcoholic liver disease mouse model.

    PubMed

    Zhang, Pengcheng; Qiang, Xiaoyan; Zhang, Miao; Ma, Dongshen; Zhao, Zheng; Zhou, Cuisong; Liu, Xie; Li, Ruiyan; Chen, Huan; Zhang, Yubin

    2015-01-01

    Excessive alcohol consumption induces oxidative stress and lipid accumulation in the liver. Mitochondria have long been recognized as the key target for alcoholic liver disease (ALD). Recently, the artificial mitochondria-targeted antioxidant MitoQ has been used to treat ALD effectively in mice. Here, we introduce the natural mitochondria-targeted antioxidant demethyleneberberine (DMB), which has been found in Chinese herb Cortex Phellodendri chinensis. The protective effect of DMB on ALD was evaluated with HepG2 cells and acutely/chronically ethanol-fed mice, mimicking two common patterns of drinking in human. The results showed that DMB, which is composed of a potential antioxidant structure, could penetrate the membrane of mitochondria and accumulate in mitochondria either in vitro or in vivo. Consequently, the acute drinking-caused oxidative stress and mitochondrial dysfunction were significantly ameliorated by DMB. Moreover, we also found that DMB suppressed CYP2E1, hypoxia inducible factor α, and inducible nitric oxide synthase, which contributed to oxidative stress and restored sirtuin 1/AMP-activated protein kinase/peroxisome proliferator-activated receptor-γ coactivator-1α pathway-associated fatty acid oxidation in chronic ethanol-fed mice, which in turn ameliorated lipid peroxidation and macrosteatosis in the liver. Taking these findings together, DMB could serve as a novel and potential therapy for ALD in human beings.

  15. N- and L-Type Voltage-Gated Calcium Channels Mediate Fast Calcium Transients in Axonal Shafts of Mouse Peripheral Nerve.

    PubMed

    Barzan, Ruxandra; Pfeiffer, Friederike; Kukley, Maria

    2016-01-01

    In the peripheral nervous system (PNS) a vast number of axons are accommodated within fiber bundles that constitute peripheral nerves. A major function of peripheral axons is to propagate action potentials along their length, and hence they are equipped with Na(+) and K(+) channels, which ensure successful generation, conduction and termination of each action potential. However little is known about Ca(2+) ion channels expressed along peripheral axons and their possible functional significance. The goal of the present study was to test whether voltage-gated Ca(2+) channels (VGCCs) are present along peripheral nerve axons in situ and mediate rapid activity-dependent Ca(2+) elevations under physiological circumstances. To address this question we used mouse sciatic nerve slices, Ca(2+) indicator Oregon Green BAPTA-1, and 2-photon Ca(2+) imaging in fast line scan mode (500 Hz). We report that transient increases in intra-axonal Ca(2+) concentration take place along peripheral nerve axons in situ when axons are stimulated electrically with single pulses. Furthermore, we show for the first time that Ca(2+) transients in peripheral nerves are fast, i.e., occur in a millisecond time-domain. Combining Ca(2+) imaging and pharmacology with specific blockers of different VGCCs subtypes we demonstrate that Ca(2+) transients in peripheral nerves are mediated mainly by N-type and L-type VGCCs. Discovery of fast Ca(2+) entry into the axonal shafts through VGCCs in peripheral nerves suggests that Ca(2+) may be involved in regulation of action potential propagation and/or properties in this system, or mediate neurotransmitter release along peripheral axons as it occurs in the optic nerve and white matter of the central nervous system (CNS).

  16. N- and L-Type Voltage-Gated Calcium Channels Mediate Fast Calcium Transients in Axonal Shafts of Mouse Peripheral Nerve

    PubMed Central

    Barzan, Ruxandra; Pfeiffer, Friederike; Kukley, Maria

    2016-01-01

    In the peripheral nervous system (PNS) a vast number of axons are accommodated within fiber bundles that constitute peripheral nerves. A major function of peripheral axons is to propagate action potentials along their length, and hence they are equipped with Na+ and K+ channels, which ensure successful generation, conduction and termination of each action potential. However little is known about Ca2+ ion channels expressed along peripheral axons and their possible functional significance. The goal of the present study was to test whether voltage-gated Ca2+ channels (VGCCs) are present along peripheral nerve axons in situ and mediate rapid activity-dependent Ca2+ elevations under physiological circumstances. To address this question we used mouse sciatic nerve slices, Ca2+ indicator Oregon Green BAPTA-1, and 2-photon Ca2+ imaging in fast line scan mode (500 Hz). We report that transient increases in intra-axonal Ca2+ concentration take place along peripheral nerve axons in situ when axons are stimulated electrically with single pulses. Furthermore, we show for the first time that Ca2+ transients in peripheral nerves are fast, i.e., occur in a millisecond time-domain. Combining Ca2+ imaging and pharmacology with specific blockers of different VGCCs subtypes we demonstrate that Ca2+ transients in peripheral nerves are mediated mainly by N-type and L-type VGCCs. Discovery of fast Ca2+ entry into the axonal shafts through VGCCs in peripheral nerves suggests that Ca2+ may be involved in regulation of action potential propagation and/or properties in this system, or mediate neurotransmitter release along peripheral axons as it occurs in the optic nerve and white matter of the central nervous system (CNS). PMID:27313508

  17. In Vivo Voltage-Sensitive Dye Study of Lateral Spreading of Cortical Activity in Mouse Primary Visual Cortex Induced by a Current Impulse

    PubMed Central

    Fehérvári, Tamás Dávid; Sawai, Hajime; Yagi, Tetsuya

    2015-01-01

    In the mammalian primary visual cortex (V1), lateral spreading of excitatory potentials is believed to be involved in spatial integrative functions, but the underlying cortical mechanism is not well understood. Visually-evoked population-level responses have been shown to propagate beyond the V1 initial activation site in mouse, similar to higher mammals. Visually-evoked responses are, however, affected by neuronal circuits prior to V1 (retina, LGN), making the separate analysis of V1 difficult. Intracortical stimulation eliminates these initial processing steps. We used in vivo RH1691 voltage-sensitive dye (VSD) imaging and intracortical microstimulation in adult C57BL/6 mice to elucidate the spatiotemporal properties of population-level signal spreading in V1 cortical circuits. The evoked response was qualitatively similar to that measured in single-cell electrophysiological experiments in rodents: a fast transient fluorescence peak followed by a fast and a slow decrease or hyperpolarization, similar to EPSP and fast and slow IPSPs in single cells. The early cortical response expanded at speeds commensurate with long horizontal projections (at 5% of the peak maximum, 0.08–0.15 m/s) however, the bulk of the VSD signal propagated slowly (at half-peak maximum, 0.05–0.08 m/s) suggesting an important role of regenerative multisynaptic transmission through short horizontal connections in V1 spatial integrative functions. We also found a tendency for a widespread and fast cortical response suppression in V1, which was eliminated by GABAA-antagonists gabazine and bicuculline methiodide. Our results help understand the neuronal circuitry involved in lateral spreading in V1. PMID:26230520

  18. Reduced availability of voltage-gated sodium channels by depolarization or blockade by tetrodotoxin boosts burst firing and catecholamine release in mouse chromaffin cells

    PubMed Central

    Vandael, David H F; Ottaviani, Matteo M; Legros, Christian; Lefort, Claudie; Guérineau, Nathalie C; Allio, Arianna; Carabelli, Valentina; Carbone, Emilio

    2015-01-01

    Action potential (AP) firing in mouse chromaffin cells (MCCs) is mainly sustained by Cav1.3 L-type channels that drive BK and SK currents and regulate the pacemaking cycle. As secretory units, CCs optimally recruit Ca2+ channels when stimulated, a process potentially dependent on the modulation of the AP waveform. Our previous work has shown that a critical determinant of AP shape is voltage-gated sodium channel (Nav) channel availability. Here, we studied the contribution of Nav channels to firing patterns and AP shapes at rest (−50 mV) and upon stimulation (−40 mV). Using quantitative RT-PCR and immunoblotting, we show that MCCs mainly express tetrodotoxin (TTX)-sensitive, fast-inactivating Nav1.3 and Nav1.7 channels that carry little or no Na+ current during slow ramp depolarizations. Time constants and the percentage of recovery from fast inactivation and slow entry into closed-state inactivation are similar to that of brain Nav1.3 and Nav1.7 channels. The fraction of available Nav channels is reduced by half after 10 mV depolarization from −50 to −40 mV. This leads to low amplitude spikes and a reduction in repolarizing K+ currents inverting the net current from outward to inward during the after-hyperpolarization. When Nav channel availability is reduced by up to 20% of total, either by TTX block or steady depolarization, a switch from tonic to burst firing is observed. The spontaneous occurrence of high frequency bursts is rare under control conditions (14% of cells) but leads to major Ca2+-entry and increased catecholamine release. Thus, Nav1.3/Nav1.7 channel availability sets the AP shape, burst-firing initiation and regulates catecholamine secretion in MCCs. Nav channel inactivation becomes important during periods of high activity, mimicking stress responses. PMID:25620605

  19. Mitochondrial E3 ligase March5 maintains stemness of mouse ES cells via suppression of ERK signalling.

    PubMed

    Gu, Hao; Li, Qidong; Huang, Shan; Lu, Weiguang; Cheng, Fangyuan; Gao, Ping; Wang, Chen; Miao, Lin; Mei, Yide; Wu, Mian

    2015-01-01

    Embryonic stem cells (ESCs) possess pluripotency, which is the capacity of cells to differentiate into all lineages of the mature organism. Increasing evidence suggests that the pluripotent state of ESCs is regulated by a combination of extrinsic and intrinsic factors. The underlying mechanisms, however, are not completely understood. Here, we show that March5, an E3 ubiquitin ligase, is involved in maintaining mouse-ESC (mESC) pluripotency. Knockdown of March5 in mESCs led to differentiation from naive pluripotency. Mechanistically, as a transcriptional target of Klf4, March5 catalyses K63-linked polyubiquitination of Prkar1a, a negative regulatory subunit of PKA, to activate PKA, thereby inhibiting the Raf/MEK/ERK pathway. Moreover, March5 is able to replace a MEK/ERK inhibitor to maintain mESC pluripotency under serum-free culture conditions. In addition, March5 can partially replace the use of Klf4 for somatic cell reprogramming. Collectively, our study uncovers a role for the Klf4-March5-PKA-ERK pathway in maintaining the stemness properties of mESCs. PMID:26033541

  20. Mitochondrial E3 ligase March5 maintains stemness of mouse ES cells via suppression of ERK signalling

    PubMed Central

    Gu, Hao; Li, Qidong; Huang, Shan; Lu, Weiguang; Cheng, Fangyuan; Gao, Ping; Miao, Lin; Mei, Yide; Wu, Mian

    2015-01-01

    Embryonic stem cells (ESCs) possess pluripotency, which is the capacity of cells to differentiate into all lineages of the mature organism. Increasing evidence suggests that the pluripotent state of ESCs is regulated by a combination of extrinsic and intrinsic factors. The underlying mechanisms, however, are not completely understood. Here, we show that March5, an E3 ubiquitin ligase, is involved in maintaining mouse-ESC (mESC) pluripotency. Knockdown of March5 in mESCs led to differentiation from naive pluripotency. Mechanistically, as a transcriptional target of Klf4, March5 catalyses K63-linked polyubiquitination of Prkar1a, a negative regulatory subunit of PKA, to activate PKA, thereby inhibiting the Raf/MEK/ERK pathway. Moreover, March5 is able to replace a MEK/ERK inhibitor to maintain mESC pluripotency under serum-free culture conditions. In addition, March5 can partially replace the use of Klf4 for somatic cell reprogramming. Collectively, our study uncovers a role for the Klf4–March5–PKA–ERK pathway in maintaining the stemness properties of mESCs. PMID:26033541

  1. Pharmacologic activation of mitochondrial biogenesis exerts widespread beneficial effects in a transgenic mouse model of Huntington's disease.

    PubMed

    Johri, Ashu; Calingasan, Noel Y; Hennessey, Thomas M; Sharma, Abhijeet; Yang, Lichuan; Wille, Elizabeth; Chandra, Abhishek; Beal, M Flint

    2012-03-01

    There is substantial evidence that impairment of peroxisome proliferator-activated receptor (PPAR)-γ-coactivator 1α (PGC-1α) levels and activity play an important role in Huntington's disease (HD) pathogenesis. We tested whether pharmacologic treatment with the pan-PPAR agonist bezafibrate would correct a deficiency of PGC-1α and exert beneficial effects in a transgenic mouse model of HD. We found that administration of bezafibrate in the diet restored levels of PGC-1α, PPARs and downstream genes to levels which occur in wild-type mice. There were significant improvements in phenotype and survival. In the striatum, astrogliosis and neuronal atrophy were attenuated and numbers of mitochondria were increased. Bezafibrate treatment prevented conversion of type I oxidative to type II glycolytic muscle fibers and increased the numbers of muscle mitochondria. Finally, bezafibrate rescued lipid accumulation and apparent vacuolization of brown adipose tissue in the HD mice. These findings provide strong evidence that treatment with bezafibrate exerts neuroprotective effects which may be beneficial in the treatment of HD. PMID:22095692

  2. Autoradiographic localization of voltage-dependent sodium channels on the mouse neuromuscular junction using /sup 125/I-alpha scorpion toxin. I. Preferential labeling of glial cells on the presynaptic side

    SciTech Connect

    Boudier, J.L.; Jover, E.; Cau, P.

    1988-05-01

    Alpha-scorpion toxins bind specifically to the voltage-sensitive sodium channel in excitable membranes, and binding is potential-dependent. The radioiodinated toxin II from the scorpion Androctonus australis Hector (alpha ScTx) was used to localize voltage-sensitive sodium channels on the presynaptic side of mouse neuromuscular junctions (NMJ) by autoradiography using both light and electron microscopy. Silver grain localization was analyzed by the cross-fire method. At the light-microscopic level, grain density over NMJ appeared 6-8x higher than over nonjunctional muscle membrane. The specificity of labeling was verified by competition/displacement with an excess of native alpha ScTx. Labeling was also inhibited by incubation in depolarizing conditions, showing its potential-dependence. At the electron-microscopic level, analysis showed that voltage-sensitive sodium channels labeled with alpha ScTx were almost exclusively localized on membranes, as expected. Due to washout after incubation, appreciable numbers of binding sites were not found on the postsynaptic membranes. However, on the presynaptic side, alpha ScTx-labeled voltage-sensitive sodium channels were localized on the membrane of non-myelin-forming Schwann cells covering NMJ. The axonal presynaptic membrane was not labeled. These results show that voltage-sensitive sodium channels are present on glial cells in vivo, as already demonstrated in vitro. It is proposed that these glial channels could be indirectly involved in the ionic homeostasis of the axonal environment.

  3. Metformin Prevents Dopaminergic Neuron Death in MPTP/P-Induced Mouse Model of Parkinson’s Disease via Autophagy and Mitochondrial ROS Clearance

    PubMed Central

    Lu, Ming; Su, Cunjin; Qiao, Chen; Bian, Yaqi; Ding, Jianhua

    2016-01-01

    Background: Our previous study demonstrated that metabolic inflammation exacerbates dopaminergic neuronal degeneration in type 2 diabetes mice. Metformin, a typical oral hypoglycemic agent for diabetes, has been regarded as an activator of AMP-activated protein kinase and a regulator of systemic energy metabolism. Although metformin plays potential protective effects in many disorders, it is unclear whether metformin has a therapeutic role in dopaminergic neuron degeneration in Parkinson’s disease. Methods: In the present study, a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine plus probenecid-induced mouse model of Parkinson’s disease was established to explore the neuroprotective effect of metformin on dopaminergic neurons in substania nigra compacta. We next cultured SH-SY5Y cells to investigate the mechanisms for the neuroprotective effect of metformin. Results: We showed that treatment with metformin (5mg/mL in drinking water) for 5 weeks significantly ameliorated the degeneration of substania nigra compacta dopaminergic neurons, increased striatal dopaminergic levels, and improved motor impairment induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine plus probenecid. We further found that metformin inhibited microglia overactivation-induced neuroinflammation in substania nigra compacta of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine plus probenecid Parkinson’s disease mice, which might contribute to the protective effect of metformin on neurodegeneration. Furthermore, metformin (2mM) activated AMP-activated protein kinase in SH-SY5Y cells, in turn inducing microtubule-associated protein 1 light chain 3-II-mediated autophagy and eliminating mitochondrial reactive oxygen species. Consequently, metformin alleviated MPP+-induced cytotoxicity and attenuated neuronal apoptosis. Conclusions: Our findings demonstrate that metformin may be a pluripotent and promising drug for dopaminergic neuron degeneration, which will give us insight into the potential of

  4. Enhancing mitochondrial calcium buffering capacity reduces aggregation of misfolded SOD1 and motor neuron cell death without extending survival in mouse models of inherited amyotrophic lateral sclerosis.

    PubMed

    Parone, Philippe A; Da Cruz, Sandrine; Han, Joo Seok; McAlonis-Downes, Melissa; Vetto, Anne P; Lee, Sandra K; Tseng, Eva; Cleveland, Don W

    2013-03-13

    Mitochondria have been proposed as targets for toxicity in amyotrophic lateral sclerosis (ALS), a progressive, fatal adult-onset neurodegenerative disorder characterized by the selective loss of motor neurons. A decrease in the capacity of spinal cord mitochondria to buffer calcium (Ca(2+)) has been observed in mice expressing ALS-linked mutants of SOD1 that develop motor neuron disease with many of the key pathological hallmarks seen in ALS patients. In mice expressing three different ALS-causing SOD1 mutants, we now test the contribution of the loss of mitochondrial Ca(2+)-buffering capacity to disease mechanism(s) by eliminating ubiquitous expression of cyclophilin D, a critical regulator of Ca(2+)-mediated opening of the mitochondrial permeability transition pore that determines mitochondrial Ca(2+) content. A chronic increase in mitochondrial buffering of Ca(2+) in the absence of cyclophilin D was maintained throughout disease course and was associated with improved mitochondrial ATP synthesis, reduced mitochondrial swelling, and retention of normal morphology. This was accompanied by an attenuation of glial activation, reduction in levels of misfolded SOD1 aggregates in the spinal cord, and a significant suppression of motor neuron death throughout disease. Despite this, muscle denervation, motor axon degeneration, and disease progression and survival were unaffected, thereby eliminating mutant SOD1-mediated loss of mitochondrial Ca(2+) buffering capacity, altered mitochondrial morphology, motor neuron death, and misfolded SOD1 aggregates, as primary contributors to disease mechanism for fatal paralysis in these models of familial ALS. PMID:23486940

  5. A conditional mouse mutant in the tumor suppressor SdhD gene unveils a link between p21(WAF1/Cip1) induction and mitochondrial dysfunction.

    PubMed

    Millán-Uclés, Africa; Díaz-Castro, Blanca; García-Flores, Paula; Báez, Alicia; Pérez-Simón, José Antonio; López-Barneo, José; Piruat, José I

    2014-01-01

    Mutations in mitochondrial complex II (MCII; succinate dehydrogenase, Sdh) genes cause familiar pheochromocytoma/paraganglioma tumors. Several mechanisms have been proposed to account for Sdh-mutation-induced tumorigenesis, the most accepted of which is based on the constitutive expression of the hypoxia-inducible factor 1α (Hif1α) at normal oxygen tension, a theory referred to as "pseudo-hypoxic drive". Other molecular processes, such as oxidative stress, apoptosis, or chromatin remodeling have been also proposed to play a causative role. Nevertheless, the actual contribution of each of these mechanisms has not been definitively established. Moreover, the biological factors that determine the tissue-specificity of these tumors have not been identified. In this work, we made use of the inducible SDHD-ESR mouse, a conditional mutant in the SdhD gene, which encodes the small subunit of MCII, and that acts as a tumor suppressor gene in humans. The analysis of the Hif1α pathway in SDHD-ESR tissues and in two newly derived cell lines after complete SdhD loss -a requirement for hereditary paraganglioma type-1 tumor formation in humans- partially recapitulated the "pseudo-hypoxic" response and rendered inconsistent results. Therefore, we performed microarray analysis of adrenal medulla and kidney in order to identify other early gene expression changes elicited by SdhD deletion. Our results revealed that each mutant tissue displayed different variations in their gene expression profiles affecting to different biological processes. However, we found that the Cdkn1a gene was up-regulated in both tissues. This gene encodes the cyclin-dependent kinase inhibitor p21(WAF1/Cip1), a factor implicated in cell cycle, senescence, and cancer. The two SDHD-ESR cell lines also showed accumulation of this protein. This new and unprecedented evidence for a link between SdhD dysfunction and p21(WAF1/Cip1) will open new avenues for the study of the mechanisms that cause tumors in Sdh

  6. Differences in cytosolic and mitochondrial 5'-nucleotidase and deoxynucleoside kinase activities in Sprague-Dawley rat and CD-1 mouse tissues: implication for the toxicity of nucleoside analogs in animal models.

    PubMed

    Mirzaee, Saeedeh; Eriksson, Staffan; Albertioni, Freidoun

    2010-01-12

    Cytosolic and mitochondrial deoxynucleoside kinases (dNKs), as well as 5'deoxynucleotidases (5'-dNTs), control intracellular and intramitochondrial phosphorylation of natural nucleotides and nucleoside analogs used in antiviral and cancer chemotherapy. The balance in the activities of these two groups of enzymes to a large extent determines both the efficacy and side effects of these drugs. Because of the broad and overlapping substrate specificities of the nucleoside kinases and 5'-NTs, their tissue distribution and roles in the metabolism of both natural nucleosides and their analogs are still not fully elucidated. Here, the activity of dNKs: dCK and TK (TK1 and TK2) as well as 5'-dNTs: CN1, CN2 and dNT (dNT1 and dNT2) were determined in 14 different adult mouse and rat tissues. In most cases tissue activities of TK1, TK2 and dCK were 2-3-fold higher in the mouse, a similar pattern was found with CN1 and dNTs although with several exceptions, e.g., TK2 activities in muscle extracts from rats were 2-10-fold higher than in the mouse. Furthermore CN1 activities in hepatic, renal and adipose extracts were 2-3-fold higher in the rat. CN2 had higher levels in the testis, spleen, pancreas and diaphragm and lower level in the lung of mouse compared to rat tissues. The result suggests that a major difference in these activity profiles between mouse and rat may account for discrepancies in pharmacological response of the two animals to certain nucleoside compounds, and may help to improve the usefulness of animal models in future efforts of drug discovery.

  7. Acetyl-L-carnitine and lipoic acid improve mitochondrial abnormalities and serum levels of liver enzymes in a mouse model of nonalcoholic fatty liver disease.

    PubMed

    Kathirvel, Elango; Morgan, Kengathevy; French, Samuel W; Morgan, Timothy R

    2013-11-01

    Mitochondrial abnormalities are suggested to be associated with the development of nonalcoholic fatty liver. Liver mitochondrial content and function have been shown to improve in oral feeding of acetyl-L-carnitine (ALC) to rodents. Carnitine is involved in the transport of acyl-coenzyme A across the mitochondrial membrane to be used in mitochondrial β-oxidation. We hypothesized that oral administration ALC with the antioxidant lipoic acid (ALC + LA) would benefit nonalcoholic fatty liver. To test our hypothesis, we fed Balb/C mice a standard diet (SF) or SF with ALC + LA or high-fat diet (HF) or HF with ALC + LA for 6 months. Acetyl-L-carnitine and LA were dissolved at 0.2:0.1% (wt/vol) in drinking water, and mice were allowed free access to food and water. Along with physical parameters, insulin resistance (blood glucose, insulin, glucose tolerance), liver function (alanine transaminase [ALT], aspartate transaminase [AST]), liver histology (hematoxylin and eosin), oxidative stress (malondialdehyde), and mitochondrial abnormalities (carbamoyl phosphate synthase 1 and electron microscopy) were done. Compared with SF, HF had higher body, liver, liver-to-body weight ratio, white adipose tissue, ALT, AST, liver fat, oxidative stress, and insulin resistance. Coadministration of ALC + LA to HF animals significantly improved the mitochondrial marker carbamoyl phosphate synthase 1 and the size of the mitochondria in liver. Alanine transaminase and AST levels were decreased. In a nonalcoholic fatty liver mice model, ALC + LA combination improved liver mitochondrial content, size, serum ALT, and AST without significant changes in oxidative stress, insulin resistance, and liver fat accumulation. PMID:24176233

  8. D-Galactose Induces a Mitochondrial Complex I Deficiency in Mouse Skeletal Muscle: Potential Benefits of Nutrient Combination in Ameliorating Muscle Impairment

    PubMed Central

    Chang, Liao; Liu, Xin; Liu, Jing; Li, Hua; Yang, Yanshen; Liu, Jia; Guo, Zihao; Xiao, Ke; Zhang, Chen; Liu, Jiankang

    2014-01-01

    Abstract Accumulating research has shown that chronic D-galactose (D-gal) exposure induces symptoms similar to natural aging in animals. Therefore, rodents chronically exposed to D-gal are increasingly used as a model for aging and delay-of-aging pharmacological research. Mitochondrial dysfunction is thought to play a vital role in aging and age-related diseases; however, whether mitochondrial dysfunction plays a significant role in mice exposed to D-gal remains unknown. In the present study, we investigated cognitive dysfunction, locomotor activity, and mitochondrial dysfunction involved in D-gal exposure in mice. We found that D-gal exposure (125 mg/kg/day, 8 weeks) resulted in a serious impairment in grip strength in mice, whereas spatial memory and locomotor coordination remained intact. Interestingly, muscular mitochondrial complex I deficiency occurred in the skeletal muscle of mice exposed to D-gal. Mitochondrial ultrastructure abnormality was implicated as a contributing factor in D-gal-induced muscular impairment. Moreover, three combinations (A, B, and C) of nutrients applied in this study effectively reversed D-gal-induced muscular impairment. Nutrient formulas B and C were especially effective in reversing complex I dysfunction in both skeletal muscle and heart muscle. These findings suggest the following: (1) chronic exposure to D-gal first results in specific muscular impairment in mice, rather than causing general, premature aging; (2) poor skeletal muscle strength induced by D-gal might be due to the mitochondrial dysfunction caused by complex I deficiency; and (3) the nutrient complexes applied in the study attenuated the skeletal muscle impairment, most likely by improving mitochondrial function. PMID:24476218

  9. Mitochondrial division inhibitor 1 (Mdivi-1) offers neuroprotection through diminishing cell death and improving functional outcome in a mouse model of traumatic brain injury.

    PubMed

    Wu, Qiong; Xia, Shui-Xiu; Li, Qian-Qian; Gao, Yuan; Shen, Xi; Ma, Lu; Zhang, Ming-Yang; Wang, Tao; Li, Yong-Sheng; Wang, Zu-Feng; Luo, Cheng-Liang; Tao, Lu-Yang

    2016-01-01

    Mitochondria dysfunction, an enormous potential crisis, has attracted increasing attention. Disturbed regulation of mitochondrial dynamics, the balance of mitochondrial fusion and fission, has been implicated in neurodegenerative diseases, such as Parkinson׳s disease and cerebral ischemia/reperfusion. However the role of mitochondrial dynamics in traumatic brain injury (TBI) has not been illuminated. The aim of the present study was to investigate the role of Mdivi-1, a small molecule inhibitor of a key mitochondrial fission protein dynamin-related protein 1 (Drp1), in TBI-induced cell death and functional outcome deficits. Protein expression of Drp1 was first investigated. Outcome parameters consist of motor test, Morris water maze, brain edema and lesion volume. Cell death was detected by propidium iodide (PI) labeling, and mitochondrial morphology was assessed using transmission electron microscopy. In addition, the expression of apoptosis-related proteins cytochrome c (cyt-c) and caspase-3 was investigated. Our findings showed that up-regulation of Drp1 expression started at 1h post-TBI and peaked at 24 h, but inhibition of Drp1 by Mdivi-1 significantly alleviated TBI-induced behavioral deficits and brain edema, reduced morphological change of mitochondria, and decreased TBI-induced cell death together with lesion volume. Moreover, treatment with Mdivi-1 remarkably inhibited TBI-induced the release of cyt-c from mitochondria to cytoplasm, and activation of caspase-3 at 24 h after TBI. Taken together, these data imply that inhibition of Drp1 may help attenuate TBI-induced functional outcome and cell death through maintaining normal mitochondrial morphology and inhibiting activation of apoptosis.

  10. Mitochondrial division inhibitor 1 (Mdivi-1) offers neuroprotection through diminishing cell death and improving functional outcome in a mouse model of traumatic brain injury.

    PubMed

    Wu, Qiong; Xia, Shui-Xiu; Li, Qian-Qian; Gao, Yuan; Shen, Xi; Ma, Lu; Zhang, Ming-Yang; Wang, Tao; Li, Yong-Sheng; Wang, Zu-Feng; Luo, Cheng-Liang; Tao, Lu-Yang

    2016-01-01

    Mitochondria dysfunction, an enormous potential crisis, has attracted increasing attention. Disturbed regulation of mitochondrial dynamics, the balance of mitochondrial fusion and fission, has been implicated in neurodegenerative diseases, such as Parkinson׳s disease and cerebral ischemia/reperfusion. However the role of mitochondrial dynamics in traumatic brain injury (TBI) has not been illuminated. The aim of the present study was to investigate the role of Mdivi-1, a small molecule inhibitor of a key mitochondrial fission protein dynamin-related protein 1 (Drp1), in TBI-induced cell death and functional outcome deficits. Protein expression of Drp1 was first investigated. Outcome parameters consist of motor test, Morris water maze, brain edema and lesion volume. Cell death was detected by propidium iodide (PI) labeling, and mitochondrial morphology was assessed using transmission electron microscopy. In addition, the expression of apoptosis-related proteins cytochrome c (cyt-c) and caspase-3 was investigated. Our findings showed that up-regulation of Drp1 expression started at 1h post-TBI and peaked at 24 h, but inhibition of Drp1 by Mdivi-1 significantly alleviated TBI-induced behavioral deficits and brain edema, reduced morphological change of mitochondria, and decreased TBI-induced cell death together with lesion volume. Moreover, treatment with Mdivi-1 remarkably inhibited TBI-induced the release of cyt-c from mitochondria to cytoplasm, and activation of caspase-3 at 24 h after TBI. Taken together, these data imply that inhibition of Drp1 may help attenuate TBI-induced functional outcome and cell death through maintaining normal mitochondrial morphology and inhibiting activation of apoptosis. PMID:26596858

  11. VOLTAGE REGULATOR

    DOEpatents

    Von Eschen, R.L.; Scheele, P.F.

    1962-04-24

    A transistorized voltage regulator which provides very close voitage regulation up to about 180 deg F is described. A diode in the positive line provides a constant voltage drop from the input to a regulating transistor emitter. An amplifier is coupled to the positive line through a resistor and is connected between a difference circuit and the regulating transistor base which is negative due to the difference in voltage drop across thc diode and the resistor so that a change in the regulator output causes the amplifier to increase or decrease the base voltage and current and incrcase or decrease the transistor impedance to return the regulator output to normal. (AEC)

  12. Mitochondrial Diseases

    MedlinePlus

    ... disorder, something goes wrong with this process. Mitochondrial diseases are a group of metabolic disorders. Mitochondria are ... cells and cause damage. The symptoms of mitochondrial disease can vary. It depends on how many mitochondria ...

  13. Acute blockage of voltage-gated K⁺ currents by 17β-estradiol in mouse neuroblastoma N2A cells.

    PubMed

    Li, Xiaoqing; Hao, Xuran; Cheng, Bo; Li, Xiantao

    2014-05-28

    In this study, whole-cell recording was carried out to explore the effects of 17β-estradiol on voltage-gated K⁺ (Kv) currents in N2A cells. The acute exposure to 17β-estradiol, in a concentration-dependent manner, significantly inhibited the peak and steady-state currents through Kv channels, showing IC50 values of 3.6 and 3.8 μM, respectively. The reduction in both the amplitude and the decay rate of Kv currents, with an increase in depolarization, suggested that it was a voltage-dependent block. The activation and inactivation experiments were conducted to determine the exact causes of the inhibitory effects. The half-maximum activation potential (V₁/₂) was +8.1 mV in control and remained stable after exposure to 10 μM 17β-estradiol. For steady-state inactivation, the half-maximum inactivation potential (V₁/₂) was -45.0 mV and shifted right to -39.7 mV without a statistical difference, and the time constants of recovery from inactivation were not altered by 17β-estradiol, suggesting that the depression was not correlated with the inactivation gate. PMID:24784585

  14. Mitochondrial and nuclear DNA-repair capacity of various brain regions in mouse is altered in an age-dependent manner.

    PubMed

    Imam, Syed Z; Karahalil, Bensu; Hogue, Barbara A; Souza-Pinto, Nadja C; Bohr, Vilhelm A

    2006-08-01

    Aging is associated with increased susceptibility to neuronal loss and disruption of cerebral function either as a component of senescence, or as a consequence of neurodegenerative disease or stroke. Here we report differential changes in the repair of oxidative DNA damage in various brain regions during aging. We evaluated mitochondrial and nuclear incision activities of oxoguanine DNA glycosylase (OGG1), uracil DNA glycosylase (UDG) and the endonuclease III homologue (NTH1) in the caudate nucleus (CN), frontal cortex (FC), hippocampus (Hip), cerebellum (CE) and brain stem (BS) of 6- and 18-month-old male C57Bl/6 mice. We observed a significant age-dependent decrease in incision activities of all three glycosylases in the mitochondria of all brain regions, whereas variable patterns of changes were seen in nuclei. No age- or region-specific changes were observed in the mitochondrial repair synthesis incorporation of uracil-initiated base-excision repair (BER). We did not observe any age or region dependent differences in levels of BER proteins among the five brain regions. In summary, our data suggest that a decreased efficiency of mitochondrial BER-glycosylases and increased oxidative damage to mitochondrial DNA might contribute to the normal aging process. These data provide a novel characterization of oxidative DNA damage processing in different brain regions implicated in various neurodegenerative disorders, and suggest that this process is regulated in an age-dependent manner. Manipulation of DNA repair mechanisms may provide a strategy to prevent neuronal loss during age-dependent neurodegenerative disorders. PMID:16005114

  15. PM2.5, SO2 and NO2 co-exposure impairs neurobehavior and induces mitochondrial injuries in the mouse brain.

    PubMed

    Ku, Tingting; Ji, Xiaotong; Zhang, Yingying; Li, Guangke; Sang, Nan

    2016-11-01

    Air pollution is a serious environmental health problem that has been previously associated with neuropathological disorders. However, current experimental evidence mainly focuses on the adverse effects of a single air pollutant, ignoring the biological responses to the co-existence of these pollutants. In the present study, we co-exposed C57BL/6 J mice to PM2.5, SO2 and NO2 and explored their neurobehavior, histopathologic abnormalities, apoptosis-related protein expression and mitochondrial dysfunction. The results indicate that co-exposure to PM2.5, SO2 and NO2 impaired spatial learning and memory and caused abnormal expression of apoptosis-related genes (p53, bax and bcl-2). Additionally, these alterations were related to morphological changes in mitochondria, a reduction of ATP, the elevation of mitochondrial fission proteins and the downregulation of fusion proteins. These findings provide a basis for the understanding of mitochondrial abnormality-related neuropathological dysfunction in response to co-exposure to ambient air pollutants, which suggests an adaptive response to the frangibility of the central nerve system. PMID:27521637

  16. PM2.5, SO2 and NO2 co-exposure impairs neurobehavior and induces mitochondrial injuries in the mouse brain.

    PubMed

    Ku, Tingting; Ji, Xiaotong; Zhang, Yingying; Li, Guangke; Sang, Nan

    2016-11-01

    Air pollution is a serious environmental health problem that has been previously associated with neuropathological disorders. However, current experimental evidence mainly focuses on the adverse effects of a single air pollutant, ignoring the biological responses to the co-existence of these pollutants. In the present study, we co-exposed C57BL/6 J mice to PM2.5, SO2 and NO2 and explored their neurobehavior, histopathologic abnormalities, apoptosis-related protein expression and mitochondrial dysfunction. The results indicate that co-exposure to PM2.5, SO2 and NO2 impaired spatial learning and memory and caused abnormal expression of apoptosis-related genes (p53, bax and bcl-2). Additionally, these alterations were related to morphological changes in mitochondria, a reduction of ATP, the elevation of mitochondrial fission proteins and the downregulation of fusion proteins. These findings provide a basis for the understanding of mitochondrial abnormality-related neuropathological dysfunction in response to co-exposure to ambient air pollutants, which suggests an adaptive response to the frangibility of the central nerve system.

  17. Blockade by ifenprodil of high voltage-activated Ca2+ channels in rat and mouse cultured hippocampal pyramidal neurones: comparison with N-methyl-D-aspartate receptor antagonist actions.

    PubMed Central

    Church, J; Fletcher, E J; Baxter, K; MacDonald, J F

    1994-01-01

    1. The block by ifenprodil of voltage-activated Ca2+ channels was investigated in intracellular free calcium concentration ([Ca2+]i) evoked by 50 mM K+ (high-[K+]o) in Fura-2-loaded rat hippocampal pyramidal neurones in culture and on currents carried by Ba2+ ions (IBa) through Ca2+ channels in mouse cultured hippocampal neurones under whole-cell voltage-clamp. The effects of ifenprodil on voltage-activated Ca2+ channels were compared with its antagonist actions on N-methyl-D-aspartate- (NMDA) evoked responses in the same neuronal preparations. 2. Rises in [Ca2+]i evoked by transient exposure to high-[K+]o in our preparation of rat cultured hippocampal pyramidal neurones are mediated predominantly by Ca2+ flux through nifedipine-sensitive Ca2+ channels, with smaller contributions from nifedipine-resistant, omega-conotoxin GVIA-sensitive Ca2+ channels and Ca2+ channels sensitive to crude funnel-web spider venom (Church et al., 1994). Ifenprodil (0.1-200 microM) reversibly attenuated high-[K+]o-evoked rises in [Ca2+]i with an IC50 value of 17 +/- 3 microM, compared with an IC50 value of 0.7 +/- 0.1 microM for the reduction of rises in [Ca2+]i evoked by 20 microM NMDA. Tested in the presence of nifedipine 10 microM, ifenprodil (1-50 microM) produced a concentration-dependent reduction of the dihydropyridine-resistant high-[K+]o-evoked rise in [Ca2+]i with an IC50 value of 13 +/- 4 microM. The results suggest that ifenprodil blocks Ca2+ flux through multiple subtypes of high voltage-activated Ca2+ channels. 3. Application of the polyamine, spermine (0.25-5 mM), produced a concentration-dependent reduction of rises in [Ca2+]i evoked by high-[K+]o.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7834201

  18. Blocking mitochondrial calcium release in Schwann cells prevents demyelinating neuropathies.

    PubMed

    Gonzalez, Sergio; Berthelot, Jade; Jiner, Jennifer; Perrin-Tricaud, Claire; Fernando, Ruani; Chrast, Roman; Lenaers, Guy; Tricaud, Nicolas

    2016-03-01

    Schwann cells produce myelin sheath around peripheral nerve axons. Myelination is critical for rapid propagation of action potentials, as illustrated by the large number of acquired and hereditary peripheral neuropathies, such as diabetic neuropathy or Charcot-Marie-Tooth diseases, that are commonly associated with a process of demyelination. However, the early molecular events that trigger the demyelination program in these diseases remain unknown. Here, we used virally delivered fluorescent probes and in vivo time-lapse imaging in a mouse model of demyelination to investigate the underlying mechanisms of the demyelination process. We demonstrated that mitochondrial calcium released by voltage-dependent anion channel 1 (VDAC1) after sciatic nerve injury triggers Schwann cell demyelination via ERK1/2, p38, JNK, and c-JUN activation. In diabetic mice, VDAC1 activity was altered, resulting in a mitochondrial calcium leak in Schwann cell cytoplasm, thereby priming the cell for demyelination. Moreover, reduction of mitochondrial calcium release, either by shRNA-mediated VDAC1 silencing or pharmacological inhibition, prevented demyelination, leading to nerve conduction and neuromuscular performance recovery in rodent models of diabetic neuropathy and Charcot-Marie-Tooth diseases. Therefore, this study identifies mitochondria as the early key factor in the molecular mechanism of peripheral demyelination and opens a potential opportunity for the treatment of demyelinating peripheral neuropathies. PMID:26878172

  19. Blocking mitochondrial calcium release in Schwann cells prevents demyelinating neuropathies

    PubMed Central

    Berthelot, Jade; Jiner, Jennifer; Perrin-Tricaud, Claire; Fernando, Ruani; Chrast, Roman; Lenaers, Guy

    2016-01-01

    Schwann cells produce myelin sheath around peripheral nerve axons. Myelination is critical for rapid propagation of action potentials, as illustrated by the large number of acquired and hereditary peripheral neuropathies, such as diabetic neuropathy or Charcot-Marie-Tooth diseases, that are commonly associated with a process of demyelination. However, the early molecular events that trigger the demyelination program in these diseases remain unknown. Here, we used virally delivered fluorescent probes and in vivo time-lapse imaging in a mouse model of demyelination to investigate the underlying mechanisms of the demyelination process. We demonstrated that mitochondrial calcium released by voltage-dependent anion channel 1 (VDAC1) after sciatic nerve injury triggers Schwann cell demyelination via ERK1/2, p38, JNK, and c-JUN activation. In diabetic mice, VDAC1 activity was altered, resulting in a mitochondrial calcium leak in Schwann cell cytoplasm, thereby priming the cell for demyelination. Moreover, reduction of mitochondrial calcium release, either by shRNA-mediated VDAC1 silencing or pharmacological inhibition, prevented demyelination, leading to nerve conduction and neuromuscular performance recovery in rodent models of diabetic neuropathy and Charcot-Marie-Tooth diseases. Therefore, this study identifies mitochondria as the early key factor in the molecular mechanism of peripheral demyelination and opens a potential opportunity for the treatment of demyelinating peripheral neuropathies. PMID:26878172

  20. Mitochondrial Cardiomyopathies

    PubMed Central

    El-Hattab, Ayman W.; Scaglia, Fernando

    2016-01-01

    Mitochondria are found in all nucleated human cells and perform various essential functions, including the generation of cellular energy. Mitochondria are under dual genome control. Only a small fraction of their proteins are encoded by mitochondrial DNA (mtDNA), whereas more than 99% of them are encoded by nuclear DNA (nDNA). Mutations in mtDNA or mitochondria-related nDNA genes result in mitochondrial dysfunction leading to insufficient energy production required to meet the needs for various organs, particularly those with high energy requirements, including the central nervous system, skeletal and cardiac muscles, kidneys, liver, and endocrine system. Because cardiac muscles are one of the high energy demanding tissues, cardiac involvement occurs in mitochondrial diseases with cardiomyopathies being one of the most frequent cardiac manifestations found in these disorders. Cardiomyopathy is estimated to occur in 20–40% of children with mitochondrial diseases. Mitochondrial cardiomyopathies can vary in severity from asymptomatic status to severe manifestations including heart failure, arrhythmias, and sudden cardiac death. Hypertrophic cardiomyopathy is the most common type; however, mitochondrial cardiomyopathies might also present as dilated, restrictive, left ventricular non-compaction, and histiocytoid cardiomyopathies. Cardiomyopathies are frequent manifestations of mitochondrial diseases associated with defects in electron transport chain complexes subunits and their assembly factors, mitochondrial transfer RNAs, ribosomal RNAs, ribosomal proteins, translation factors, mtDNA maintenance, and coenzyme Q10 synthesis. Other mitochondrial diseases with cardiomyopathies include Barth syndrome, Sengers syndrome, TMEM70-related mitochondrial complex V deficiency, and Friedreich ataxia. PMID:27504452

  1. Mitochondrial Cardiomyopathies.

    PubMed

    El-Hattab, Ayman W; Scaglia, Fernando

    2016-01-01

    Mitochondria are found in all nucleated human cells and perform various essential functions, including the generation of cellular energy. Mitochondria are under dual genome control. Only a small fraction of their proteins are encoded by mitochondrial DNA (mtDNA), whereas more than 99% of them are encoded by nuclear DNA (nDNA). Mutations in mtDNA or mitochondria-related nDNA genes result in mitochondrial dysfunction leading to insufficient energy production required to meet the needs for various organs, particularly those with high energy requirements, including the central nervous system, skeletal and cardiac muscles, kidneys, liver, and endocrine system. Because cardiac muscles are one of the high energy demanding tissues, cardiac involvement occurs in mitochondrial diseases with cardiomyopathies being one of the most frequent cardiac manifestations found in these disorders. Cardiomyopathy is estimated to occur in 20-40% of children with mitochondrial diseases. Mitochondrial cardiomyopathies can vary in severity from asymptomatic status to severe manifestations including heart failure, arrhythmias, and sudden cardiac death. Hypertrophic cardiomyopathy is the most common type; however, mitochondrial cardiomyopathies might also present as dilated, restrictive, left ventricular non-compaction, and histiocytoid cardiomyopathies. Cardiomyopathies are frequent manifestations of mitochondrial diseases associated with defects in electron transport chain complexes subunits and their assembly factors, mitochondrial transfer RNAs, ribosomal RNAs, ribosomal proteins, translation factors, mtDNA maintenance, and coenzyme Q10 synthesis. Other mitochondrial diseases with cardiomyopathies include Barth syndrome, Sengers syndrome, TMEM70-related mitochondrial complex V deficiency, and Friedreich ataxia. PMID:27504452

  2. Mitochondrial vasculopathy

    PubMed Central

    Finsterer, Josef; Zarrouk-Mahjoub, Sinda

    2016-01-01

    Mitochondrial disorders (MIDs) are usually multisystem disorders (mitochondrial multiorgan disorder syndrome) either on from onset or starting at a point during the disease course. Most frequently affected tissues are those with a high oxygen demand such as the central nervous system, the muscle, endocrine glands, or the myocardium. Recently, it has been shown that rarely also the arteries may be affected (mitochondrial arteriopathy). This review focuses on the type, diagnosis, and treatment of mitochondrial vasculopathy in MID patients. A literature search using appropriate search terms was carried out. Mitochondrial vasculopathy manifests as either microangiopathy or macroangiopathy. Clinical manifestations of mitochondrial microangiopathy include leukoencephalopathy, migraine-like headache, stroke-like episodes, or peripheral retinopathy. Mitochondrial macroangiopathy manifests as atherosclerosis, ectasia of arteries, aneurysm formation, dissection, or spontaneous rupture of arteries. The diagnosis relies on the documentation and confirmation of the mitochondrial metabolic defect or the genetic cause after exclusion of non-MID causes. Treatment is not at variance compared to treatment of vasculopathy due to non-MID causes. Mitochondrial vasculopathy exists and manifests as micro- or macroangiopathy. Diagnosing mitochondrial vasculopathy is crucial since appropriate treatment may prevent from severe complications. PMID:27231520

  3. Apaf-1-deficient fog mouse cell apoptosis involves hypo-polarization of the mitochondrial inner membrane, ATP depletion and citrate accumulation.

    PubMed

    Katoh, Iyoko; Sato, Shingo; Fukunishi, Nahoko; Yoshida, Hiroki; Imai, Takasuke; Kurata, Shun-Ichi

    2008-12-01

    To explore how the intrinsic apoptosis pathway is controlled in the spontaneous fog (forebrain overgrowth) mutant mice with an Apaf1 splicing deficiency, we examined spleen and bone marrow cells from Apaf1(+/+) (+/+) and Apaf1(fog/fog) (fog/fog) mice for initiator caspase-9 activation by cellular stresses. When the mitochondrial inner membrane potential (Deltapsim) was disrupted by staurosporine, +/+ cells but not fog/fog cells activated caspase-9 to cause apoptosis, indicating the lack of apoptosome (apoptosis protease activating factor 1 (Apaf-1)/cytochrome c/(d)ATP/procaspase-9) function in fog/fog cells. However, when a marginal ( approximately 20%) decrease in Deltapsim was caused by hydrogen peroxide (0.1 mM), peroxynitritedonor 3-morpholinosydnonimine (0.1 mM) and UV-C irradiation (20 J/m(2)), both +/+ and fog/fog cells triggered procaspase-9 auto-processing and its downstream cascade activation. Supporting our previous results, procaspase-9 pre-existing in the mitochondria induced its auto-processing before the cytosolic caspase activation regardless of the genotypes. Cellular ATP concentration significantly decreased under the hypoactive Deltapsim condition. Furthermore, we detected accumulation of citrate, a kosmotrope known to facilitate procaspase-9 dimerization, probably due to a feedback control of the Krebs cycle by the electron transfer system. Thus, mitochondrial in situ caspase-9 activation may be caused by the major metabolic reactions in response to physiological stresses, which may represent a mode of Apaf-1-independent apoptosis hypothesized from recent genetic studies.

  4. Mitochondrial Cristae: Where Beauty Meets Functionality.

    PubMed

    Cogliati, Sara; Enriquez, Jose A; Scorrano, Luca

    2016-03-01

    Mitochondrial cristae are dynamic bioenergetic compartments whose shape changes under different physiological conditions. Recent discoveries have unveiled the relation between cristae shape and oxidative phosphorylation (OXPHOS) function, suggesting that membrane morphology modulates the organization and function of the OXPHOS system, with a direct impact on cellular metabolism. As a corollary, cristae-shaping proteins have emerged as potential modulators of mitochondrial bioenergetics, a concept confirmed by genetic experiments in mouse models of respiratory chain deficiency. Here, we review our knowledge of mitochondrial ultrastructural organization and how it impacts mitochondrial metabolism.

  5. Mitochondrial Cristae: Where Beauty Meets Functionality.

    PubMed

    Cogliati, Sara; Enriquez, Jose A; Scorrano, Luca

    2016-03-01

    Mitochondrial cristae are dynamic bioenergetic compartments whose shape changes under different physiological conditions. Recent discoveries have unveiled the relation between cristae shape and oxidative phosphorylation (OXPHOS) function, suggesting that membrane morphology modulates the organization and function of the OXPHOS system, with a direct impact on cellular metabolism. As a corollary, cristae-shaping proteins have emerged as potential modulators of mitochondrial bioenergetics, a concept confirmed by genetic experiments in mouse models of respiratory chain deficiency. Here, we review our knowledge of mitochondrial ultrastructural organization and how it impacts mitochondrial metabolism. PMID:26857402

  6. Mitochondrial biogenesis: pharmacological approaches.

    PubMed

    Valero, Teresa

    2014-01-01

    neurodevelopmental disorders. In the context of neural differentiation, Martine Uittenbogaard and Anne Chiaramello (Department of Anatomy and Regenerative Biology, George Washington University School of Medicine and Health Sciences, USA) [7] thoroughly describe the implication of mitochondrial biogenesis on neuronal differentiation, its timing, its regulation by specific signaling pathways and new potential therapeutic strategies. The maintenance of mitochondrial homeostasis is crucial for neuronal development. A mitochondrial dynamic balance is necessary between mitochondrial fusion, fission and quality control systems and mitochondrial biogenesis. Concerning the signaling pathways leading to mitochondrial biogenesis this review highlights the implication of different regulators such as AMPK, SIRT1, PGC-1α, NRF1, NRF2, Tfam, etc. on the specific case of neuronal development, providing examples of diseases in which these pathways are altered and transgenic mouse models lacking these regulators. A common hallmark of several neurodegenerative diseases (Huntington´s Disease, Alzheimer´s Disease and Parkinson´s Disease) is the impaired function or expression of PGC-1α, the master regulator of mitochondrial biogenesis. Among the promising strategies to ameliorate mitochondrial-based diseases these authors highlight the induction of PGC-1α via activation of PPAR receptors (rosiglitazone, bezafibrate) or modulating its activity by AMPK (AICAR, metformin, resveratrol) or SIRT1 (SRT1720 and several isoflavone-derived compounds). This article also presents a review of the current animal and cellular models useful to study mitochondriogenesis. Although it is known that many neurodegenerative and neurodevelopmental diseases are originated in mitochondria, the regulation of mitochondrial biogenesis has never been extensively studied. (ABSTRACT TRUNCATED)

  7. Mitochondrial biogenesis: pharmacological approaches.

    PubMed

    Valero, Teresa

    2014-01-01

    neurodevelopmental disorders. In the context of neural differentiation, Martine Uittenbogaard and Anne Chiaramello (Department of Anatomy and Regenerative Biology, George Washington University School of Medicine and Health Sciences, USA) [7] thoroughly describe the implication of mitochondrial biogenesis on neuronal differentiation, its timing, its regulation by specific signaling pathways and new potential therapeutic strategies. The maintenance of mitochondrial homeostasis is crucial for neuronal development. A mitochondrial dynamic balance is necessary between mitochondrial fusion, fission and quality control systems and mitochondrial biogenesis. Concerning the signaling pathways leading to mitochondrial biogenesis this review highlights the implication of different regulators such as AMPK, SIRT1, PGC-1α, NRF1, NRF2, Tfam, etc. on the specific case of neuronal development, providing examples of diseases in which these pathways are altered and transgenic mouse models lacking these regulators. A common hallmark of several neurodegenerative diseases (Huntington´s Disease, Alzheimer´s Disease and Parkinson´s Disease) is the impaired function or expression of PGC-1α, the master regulator of mitochondrial biogenesis. Among the promising strategies to ameliorate mitochondrial-based diseases these authors highlight the induction of PGC-1α via activation of PPAR receptors (rosiglitazone, bezafibrate) or modulating its activity by AMPK (AICAR, metformin, resveratrol) or SIRT1 (SRT1720 and several isoflavone-derived compounds). This article also presents a review of the current animal and cellular models useful to study mitochondriogenesis. Although it is known that many neurodegenerative and neurodevelopmental diseases are originated in mitochondria, the regulation of mitochondrial biogenesis has never been extensively studied. (ABSTRACT TRUNCATED) PMID:24606795

  8. Glucocorticoid Modulation of Mitochondrial Function in Hepatoma Cells Requires the Mitochondrial Fission Protein Drp1

    PubMed Central

    Hernández-Alvarez, María Isabel; Paz, José C.; Sebastián, David; Muñoz, Juan Pablo; Liesa, Marc; Segalés, Jessica; Palacín, Manuel

    2013-01-01

    Abstract Aims: Glucocorticoids, such as dexamethasone, enhance hepatic energy metabolism and gluconeogenesis partly through changes in mitochondrial function. Mitochondrial function is influenced by the balance between mitochondrial fusion and fission events. However, whether glucocorticoids modulate mitochondrial function through the regulation of mitochondrial dynamics is currently unknown. Results: Here, we report that the effects of dexamethasone on mitochondrial function and gluconeogenesis in hepatoma cells are dependent on the mitochondrial fission protein dynamin-related protein 1 (Drp1). Dexamethasone increased routine oxygen consumption, maximal respiratory capacity, superoxide anion, proton leak, and gluconeogenesis in hepatoma cells. Under these conditions, dexamethasone altered mitochondrial morphology, which was paralleled by a large increase in Drp1 expression, and reduced mitofusin 1 (Mfn1) and Mfn2. In vivo dexamethasone treatment also enhanced Drp1 expression in mouse liver. On the basis of these observations, we analyzed the dependence on the Drp1 function of dexamethasone effects on mitochondrial respiration and gluconeogenesis. We show that the increase in mitochondrial respiration and gluconeogenesis induced by dexamethasone are hampered by the inhibition of Drp1 function. Innovation: Our findings provide the first evidence that the effects of glucocorticoids on hepatic metabolism require the mitochondrial fission protein Drp1. Conclusion: In summary, we demonstrate that the mitochondrial effects of dexamethasone both on mitochondrial respiration and on the gluconeogenic pathway depend on Drp1. Antioxid. Redox Signal. 19, 366–378. PMID:22703557

  9. Citrinin-generated reactive oxygen species cause cell cycle arrest leading to apoptosis via the intrinsic mitochondrial pathway in mouse skin.

    PubMed

    Kumar, Rahul; Dwivedi, Premendra D; Dhawan, Alok; Das, Mukul; Ansari, Kausar M

    2011-08-01

    The mycotoxin, citrinin (CTN), is a contaminant of various food and feed materials. Several in vivo and in vitro studies have demonstrated that CTN has broad toxicity spectra; however, dermal toxicity is not known. In the present investigation, dermal exposure to CTN was undertaken to study oxidative stress, DNA damage, cell cycle arrest, and apoptosis in mouse skin. A single topical application of CTN caused significant change in oxidative stress markers, such as lipid peroxidation, protein carbonyl content, glutathione (GSH) content, and antioxidant enzymes in a dose-dependent (25-100 μg/mouse) and time-dependent (12-72 h) manner. Single topical application of CTN (50 μg/mouse) for 12-72 h caused significant enhancement in (1) reactive oxygen species (ROS); (2) cell cycle arrest at the G0/G1 phase (30-71%) and G2/M phase (56-65%) along with the induction of apoptosis (3.6-27%); (3) expression of p53, p21/waf1; (4) Bax/Bcl₂ ratio and cytochome c release; and (5) activities of caspase 9 (22-46%) and 3 (42-54%) as well as increased poly(ADP-ribose) polymerase cleavage. It was also observed that pretreatment with bio-antioxidants viz butylated hydroxyanisole (55 μmol/100 μl), quercetin (10 μmol/100 μl), or α-tocopherol (40 μmol/100 μl) resulted in decreases of ROS generation, arrest in the G0/G1 phase of the cell cycle, and apoptosis. These data confirm the involvement of ROS in apoptosis and suggest that these bio-antioxidants may be useful in the prevention of CTN-induced dermal toxicity.

  10. The ubiquitous mitochondrial creatine kinase gene maps to a conserved region on human chromosome 15q15 and mouse chromosome 2 bands F1-F3

    SciTech Connect

    Steeghs, K.; Wieringa, B.; Merkx, G.

    1994-11-01

    Members of the creatine kinase isoenzyme family (CKs; EC 2.7.3.2) are found in mitochondria and specialized subregions of the cytoplasm and catalyze the reversible exchange of high-energy phosphoryl between ATP and phosphocreatine. At least four functionally active genes, which encode the distinct CK subunits CKB, CKM, CKMT1 (ubiquitous), and CKMT2 (sarcomeric), and a variable number of CKB pseudogenes have been identified. Here, we report the use of a CKMT1 containing phage to map the CKMT1 gene by in situ hybridization on both human and mouse chromosomes.

  11. Mitochondrial HMG to CoA synthase (mHS): cDNA cloning in human, mouse and C. elegans, mapping to human chromosome 1p12-13 and partial human genomic cloning

    SciTech Connect

    Boukaftane, Y.; Robert, M.F.; Mitchell, G.A. |

    1994-09-01

    mHS catalyzes the rate-limiting first step of ketogenesis in the liver. A cytoplasmic HS isozyme, encoded by another gene, catalyzes an early step in cholesterol synthesis. Starting from a rat mHS cDNA obtained by RT-PCR from the published rat cDNA sequence, we obtained and sequenced human and mouse cDNAs spanning the entire coding sequence of natural human and mouse mHS, as well as sequencing C. elegans HS-like cDNA. Consensus sequences for 3 mitochondrial and 4 cytoplasmic HSs were created and compared to invertebrate HS sequences. We found high conversation in the active site and at other regions presumably important for HS function. We mapped the mHS locus, HMGCS2 by in situ hybridization to chromosome 1P12-13, in contrast to the human cHS locus (HMGCS1) known to be on chromosome 5p13. Comparative mapping results suggest that these two chromosomal regions may be contiguous in other species, constant with a recent gene duplication event. Furthermore, we have characterized a human genomic mHS subclone containing 4 mHS exons, and found the position of all splice junctions to be identical to that of the hamster cHS gene except for one site in the 3{prime} nontranslated region. We calculate that the mHS and cHS genes were derived from a common ancestor 400-700 Myrs ago, implying that ketogenesis from fat may have become possible around the time of emergence of vertebrates ({approximately}500 Myr ago). Ketogenesis has evolved into an important pathway of energy metabolism, and we predict the mHS deficiency may prove to be responsible for some as yet explained cases of Reye-like syndromes in humans. This hypothesis can now be tested at the molecular level without the necessity of obtaining hepatic tissue.

  12. Mitochondrial cytopathies.

    PubMed

    El-Hattab, Ayman W; Scaglia, Fernando

    2016-09-01

    Mitochondria are found in all nucleated human cells and perform a variety of essential functions, including the generation of cellular energy. Most of mitochondrial proteins are encoded by the nuclear DNA (nDNA) whereas a very small fraction is encoded by the mitochondrial DNA (mtDNA). Mutations in mtDNA or mitochondria-related nDNA genes can result in mitochondrial dysfunction which leads to a wide range of cellular perturbations including aberrant calcium homeostasis, excessive reactive oxygen species production, dysregulated apoptosis, and insufficient energy generation to meet the needs of various organs, particularly those with high energy demand. Impaired mitochondrial function in various tissues and organs results in the multi-organ manifestations of mitochondrial diseases including epilepsy, intellectual disability, skeletal and cardiac myopathies, hepatopathies, endocrinopathies, and nephropathies. Defects in nDNA genes can be inherited in an autosomal or X-linked manners, whereas, mtDNA is maternally inherited. Mitochondrial diseases can result from mutations of nDNA genes encoding subunits of the electron transport chain complexes or their assembly factors, proteins associated with the mitochondrial import or networking, mitochondrial translation factors, or proteins involved in mtDNA maintenance. MtDNA defects can be either point mutations or rearrangements. The diagnosis of mitochondrial disorders can be challenging in many cases and is based on clinical recognition, biochemical screening, histopathological studies, functional studies, and molecular genetic testing. Currently, there are no satisfactory therapies available for mitochondrial disorders that significantly alter the course of the disease. Therapeutic options include symptomatic treatment, cofactor supplementation, and exercise. PMID:26996063

  13. The ins and outs of mitochondrial calcium.

    PubMed

    Finkel, Toren; Menazza, Sara; Holmström, Kira M; Parks, Randi J; Liu, Julia; Sun, Junhui; Liu, Jie; Pan, Xin; Murphy, Elizabeth

    2015-05-22

    Calcium is thought to play an important role in regulating mitochondrial function. Evidence suggests that an increase in mitochondrial calcium can augment ATP production by altering the activity of calcium-sensitive mitochondrial matrix enzymes. In contrast, the entry of large amounts of mitochondrial calcium in the setting of ischemia-reperfusion injury is thought to be a critical event in triggering cellular necrosis. For many decades, the details of how calcium entered the mitochondria remained a biological mystery. In the past few years, significant progress has been made in identifying the molecular components of the mitochondrial calcium uniporter complex. Here, we review how calcium enters and leaves the mitochondria, the growing insight into the topology, stoichiometry and function of the uniporter complex, and the early lessons learned from some initial mouse models that genetically perturb mitochondrial calcium homeostasis.

  14. Targeting a mitochondrial potassium channel to fight cancer.

    PubMed

    Leanza, Luigi; Venturini, Elisa; Kadow, Stephanie; Carpinteiro, Alexander; Gulbins, Erich; Becker, Katrin Anne

    2015-07-01

    Although chemotherapy is able to cure many patients with malignancies, it still also often fails. Therefore, novel approaches and targets for chemotherapeutic treatment of malignancies are urgently required. Recent studies demonstrated the expression of several potassium channels in the inner mitochondrial membrane. Among them the voltage gated potassium channel Kv1.3 and the big-potassium (BK) channel were shown to directly function in cell death by serving as target for pro-apoptotic Bax and Bak proteins. Here, we discuss the role of mitochondrial potassium channel Kv1.3 (mitoKv1.3) in cell death and its potential function in treatment of solid tumors, leukemia and lymphoma. Bax and Bak inhibit mitoKv1.3 by directly binding into the pore of the channel, by a toxin-like mechanism. Inhibition of mitoKv1.3 results in an initial hyperpolarization of the inner mitochondrial membrane that triggers the production of reactive oxygen species (ROS). ROS in turn induce a release of cytochrome c from the cristae of the inner mitochondrial membrane and an activation of the permeability transition pore, resulting in opening of the intrinsic apoptotic cell death. Since mitoKv1.3 functions downstream of pro-apoptotic Bax and Bak, compounds that directly inhibit mitoKv1.3 may serve as a new class of drugs for treatment of tumors, even with an altered expression of either pro- or anti-apoptotic Bcl-2 protein family members. This was successfully proven by the in vivo treatment of mouse melanoma and ex vivo human chronic leukemia B cells with inhibitors of mitoKv1.3.

  15. Mitochondrial Myopathy

    MedlinePlus

    ... with ragged-red fibers, and mitochondrial encephalomyopathy with lactic acidosis and stroke-like episodes. The symptoms of ... riboflavin, coenzyme Q, and carnitine (a specialized amino acid) may provide subjective improvement in fatigue and energy ...

  16. Mitochondrial DNA.

    ERIC Educational Resources Information Center

    Wright, Russell G.; Bottino, Paul J.

    1986-01-01

    Provides background information for teachers on mitochondrial DNA, pointing out that it may have once been a free-living organism. Includes a ready-to-duplicate exercise titled "Using Microchondrial DNA to Measure Evolutionary Distance." (JN)

  17. Mitochondrial Myopathies

    MedlinePlus

    ... line and are therefore called the electron transport chain, and complex V actually churns out ATP, so ... coQ10 , is a component of the electron transport chain, which uses oxygen to manufacture ATP. Some mitochondrial ...

  18. Mitochondrial impairment induced by postnatal ActRIIB blockade does not alter function and energy status in exercising mouse glycolytic muscle in vivo.

    PubMed

    Béchir, Nelly; Pecchi, Émilie; Relizani, Karima; Vilmen, Christophe; Le Fur, Yann; Bernard, Monique; Amthor, Helge; Bendahan, David; Giannesini, Benoît

    2016-04-01

    Because it leads to a rapid and massive muscle hypertrophy, postnatal blockade of the activin type IIB receptor (ActRIIB) is a promising therapeutic strategy for counteracting muscle wasting. However, the functional consequences remain very poorly documented in vivo. Here, we have investigated the impact of 8-wk ActRIIB blockade with soluble receptor (sActRIIB-Fc) on gastrocnemius muscle anatomy, energy metabolism, and force-generating capacity in wild-type mice, using totally noninvasive magnetic resonance imaging (MRI) and dynamic(31)P-MRS. Compared with vehicle (PBS) control, sActRIIB-Fc treatment resulted in a dramatic increase in body weight (+29%) and muscle volume (+58%) calculated from hindlimb MR imaging, but did not alter fiber type distribution determined via myosin heavy chain isoform analysis. In resting muscle, sActRIIB-Fc treatment induced acidosis and PCr depletion, thereby suggesting reduced tissue oxygenation. During an in vivo fatiguing exercise (6-min repeated maximal isometric contraction electrically induced at 1.7 Hz), maximal and total absolute forces were larger in sActRIIB-Fc treated animals (+26 and +12%, respectively), whereas specific force and fatigue resistance were lower (-30 and -37%, respectively). Treatment with sActRIIB-Fc further decreased the maximal rate of oxidative ATP synthesis (-42%) and the oxidative capacity (-34%), but did not alter the bioenergetics status in contracting muscle. Our findings demonstrate in vivo that sActRIIB-Fc treatment increases absolute force-generating capacity and reduces mitochondrial function in glycolytic gastrocnemius muscle, but this reduction does not compromise energy status during sustained activity. Overall, these data support the clinical interest of postnatal ActRIIB blockade.

  19. Validation of Mitochondrial Gene Delivery in Liver and Skeletal Muscle via Hydrodynamic Injection Using an Artificial Mitochondrial Reporter DNA Vector.

    PubMed

    Yasuzaki, Yukari; Yamada, Yuma; Ishikawa, Takuya; Harashima, Hideyoshi

    2015-12-01

    For successful mitochondrial transgene expression, two independent processes, i.e., developing a mitochondrial gene delivery system and construction of DNA vector to achieve mitochondrial gene expression, are required. To date, very few studies dealing with mitochondrial gene delivery have been reported and, in most cases, transgene expression was not validated, because the construction of a reporter DNA vector for mitochondrial gene expression is the bottleneck. In this study, mitochondrial transgene expression by the in vivo mitochondrial gene delivery of an artificial mitochondrial reporter DNA vector via hydrodynamic injection is demonstrated. In the procedure, a large volume of naked plasmid DNA (pDNA) is rapidly injected. We designed and constructed pHSP-mtLuc (CGG) as a mitochondrial reporter DNA vector that possesses a mitochondrial heavy strand promoter (HSP) and an artificial mitochondrial genome with the reporter NanoLuc (Nluc) luciferase gene that records adjustments to the mitochondrial codon system. We delivered the pDNA into mouse liver mitochondria by hydrodynamic injection, and detected exogenous mRNA in the liver using reverse transcription PCR analysis. The hydrodynamic injection of pHSP-mtLuc (CGG) resulted in the expression of the Nluc luciferase protein in liver and skeletal muscle. Our mitochondrial transgene expression reporter system would contribute to mitochondrial gene therapy and further studies directed at mitochondrial molecular biology.

  20. Mitochondrial genetics

    PubMed Central

    Chinnery, Patrick Francis; Hudson, Gavin

    2013-01-01

    Introduction In the last 10 years the field of mitochondrial genetics has widened, shifting the focus from rare sporadic, metabolic disease to the effects of mitochondrial DNA (mtDNA) variation in a growing spectrum of human disease. The aim of this review is to guide the reader through some key concepts regarding mitochondria before introducing both classic and emerging mitochondrial disorders. Sources of data In this article, a review of the current mitochondrial genetics literature was conducted using PubMed (http://www.ncbi.nlm.nih.gov/pubmed/). In addition, this review makes use of a growing number of publically available databases including MITOMAP, a human mitochondrial genome database (www.mitomap.org), the Human DNA polymerase Gamma Mutation Database (http://tools.niehs.nih.gov/polg/) and PhyloTree.org (www.phylotree.org), a repository of global mtDNA variation. Areas of agreement The disruption in cellular energy, resulting from defects in mtDNA or defects in the nuclear-encoded genes responsible for mitochondrial maintenance, manifests in a growing number of human diseases. Areas of controversy The exact mechanisms which govern the inheritance of mtDNA are hotly debated. Growing points Although still in the early stages, the development of in vitro genetic manipulation could see an end to the inheritance of the most severe mtDNA disease. PMID:23704099

  1. Increased mitochondrial ATP production capacity in brain of healthy mice and a mouse model of isolated complex I deficiency after isoflurane anesthesia.

    PubMed

    Manjeri, Ganesh R; Rodenburg, Richard J; Blanchet, Lionel; Roelofs, Suzanne; Nijtmans, Leo G; Smeitink, Jan A; Driessen, Jacques J; Koopman, Werner J H; Willems, Peter H

    2016-01-01

    We reported before that the minimal alveolar concentration (MAC) of isoflurane is decreased in complex I-deficient mice lacking the NDUFS4 subunit of the respiratory chain (RC) (1.55 and 0.81% at postnatal (PN) 22-25 days and 1.68 and 0.65% at PN 31-34 days for wildtype (WT) and CI-deficient KO, respectively). A more severe respiratory depression was caused by 1.0 MAC isoflurane in KO mice (respiratory rate values of 86 and 45 at PN 22-25 days and 69 and 29 at PN 31-34 days for anesthetized WT and KO, respectively). Here, we address the idea that isoflurane anesthesia causes a much larger decrease in brain mitochondrial ATP production in KO mice thus explaining their increased sensitivity to this anesthetic. Brains from WT and KO mice of the above study were removed immediately after MAC determination at PN 31-34 days and a mitochondria-enriched fraction was prepared. Aliquots were used for measurement of maximal ATP production in the presence of pyruvate, malate, ADP and creatine and, after freeze-thawing, the maximal activity of the individual RC complexes in the presence of complex-specific substrates. CI activity was dramatically decreased in KO, whereas ATP production was decreased by only 26% (p < 0.05). The activities of CII, CIII, and CIV were the same for WT and KO. Isoflurane anesthesia decreased the activity of CI by 30% (p < 0.001) in WT. In sharp contrast, it increased the activity of CII by 37% (p < 0.001) and 50% (p < 0.001) and that of CIII by 37% (p < 0.001) and 40% (p < 0.001) in WT and KO, respectively, whereas it tended to increase that of CIV in both WT and KO. Isoflurane anesthesia increased ATP production by 52 and 69% in WT (p < 0.05) and KO (p < 0.01), respectively. Together these findings indicate that isoflurane anesthesia interferes positively rather than negatively with the ability of CI-deficient mice brain mitochondria to convert their main substrate pyruvate into ATP.

  2. Photoacoustic imaging of voltage signals (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Rao, Bin; Zhang, Ruiying; Wang, Lihong V.

    2016-03-01

    Optical imaging of brain voltage signals is significantly limited in depth due to optical scattering and the absorptive property of brain tissue. Photoacoustic (PA) imaging promises to break this hard limit by utilizing both ballistic and diffused photons. To demonstrate the feasibility of PA, we used an in vivo mouse model. The brain cortex tissue was stained with dipicrylamine dye, electrically stimulated, and imaged with a customized dual-isosbestic-wavelength PA microscope (DIW-PAM). DIW-PAM separates voltage-induced PA signals from blood-induced PA signals and thereby allows recording the voltage response of mouse cortex tissue without interference from hemoglobin responses. The resting state PA voltage response signal exhibited a noise-like signal in the frequency domain. Upon 3 Hz electrical stimulation, the PA voltage response signal showed frequency peaks of 3.2 Hz and 6.3 Hz (Fig. 1). Although dipicrylamine dye is not fast enough for recording neuron action potentials, it served well for the purpose of this feasibility study. In conclusion, we successfully demonstrated in vivo photoacoustic imaging of mouse brain voltage signals for the first time. If a fast voltage-sensitive dye is available, using photoacoustic computed tomography (PACT) instead of PA microscopy could allow acquiring full-field PA action potential images at a speed limited only by the laser pulse repetition rate.

  3. Nicotinamide riboside restores cognition through an upregulation of proliferator-activated receptor-γ coactivator 1α regulated β-secretase 1 degradation and mitochondrial gene expression in Alzheimer's mouse models.

    PubMed

    Gong, Bing; Pan, Yong; Vempati, Prashant; Zhao, Wei; Knable, Lindsay; Ho, Lap; Wang, Jun; Sastre, Magdalena; Ono, Kenjiro; Sauve, Anthony A; Pasinetti, Giulio M

    2013-06-01

    Nicotinamide adenine dinucleotide (NAD)(+), a coenzyme involved in redox activities in the mitochondrial electron transport chain, has been identified as a key regulator of the lifespan-extending effects, and the activation of NAD(+) expression has been linked with a decrease in beta-amyloid (Aβ) toxicity in Alzheimer's disease (AD). Nicotinamide riboside (NR) is a NAD(+) precursor, it promotes peroxisome proliferator-activated receptor-γ coactivator 1 (PGC)-1α expression in the brain. Evidence has shown that PGC-1α is a crucial regulator of Aβ generation because it affects β-secretase (BACE1) degradation. In this study we tested the hypothesis that NR treatment in an AD mouse model could attenuate Aβ toxicity through the activation of PGC-1α-mediated BACE1 degradation. Using the Tg2576 AD mouse model, using in vivo behavioral analyses, biochemistry assays, small hairpin RNA (shRNA) gene silencing and electrophysiological recording, we found (1) dietary treatment of Tg2576 mice with 250 mg/kg/day of NR for 3 months significantly attenuates cognitive deterioration in Tg2576 mice and coincides with an increase in the steady-state levels of NAD(+) in the cerebral cortex; (2) application of NR to hippocampal slices (10 μM) for 4 hours abolishes the deficits in long-term potentiation recorded in the CA1 region of Tg2576 mice; (3) NR treatment promotes PGC-1α expression in the brain coinciding with enhanced degradation of BACE1 and the reduction of Aβ production in Tg2576 mice. Further in vitro studies confirmed that BACE1 protein content is decreased by NR treatment in primary neuronal cultures derived from Tg2576 embryos, in which BACE1 degradation was prevented by PGC-1α-shRNA gene silencing; and (4) NR treatment and PGC-1α overexpression enhance BACE1 ubiquitination and proteasomal degradation. Our studies suggest that dietary treatment with NR might benefit AD cognitive function and synaptic plasticity, in part by promoting PGC-1α-mediated BACE1

  4. Protective effects of 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside in the MPTP-induced mouse model of Parkinson's disease: Involvement of reactive oxygen species-mediated JNK, P38 and mitochondrial pathways.

    PubMed

    He, Hong; Wang, Songhai; Tian, Jiyu; Chen, Lei; Zhang, Wei; Zhao, Junjie; Tang, Haifeng; Zhang, Xiaojun; Chen, Jianzong

    2015-11-15

    Parkinson's disease (PD) is characterized by the selective death of dopaminergic neurons in the substantia nigra pars compacta. Oxidative stress-induced neuron loss is thought to play a crucial role in the pathogenesis of PD. Previous work from our group suggests that 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component extracted from a traditional Chinese herb, Polygonum multiflorum thunb, can attenuate 1-methyl-4-phenyl pyridium-induced apoptosis in the neuronal cell line PC12, by inhibiting reactive oxygen species generation and modulating c-Jun N-terminal kinases (JNK) activation. Here, we investigated the protective effects of TSG against 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP)-induced loss of tyrosine hydroxylase positive cells in mice and the underlying mechanisms. The results showed that MPTP-induced loss of tyrosine hydroxylase positive cells and reactive oxygen species generation were prevented by TSG in a dose-dependent manner. The reactive oxygen species scavenger N-acetylcysteine could also mitigate reactive oxygen species generation. Moreover, JNK and P38 were activated by MPTP, but extracellular signal-regulated protein kinases phosphorylation did not change after MPTP treatment. TSG at different doses blocked the activation of JNK and P38. The protective effect of TSG was also associated with downregulation of the bax/bcl-2 ratio, reversed the release of cytochrome c and smac, and inhibited the activation of caspase-3, -6, and -9 induced by MPTP. In conclusion, our studies demonstrated that the protective effects of TSG in the MPTP-induced mouse model of PD are involved, at least in part, in controlling reactive oxygen species-mediated JNK, P38, and mitochondrial pathways.

  5. Protective effects of 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside in the MPTP-induced mouse model of Parkinson's disease: Involvement of reactive oxygen species-mediated JNK, P38 and mitochondrial pathways.

    PubMed

    He, Hong; Wang, Songhai; Tian, Jiyu; Chen, Lei; Zhang, Wei; Zhao, Junjie; Tang, Haifeng; Zhang, Xiaojun; Chen, Jianzong

    2015-11-15

    Parkinson's disease (PD) is characterized by the selective death of dopaminergic neurons in the substantia nigra pars compacta. Oxidative stress-induced neuron loss is thought to play a crucial role in the pathogenesis of PD. Previous work from our group suggests that 2,3,5,4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG), an active component extracted from a traditional Chinese herb, Polygonum multiflorum thunb, can attenuate 1-methyl-4-phenyl pyridium-induced apoptosis in the neuronal cell line PC12, by inhibiting reactive oxygen species generation and modulating c-Jun N-terminal kinases (JNK) activation. Here, we investigated the protective effects of TSG against 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP)-induced loss of tyrosine hydroxylase positive cells in mice and the underlying mechanisms. The results showed that MPTP-induced loss of tyrosine hydroxylase positive cells and reactive oxygen species generation were prevented by TSG in a dose-dependent manner. The reactive oxygen species scavenger N-acetylcysteine could also mitigate reactive oxygen species generation. Moreover, JNK and P38 were activated by MPTP, but extracellular signal-regulated protein kinases phosphorylation did not change after MPTP treatment. TSG at different doses blocked the activation of JNK and P38. The protective effect of TSG was also associated with downregulation of the bax/bcl-2 ratio, reversed the release of cytochrome c and smac, and inhibited the activation of caspase-3, -6, and -9 induced by MPTP. In conclusion, our studies demonstrated that the protective effects of TSG in the MPTP-induced mouse model of PD are involved, at least in part, in controlling reactive oxygen species-mediated JNK, P38, and mitochondrial pathways. PMID:26477638

  6. Mitochondrial Evolution

    PubMed Central

    Gray, Michael W.

    2012-01-01

    Viewed through the lens of the genome it contains, the mitochondrion is of unquestioned bacterial ancestry, originating from within the bacterial phylum α-Proteobacteria (Alphaproteobacteria). Accordingly, the endosymbiont hypothesis—the idea that the mitochondrion evolved from a bacterial progenitor via symbiosis within an essentially eukaryotic host cell—has assumed the status of a theory. Yet mitochondrial genome evolution has taken radically different pathways in diverse eukaryotic lineages, and the organelle itself is increasingly viewed as a genetic and functional mosaic, with the bulk of the mitochondrial proteome having an evolutionary origin outside Alphaproteobacteria. New data continue to reshape our views regarding mitochondrial evolution, particularly raising the question of whether the mitochondrion originated after the eukaryotic cell arose, as assumed in the classical endosymbiont hypothesis, or whether this organelle had its beginning at the same time as the cell containing it. PMID:22952398

  7. Parkin suppresses Drp1-independent mitochondrial division.

    PubMed

    Roy, Madhuparna; Itoh, Kie; Iijima, Miho; Sesaki, Hiromi

    2016-07-01

    The cycle of mitochondrial division and fusion disconnect and reconnect individual mitochondria in cells to remodel this energy-producing organelle. Although dynamin-related protein 1 (Drp1) plays a major role in mitochondrial division in cells, a reduced level of mitochondrial division still persists even in the absence of Drp1. It is unknown how much Drp1-mediated mitochondrial division accounts for the connectivity of mitochondria. The role of a Parkinson's disease-associated protein-parkin, which biochemically and genetically interacts with Drp1-in mitochondrial connectivity also remains poorly understood. Here, we quantified the number and connectivity of mitochondria using mitochondria-targeted photoactivatable GFP in cells. We show that the loss of Drp1 increases the connectivity of mitochondria by 15-fold in mouse embryonic fibroblasts (MEFs). While a single loss of parkin does not affect the connectivity of mitochondria, the connectivity of mitochondria significantly decreased compared with a single loss of Drp1 when parkin was lost in the absence of Drp1. Furthermore, the loss of parkin decreased the frequency of depolarization of the mitochondrial inner membrane that is caused by increased mitochondrial connectivity in Drp1-knockout MEFs. Therefore, our data suggest that parkin negatively regulates Drp1-indendent mitochondrial division. PMID:27181353

  8. Parkin suppresses Drp1-independent mitochondrial division.

    PubMed

    Roy, Madhuparna; Itoh, Kie; Iijima, Miho; Sesaki, Hiromi

    2016-07-01

    The cycle of mitochondrial division and fusion disconnect and reconnect individual mitochondria in cells to remodel this energy-producing organelle. Although dynamin-related protein 1 (Drp1) plays a major role in mitochondrial division in cells, a reduced level of mitochondrial division still persists even in the absence of Drp1. It is unknown how much Drp1-mediated mitochondrial division accounts for the connectivity of mitochondria. The role of a Parkinson's disease-associated protein-parkin, which biochemically and genetically interacts with Drp1-in mitochondrial connectivity also remains poorly understood. Here, we quantified the number and connectivity of mitochondria using mitochondria-targeted photoactivatable GFP in cells. We show that the loss of Drp1 increases the connectivity of mitochondria by 15-fold in mouse embryonic fibroblasts (MEFs). While a single loss of parkin does not affect the connectivity of mitochondria, the connectivity of mitochondria significantly decreased compared with a single loss of Drp1 when parkin was lost in the absence of Drp1. Furthermore, the loss of parkin decreased the frequency of depolarization of the mitochondrial inner membrane that is caused by increased mitochondrial connectivity in Drp1-knockout MEFs. Therefore, our data suggest that parkin negatively regulates Drp1-indendent mitochondrial division.

  9. Salvaging hope: Is increasing NAD(+) a key to treating mitochondrial myopathy?

    PubMed

    Lightowlers, Robert N; Chrzanowska-Lightowlers, Zofia M A

    2014-06-01

    Mitochondrial diseases can arise from mutations either in mitochondrial DNA or in nuclear DNA encoding mitochondrially destined proteins. Currently, there is no cure for these diseases although treatments to ameliorate a subset of the symptoms are being developed. In this issue of EMBO Molecular Medicine, Khan et al (2014) use a mouse model to test the efficacy of a simple dietary supplement of nicotinamide riboside to treat and prevent mitochondrial myopathies. PMID:24838280

  10. What Is Mitochondrial DNA?

    MedlinePlus

    ... DNA What is mitochondrial DNA? What is mitochondrial DNA? Although most DNA is packaged in chromosomes within ... proteins. For more information about mitochondria and mitochondrial DNA: Molecular Expressions, a web site from the Florida ...

  11. Dynamics of Mitochondrial Transport in Axons

    PubMed Central

    Niescier, Robert F.; Kwak, Sang Kyu; Joo, Se Hun; Chang, Karen T.; Min, Kyung-Tai

    2016-01-01

    The polarized structure and long neurites of neurons pose a unique challenge for proper mitochondrial distribution. It is widely accepted that mitochondria move from the cell body to axon ends and vice versa; however, we have found that mitochondria originating from the axon ends moving in the retrograde direction never reach to the cell body, and only a limited number of mitochondria moving in the anterograde direction from the cell body arrive at the axon ends of mouse hippocampal neurons. Furthermore, we have derived a mathematical formula using the Fokker-Planck equation to characterize features of mitochondrial transport, and the equation could determine altered mitochondrial transport in axons overexpressing parkin. Our analysis will provide new insights into the dynamics of mitochondrial transport in axons of normal and unhealthy neurons. PMID:27242435

  12. Hypoxia as a therapy for mitochondrial disease.

    PubMed

    Jain, Isha H; Zazzeron, Luca; Goli, Rahul; Alexa, Kristen; Schatzman-Bone, Stephanie; Dhillon, Harveen; Goldberger, Olga; Peng, Jun; Shalem, Ophir; Sanjana, Neville E; Zhang, Feng; Goessling, Wolfram; Zapol, Warren M; Mootha, Vamsi K

    2016-04-01

    Defects in the mitochondrial respiratory chain (RC) underlie a spectrum of human conditions, ranging from devastating inborn errors of metabolism to aging. We performed a genome-wide Cas9-mediated screen to identify factors that are protective during RC inhibition. Our results highlight the hypoxia response, an endogenous program evolved to adapt to limited oxygen availability. Genetic or small-molecule activation of the hypoxia response is protective against mitochondrial toxicity in cultured cells and zebrafish models. Chronic hypoxia leads to a marked improvement in survival, body weight, body temperature, behavior, neuropathology, and disease biomarkers in a genetic mouse model of Leigh syndrome, the most common pediatric manifestation of mitochondrial disease. Further preclinical studies are required to assess whether hypoxic exposure can be developed into a safe and effective treatment for human diseases associated with mitochondrial dysfunction.

  13. Hypoxia as a Therapy for Mitochondrial Disease

    PubMed Central

    Jain, Isha H.; Zazzeron, Luca; Goli, Rahul; Alexa, Kristen; Schatzman-Bone, Stephanie; Dhillon, Harveen; Goldberger, Olga; Peng, Jun; Shalem, Ophir; Sanjana, Neville E.; Zhang, Feng; Goessling, Wolfram; Zapol, Warren M.; Mootha, Vamsi K.

    2016-01-01

    Defects in the mitochondrial respiratory chain (RC) underlie a spectrum of human conditions, ranging from devastating inborn errors of metabolism to aging. We performed a genome-wide, Cas9-mediated screen to identify factors that are protective during RC inhibition. Our results highlight the hypoxia response, an endogenous program evolved to adapt to limiting oxygen availability. Genetic or small molecule activation of the hypoxia response is protective against mitochondrial toxicity in cultured cells and zebrafish models. Chronic hypoxia leads to a marked improvement in survival, body weight, body temperature, behavior, neuropathology and disease biomarkers in a genetic mouse model of Leigh syndrome, the most common pediatric manifestation of mitochondrial disease. Further preclinical studies are required to assess whether hypoxic exposure can be developed into a safe and effective treatment for human diseases associated with mitochondrial dysfunction. PMID:26917594

  14. Mitochondrial respiration controls lysosomal function during inflammatory T cell responses

    PubMed Central

    Baixauli, Francesc; Acín-Pérez, Rebeca; Villarroya-Beltrí, Carolina; Mazzeo, Carla; Nuñez-Andrade, Norman; Gabandé-Rodriguez, Enrique; Dolores Ledesma, Maria; Blázquez, Alberto; Martin, Miguel Angel; Falcón-Pérez, Juan Manuel; Redondo, Juan Miguel; Enríquez, Jose Antonio; Mittelbrunn, Maria

    2016-01-01

    Summary The endolysosomal system is critical for the maintenance of cellular homeostasis. However, how endolysosomal compartment is regulated by mitochondrial function is largely unknown. We have generated a mouse model with defective mitochondrial function in CD4+ T lymphocytes by genetic deletion of the mitochondrial transcription factor A (Tfam). Mitochondrial respiration-deficiency impairs lysosome function, promotes p62 and sphingomyelin accumulation and disrupts endolysosomal trafficking pathways and autophagy, thus linking a primary mitochondrial dysfunction to a lysosomal storage disorder. The impaired lysosome function in Tfam-deficient cells subverts T cell differentiation toward pro-inflammatory subsets and exacerbates the in vivo inflammatory response. Restoration of NAD+ levels improves lysosome function and corrects the inflammatory defects in Tfam-deficient T cells. Our results uncover a mechanism by which mitochondria regulate lysosome function to preserve T cell differentiation and effector functions, and identify novel strategies for intervention in mitochondrial-related diseases. PMID:26299452

  15. Mitochondrial Respiration Controls Lysosomal Function during Inflammatory T Cell Responses.

    PubMed

    Baixauli, Francesc; Acín-Pérez, Rebeca; Villarroya-Beltrí, Carolina; Mazzeo, Carla; Nuñez-Andrade, Norman; Gabandé-Rodriguez, Enrique; Ledesma, Maria Dolores; Blázquez, Alberto; Martin, Miguel Angel; Falcón-Pérez, Juan Manuel; Redondo, Juan Miguel; Enríquez, Jose Antonio; Mittelbrunn, Maria

    2015-09-01

    The endolysosomal system is critical for the maintenance of cellular homeostasis. However, how endolysosomal compartment is regulated by mitochondrial function is largely unknown. We have generated a mouse model with defective mitochondrial function in CD4(+) T lymphocytes by genetic deletion of the mitochondrial transcription factor A (Tfam). Mitochondrial respiration deficiency impairs lysosome function, promotes p62 and sphingomyelin accumulation, and disrupts endolysosomal trafficking pathways and autophagy, thus linking a primary mitochondrial dysfunction to a lysosomal storage disorder. The impaired lysosome function in Tfam-deficient cells subverts T cell differentiation toward proinflammatory subsets and exacerbates the in vivo inflammatory response. Restoration of NAD(+) levels improves lysosome function and corrects the inflammatory defects in Tfam-deficient T cells. Our results uncover a mechanism by which mitochondria regulate lysosome function to preserve T cell differentiation and effector functions, and identify strategies for intervention in mitochondrial-related diseases.

  16. Mitochondrial Dynamics and Mitochondrial Dysfunction in Diabetes.

    PubMed

    Wada, Jun; Nakatsuka, Atsuko

    2016-06-01

    The mitochondria are involved in active and dynamic processes, such as mitochondrial biogenesis, fission, fusion and mitophagy to maintain mitochondrial and cellular functions. In obesity and type 2 diabetes, impaired oxidation, reduced mitochondrial contents, lowered rates of oxidative phosphorylation and excessive reactive oxygen species (ROS) production have been reported. Mitochondrial biogenesis is regulated by various transcription factors such as peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), peroxisome proliferator-activated receptors (PPARs), estrogen-related receptors (ERRs), and nuclear respiratory factors (NRFs). Mitochondrial fusion is promoted by mitofusin 1 (MFN1), mitofusin 2 (MFN2) and optic atrophy 1 (OPA1), while fission is governed by the recruitment of dynamin-related protein 1 (DRP1) by adaptor proteins such as mitochondrial fission factor (MFF), mitochondrial dynamics proteins of 49 and 51 kDa (MiD49 and MiD51), and fission 1 (FIS1). Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) and PARKIN promote DRP1-dependent mitochondrial fission, and the outer mitochondrial adaptor MiD51 is required in DRP1 recruitment and PARKIN-dependent mitophagy. This review describes the molecular mechanism of mitochondrial dynamics, its abnormality in diabetes and obesity, and pharmaceuticals targeting mitochondrial biogenesis, fission, fusion and mitophagy. PMID:27339203

  17. Nitric oxide regulates vascular adaptive mitochondrial dynamics.

    PubMed

    Miller, Matthew W; Knaub, Leslie A; Olivera-Fragoso, Luis F; Keller, Amy C; Balasubramaniam, Vivek; Watson, Peter A; Reusch, Jane E B

    2013-06-15

    Cardiovascular disease risk factors, such as diabetes, hypertension, dyslipidemia, obesity, and physical inactivity, are all correlated with impaired endothelial nitric oxide synthase (eNOS) function and decreased nitric oxide (NO) production. NO-mediated regulation of mitochondrial biogenesis has been established in many tissues, yet the role of eNOS in vascular mitochondrial biogenesis and dynamics is unclear. We hypothesized that genetic eNOS deletion and 3-day nitric oxide synthase (NOS) inhibition in rodents would result in impaired mitochondrial biogenesis and defunct fission/fusion and autophagy profiles within the aorta. We observed a significant, eNOS expression-dependent decrease in mitochondrial electron transport chain (ETC) protein subunits from complexes I, II, III, and V in eNOS heterozygotes and eNOS null mice compared with age-matched controls. In response to NOS inhibition with NG-nitro-L-arginine methyl ester (L-NAME) treatment in Sprague Dawley rats, significant decreases were observed in ETC protein subunits from complexes I, III, and IV as well as voltage-dependent anion channel 1. Decreased protein content of upstream regulators of mitochondrial biogenesis, cAMP response element-binding protein and peroxisome proliferator-activated receptor-γ coactivator-1α, were observed in response to 3-day L-NAME treatment. Both genetic eNOS deletion and NOS inhibition resulted in decreased manganese superoxide dismutase protein. L-NAME treatment resulted in significant changes to mitochondrial dynamic protein profiles with decreased fusion, increased fission, and minimally perturbed autophagy. In addition, L-NAME treatment blocked mitochondrial adaptation to an exercise intervention in the aorta. These results suggest that eNOS/NO play a role in basal and adaptive mitochondrial biogenesis in the vasculature and regulation of mitochondrial turnover. PMID:23585138

  18. Alterations in Mitochondrial Quality Control in Alzheimer’s Disease

    PubMed Central

    Cai, Qian; Tammineni, Prasad

    2016-01-01

    Mitochondrial dysfunction is one of the earliest and most prominent features in the brains of Alzheimer’s disease (AD) patients. Recent studies suggest that mitochondrial dysfunction plays a pivotal role in the pathogenesis of AD. Neurons are metabolically active cells, causing them to be particularly dependent on mitochondrial function for survival and maintenance. As highly dynamic organelles, mitochondria are characterized by a balance of fusion and fission, transport, and mitophagy, all of which are essential for maintaining mitochondrial integrity and function. Mitochondrial dynamics and mitophagy can therefore be identified as key pathways in mitochondrial quality control. Tremendous progress has been made in studying changes in these key aspects of mitochondrial biology in the vulnerable neurons of AD brains and mouse models, and the potential underlying mechanisms of such changes. This review highlights recent findings on alterations in the mitochondrial dynamics and mitophagy in AD and discusses how these abnormalities impact mitochondrial quality control and thus contribute to mitochondrial dysfunction in AD. PMID:26903809

  19. Batteries: Widening voltage windows

    NASA Astrophysics Data System (ADS)

    Xu, Kang; Wang, Chunsheng

    2016-10-01

    The energy output of aqueous batteries is largely limited by the narrow voltage window of their electrolytes. Now, a hydrate melt consisting of lithium salts is shown to expand such voltage windows, leading to a high-energy aqueous battery.

  20. Automatic voltage imbalance detector

    DOEpatents

    Bobbett, Ronald E.; McCormick, J. Byron; Kerwin, William J.

    1984-01-01

    A device for indicating and preventing damage to voltage cells such as galvanic cells and fuel cells connected in series by detecting sequential voltages and comparing these voltages to adjacent voltage cells. The device is implemented by using operational amplifiers and switching circuitry is provided by transistors. The device can be utilized in battery powered electric vehicles to prevent galvanic cell damage and also in series connected fuel cells to prevent fuel cell damage.

  1. Autophagy plays a role in skeletal muscle mitochondrial biogenesis in an endurance exercise-trained condition.

    PubMed

    Ju, Jeong-Sun; Jeon, Sei-Il; Park, Je-Young; Lee, Jong-Young; Lee, Seong-Cheol; Cho, Ki-Jung; Jeong, Jong-Moon

    2016-09-01

    Mitochondrial homeostasis is tightly regulated by two major processes: mitochondrial biogenesis and mitochondrial degradation by autophagy (mitophagy). Research in mitochondrial biogenesis in skeletal muscle in response to endurance exercise training has been well established, while the mechanisms regulating mitophagy and the interplay between mitochondrial biogenesis and degradation following endurance exercise training are not yet well defined. The purpose of this study was to examine the effects of a short-term inhibition of autophagy in response to acute endurance exercise on skeletal muscle mitochondrial biogenesis and dynamics in an exercise-trained condition. Male wild-type C57BL/6 mice performed five daily bouts of 1-h swimming per week for 8 weeks. In order to measure autophagy flux in mouse skeletal muscle, mice were treated with or without 2 days of 0.4 mg/kg/day intraperitoneal colchicine (blocking the degradation of autophagosomes) following swimming exercise training. The autophagic flux assay demonstrated that swimming training resulted in an increase in the autophagic flux (~100 % increase in LC3-II) in mouse skeletal muscle. Mitochondrial fusion proteins, Opa1 and MFN2, were significantly elevated, and mitochondrial fission protein, Drp1, was also increased in trained mouse skeletal muscle, suggesting that endurance exercise training promotes both mitochondrial fusion and fission processes. A mitochondrial receptor, Bnip3, was further increased in exercised muscle when treated with colchicine while Pink/Parkin protein levels were unchanged. The endurance exercise training induced increases in mitochondrial biogenesis marker proteins, SDH, COX IV, and a mitochondrial biogenesis promoting factor, PGC-1α but this effect was abolished in colchicine-treated mouse skeletal muscle. This suggests that autophagy plays an important role in mitochondrial biogenesis and this coordination between these opposing processes is involved in the cellular

  2. High Voltage SPT Performance

    NASA Technical Reports Server (NTRS)

    Manzella, David; Jacobson, David; Jankovsky, Robert

    2001-01-01

    A 2.3 kW stationary plasma thruster designed to operate at high voltage was tested at discharge voltages between 300 and 1250 V. Discharge specific impulses between 1600 and 3700 sec were demonstrated with thrust between 40 and 145 mN. Test data indicated that discharge voltage can be optimized for maximum discharge efficiency. The optimum discharge voltage was between 500 and 700 V for the various anode mass flow rates considered. The effect of operating voltage on optimal magnet field strength was investigated. The effect of cathode flow rate on thruster efficiency was considered for an 800 V discharge.

  3. The structure of the mitochondrial cloud of Xenopus laevis oocytes.

    PubMed

    Billett, F S; Adam, E

    1976-12-01

    The ultrastructure of the mitochondrial cloud (Balbiani body) of the pre-vitellogenic oocytes of Xenopus laevis has been examined using transmission and stereoscan electron microscopy. Examination of conventional thin sections confirm previous observations which suggest that the cloud consists essentially of many thousands mitochondria and numerous small vesicles; larger clouds, in oocytes greater than 200 mum in diameter, contain relatively more vesicles. Using a standard electron microscope at 100 kV very long and coursing arrays of mitochondrial profiles can be detected. The presence of very long mitochondrial elements has been confirmed using a high voltage microscope operating at 500-1000 kV. Stereoscan preparations, isolated from pre-vitellogenic oocytes, lend some support to the view that the mitochondrial cloud amy consist of a mass of long filamentous mitochondria and the possibility that there are large continuous regions of mitochondrial material cannot be ruled out.

  4. Mitochondrial RNA granules: Compartmentalizing mitochondrial gene expression

    PubMed Central

    Jourdain, Alexis A.; Boehm, Erik; Maundrell, Kinsey

    2016-01-01

    In mitochondria, DNA replication, gene expression, and RNA degradation machineries coexist within a common nondelimited space, raising the question of how functional compartmentalization of gene expression is achieved. Here, we discuss the recently characterized “mitochondrial RNA granules,” mitochondrial subdomains with an emerging role in the regulation of gene expression. PMID:26953349

  5. Tissue-specific modulation of mitochondrial DNA segregation by a defect in mitochondrial division.

    PubMed

    Jokinen, Riikka; Marttinen, Paula; Stewart, James B; Neil Dear, T; Battersby, Brendan J

    2016-02-15

    Mitochondria are dynamic organelles that divide and fuse by remodeling an outer and inner membrane in response to developmental, physiological and stress stimuli. These events are coordinated by conserved dynamin-related GTPases. The dynamics of mitochondrial morphology require coordination with mitochondrial DNA (mtDNA) to ensure faithful genome transmission, however, this process remains poorly understood. Mitochondrial division is linked to the segregation of mtDNA but how it affects cases of mtDNA heteroplasmy, where two or more mtDNA variants/mutations co-exist in a cell, is unknown. Segregation of heteroplasmic human pathogenic mtDNA mutations is a critical factor in the onset and severity of human mitochondrial diseases. Here, we investigated the coupling of mitochondrial morphology to the transmission and segregation of mtDNA in mammals by taking advantage of two genetically modified mouse models: one with a dominant-negative mutation in the dynamin-related protein 1 (Drp1 or Dnm1l) that impairs mitochondrial fission and the other, heteroplasmic mice segregating two neutral mtDNA haplotypes (BALB and NZB). We show a tissue-specific response to mtDNA segregation from a defect in mitochondrial fission. Only mtDNA segregation in the hematopoietic compartment is modulated from impaired Dnm1l function. In contrast, no effect was observed in other tissues arising from the three germ layers during development and in mtDNA transmission through the female germline. Our data suggest a robust organization of a heteroplasmic mtDNA segregating unit across mammalian cell types that can overcome impaired mitochondrial division to ensure faithful transmission of the mitochondrial genome. PMID:26681804

  6. Mitochondrial biogenesis in the pulmonary vasculature during inhalation lung injury and fibrosis

    EPA Science Inventory

    Cell survival and injury repair is facilitated by mitochondrial biogenesis; however, the role of this process in lung repair is unknown. We evaluated mitochondrial biogenesis in the mouse lung in two injuries that cause acute inflammation and in two that cause chronic inflammatio...

  7. Eliminate mitochondrial diseases by gene editing in germ-line cells and embryos.

    PubMed

    Wang, Si; Yi, Fei; Qu, Jing

    2015-07-01

    Nuclease-based gene editing technologies have opened up opportunities for correcting human genetic diseases. For the first time, scientists achieved targeted gene editing of mitochondrial DNA in mouse oocytes fused with patient cells. This fascinating progression may encourage the development of novel therapy for human maternally inherent mitochondrial diseases. PMID:26081469

  8. Optical voltage reference

    DOEpatents

    Rankin, R.; Kotter, D.

    1994-04-26

    An optical voltage reference for providing an alternative to a battery source is described. The optical reference apparatus provides a temperature stable, high precision, isolated voltage reference through the use of optical isolation techniques to eliminate current and impedance coupling errors. Pulse rate frequency modulation is employed to eliminate errors in the optical transmission link while phase-lock feedback is employed to stabilize the frequency to voltage transfer function. 2 figures.

  9. Human Mitochondrial Protein Database

    National Institute of Standards and Technology Data Gateway

    SRD 131 Human Mitochondrial Protein Database (Web, free access)   The Human Mitochondrial Protein Database (HMPDb) provides comprehensive data on mitochondrial and human nuclear encoded proteins involved in mitochondrial biogenesis and function. This database consolidates information from SwissProt, LocusLink, Protein Data Bank (PDB), GenBank, Genome Database (GDB), Online Mendelian Inheritance in Man (OMIM), Human Mitochondrial Genome Database (mtDB), MITOMAP, Neuromuscular Disease Center and Human 2-D PAGE Databases. This database is intended as a tool not only to aid in studying the mitochondrion but in studying the associated diseases.

  10. Loss of p53 causes mitochondrial DNA depletion and altered mitochondrial reactive oxygen species homeostasis

    PubMed Central

    Lebedeva, Maria A.; Eaton, Jana S.; Shadel, Gerald S.

    2009-01-01

    In addition to its central role in cellular stress signaling, the tumor suppressor p53 modulates mitochondrial respiration through its nuclear transcription factor activity and localizes to mitochondria where it enhances apoptosis and suppresses mitochondrial DNA (mtDNA) mutagenesis. Here we demonstrate a new conserved role for p53 in mtDNA copy number maintenance and mitochondrial reactive oxygen species (ROS) homeostasis. In mammals, mtDNA is present in thousands of copies per cell and is essential for normal development and cell function. We show that p53 null mouse and p53 knock-down human primary fibroblasts exhibit mtDNA depletion and decreased mitochondrial mass under normal culture growth conditions. This is accompanied by a reduction of the p53R2 subunit of ribonucleotide reductase mRNA and protein and of mitochondrial transcription factor A (mtTFA) at the protein level only. Finally, p53-depleted cells exhibit significant disruption of cellular ROS homeostasis, characterized by reduced mitochondrial and cellular superoxide levels and increased cellular hydrogen peroxide. Altogether, these results elucidate additional mitochondria-related functions for p53 and implicate mtDNA depletion and ROS alterations as potentially relevant to cellular transformation, cancer cell phenotypes, and the Warburg Effect. PMID:19413947

  11. Mitochondrial regulation of macrophage cholesterol homeostasis.

    PubMed

    Graham, Annette

    2015-12-01

    This review explores the relationship between mitochondrial structure and function in the regulation of macrophage cholesterol metabolism and proposes that mitochondrial dysfunction contributes to loss of the elegant homeostatic mechanisms which normally maintain cellular sterol levels within defined limits. Mitochondrial sterol 27-hydroxylase (CYP27A1) can generate oxysterol activators of liver X receptors which heterodimerise with retinoid X receptors, enhancing the transcription of ATP binding cassette transporters (ABCA1, ABCG1, and ABCG4), that can remove excess cholesterol via efflux to apolipoproteins A-1, E, and high density lipoprotein, and inhibit inflammation. The activity of CYP27A1 is regulated by the rate of supply of cholesterol substrate to the inner mitochondrial membrane, mediated by a complex of proteins. The precise identity of this dynamic complex remains controversial, even in steroidogenic tissues, but may include steroidogenic acute regulatory protein and the 18 kDa translocator protein, together with voltage-dependent anion channels, ATPase AAA domain containing protein 3A, and optic atrophy type 1 proteins. Certainly, overexpression of StAR and TSPO proteins can enhance macrophage cholesterol efflux to apoA-I and/or HDL, while perturbations in mitochondrial function, or changes in the expression of mitochondrial fusion proteins, alter the efficiency of cholesterol efflux. Molecules which can sustain or improve mitochondrial function or increase the activity of the protein complex involved in cholesterol transfer may have utility in resolving the problem of dysregulated macrophage cholesterol homeostasis, a condition which may contribute to inflammation, atherosclerosis, nonalcoholic steatohepatitis, osteoblastic bone resorption, and some disorders of the central nervous system.

  12. Voltage verification unit

    DOEpatents

    Martin, Edward J.

    2008-01-15

    A voltage verification unit and method for determining the absence of potentially dangerous potentials within a power supply enclosure without Mode 2 work is disclosed. With this device and method, a qualified worker, following a relatively simple protocol that involves a function test (hot, cold, hot) of the voltage verification unit before Lock Out/Tag Out and, and once the Lock Out/Tag Out is completed, testing or "trying" by simply reading a display on the voltage verification unit can be accomplished without exposure of the operator to the interior of the voltage supply enclosure. According to a preferred embodiment, the voltage verification unit includes test leads to allow diagnostics with other meters, without the necessity of accessing potentially dangerous bus bars or the like.

  13. TMEM16A is associated with voltage-gated calcium channels in mouse retina and its function is disrupted upon mutation of the auxiliary α2δ4 subunit

    PubMed Central

    Caputo, Antonella; Piano, Ilaria; Demontis, Gian Carlo; Bacchi, Niccolò; Casarosa, Simona; Santina, Luca Della; Gargini, Claudia

    2015-01-01

    Photoreceptors rely upon highly specialized synapses to efficiently transmit signals to multiple postsynaptic targets. Calcium influx in the presynaptic terminal is mediated by voltage-gated calcium channels (VGCC). This event triggers neurotransmitter release, but also gates calcium-activated chloride channels (TMEM), which in turn regulate VGCC activity. In order to investigate the relationship between VGCC and TMEM channels, we analyzed the retina of wild type (WT) and Cacna2d4 mutant mice, in which the VGCC auxiliary α2δ4 subunit carries a nonsense mutation, disrupting the normal channel function. Synaptic terminals of mutant photoreceptors are disarranged and synaptic proteins as well as TMEM16A channels lose their characteristic localization. In parallel, calcium-activated chloride currents are impaired in rods, despite unaltered TMEM16A protein levels. Co-immunoprecipitation revealed the interaction between VGCC and TMEM16A channels in the retina. Heterologous expression of these channels in tsA-201 cells showed that TMEM16A associates with the CaV1.4 subunit, and the association persists upon expression of the mutant α2δ4 subunit. Collectively, our experiments show association between TMEM16A and the α1 subunit of VGCC. Close proximity of these channels allows optimal function of the photoreceptor synaptic terminal under physiological conditions, but also makes TMEM16A channels susceptible to changes occurring to calcium channels. PMID:26557056

  14. The physiological role of mitochondrial calcium revealed by mice lacking the mitochondrial calcium uniporter.

    PubMed

    Pan, Xin; Liu, Jie; Nguyen, Tiffany; Liu, Chengyu; Sun, Junhui; Teng, Yanjie; Fergusson, Maria M; Rovira, Ilsa I; Allen, Michele; Springer, Danielle A; Aponte, Angel M; Gucek, Marjan; Balaban, Robert S; Murphy, Elizabeth; Finkel, Toren

    2013-12-01

    Mitochondrial calcium has been postulated to regulate a wide range of processes from bioenergetics to cell death. Here, we characterize a mouse model that lacks expression of the recently discovered mitochondrial calcium uniporter (MCU). Mitochondria derived from MCU(-/-) mice have no apparent capacity to rapidly uptake calcium. Whereas basal metabolism seems unaffected, the skeletal muscle of MCU(-/-) mice exhibited alterations in the phosphorylation and activity of pyruvate dehydrogenase. In addition, MCU(-/-) mice exhibited marked impairment in their ability to perform strenuous work. We further show that mitochondria from MCU(-/-) mice lacked evidence for calcium-induced permeability transition pore (PTP) opening. The lack of PTP opening does not seem to protect MCU(-/-) cells and tissues from cell death, although MCU(-/-) hearts fail to respond to the PTP inhibitor cyclosporin A. Taken together, these results clarify how acute alterations in mitochondrial matrix calcium can regulate mammalian physiology.

  15. Mitochondrial DNA polymorphisms specifically modify cerebral β-amyloid proteostasis.

    PubMed

    Scheffler, Katja; Krohn, Markus; Dunkelmann, Tina; Stenzel, Jan; Miroux, Bruno; Ibrahim, Saleh; von Bohlen Und Halbach, Oliver; Heinze, Hans-Jochen; Walker, Lary C; Gsponer, Jörg A; Pahnke, Jens

    2012-08-01

    Several lines of evidence link mutations and deletions in mitochondrial DNA (mtDNA) and its maternal inheritance to neurodegenerative diseases in the elderly. Age-related mutations of mtDNA modulate the tricarboxylic cycle enzyme activity, mitochondrial oxidative phosphorylation capacity and oxidative stress response. To investigate the functional relevance of specific mtDNA polymorphisms of inbred mouse strains in the proteostasis regulation of the brain, we established novel mitochondrial congenic mouse lines of Alzheimer's disease (AD). We crossed females from inbred strains (FVB/N, AKR/J, NOD/LtJ) with C57BL/6 males for at least ten generations to gain specific mitochondrial conplastic strains with pure C57BL/6 nuclear backgrounds. We show that specific mtDNA polymorphisms originating from the inbred strains differentially influence mitochondrial energy metabolism, ATP production and ATP-driven microglial activity, resulting in alterations of cerebral β-amyloid (Aβ) accumulation. Our findings demonstrate that mtDNA-related increases in ATP levels and subsequently in microglial activity are directly linked to decreased Aβ accumulation in vivo, implicating reduced mitochondrial function in microglia as a causative factor in the development of age-related cerebral proteopathies such as AD.

  16. Voltage balanced multilevel voltage source converter system

    DOEpatents

    Peng, Fang Zheng; Lai, Jih-Sheng

    1997-01-01

    A voltage balanced multilevel converter for high power AC applications such as adjustable speed motor drives and back-to-back DC intertie of adjacent power systems. This converter provides a multilevel rectifier, a multilevel inverter, and a DC link between the rectifier and the inverter allowing voltage balancing between each of the voltage levels within the multilevel converter. The rectifier is equipped with at least one phase leg and a source input node for each of the phases. The rectifier is further equipped with a plurality of rectifier DC output nodes. The inverter is equipped with at least one phase leg and a load output node for each of the phases. The inverter is further equipped with a plurality of inverter DC input nodes. The DC link is equipped with a plurality of rectifier charging means and a plurality of inverter discharging means. The plurality of rectifier charging means are connected in series with one of the rectifier charging means disposed between and connected in an operable relationship with each adjacent pair of rectifier DC output nodes. The plurality of inverter discharging means are connected in series with one of the inverter discharging means disposed between and connected in an operable relationship with each adjacent pair of inverter DC input nodes. Each of said rectifier DC output nodes are individually electrically connected to the respective inverter DC input nodes. By this means, each of the rectifier DC output nodes and each of the inverter DC input nodes are voltage balanced by the respective charging and discharging of the rectifier charging means and the inverter discharging means.

  17. Voltage balanced multilevel voltage source converter system

    DOEpatents

    Peng, F.Z.; Lai, J.S.

    1997-07-01

    Disclosed is a voltage balanced multilevel converter for high power AC applications such as adjustable speed motor drives and back-to-back DC intertie of adjacent power systems. This converter provides a multilevel rectifier, a multilevel inverter, and a DC link between the rectifier and the inverter allowing voltage balancing between each of the voltage levels within the multilevel converter. The rectifier is equipped with at least one phase leg and a source input node for each of the phases. The rectifier is further equipped with a plurality of rectifier DC output nodes. The inverter is equipped with at least one phase leg and a load output node for each of the phases. The inverter is further equipped with a plurality of inverter DC input nodes. The DC link is equipped with a plurality of rectifier charging means and a plurality of inverter discharging means. The plurality of rectifier charging means are connected in series with one of the rectifier charging means disposed between and connected in an operable relationship with each adjacent pair of rectifier DC output nodes. The plurality of inverter discharging means are connected in series with one of the inverter discharging means disposed between and connected in an operable relationship with each adjacent pair of inverter DC input nodes. Each of said rectifier DC output nodes are individually electrically connected to the respective inverter DC input nodes. By this means, each of the rectifier DC output nodes and each of the inverter DC input nodes are voltage balanced by the respective charging and discharging of the rectifier charging means and the inverter discharging means. 15 figs.

  18. Low voltage to high voltage level shifter and related methods

    NASA Technical Reports Server (NTRS)

    Mentze, Erik J. (Inventor); Hess, Herbert L. (Inventor); Buck, Kevin M. (Inventor); Cox, David F. (Inventor)

    2006-01-01

    A shifter circuit comprises a high and low voltage buffer stages and an output buffer stage. The high voltage buffer stage comprises multiple transistors arranged in a transistor stack having a plurality of intermediate nodes connecting individual transistors along the stack. The transistor stack is connected between a voltage level being shifted to and an input voltage. An inverter of this stage comprises multiple inputs and an output. Inverter inputs are connected to a respective intermediate node of the transistor stack. The low voltage buffer stage has an input connected to the input voltage and an output, and is operably connected to the high voltage buffer stage. The low voltage buffer stage is connected between a voltage level being shifted away from and a lower voltage. The output buffer stage is driven by the outputs of the high voltage buffer stage inverter and the low voltage buffer stage.

  19. Trypanosomal TAC40 constitutes a novel subclass of mitochondrial β-barrel proteins specialized in mitochondrial genome inheritance

    PubMed Central

    Schnarwiler, Felix; Niemann, Moritz; Doiron, Nicholas; Harsman, Anke; Käser, Sandro; Mani, Jan; Chanfon, Astrid; Dewar, Caroline E.; Oeljeklaus, Silke; Jackson, Christopher B.; Pusnik, Mascha; Schmidt, Oliver; Meisinger, Chris; Hiller, Sebastian; Warscheid, Bettina; Schnaufer, Achim C.; Ochsenreiter, Torsten; Schneider, André

    2014-01-01

    Mitochondria cannot form de novo but require mechanisms allowing their inheritance to daughter cells. In contrast to most other eukaryotes Trypanosoma brucei has a single mitochondrion whose single-unit genome is physically connected to the flagellum. Here we identify a β-barrel mitochondrial outer membrane protein, termed tripartite attachment complex 40 (TAC40), that localizes to this connection. TAC40 is essential for mitochondrial DNA inheritance and belongs to the mitochondrial porin protein family. However, it is not specifically related to any of the three subclasses of mitochondrial porins represented by the metabolite transporter voltage-dependent anion channel (VDAC), the protein translocator of the outer membrane 40 (TOM40), or the fungi-specific MDM10, a component of the endoplasmic reticulum–mitochondria encounter structure (ERMES). MDM10 and TAC40 mediate cellular architecture and participate in transmembrane complexes that are essential for mitochondrial DNA inheritance. In yeast MDM10, in the context of the ERMES, is postulated to connect the mitochondrial genomes to actin filaments, whereas in trypanosomes TAC40 mediates the linkage of the mitochondrial DNA to the basal body of the flagellum. However, TAC40 does not colocalize with trypanosomal orthologs of ERMES components and, unlike MDM10, it regulates neither mitochondrial morphology nor the assembly of the protein translocase. TAC40 therefore defines a novel subclass of mitochondrial porins that is distinct from VDAC, TOM40, and MDM10. However, whereas the architecture of the TAC40-containing complex in trypanosomes and the MDM10-containing ERMES in yeast is very different, both are organized around a β-barrel protein of the mitochondrial porin family that mediates a DNA–cytoskeleton linkage that is essential for mitochondrial DNA inheritance. PMID:24821793

  20. Mitochondrial Dysfunction Meets Senescence.

    PubMed

    Gallage, Suchira; Gil, Jesús

    2016-03-01

    Cellular senescence and mitochondrial dysfunction are hallmarks of ageing, but until now their relationship has not been clear. Recent work by Wiley et al. shows that mitochondrial defects can cause a distinct senescence phenotype termed MiDAS (mitochondrial dysfunction-associated senescence). MiDAS has a specific secretome that is able to drive some of the aging phenotypes. These findings suggest novel therapeutic opportunities for treating age-related pathologies. PMID:26874922

  1. MYC and Mitochondrial Biogenesis

    PubMed Central

    Morrish, Fionnuala; Hockenbery, David

    2014-01-01

    Mitochondria, the powerhouses of the cell, face two imperatives concerning biogenesis. The first is the requirement for dividing cells to replicate their mitochondrial content by growth of existing mitochondria. The second is the dynamic regulation of mitochondrial content in response to organismal and cellular cues (e.g., exercise, caloric restriction, energy status, temperature). MYC provides the clearest example of a programmed expansion of mitochondrial content linked to the cell cycle. As an oncogene, MYC also presents intriguing questions about the role of its mitochondrial targets in cancer-related phenotypes, such as the Warburg effect and MYC-dependent apoptosis. PMID:24789872

  2. High voltage power supply

    NASA Technical Reports Server (NTRS)

    Ruitberg, A. P.; Young, K. M. (Inventor)

    1985-01-01

    A high voltage power supply is formed by three discrete circuits energized by a battery to provide a plurality of concurrent output signals floating at a high output voltage on the order of several tens of kilovolts. In the first two circuits, the regulator stages are pulse width modulated and include adjustable ressistances for varying the duty cycles of pulse trains provided to corresponding oscillator stages while the third regulator stage includes an adjustable resistance for varying the amplitude of a steady signal provided to a third oscillator stage. In the first circuit, the oscillator, formed by a constant current drive network and a tuned resonant network included a step up transformer, is coupled to a second step up transformer which, in turn, supplies an amplified sinusoidal signal to a parallel pair of complementary poled rectifying, voltage multiplier stages to generate the high output voltage.

  3. Imaging voltage in neurons

    PubMed Central

    Peterka, Darcy S.; Takahashi, Hiroto; Yuste, Rafael

    2011-01-01

    In the last decades, imaging membrane potential has become a fruitful approach to study neural circuits, especially in invertebrate preparations with large, resilient neurons. At the same time, particularly in mammalian preparations, voltage imaging methods suffer from poor signal to noise and secondary side effects, and they fall short of providing single-cell resolution when imaging of the activity of neuronal populations. As an introduction to these techniques, we briefly review different voltage imaging methods (including organic fluorophores, SHG chromophores, genetic indicators, hybrid, nanoparticles and intrinsic approaches), and illustrate some of their applications to neuronal biophysics and mammalian circuit analysis. We discuss their mechanisms of voltage sensitivity, from reorientation, electrochromic or electro-optical phenomena, to interaction among chromophores or membrane scattering, and highlight their advantages and shortcomings, commenting on the outlook for development of novel voltage imaging methods. PMID:21220095

  4. High voltage DC power supply

    DOEpatents

    Droege, T.F.

    1989-12-19

    A high voltage DC power supply having a first series resistor at the output for limiting current in the event of a short-circuited output, a second series resistor for sensing the magnitude of output current, and a voltage divider circuit for providing a source of feedback voltage for use in voltage regulation is disclosed. The voltage divider circuit is coupled to the second series resistor so as to compensate the feedback voltage for a voltage drop across the first series resistor. The power supply also includes a pulse-width modulated control circuit, having dual clock signals, which is responsive to both the feedback voltage and a command voltage, and also includes voltage and current measuring circuits responsive to the feedback voltage and the voltage developed across the second series resistor respectively. 7 figs.

  5. High voltage DC power supply

    DOEpatents

    Droege, Thomas F.

    1989-01-01

    A high voltage DC power supply having a first series resistor at the output for limiting current in the event of a short-circuited output, a second series resistor for sensing the magnitude of output current, and a voltage divider circuit for providing a source of feedback voltage for use in voltage regulation is disclosed. The voltage divider circuit is coupled to the second series resistor so as to compensate the feedback voltage for a voltage drop across the first series resistor. The power supply also includes a pulse-width modulated control circuit, having dual clock signals, which is responsive to both the feedback voltage and a command voltage, and also includes voltage and current measuring circuits responsive to the feedback voltage and the voltage developed across the second series resistor respectively.

  6. Mitochondrial protein hyperacetylation in the failing heart

    PubMed Central

    Horton, Julie L.; Martin, Ola J.; Lai, Ling; Richards, Alicia L.; Vega, Rick B.; Leone, Teresa C.; Pagliarini, David J.; Muoio, Deborah M.; Bedi, Kenneth C.; Coon, Joshua J.

    2016-01-01

    Myocardial fuel and energy metabolic derangements contribute to the pathogenesis of heart failure. Recent evidence implicates posttranslational mechanisms in the energy metabolic disturbances that contribute to the pathogenesis of heart failure. We hypothesized that accumulation of metabolite intermediates of fuel oxidation pathways drives posttranslational modifications of mitochondrial proteins during the development of heart failure. Myocardial acetylproteomics demonstrated extensive mitochondrial protein lysine hyperacetylation in the early stages of heart failure in well-defined mouse models and the in end-stage failing human heart. To determine the functional impact of increased mitochondrial protein acetylation, we focused on succinate dehydrogenase A (SDHA), a critical component of both the tricarboxylic acid (TCA) cycle and respiratory complex II. An acetyl-mimetic mutation targeting an SDHA lysine residue shown to be hyperacetylated in the failing human heart reduced catalytic function and reduced complex II–driven respiration. These results identify alterations in mitochondrial acetyl-CoA homeostasis as a potential driver of the development of energy metabolic derangements that contribute to heart failure. PMID:26998524

  7. Twin Mitochondrial Sequence Analysis.

    PubMed

    Bouhlal, Yosr; Martinez, Selena; Gong, Henry; Dumas, Kevin; Shieh, Joseph T C

    2013-09-01

    When applying genome-wide sequencing technologies to disease investigation, it is increasingly important to resolve sequence variation in regions of the genome that may have homologous sequences. The human mitochondrial genome challenges interpretation given the potential for heteroplasmy, somatic variation, and homologous nuclear mitochondrial sequences (numts). Identical twins share the same mitochondrial DNA (mtDNA) from early life, but whether the mitochondrial sequence remains similar is unclear. We compared an adult monozygotic twin pair using high throughput-sequencing and evaluated variants with primer extension and mitochondrial pre-enrichment. Thirty-seven variants were shared between the twin individuals, and the variants were verified on the original genomic DNA. These studies support highly identical genetic sequence in this case. Certain low-level variant calls were of high quality and homology to the mitochondrial DNA, and they were further evaluated. When we assessed calls in pre-enriched mitochondrial DNA templates, we found that these may represent numts, which can be differentiated from mtDNA variation. We conclude that twin identity extends to mitochondrial DNA, and it is critical to differentiate between numts and mtDNA in genome sequencing, particularly since significant heteroplasmy could influence genome interpretation. Further studies on mtDNA and numts will aid in understanding how variation occurs and persists. PMID:24040623

  8. Clinical mitochondrial genetics

    PubMed Central

    Chinnery, P.; Howell, N.; Andrews, R.; Turnbull, D.

    1999-01-01

    The last decade has been an age of enlightenment as far as mitochondrial pathology is concerned. Well established nuclear genetic diseases, such as Friedreich's ataxia,12 Wilson disease,3 and autosomal recessive hereditary spastic paraplegia,4 have been shown to have a mitochondrial basis, and we are just starting to unravel the complex nuclear genetic disorders which directly cause mitochondrial dysfunction (table 1). However, in addition to the 3 billion base pair nuclear genome, each human cell typically contains thousands of copies of a small, 16.5 kb circular molecule of double stranded DNA (fig 1). Mitochondrial DNA (mtDNA) accounts for only 1% of the total cellular nucleic acid content. It encodes for 13 polypeptides which are essential for aerobic metabolism and defects of the mitochondrial genome are an important cause of human disease.9293 Since the characterisation of the first pathogenic mtDNA defects in 1988,513 over 50 point mutations and well over 100 rearrangements of the mitochondrial genome have been associated with human disease9495 (http://www.gen.emory.edu/mitomap.html). These disorders form the focus of this article.


Keywords: mitochondrial DNA; mitochondrial disease; heteroplasmy; genetic counselling PMID:10874629

  9. Mitochondrial regulation of apoptosis.

    PubMed

    Mayer, Bernd; Oberbauer, Rainer

    2003-06-01

    Mitochondria play a central part in cellular survival and apoptotic death. These processes are highly regulated by pro- and antiapoptotic Bcl-2 superfamily members. A key feature within apoptosis cascades is disruption of mitochondrial transmembrane potential and apoptogenic protein release, caused by opening of the permeability transition pore (PT). New data, however, indicate that mitochondrial apoptosis may occur without PT involvement.

  10. Mitochondrial and Cell Death Mechanisms in Neurodegenerative Diseases

    PubMed Central

    Martin, Lee J.

    2010-01-01

    Alzheimer’s disease (AD), Parkinson’s disease (PD) and amyotrophic lateral sclerosis (ALS) are the most common human adult-onset neurodegenerative diseases. They are characterized by prominent age-related neurodegeneration in selectively vulnerable neural systems. Some forms of AD, PD, and ALS are inherited, and genes causing these diseases have been identified. Nevertheless, the mechanisms of the neuronal cell death are unresolved. Morphological, biochemical, genetic, as well as cell and animal model studies reveal that mitochondria could have roles in this neurodegeneration. The functions and properties of mitochondria might render subsets of selectively vulnerable neurons intrinsically susceptible to cellular aging and stress and overlying genetic variations, triggering neurodegeneration according to a cell death matrix theory. In AD, alterations in enzymes involved in oxidative phosphorylation, oxidative damage, and mitochondrial binding of Aβ and amyloid precursor protein have been reported. In PD, mutations in putative mitochondrial proteins have been identified and mitochondrial DNA mutations have been found in neurons in the substantia nigra. In ALS, changes occur in mitochondrial respiratory chain enzymes and mitochondrial cell death proteins. Transgenic mouse models of human neurodegenerative disease are beginning to reveal possible principles governing the biology of selective neuronal vulnerability that implicate mitochondria and the mitochondrial permeability transition pore. This review summarizes how mitochondrial pathobiology might contribute to neuronal death in AD, PD, and ALS and could serve as a target for drug therapy. PMID:21258649

  11. High voltage pulse generator

    DOEpatents

    Fasching, George E.

    1977-03-08

    An improved high-voltage pulse generator has been provided which is especially useful in ultrasonic testing of rock core samples. An N number of capacitors are charged in parallel to V volts and at the proper instance are coupled in series to produce a high-voltage pulse of N times V volts. Rapid switching of the capacitors from the paralleled charging configuration to the series discharging configuration is accomplished by using silicon-controlled rectifiers which are chain self-triggered following the initial triggering of a first one of the rectifiers connected between the first and second of the plurality of charging capacitors. A timing and triggering circuit is provided to properly synchronize triggering pulses to the first SCR at a time when the charging voltage is not being applied to the parallel-connected charging capacitors. Alternate circuits are provided for controlling the application of the charging voltage from a charging circuit to be applied to the parallel capacitors which provides a selection of at least two different intervals in which the charging voltage is turned "off" to allow the SCR's connecting the capacitors in series to turn "off" before recharging begins. The high-voltage pulse-generating circuit including the N capacitors and corresponding SCR's which connect the capacitors in series when triggered "on" further includes diodes and series-connected inductors between the parallel-connected charging capacitors which allow sufficiently fast charging of the capacitors for a high pulse repetition rate and yet allow considerable control of the decay time of the high-voltage pulses from the pulse-generating circuit.

  12. Device for monitoring cell voltage

    DOEpatents

    Doepke, Matthias; Eisermann, Henning

    2012-08-21

    A device for monitoring a rechargeable battery having a number of electrically connected cells includes at least one current interruption switch for interrupting current flowing through at least one associated cell and a plurality of monitoring units for detecting cell voltage. Each monitoring unit is associated with a single cell and includes a reference voltage unit for producing a defined reference threshold voltage and a voltage comparison unit for comparing the reference threshold voltage with a partial cell voltage of the associated cell. The reference voltage unit is electrically supplied from the cell voltage of the associated cell. The voltage comparison unit is coupled to the at least one current interruption switch for interrupting the current of at least the current flowing through the associated cell, with a defined minimum difference between the reference threshold voltage and the partial cell voltage.

  13. What do we not know about mitochondrial potassium channels?

    PubMed

    Laskowski, Michał; Augustynek, Bartłomiej; Kulawiak, Bogusz; Koprowski, Piotr; Bednarczyk, Piotr; Jarmuszkiewicz, Wieslawa; Szewczyk, Adam

    2016-08-01

    In this review, we summarize our knowledge about mitochondrial potassium channels, with a special focus on unanswered questions in this field. The following potassium channels have been well described in the inner mitochondrial membrane: ATP-regulated potassium channel, Ca(2+)-activated potassium channel, the voltage-gated Kv1.3 potassium channel, and the two-pore domain TASK-3 potassium channel. The primary functional roles of these channels include regulation of mitochondrial respiration and the alteration of membrane potential. Additionally, they modulate the mitochondrial matrix volume and the synthesis of reactive oxygen species by mitochondria. Mitochondrial potassium channels are believed to contribute to cytoprotection and cell death. In this paper, we discuss fundamental issues concerning mitochondrial potassium channels: their molecular identity, channel pharmacology and functional properties. Attention will be given to the current problems present in our understanding of the nature of mitochondrial potassium channels. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016', edited by Prof. Paolo Bernardi.

  14. Cardioprotection by modulation of mitochondrial respiration during ischemia–reperfusion: Role of apoptosis-inducing factor

    SciTech Connect

    Xu, Aijun; Szczepanek, Karol; Hu, Ying; Lesnefsky, Edward J.; Chen, Qun

    2013-06-14

    Highlights: •Blockade of electron transport prevents the loss of AIF from mitochondria during IR. •Blockade of electron transport decreases caspase-independent cell death during IR. •Mitochondrial AIF content is down-regulated in Harlequin mice. •Blockade of electron transport protects Harlequin mouse hearts during IR. •Amobarbital protection is partially dependent on mitochondrial AIF content. -- Abstract: The transient, reversible blockade of electron transport (BET) during ischemia or at the onset of reperfusion protects mitochondria and decreases cardiac injury. Apoptosis inducing factor (AIF) is located within the mitochondrial intermembrane space. A release of AIF from mitochondria into cytosol and nucleus triggers caspase-independent cell death. We asked if BET prevents the loss of AIF from mitochondria as a mechanism of protection in the buffer perfused heart. BET during ischemia with amobarbital, a rapidly reversible inhibitor of mitochondrial complex I, attenuated a release of AIF from mitochondria into cytosol, in turn decreasing the formation of cleaved and activated PARP-1. These results suggest that BET-mediated protection may occur through prevention of the loss of AIF from mitochondria during ischemia–reperfusion. In order to further clarify the role of mitochondrial AIF in BET-mediated protection, Harlequin (Hq) mice, a genetic model with mitochondrial AIF deficiency, were used to test whether BET could still decrease cell injury in Hq mouse hearts during reperfusion. BET during ischemia protected Hq mouse hearts against ischemia–reperfusion injury and improved mitochondrial function in these hearts during reperfusion. Thus, cardiac injury can still be decreased in the presence of down-regulated mitochondrial AIF content. Taken together, BET during ischemia protects both hearts with normal mitochondrial AIF content and hearts with mitochondrial AIF deficiency. Although preservation of mitochondrial AIF content plays a key role in

  15. Mitochondrial Therapeutics for Cardioprotection

    PubMed Central

    Carreira, Raquel S.; Lee, Pamela; Gottlieb, Roberta A.

    2013-01-01

    Mitochondria represent approximately one-third of the mass of the heart and play a critical role in maintaining cellular function—however, they are also a potent source of free radicals and pro-apoptotic factors. As such, maintaining mitochondrial homeostasis is essential to cell survival. As the dominant source of ATP, continuous quality control is mandatory to ensure their ongoing optimal function. Mitochondrial quality control is accomplished by the dynamic interplay of fusion, fission, autophagy, and mitochondrial biogenesis. This review examines these processes in the heart and considers their role in the context of ischemia-reperfusion injury. Interventions that modulate mitochondrial turnover, including pharmacologic agents, exercise, and caloric restriction are discussed as a means to improve mitochondrial quality control, ameliorate cardiovascular dysfunction, and enhance longevity. PMID:21718247

  16. Voltage controlled current source

    DOEpatents

    Casne, Gregory M.

    1992-01-01

    A seven decade, voltage controlled current source is described for use in testing intermediate range nuclear instruments that covers the entire test current range of from 10 picoamperes to 100 microamperes. High accuracy is obtained throughout the entire seven decades of output current with circuitry that includes a coordinated switching scheme responsive to the input signal from a hybrid computer to control the input voltage to an antilog amplifier, and to selectively connect a resistance to the antilog amplifier output to provide a continuous output current source as a function of a preset range of input voltage. An operator controlled switch provides current adjustment for operation in either a real-time simulation test mode or a time response test mode.

  17. Electron launching voltage monitor

    DOEpatents

    Mendel, Clifford W.; Savage, Mark E.

    1992-01-01

    An electron launching voltage monitor measures MITL voltage using a relationship between anode electric field and electron current launched from a cathode-mounted perturbation. An electron launching probe extends through and is spaced from the edge of an opening in a first MITL conductor, one end of the launching probe being in the gap between the MITL conductor, the other end being adjacent a first side of the first conductor away from the second conductor. A housing surrounds the launching probe and electrically connects the first side of the first conductor to the other end of the launching probe. A detector detects the current passing through the housing to the launching probe, the detected current being representative of the voltage between the conductors.

  18. Electron launching voltage monitor

    DOEpatents

    Mendel, C.W.; Savage, M.E.

    1992-03-17

    An electron launching voltage monitor measures MITL voltage using a relationship between anode electric field and electron current launched from a cathode-mounted perturbation. An electron launching probe extends through and is spaced from the edge of an opening in a first MITL conductor, one end of the launching probe being in the gap between the MITL conductor, the other end being adjacent a first side of the first conductor away from the second conductor. A housing surrounds the launching probe and electrically connects the first side of the first conductor to the other end of the launching probe. A detector detects the current passing through the housing to the launching probe, the detected current being representative of the voltage between the conductors. 5 figs.

  19. High voltage variable diameter insulator

    DOEpatents

    Vanacek, D.L.; Pike, C.D.

    1982-07-13

    A high voltage feedthrough assembly having a tubular insulator extending between the ground plane ring and the high voltage ring. The insulator is made of Pyrex and decreases in diameter from the ground plane ring to the high voltage ring, producing equipotential lines almost perpendicular to the wall of the insulator to optimize the voltage-holding capability of the feedthrough assembly.

  20. Expression and Purification of Mitochondrial RNA Polymerase and Transcription Factor A from Drosophila melanogaster.

    PubMed

    Gajewski, John P; Arnold, Jamie J; Salminen, Tiina S; Kaguni, Laurie S; Cameron, Craig E

    2016-01-01

    Mitochondrial gene expression is essential in all organisms. Our understanding of mitochondrial transcription on a biochemical level has been limited by the inability to purify the individual protein components involved in mitochondrial gene expression. Recently, new systems have been identified that permit purification of these proteins from bacteria. However, the generalizability of these systems is not clear. Here, we have applied the technology from the Cameron lab to express and purify mitochondrial RNA polymerase and transcription factor A from Drosophila melanogaster. We show that the use of SUMO system to produce SUMO fusion proteins in bacteria is effective not only for the human and mouse proteins, but also for the fly proteins. The application of this system to produce the mitochondrial proteins from other organisms should permit detailed understanding of mitochondrial transcription from any organism.

  1. Voltage Regulators for Photovoltaic Systems

    NASA Technical Reports Server (NTRS)

    Delombard, R.

    1986-01-01

    Two simple circuits developed to provide voltage regulation for highvoltage (i.e., is greater than 75 volts) and low-voltage (i.e., is less than 36 volts) photovoltaic/battery power systems. Use of these circuits results in voltage regulator small, low-cost, and reliable, with very low power dissipation. Simple oscillator circuit controls photovoltaic-array current to regulate system voltage and control battery charging. Circuit senses battery (and system) voltage and adjusts array current to keep battery voltage from exceeding maximum voltage.

  2. Voltage-Controlled Oscillator

    NASA Technical Reports Server (NTRS)

    1995-01-01

    Integrated Component Systems, Inc. incorporated information from a NASA Tech Briefs article into a voltage-controlled oscillator it designed for a customer. The company then applied the technology to its series of phase-locked loop synthesizers, which offer superior phase noise performance.

  3. High Voltage Insulation Technology

    NASA Astrophysics Data System (ADS)

    Scherb, V.; Rogalla, K.; Gollor, M.

    2008-09-01

    In preparation of new Electronic Power Conditioners (EPC's) for Travelling Wave Tub Amplifiers (TWTA's) on telecom satellites a study for the development of new high voltage insulation technology is performed. The initiative is mandatory to allow compact designs and to enable higher operating voltages. In a first task a market analysis was performed, comparing different materials with respect to their properties and processes. A hierarchy of selection criteria was established and finally five material candidates (4 Epoxy resins and 1 Polyurethane resin) were selected to be further investigated in the test program. Samples for the test program were designed to represent core elements of an EPC, the high voltage transformer and Printed Circuit Boards of the high voltage section. All five materials were assessed in the practical work flow of the potting process and electrical, mechanical, thermal and lifetime testing was performed. Although the lifetime tests results were overlayed by a larges scatter, finally two candidates have been identified for use in a subsequent qualification program. This activity forms part of element 5 of the ESA ARTES Programme.

  4. Geomagnetism and Induced Voltage

    ERIC Educational Resources Information Center

    Abdul-Razzaq, W.; Biller, R. D.

    2010-01-01

    Introductory physics laboratories have seen an influx of "conceptual integrated science" over time in their classrooms with elements of other sciences such as chemistry, biology, Earth science, and astronomy. We describe a laboratory to introduce this development, as it attracts attention to the voltage induced in the human brain as it is…

  5. Measuring Breakdown Voltage.

    ERIC Educational Resources Information Center

    Auer, Herbert J.

    1978-01-01

    The article discusses an aspect of conductivity, one of the electrical properties subdivisions, and describes a tester that can be shop-built. Breakdown voltage of an insulation material is specifically examined. Test procedures, parts lists, diagrams, and test data form are included. (MF)

  6. United Mitochondrial Disease Foundation

    MedlinePlus

    ... to Mitochondrial Disease FAQ's MitoFirst Handbook More Information Mito 101 Symposium Archives Get Connected Find an Event Adult Advisory Council Team Ask The Mito Doc Grand Rounds Kids & Teens Medical Child Abuse ...

  7. Defects in mitochondrial DNA replication and oxidative damage in muscle of mtDNA mutator mice.

    PubMed

    Kolesar, Jill E; Safdar, Adeel; Abadi, Arkan; MacNeil, Lauren G; Crane, Justin D; Tarnopolsky, Mark A; Kaufman, Brett A

    2014-10-01

    A causal role for mitochondrial dysfunction in mammalian aging is supported by recent studies of the mtDNA mutator mouse ("PolG" mouse), which harbors a defect in the proofreading-exonuclease activity of mitochondrial DNA polymerase gamma. These mice exhibit accelerated aging phenotypes characteristic of human aging, including systemic mitochondrial dysfunction, exercise intolerance, alopecia and graying of hair, curvature of the spine, and premature mortality. While mitochondrial dysfunction has been shown to cause increased oxidative stress in many systems, several groups have suggested that PolG mutator mice show no markers of oxidative damage. These mice have been presented as proof that mitochondrial dysfunction is sufficient to accelerate aging without oxidative stress. In this study, by normalizing to mitochondrial content in enriched fractions we detected increased oxidative modification of protein and DNA in PolG skeletal muscle mitochondria. We separately developed novel methods that allow simultaneous direct measurement of mtDNA replication defects and oxidative damage. Using this approach, we find evidence that suggests PolG muscle mtDNA is indeed oxidatively damaged. We also observed a significant decrease in antioxidants and expression of mitochondrial biogenesis pathway components and DNA repair enzymes in these mice, indicating an association of maladaptive gene expression with the phenotypes observed in PolG mice. Together, these findings demonstrate the presence of oxidative damage associated with the premature aging-like phenotypes induced by mitochondrial dysfunction.

  8. Fast kinase domain-containing protein 3 is a mitochondrial protein essential for cellular respiration

    SciTech Connect

    Simarro, Maria; Gimenez-Cassina, Alfredo; Kedersha, Nancy; Lazaro, Jean-Bernard; Adelmant, Guillaume O.; Marto, Jarrod A.; Rhee, Kirsten; Tisdale, Sarah; Danial, Nika; Benarafa, Charaf; Orduna, Anonio; Anderson, Paul

    2010-10-22

    Research highlights: {yields} Five members of the FAST kinase domain-containing proteins are localized to mitochondria in mammalian cells. {yields} The FASTKD3 interactome includes proteins involved in various aspects of mitochondrial metabolism. {yields} Targeted knockdown of FASTKD3 significantly reduces basal and maximal mitochondrial oxygen consumption. -- Abstract: Fas-activated serine/threonine phosphoprotein (FAST) is the founding member of the FAST kinase domain-containing protein (FASTKD) family that includes FASTKD1-5. FAST is a sensor of mitochondrial stress that modulates protein translation to promote the survival of cells exposed to adverse conditions. Mutations in FASTKD2 have been linked to a mitochondrial encephalomyopathy that is associated with reduced cytochrome c oxidase activity, an essential component of the mitochondrial electron transport chain. We have confirmed the mitochondrial localization of FASTKD2 and shown that all FASTKD family members are found in mitochondria. Although human and mouse FASTKD1-5 genes are expressed ubiquitously, some of them are most abundantly expressed in mitochondria-enriched tissues. We have found that RNA interference-mediated knockdown of FASTKD3 severely blunts basal and stress-induced mitochondrial oxygen consumption without disrupting the assembly of respiratory chain complexes. Tandem affinity purification reveals that FASTKD3 interacts with components of mitochondrial respiratory and translation machineries. Our results introduce FASTKD3 as an essential component of mitochondrial respiration that may modulate energy balance in cells exposed to adverse conditions by functionally coupling mitochondrial protein synthesis to respiration.

  9. SDF-1/CXCL12 modulates mitochondrial respiration of immature blood cells in a bi-phasic manner.

    PubMed

    Messina-Graham, Steven; Broxmeyer, Hal

    2016-05-01

    SDF-1/CXCL12 is a potent chemokine required for the homing and engraftment of hematopoietic stem and progenitor cells. Previous data from our group has shown that in an SDF-1/CXCL12 transgenic mouse model, lineage(-) Sca-1(+) c-Kit(+) (LSK) bone marrow cells have reduced mitochondrial membrane potential versus wild-type. These results suggested that SDF-1/CXCL12 may function to keep mitochondrial respiration low in immature blood cells in the bone marrow. Low mitochondrial metabolism helps to maintain low levels of reactive oxygen species (ROS), which can influence differentiation. To test whether SDF-1/CXCL12 regulates mitochondrial metabolism, we employed the human leukemia cell line HL-60, that expresses high levels of the SDF-1/CXCL12 receptor, CXCR4, as a model of hematopoietic progenitor cells in vitro. We treated HL-60 cells with SDF-1/CXCL12 for 2 and 24h. Oxygen consumption rates (OCR), mitochondrial-associated ATP production, mitochondrial mass, and mitochondrial membrane potential of HL-60 cells were significantly reduced at 2h and increased at 24h as compared to untreated control cells. These biphasic effects of SDF-1/CXCL12 were reproduced with lineage negative primary mouse bone marrow cells, suggesting a novel function of SDF-1/CXCL12 in modulating mitochondrial respiration by regulating mitochondrial oxidative phosphorylation, ATP production and mitochondrial content. PMID:27067482

  10. Conductance hysteresis in the voltage dependent anion-selective channel

    PubMed Central

    Hoogerheide, David P.; Rostovtseva, Tatiana K.; Berezhkovskii, Alexander M.; Bezrukov, Sergey M.

    2015-01-01

    When the transmembrane voltage is periodically varied with time, the conductance of voltage-sensitive ion channels shows hysteretic behavior. Although this phenomenon has been used in studies of gating of the voltage-dependent anion channel, VDAC, from the outer mitochondrial membrane for nearly four decades, full hysteresis curves have never been reported, since the focus was only on the channel opening branches of the hysteresis loops. Here we study hysteretic response of a multichannel VDAC system to a triangular voltage ramp whose frequency varies within three orders of magnitude, ranging from 0.5 mHz to 0.2 Hz. We find that in this wide frequency range the area encircled by the hysteresis curves changes by less than a factor of three, thus suggesting a broad distribution of the characteristic times and strongly non-equilibrium behavior. At the same time, hysteresis branches corresponding to VDAC opening show quasi-equilibrium two-state behavior. This allows calculating usual equilibrium gating parameters, the gating charge and voltage of equipartitioning, which turn out to be virtually insensitive to the ramp frequency. To rationalize this peculiarity, we hypothesize that during voltage-induced closure and opening the system explores different regions of the complex free energy landscape, where, in the opening branch, it follows quasi-equilibrium paths. PMID:26094068

  11. Conductance hysteresis in the voltage-dependent anion channel.

    PubMed

    Rappaport, Shay M; Teijido, Oscar; Hoogerheide, David P; Rostovtseva, Tatiana K; Berezhkovskii, Alexander M; Bezrukov, Sergey M

    2015-09-01

    Hysteresis in the conductance of voltage-sensitive ion channels is observed when the transmembrane voltage is periodically varied with time. Although this phenomenon has been used in studies of gating of the voltage-dependent anion channel, VDAC, from the outer mitochondrial membrane for nearly four decades, full hysteresis curves have never been reported, because the focus was solely on the channel opening branches of the hysteresis loops. We studied the hysteretic response of a multichannel VDAC system to a triangular voltage ramp the frequency of which was varied over three orders of magnitude, from 0.5 mHz to 0.2 Hz. We found that in this wide frequency range the area encircled by the hysteresis curves changes by less than a factor of three, suggesting broad distribution of the characteristic times and strongly non-equilibrium behavior. At the same time, quasi-equilibrium two-state behavior is observed for hysteresis branches corresponding to VDAC opening. This enables calculation of the usual equilibrium gating parameters, gating charge and voltage of equipartitioning, which were found to be almost insensitive to the ramp frequency. To rationalize this peculiarity, we hypothesize that during voltage-induced closure and opening the system explores different regions of the complex free energy landscape, and, in the opening branch, follows quasi-equilibrium paths.

  12. A low voltage ``railgun''

    NASA Astrophysics Data System (ADS)

    Starr, Stanley O.; Youngquist, Robert C.; Cox, Robert B.

    2013-01-01

    Due to recent advances in solid-state switches and ultra-capacitors, it is now possible to construct a "railgun" that can operate at voltages below 20 V. Railguns typically operate above a thousand volts, generating huge currents for a few milliseconds to provide thousands of g's of acceleration to a small projectile. The low voltage railgun described herein operates for much longer time periods (tenths of seconds to seconds), has far smaller acceleration and speed, but can potentially propel a much larger object. The impetus for this development is to lay the groundwork for a possible ground-based supersonic launch track, but the resulting system may also have applications as a simple linear motor. The system would also be a useful teaching tool, requiring concepts from electrodynamics, mechanics, and electronics for its understanding, and is relatively straightforward to construct.

  13. Nickel inhibits mitochondrial fatty acid oxidation.

    PubMed

    Uppala, Radha; McKinney, Richard W; Brant, Kelly A; Fabisiak, James P; Goetzman, Eric S

    2015-08-01

    Nickel exposure is associated with changes in cellular energy metabolism which may contribute to its carcinogenic properties. Here, we demonstrate that nickel strongly represses mitochondrial fatty acid oxidation-the pathway by which fatty acids are catabolized for energy-in both primary human lung fibroblasts and mouse embryonic fibroblasts. At the concentrations used, nickel suppresses fatty acid oxidation without globally suppressing mitochondrial function as evidenced by increased glucose oxidation to CO2. Pre-treatment with l-carnitine, previously shown to prevent nickel-induced mitochondrial dysfunction in neuroblastoma cells, did not prevent the inhibition of fatty acid oxidation. The effect of nickel on fatty acid oxidation occurred only with prolonged exposure (>5 h), suggesting that direct inhibition of the active sites of metabolic enzymes is not the mechanism of action. Nickel is a known hypoxia-mimetic that activates hypoxia inducible factor-1α (HIF1α). Nickel-induced inhibition of fatty acid oxidation was blunted in HIF1α knockout fibroblasts, implicating HIF1α as one contributor to the mechanism. Additionally, nickel down-regulated the protein levels of the key fatty acid oxidation enzyme very long-chain acyl-CoA dehydrogenase (VLCAD) in a dose-dependent fashion. In conclusion, inhibition of fatty acid oxidation by nickel, concurrent with increased glucose metabolism, represents a form of metabolic reprogramming that may contribute to nickel-induced carcinogenesis.

  14. Nickel Inhibits Mitochondrial Fatty Acid Oxidation

    PubMed Central

    Uppala, Radha; McKinney, Richard W.; Brant, Kelly A.; Fabisiak, James P.; Goetzman, Eric S.

    2015-01-01

    Nickel exposure is associated with changes in cellular energy metabolism which may contribute to its carcinogenic properties. Here, we demonstrate that nickel strongly represses mitochondrial fatty acid oxidation—the pathway by which fatty acids are catabolized for energy—in both primary human lung fibroblasts and mouse embryonic fibroblasts. At the concentrations used, nickel suppresses fatty acid oxidation without globally suppressing mitochondrial function as evidenced by increased glucose oxidation to CO2. Pre-treatment with L-carnitine, previously shown to prevent nickel-induced mitochondrial dysfunction in neuroblastoma cells, did not prevent the inhibition of fatty acid oxidation. The effect of nickel on fatty acid oxidation occurred only with prolonged exposure (>5 hr), suggesting that direct inhibition of the active sites of metabolic enzymes is not the mechanism of action. Nickel is a known hypoxia-mimetic that activates hypoxia inducible factor-1α (HIF1α). Nickel-induced inhibition of fatty acid oxidation was blunted in HIF1α knockout fibroblasts, implicating HIF1α as one contributor to the mechanism. Additionally, nickel down-regulated the protein levels of the key fatty acid oxidation enzyme very long-chain acyl-CoA dehydrogenase (VLCAD) in a dose-dependent fashion. In conclusion, inhibition of fatty acid oxidation by nickel, concurrent with increased glucose metabolism, represents a form of metabolic reprogramming that may contribute to nickel-induced carcinogenesis. PMID:26051273

  15. Mitochondrial benzodiazepine receptors regulate steroid biosynthesis

    SciTech Connect

    Mukhin, A.G.; Papadopoulos, V.; Costa, E.; Krueger, K.E. )

    1989-12-01

    Recent observations on the steroid synthetic capability within the brain open the possibility that benzodiazepines may influence steroid synthesis in nervous tissue through interactions with peripheral-type benzodiazepine recognition sites, which are highly expressed in steroidogenic cells and associated with the outer mitochondrial membrane. To examine this possibility nine molecules that exhibit a greater than 10,000-fold difference in their affinities for peripheral-type benzodiazepine binding sites were tested for their effects on a well-established steroidogenic model system, the Y-1 mouse adrenal tumor cell line. 4{prime}-Chlorodiazepam, PK 11195, and PK 14067 stimulated steroid production by 2-fold in Y-1 cells, whereas diazepam, flunitrazepam, zolpidem, and PK 14068 displayed a lower (1.2- to 1.5-fold) maximal stimulation. In contrast, clonazepam and flumazenil did not stimulate steroid synthesis. The potencies of these compounds to inhibit {sup 3}H-labeled PK 11195 binding to peripheral-type benzodiazepine recognition sites correlated with their potencies to stimulate steroid production. Similar findings were observed in bovine and rat adrenocortical cell preparations. These results suggest that ligands of the peripheral-type benzodiazepine recognition site acting on this mitochondrial receptor can enhance steroid production. This action may contribute specificity to the pharmacological profile of drugs preferentially acting on the benzodiazepine recognition site associated with the outer membrane of certain mitochondrial populations.

  16. Increased voltage photovoltaic cell

    NASA Technical Reports Server (NTRS)

    Ross, B.; Bickler, D. B.; Gallagher, B. D. (Inventor)

    1985-01-01

    A photovoltaic cell, such as a solar cell, is provided which has a higher output voltage than prior cells. The improved cell includes a substrate of doped silicon, a first layer of silicon disposed on the substrate and having opposite doping, and a second layer of silicon carbide disposed on the first layer. The silicon carbide preferably has the same type of doping as the first layer.

  17. Insulators for high voltages

    SciTech Connect

    Looms, J.S.T.

    1987-01-01

    This book describes electrical insulators for high voltage applications. Topics considered include the insulating materials, the manufacture of wet process porcelain, the manufacture of tempered glass, the glass-fibre core, the polymeric housing, the common problem - terminating an insulator, mechanical constraints, the physics of pollution flashover, the physics of contamination, testing of insulators, conclusions from testing, remedies for flashover, insulators for special cases, interference and noise, and the insulator of the future.

  18. High voltage pulse conditioning

    DOEpatents

    Springfield, Ray M.; Wheat, Jr., Robert M.

    1990-01-01

    Apparatus for conditioning high voltage pulses from particle accelerators in order to shorten the rise times of the pulses. Flashover switches in the cathode stalk of the transmission line hold off conduction for a determinable period of time, reflecting the early portion of the pulses. Diodes upstream of the switches divert energy into the magnetic and electrostatic storage of the capacitance and inductance inherent to the transmission line until the switches close.

  19. High voltage generator

    DOEpatents

    Schwemin, A. J.

    1959-03-17

    A generator for producing relatively large currents at high voltages is described. In general, the invention comprises a plurality of capacitors connected in series by a plurality of switches alternately disposed with the capacitors. The above-noted circuit is mounted for movement with respect to contact members and switch closure means so that a load device and power supply are connected across successive numbers of capacitors, while the other capacitors are successively charged with the same power supply.

  20. High Voltage Seismic Generator

    NASA Astrophysics Data System (ADS)

    Bogacz, Adrian; Pala, Damian; Knafel, Marcin

    2015-04-01

    This contribution describes the preliminary result of annual cooperation of three student research groups from AGH UST in Krakow, Poland. The aim of this cooperation was to develop and construct a high voltage seismic wave generator. Constructed device uses a high-energy electrical discharge to generate seismic wave in ground. This type of device can be applied in several different methods of seismic measurement, but because of its limited power it is mainly dedicated for engineering geophysics. The source operates on a basic physical principles. The energy is stored in capacitor bank, which is charged by two stage low to high voltage converter. Stored energy is then released in very short time through high voltage thyristor in spark gap. The whole appliance is powered from li-ion battery and controlled by ATmega microcontroller. It is possible to construct larger and more powerful device. In this contribution the structure of device with technical specifications is resented. As a part of the investigation the prototype was built and series of experiments conducted. System parameter was measured, on this basis specification of elements for the final device were chosen. First stage of the project was successful. It was possible to efficiently generate seismic waves with constructed device. Then the field test was conducted. Spark gap wasplaced in shallowborehole(0.5 m) filled with salt water. Geophones were placed on the ground in straight line. The comparison of signal registered with hammer source and sparker source was made. The results of the test measurements are presented and discussed. Analysis of the collected data shows that characteristic of generated seismic signal is very promising, thus confirms possibility of practical application of the new high voltage generator. The biggest advantage of presented device after signal characteristics is its size which is 0.5 x 0.25 x 0.2 m and weight approximately 7 kg. This features with small li-ion battery makes

  1. High Voltage Connector

    SciTech Connect

    Kurita, C.H.; /Fermilab

    1987-03-06

    The originally designed high voltage connectors were to be made of brass. However, if treated like a Bellevile spring with the initially given dimensions, the stresses of the connector when crimped were calculated to be much higher than the yield stress of brass. Since the flange and outer diameters of the connector are to remain small, it was necessary to alter the other dimensions and choice of material in order to bring down the stresses applied to the connector.

  2. HIGH VOLTAGE GENERATOR

    DOEpatents

    Schwemin, A.J.

    1959-03-17

    A generator is presented for producing relatively large currents at high voltages. In general, the invention comprises a plurality of capacitors connected in series by a plurality of switches alternately disposed with the capacitors. The circuit is mounted for movement with respect to contact members and switch closure means so that a load device and power supply are connected across successive numbers of capacitors, while the other capacitors are successively charged with the same power supply.

  3. An Outer Mitochondrial Translocase, Tom22, Is Crucial for Inner Mitochondrial Steroidogenic Regulation in Adrenal and Gonadal Tissues

    PubMed Central

    Rajapaksha, Maheshinie; Kaur, Jasmeet; Prasad, Manoj; Pawlak, Kevin J.; Marshall, Brendan; Perry, Elizabeth W.; Whittal, Randy M.

    2016-01-01

    After cholesterol is transported into the mitochondria of steroidogenic tissues, the first steroid, pregnenolone, is synthesized in adrenal and gonadal tissues to initiate steroid synthesis by catalyzing the conversion of pregnenolone to progesterone, which is mediated by the inner mitochondrial enzyme 3β-hydroxysteroid dehydrogenase 2 (3βHSD2). We report that the mitochondrial translocase Tom22 is essential for metabolic conversion, as its knockdown by small interfering RNA (siRNA) completely ablated progesterone conversion in both steroidogenic mouse Leydig MA-10 and human adrenal NCI cells. Tom22 forms a 500-kDa complex with mitochondrial proteins associated with 3βHSD2. Although the absence of Tom22 did not inhibit mitochondrial import of cytochrome P450scc (cytochrome P450 side chain cleavage enzyme) and aldosterone synthase, it did inhibit 3βHSD2 expression. Electron microscopy showed that Tom22 is localized at the outer mitochondrial membrane (OMM), while 3βHSD2 is localized at the inner mitochondrial space (IMS), where it interacts through a specific region with Tom22 with its C-terminal amino acids and a small amino acid segment of Tom22 exposed to the IMS. Therefore, Tom22 is a critical regulator of steroidogenesis, and thus, it is essential for mammalian survival. PMID:26787839

  4. A mitochondrial location for haemoglobins--dynamic distribution in ageing and Parkinson's disease.

    PubMed

    Shephard, Freya; Greville-Heygate, Oliver; Marsh, Oliver; Anderson, Susan; Chakrabarti, Lisa

    2014-01-01

    Haemoglobins are iron-containing proteins that transport oxygen in the blood of most vertebrates. The mitochondrion is the cellular organelle which consumes oxygen in order to synthesise ATP. Mitochondrial dysfunction is implicated in neurodegeneration and ageing. We find that α and β haemoglobin (Hba and Hbb) proteins are altered in their distribution in mitochondrial fractions from degenerating brain. We demonstrate that both Hba and Hbb are co-localised with the mitochondrion in mammalian brain. The precise localisation of the Hbs is within the inner membrane space and associated with inner mitochondrial membrane. Relative mitochondrial to cytoplasmic ratios of Hba and Hbb show changing distributions of these proteins during the process of neurodegeneration in the pcd(5j) mouse brain. A significant difference in mitochondrial Hba and Hbb content in the mitochondrial fraction is seen at 31 days after birth, this corresponds to a stage when dynamic neuronal loss is measured to be greatest in the Purkinje Cell Degeneration mouse. We also report changes in mitochondrial Hba and Hbb levels in ageing brain and muscle. Significant differences in mitochondrial Hba and Hbb can be seen when comparing aged brain to muscle, suggesting tissue specific functions of these proteins in the mitochondrion. In muscle there are significant differences between Hba levels in old and young mitochondria. To understand whether the changes detected in mitochondrial Hbs are of clinical significance, we examined Parkinson's disease brain, immunohistochemistry studies suggest that cell bodies in the substantia nigra accumulate mitochondrial Hb. However, western blotting of mitochondrial fractions from PD and control brains indicates significantly less Hb in PD brain mitochondria. One explanation could be a specific loss of cells containing mitochondria loaded with Hb proteins. Our study opens the door to an examination of the role of Hb function, within the context of the mitochondrion

  5. HIGH VOLTAGE ION SOURCE

    DOEpatents

    Luce, J.S.

    1960-04-19

    A device is described for providing a source of molecular ions having a large output current and with an accelerated energy of the order of 600 kv. Ions are produced in an ion source which is provided with a water-cooled source grid of metal to effect maximum recombination of atomic ions to molecular ions. A very high accelerating voltage is applied to withdraw and accelerate the molecular ions from the source, and means are provided for dumping the excess electrons at the lowest possible potentials. An accelerating grid is placed adjacent to the source grid and a slotted, grounded accelerating electrode is placed adjacent to the accelerating grid. A potential of about 35 kv is maintained between the source grid and accelerating grid, and a potential of about 600 kv is maintained between the accelerating grid and accelerating electrode. In order to keep at a minimum the large number of oscillating electrons which are created when such high voltages are employed in the vicinity of a strong magnetic field, a plurality of high voltage cascaded shields are employed with a conventional electron dumping system being employed between each shield so as to dump the electrons at the lowest possible potential rather than at 600 kv.

  6. APPARATUS FOR REGULATING HIGH VOLTAGE

    DOEpatents

    Morrison, K.G.

    1951-03-20

    This patent describes a high-voltage regulator of the r-f type wherein the modulation of the r-f voltage is accomplished at a high level, resulting in good stabilization over a large range of load conditions.

  7. The Mitochondrial Permeability Transition Pore Regulates Nitric Oxide-Mediated Apoptosis of Neurons Induced by Target Deprivation

    PubMed Central

    Martin, Lee J.; Adams, Neal A.; Pan, Yan; Price, Ann; Wong, Margaret

    2011-01-01

    Ablation of mouse occipital cortex induces precisely timed and uniform p53-modulated and Bax-dependent apoptosis of thalamocortical projection neurons in the dorsal lateral geniculate nucleus (LGN) by 7 days postlesion. We tested the hypothesis that this neuronal apoptosis is initiated by oxidative stress and the mitochondrial permeability transition pore (mPTP). Pre-apoptotic LGN neurons accumulate mitochondria, Zn2+ and Ca2+, and generate higher levels of reactive oxygen species (ROS), including superoxide, nitric oxide (NO) and peroxynitrite, than LGN neurons with an intact cortical target. Pre-apoptosis of LGN neurons is associated with increased formation of protein carbonyls, protein nitration, and protein S-nitrosylation. Genetic deletion of nitric oxide synthase 1 (nos1) and inhibition of NOS1 with nitroindazole protected LGN neurons from apoptosis, revealing NO as a mediator. Putative components of the mPTP are expressed in mouse LGN, including the voltage-dependent anion channel (VDAC), adenine nucleotide translocator (ANT), and cyclophilin D (CyPD). Nitration of CyPD and ANT in LGN mitochondria occurs by 2 days after cortical injury. Chemical cross-linking showed that LGN neuron pre-apoptosis is associated with formation of CyPD and VDAC oligomers, consistent with mPTP formation. Mice without CyPD are rescued from neuron apoptosis as are mice treated with the mPTP inhibitors TRO-19622 and TAT-Bcl-XL-BH4. Manipulation of the mPTP markedly attenuated the early pre-apoptotic production of reactive oxygen/nitrogen species in target-deprived neurons. Our results demonstrate in adult mouse brain neurons that the mPTP functions to enhance ROS production and the mPTP and NO trigger apoptosis; thus, the mPTP is a target for neuroprotection in vivo. PMID:21209222

  8. Automatic voltage-imbalance detector

    SciTech Connect

    Bobbett, R.E.; McCormick, J.B.; Kerwin, W.J.

    1981-05-20

    A device is described for indicating and preventing damage to voltage cells such as galvanic cells and fuel cells connected in series by detecting sequential voltages and comparing these voltages to adjacent voltage cells. The device is implemented by using operational amplifiers and switching circuitry is provided by transistors. The device can be utilized in battery powered electric vehicles to prevent galvanic cell damage and also in series connected fuel cells to prevent fuel cell damage.

  9. High voltage variable diameter insulator

    DOEpatents

    Vanecek, David L.; Pike, Chester D.

    1984-01-01

    A high voltage feedthrough assembly (10) having a tubular insulator (15) extending between the ground plane ring (16) and the high voltage ring (30). The insulator (15) is made of Pyrex and decreases in diameter from the ground plane ring (16) to the high voltage ring (30), producing equipotential lines almost perpendicular to the wall (27) of the insulator (15) to optimize the voltage-holding capability of the feedthrough assembly (10).

  10. The Spectrum of Mitochondrial Ultrastructural Defects in Mitochondrial Myopathy.

    PubMed

    Vincent, Amy E; Ng, Yi Shiau; White, Kathryn; Davey, Tracey; Mannella, Carmen; Falkous, Gavin; Feeney, Catherine; Schaefer, Andrew M; McFarland, Robert; Gorman, Grainne S; Taylor, Robert W; Turnbull, Doug M; Picard, Martin

    2016-01-01

    Mitochondrial functions are intrinsically linked to their morphology and membrane ultrastructure. Characterizing abnormal mitochondrial structural features may thus provide insight into the underlying pathogenesis of inherited and acquired mitochondrial diseases. Following a systematic literature review on ultrastructural defects in mitochondrial myopathy, we investigated skeletal muscle biopsies from seven subjects with genetically defined mtDNA mutations. Mitochondrial ultrastructure and morphology were characterized using two complimentary approaches: transmission electron microscopy (TEM) and serial block face scanning EM (SBF-SEM) with 3D reconstruction. Six ultrastructural abnormalities were identified including i) paracrystalline inclusions, ii) linearization of cristae and abnormal angular features, iii) concentric layering of cristae membranes, iv) matrix compartmentalization, v) nanotunelling, and vi) donut-shaped mitochondria. In light of recent molecular advances in mitochondrial biology, these findings reveal novel aspects of mitochondrial ultrastructure and morphology in human tissues with implications for understanding the mechanisms linking mitochondrial dysfunction to disease. PMID:27506553

  11. The Spectrum of Mitochondrial Ultrastructural Defects in Mitochondrial Myopathy

    PubMed Central

    Vincent, Amy E.; Ng, Yi Shiau; White, Kathryn; Davey, Tracey; Mannella, Carmen; Falkous, Gavin; Feeney, Catherine; Schaefer, Andrew M.; McFarland, Robert; Gorman, Grainne S.; Taylor, Robert W.; Turnbull, Doug M.; Picard, Martin

    2016-01-01

    Mitochondrial functions are intrinsically linked to their morphology and membrane ultrastructure. Characterizing abnormal mitochondrial structural features may thus provide insight into the underlying pathogenesis of inherited and acquired mitochondrial diseases. Following a systematic literature review on ultrastructural defects in mitochondrial myopathy, we investigated skeletal muscle biopsies from seven subjects with genetically defined mtDNA mutations. Mitochondrial ultrastructure and morphology were characterized using two complimentary approaches: transmission electron microscopy (TEM) and serial block face scanning EM (SBF-SEM) with 3D reconstruction. Six ultrastructural abnormalities were identified including i) paracrystalline inclusions, ii) linearization of cristae and abnormal angular features, iii) concentric layering of cristae membranes, iv) matrix compartmentalization, v) nanotunelling, and vi) donut-shaped mitochondria. In light of recent molecular advances in mitochondrial biology, these findings reveal novel aspects of mitochondrial ultrastructure and morphology in human tissues with implications for understanding the mechanisms linking mitochondrial dysfunction to disease. PMID:27506553

  12. α-Synuclein amino terminus regulates mitochondrial membrane permeability.

    PubMed

    Shen, Jiamei; Du, Tingting; Wang, Xue; Duan, Chunli; Gao, Ge; Zhang, Jianliang; Lu, Lingling; Yang, Hui

    2014-12-01

    Parkinson's disease (PD) is a common neurodegenerative movement disorder affecting an increasing number of elderly. Various studies have shown that mitochondrial dysfunction and abnormal protein aggregation are two major contributors to the progression of PD. The N terminus of α-synuclein (α-Syn/N), which adopts an α-helical conformation upon lipid binding, is essential for membrane interaction; yet its role in mitochondria remains poorly defined. A functional characterization of the α-Syn N-terminal domain and investigation of its effect on mitochondrial membrane permeability were undertaken in this study. α-Syn/N and α-Syn/delN (amino acids 1-65 and 61-140, respectively) constructs were overexpressed in dopaminergic MN9D cells and primary cortical neurons. A decrease in cell viability was observed in cells transfected with α-Syn/N but not α-Syn/delN. In addition, an α-Syn/N-induced increase in the level of intracellular reactive oxygen species, alteration in mitochondrial morphology, and decrease in mitochondrial membrane potential were accompanied by the activation of mitochondrial permeability transition pores (mPTP). These changes were also associated with a decline in mitochondrial cardiolipin content and interaction with the voltage-dependent anion channel and adenine nucleotide translocator in the mitochondrial membrane. The activation of mPTPs and reduction in cell viability were partially reversed by bongkrekic acid, an inhibitor of adenine nucleotide translocator (ANT), suggesting that the interaction between α-Syn and ANT promoted mPTP activation and was toxic to cells. BKA treatment reduced interaction of α-Syn/N with ANT and VDAC. These results suggest that the N terminus of α-Syn is essential for the regulation of mitochondrial membrane permeability and is a likely factor in the neurodegeneration associated with PD.

  13. Charge-pump voltage converter

    DOEpatents

    Brainard, John P.; Christenson, Todd R.

    2009-11-03

    A charge-pump voltage converter for converting a low voltage provided by a low-voltage source to a higher voltage. Charge is inductively generated on a transfer rotor electrode during its transit past an inductor stator electrode and subsequently transferred by the rotating rotor to a collector stator electrode for storage or use. Repetition of the charge transfer process leads to a build-up of voltage on a charge-receiving device. Connection of multiple charge-pump voltage converters in series can generate higher voltages, and connection of multiple charge-pump voltage converters in parallel can generate higher currents. Microelectromechanical (MEMS) embodiments of this invention provide a small and compact high-voltage (several hundred V) voltage source starting with a few-V initial voltage source. The microscale size of many embodiments of this invention make it ideally suited for MEMS- and other micro-applications where integration of the voltage or charge source in a small package is highly desirable.

  14. Mitochondrial Dysfunction in Cancer

    PubMed Central

    Boland, Michelle L.; Chourasia, Aparajita H.; Macleod, Kay F.

    2013-01-01

    A mechanistic understanding of how mitochondrial dysfunction contributes to cell growth and tumorigenesis is emerging beyond Warburg as an area of research that is under-explored in terms of its significance for clinical management of cancer. Work discussed in this review focuses less on the Warburg effect and more on mitochondria and how dysfunctional mitochondria modulate cell cycle, gene expression, metabolism, cell viability, and other established aspects of cell growth and stress responses. There is increasing evidence that key oncogenes and tumor suppressors modulate mitochondrial dynamics through important signaling pathways and that mitochondrial mass and function vary between tumors and individuals but the significance of these events for cancer are not fully appreciated. We explore the interplay between key molecules involved in mitochondrial fission and fusion and in apoptosis, as well as in mitophagy, biogenesis, and spatial dynamics of mitochondria and consider how these distinct mechanisms are coordinated in response to physiological stresses such as hypoxia and nutrient deprivation. Importantly, we examine how deregulation of these processes in cancer has knock on effects for cell proliferation and growth. We define major forms of mitochondrial dysfunction and address the extent to which the functional consequences of such dysfunction can be determined and exploited for cancer diagnosis and treatment. PMID:24350057

  15. Mitochondrial fusion and inheritance of the mitochondrial genome.

    PubMed

    Takano, Hiroyoshi; Onoue, Kenta; Kawano, Shigeyuki

    2010-03-01

    Although maternal or uniparental inheritance of mitochondrial genomes is a general rule, biparental inheritance is sometimes observed in protists and fungi,including yeasts. In yeast, recombination occurs between the mitochondrial genomes inherited from both parents.Mitochondrial fusion observed in yeast zygotes is thought to set up a space for DNA recombination. In the last decade,a universal mitochondrial fusion mechanism has been uncovered, using yeast as a model. On the other hand, an alternative mitochondrial fusion mechanism has been identified in the true slime mold Physarum polycephalum.A specific mitochondrial plasmid, mF, has been detected as the genetic material that causes mitochondrial fusion in P. polycephalum. Without mF, fusion of the mitochondria is not observed throughout the life cycle, suggesting that Physarum has no constitutive mitochondrial fusion mechanism.Conversely, mitochondria fuse in zygotes and during sporulation with mF. The complete mF sequence suggests that one gene, ORF640, encodes a fusogen for Physarum mitochondria. Although in general, mitochondria are inherited uniparentally, biparental inheritance occurs with specific sexual crossing in P. polycephalum.An analysis of the transmission of mitochondrial genomes has shown that recombinations between two parental mitochondrial genomes require mitochondrial fusion,mediated by mF. Physarum is a unique organism for studying mitochondrial fusion. PMID:20196232

  16. Mitochondrial fusion and inheritance of the mitochondrial genome.

    PubMed

    Takano, Hiroyoshi; Onoue, Kenta; Kawano, Shigeyuki

    2010-03-01

    Although maternal or uniparental inheritance of mitochondrial genomes is a general rule, biparental inheritance is sometimes observed in protists and fungi,including yeasts. In yeast, recombination occurs between the mitochondrial genomes inherited from both parents.Mitochondrial fusion observed in yeast zygotes is thought to set up a space for DNA recombination. In the last decade,a universal mitochondrial fusion mechanism has been uncovered, using yeast as a model. On the other hand, an alternative mitochondrial fusion mechanism has been identified in the true slime mold Physarum polycephalum.A specific mitochondrial plasmid, mF, has been detected as the genetic material that causes mitochondrial fusion in P. polycephalum. Without mF, fusion of the mitochondria is not observed throughout the life cycle, suggesting that Physarum has no constitutive mitochondrial fusion mechanism.Conversely, mitochondria fuse in zygotes and during sporulation with mF. The complete mF sequence suggests that one gene, ORF640, encodes a fusogen for Physarum mitochondria. Although in general, mitochondria are inherited uniparentally, biparental inheritance occurs with specific sexual crossing in P. polycephalum.An analysis of the transmission of mitochondrial genomes has shown that recombinations between two parental mitochondrial genomes require mitochondrial fusion,mediated by mF. Physarum is a unique organism for studying mitochondrial fusion.

  17. Adult-onset mitochondrial myopathy.

    PubMed Central

    Fernandez-Sola, J.; Casademont, J.; Grau, J. M.; Graus, F.; Cardellach, F.; Pedrol, E.; Urbano-Marquez, A.

    1992-01-01

    Mitochondrial diseases are polymorphic entities which may affect many organs and systems. Skeletal muscle involvement is frequent in the context of systemic mitochondrial disease, but adult-onset pure mitochondrial myopathy appears to be rare. We report 3 patients with progressive skeletal mitochondrial myopathy starting in adult age. In all cases, the proximal myopathy was the only clinical feature. Mitochondrial pathology was confirmed by evidence of ragged-red fibres in muscle histochemistry, an abnormal mitochondrial morphology in electron microscopy and by exclusion of other underlying diseases. No deletions of mitochondrial DNA were found. We emphasize the need to look for a mitochondrial disorder in some non-specific myopathies starting in adult life. Images Figure 1 Figure 2 PMID:1589382

  18. Renal Mitochondrial Cytopathies

    PubMed Central

    Emma, Francesco; Montini, Giovanni; Salviati, Leonardo; Dionisi-Vici, Carlo

    2011-01-01

    Renal diseases in mitochondrial cytopathies are a group of rare diseases that are characterized by frequent multisystemic involvement and extreme variability of phenotype. Most frequently patients present a tubular defect that is consistent with complete De Toni-Debré-Fanconi syndrome in most severe forms. More rarely, patients present with chronic tubulointerstitial nephritis, cystic renal diseases, or primary glomerular involvement. In recent years, two clearly defined entities, namely 3243 A > G tRNALEU mutations and coenzyme Q10 biosynthesis defects, have been described. The latter group is particularly important because it represents the only treatable renal mitochondrial defect. In this paper, the physiopathologic bases of mitochondrial cytopathies, the diagnostic approaches, and main characteristics of related renal diseases are summarized. PMID:21811680

  19. The role of myeloid differentiation factor 88 on mitochondrial dysfunction of peritoneal leukocytes during polymicrobial sepsis

    PubMed Central

    Zou, Lin; Chen, Dunjin; Chao, Wei

    2016-01-01

    Objective To investigate the role of myeloid differentiation factor 88 (MyD88) on mitochondrial dysfunction of peritoneal leukocytes during polymicrobial sepsis. Material and methods Polymicrobial peritonitis, a clinically relevant mouse model of sepsis, was generated by cecum ligation and puncture (CLP) in both male C57BL/6J wild-type (WT) and MyD88 knockout (MyD88–/–) mice. Twenty-four hours after surgeries, peritoneal leukocytes were collected and four parameters of mitochondrial function, including total intracellular and mitochondrial ROS burst, mitochondrial membrane depolarization and ATP depletion, were measured by flow cytometry or ATP assay, and then compared. Results Polymicrobial sepsis led to a marked mitochondrial dysfunction of peritoneal leukocytes with total intracellular and mitochondrial ROS overproduction, decreased mitochondrial membrane potential and reduced intracellular ATP production. In comparison, there was no significant difference in the extent of mitochondrial dysfunction of peritoneal leukocytes between WT and MyD88–/– septic mice. Conclusions MyD88 may be not sufficient to regulate mitochondrial dysfunction of peritoneal leukocytes during polymicrobial sepsis. PMID:27536200

  20. Loss of parkin or PINK1 function increases Drp1-dependent mitochondrial fragmentation.

    PubMed

    Lutz, A Kathrin; Exner, Nicole; Fett, Mareike E; Schlehe, Julia S; Kloos, Karina; Lämmermann, Kerstin; Brunner, Bettina; Kurz-Drexler, Annerose; Vogel, Frank; Reichert, Andreas S; Bouman, Lena; Vogt-Weisenhorn, Daniela; Wurst, Wolfgang; Tatzelt, Jörg; Haass, Christian; Winklhofer, Konstanze F

    2009-08-21

    Loss-of-function mutations in the parkin gene (PARK2) and PINK1 gene (PARK6) are associated with autosomal recessive parkinsonism. PINK1 deficiency was recently linked to mitochondrial pathology in human cells and Drosophila melanogaster, which can be rescued by parkin, suggesting that both genes play a role in maintaining mitochondrial integrity. Here we demonstrate that an acute down-regulation of parkin in human SH-SY5Y cells severely affects mitochondrial morphology and function, a phenotype comparable with that induced by PINK1 deficiency. Alterations in both mitochondrial morphology and ATP production caused by either parkin or PINK1 loss of function could be rescued by the mitochondrial fusion proteins Mfn2 and OPA1 or by a dominant negative mutant of the fission protein Drp1. Both parkin and PINK1 were able to suppress mitochondrial fragmentation induced by Drp1. Moreover, in Drp1-deficient cells the parkin/PINK1 knockdown phenotype did not occur, indicating that mitochondrial alterations observed in parkin- or PINK1-deficient cells are associated with an increase in mitochondrial fission. Notably, mitochondrial fragmentation is an early phenomenon upon PINK1/parkin silencing that also occurs in primary mouse neurons and Drosophila S2 cells. We propose that the discrepant findings in adult flies can be explained by the time of phenotype analysis and suggest that in mammals different strategies may have evolved to cope with dysfunctional mitochondria.

  1. Loss of Parkin or PINK1 Function Increases Drp1-dependent Mitochondrial Fragmentation*

    PubMed Central

    Lutz, A. Kathrin; Exner, Nicole; Fett, Mareike E.; Schlehe, Julia S.; Kloos, Karina; Lämmermann, Kerstin; Brunner, Bettina; Kurz-Drexler, Annerose; Vogel, Frank; Reichert, Andreas S.; Bouman, Lena; Vogt-Weisenhorn, Daniela; Wurst, Wolfgang; Tatzelt, Jörg; Haass, Christian; Winklhofer, Konstanze F.

    2009-01-01

    Loss-of-function mutations in the parkin gene (PARK2) and PINK1 gene (PARK6) are associated with autosomal recessive parkinsonism. PINK1 deficiency was recently linked to mitochondrial pathology in human cells and Drosophila melanogaster, which can be rescued by parkin, suggesting that both genes play a role in maintaining mitochondrial integrity. Here we demonstrate that an acute down-regulation of parkin in human SH-SY5Y cells severely affects mitochondrial morphology and function, a phenotype comparable with that induced by PINK1 deficiency. Alterations in both mitochondrial morphology and ATP production caused by either parkin or PINK1 loss of function could be rescued by the mitochondrial fusion proteins Mfn2 and OPA1 or by a dominant negative mutant of the fission protein Drp1. Both parkin and PINK1 were able to suppress mitochondrial fragmentation induced by Drp1. Moreover, in Drp1-deficient cells the parkin/PINK1 knockdown phenotype did not occur, indicating that mitochondrial alterations observed in parkin- or PINK1-deficient cells are associated with an increase in mitochondrial fission. Notably, mitochondrial fragmentation is an early phenomenon upon PINK1/parkin silencing that also occurs in primary mouse neurons and Drosophila S2 cells. We propose that the discrepant findings in adult flies can be explained by the time of phenotype analysis and suggest that in mammals different strategies may have evolved to cope with dysfunctional mitochondria. PMID:19546216

  2. Cancer: Mitochondrial Origins.

    PubMed

    Stefano, George B; Kream, Richard M

    2015-12-01

    The primacy of glucose derived from photosynthesis as an existential source of chemical energy across plant and animal phyla is universally accepted as a core principle in the biological sciences. In mammalian cells, initial processing of glucose to triose phosphate intermediates takes place within the cytosolic glycolytic pathway and terminates with temporal transport of reducing equivalents derived from pyruvate metabolism by membrane-associated respiratory complexes in the mitochondrial matrix. The intra-mitochondrial availability of molecular oxygen as the ultimate electron acceptor drives the evolutionary fashioned chemiosmotic production of ATP as a high-efficiency biological process. The mechanistic bases of carcinogenesis have demonstrated profound alteration of normative mitochondrial function, notably dysregulated respiratory processes. Accordingly, the classic Warburg effect functionally links aerobic glycolysis, aberrant production and release of lactate, and metabolic down-regulation of mitochondrial oxidative processes with the carcinogenetic phenotype. We surmise, however, that aerobic fermentation by cancer cells may also represent a developmental re-emergence of an evolutionarily conserved early phenotype, which was "sidelined" with the emergence of mitochondrial oxidative phosphorylation as a primary mechanism for ATP production in normal cells. Regardless of state-dependent physiological status in mixed populations of cancer cells, it has been established that mitochondria are functionally linked to the initiation of cancer and its progression. Biochemical, molecular, and physiological differences in cancer cell mitochondria, notably mtDNA heteroplasmy and allele-specific expression of selected nuclear genes, may represent major focal points for novel targeting and elimination of cancer cells in metastatic disease afflicting human populations. To date, and despite considerable research efforts, the practical realization of advanced mitochondrial

  3. Cancer: Mitochondrial Origins

    PubMed Central

    Stefano, George B.; Kream, Richard M.

    2015-01-01

    The primacy of glucose derived from photosynthesis as an existential source of chemical energy across plant and animal phyla is universally accepted as a core principle in the biological sciences. In mammalian cells, initial processing of glucose to triose phosphate intermediates takes place within the cytosolic glycolytic pathway and terminates with temporal transport of reducing equivalents derived from pyruvate metabolism by membrane-associated respiratory complexes in the mitochondrial matrix. The intra-mitochondrial availability of molecular oxygen as the ultimate electron acceptor drives the evolutionary fashioned chemiosmotic production of ATP as a high-efficiency biological process. The mechanistic bases of carcinogenesis have demonstrated profound alteration of normative mitochondrial function, notably dysregulated respiratory processes. Accordingly, the classic Warburg effect functionally links aerobic glycolysis, aberrant production and release of lactate, and metabolic down-regulation of mitochondrial oxidative processes with the carcinogenetic phenotype. We surmise, however, that aerobic fermentation by cancer cells may also represent a developmental re-emergence of an evolutionarily conserved early phenotype, which was “sidelined” with the emergence of mitochondrial oxidative phosphorylation as a primary mechanism for ATP production in normal cells. Regardless of state-dependent physiological status in mixed populations of cancer cells, it has been established that mitochondria are functionally linked to the initiation of cancer and its progression. Biochemical, molecular, and physiological differences in cancer cell mitochondria, notably mtDNA heteroplasmy and allele-specific expression of selected nuclear genes, may represent major focal points for novel targeting and elimination of cancer cells in metastatic disease afflicting human populations. To date, and despite considerable research efforts, the practical realization of advanced

  4. Mitochondrial calcium uptake.

    PubMed

    Williams, George S B; Boyman, Liron; Chikando, Aristide C; Khairallah, Ramzi J; Lederer, W J

    2013-06-25

    Calcium (Ca(2+)) uptake into the mitochondrial matrix is critically important to cellular function. As a regulator of matrix Ca(2+) levels, this flux influences energy production and can initiate cell death. If large, this flux could potentially alter intracellular Ca(2+) ([Ca(2+)]i) signals. Despite years of study, fundamental disagreements on the extent and speed of mitochondrial Ca(2+) uptake still exist. Here, we review and quantitatively analyze mitochondrial Ca(2+) uptake fluxes from different tissues and interpret the results with respect to the recently proposed mitochondrial Ca(2+) uniporter (MCU) candidate. This quantitative analysis yields four clear results: (i) under physiological conditions, Ca(2+) influx into the mitochondria via the MCU is small relative to other cytosolic Ca(2+) extrusion pathways; (ii) single MCU conductance is ∼6-7 pS (105 mM [Ca(2+)]), and MCU flux appears to be modulated by [Ca(2+)]i, suggesting Ca(2+) regulation of MCU open probability (P(O)); (iii) in the heart, two features are clear: the number of MCU channels per mitochondrion can be calculated, and MCU probability is low under normal conditions; and (iv) in skeletal muscle and liver cells, uptake per mitochondrion varies in magnitude but total uptake per cell still appears to be modest. Based on our analysis of available quantitative data, we conclude that although Ca(2+) critically regulates mitochondrial function, the mitochondria do not act as a significant dynamic buffer of cytosolic Ca(2+) under physiological conditions. Nevertheless, with prolonged (superphysiological) elevations of [Ca(2+)]i, mitochondrial Ca(2+) uptake can increase 10- to 1,000-fold and begin to shape [Ca(2+)]i dynamics.

  5. Mitochondrial carrier homolog 2 (MTCH2): the recruitment and evolution of a mitochondrial carrier protein to a critical player in apoptosis.

    PubMed

    Robinson, Alan J; Kunji, Edmund R S; Gross, Atan

    2012-07-01

    Recent studies report mitochondrial carrier homolog 2 (MTCH2) as a novel and uncharacterized protein that acts as a receptor-like protein for the truncated BH3-interacting domain death agonist (tBID) protein in the outer membrane of mitochondria. These studies, using mouse embryonic stem cells and fibroblasts as well as mice with a conditional knockout of MTCH2 in the liver, showed that deletion of MTCH2 hindered recruitment of tBID to the mitochondria with subsequent reductions in the activation of pro-apoptotic proteins, mitochondrial outer membrane permeabilization and apoptosis. Sequence analysis shows that MTCH2 is present in all examined multicellular Metazoa as well as unicellular Choanoflagellata, and is a highly derived member of the mitochondrial carrier family. Mitochondrial carriers are monomeric transport proteins that are usually found in the inner mitochondrial membrane, where they exchange small substrates between the mitochondrial matrix and intermembrane space. There are extensive differences between the protein sequences of MTCH2 and other mitochondrial carriers that may explain the ability of MTCH2 to associate with tBID and thus its role in apoptosis. We review the experimental evidence for the role of MTCH2 in apoptosis and suggest that the original transport function of the ancestral MTCH2 mitochondrial carrier has been co-opted by the apoptotic machinery to provide a receptor and signaling mechanism. PMID:22326460

  6. Dynamics of enhanced mitochondrial respiration in female compared with male rat cerebral arteries.

    PubMed

    Rutkai, Ibolya; Dutta, Somhrita; Katakam, Prasad V; Busija, David W

    2015-11-01

    Mitochondrial respiration has never been directly examined in intact cerebral arteries. We tested the hypothesis that mitochondrial energetics of large cerebral arteries ex vivo are sex dependent. The Seahorse XFe24 analyzer was used to examine mitochondrial respiration in isolated cerebral arteries from adult male and female Sprague-Dawley rats. We examined the role of nitric oxide (NO) on mitochondrial respiration under basal conditions, using N(ω)-nitro-l-arginine methyl ester, and following pharmacological challenge using diazoxide (DZ), and also determined levels of mitochondrial and nonmitochondrial proteins using Western blot, and vascular diameter responses to DZ. The components of mitochondrial respiration including basal respiration, ATP production, proton leak, maximal respiration, and spare respiratory capacity were elevated in females compared with males, but increased in both male and female arteries in the presence of the NOS inhibitor. Although acute DZ treatment had little effect on mitochondrial respiration of male arteries, it decreased the respiration in female arteries. Levels of mitochondrial proteins in Complexes I-V and the voltage-dependent anion channel protein were elevated in female compared with male cerebral arteries. The DZ-induced vasodilation was greater in females than in males. Our findings show that substantial sex differences in mitochondrial respiratory dynamics exist in large cerebral arteries and may provide the mechanistic basis for observations that the female cerebral vasculature is more adaptable after injury.

  7. TSPO interacts with VDAC1 and triggers a ROS-mediated inhibition of mitochondrial quality control

    PubMed Central

    Gatliff, Jemma; East, Daniel; Crosby, James; Abeti, Rosella; Harvey, Robert; Craigen, William; Parker, Peter; Campanella, Michelangelo

    2015-01-01

    The 18-kDa TSPO (translocator protein) localizes on the outer mitochondrial membrane (OMM) and participates in cholesterol transport. Here, we report that TSPO inhibits mitochondrial autophagy downstream of the PINK1-PARK2 pathway, preventing essential ubiquitination of proteins. TSPO abolishes mitochondrial relocation of SQSTM1/p62 (sequestosome 1), and consequently that of the autophagic marker LC3 (microtubule-associated protein 1 light chain 3), thus leading to an accumulation of dysfunctional mitochondria, altering the appearance of the network. Independent of cholesterol regulation, the modulation of mitophagy by TSPO is instead dependent on VDAC1 (voltage-dependent anion channel 1), to which TSPO binds, reducing mitochondrial coupling and promoting an overproduction of reactive oxygen species (ROS) that counteracts PARK2-mediated ubiquitination of proteins. These data identify TSPO as a novel element in the regulation of mitochondrial quality control by autophagy, and demonstrate the importance for cell homeostasis of its expression ratio with VDAC1. PMID:25470454

  8. Mitochondrial DNA with a large-scale deletion causes two distinct mitochondrial disease phenotypes in mice.

    PubMed

    Katada, Shun; Mito, Takayuki; Ogasawara, Emi; Hayashi, Jun-Ichi; Nakada, Kazuto

    2013-09-01

    Studies in patients have suggested that the clinical phenotypes of some mitochondrial diseases might transit from one disease to another (e.g., Pearson syndrome [PS] to Kearns-Sayre syndrome) in single individuals carrying mitochondrial (mt) DNA with a common deletion (ΔmtDNA), but there is no direct experimental evidence for this. To determine whether ΔmtDNA has the pathologic potential to induce multiple mitochondrial disease phenotypes, we used trans-mitochondrial mice with a heteroplasmic state of wild-type mtDNA and ΔmtDNA (mito-miceΔ). Late-stage embryos carrying ≥50% ΔmtDNA showed abnormal hematopoiesis and iron metabolism in livers that were partly similar to PS (PS-like phenotypes), although they did not express sideroblastic anemia that is a typical symptom of PS. More than half of the neonates with PS-like phenotypes died by 1 month after birth, whereas the rest showed a decrease of ΔmtDNA load in the affected tissues, peripheral blood and liver, and they recovered from PS-like phenotypes. The proportion of ΔmtDNA in various tissues of the surviving mito-miceΔ increased with time, and Kearns-Sayre syndrome-like phenotypes were expressed when the proportion of mtDNA in various tissues reached >70-80%. Our model mouse study clearly showed that a single ΔmtDNA was responsible for at least two distinct disease phenotypes at different ages and suggested that the level and dynamics of mtDNA load in affected tissues would be important for the onset and transition of mitochondrial disease phenotypes in mice. PMID:23853091

  9. Impaired mitochondrial respiration and decreased fatigue resistance followed by severe muscle weakness in skeletal muscle of mitochondrial DNA mutator mice.

    PubMed

    Yamada, Takashi; Ivarsson, Niklas; Hernández, Andrés; Fahlström, Andreas; Cheng, Arthur J; Zhang, Shi-Jin; Bruton, Joseph D; Ulfhake, Brun; Westerblad, Håkan

    2012-12-01

    Mitochondrial dysfunction can drastically impair muscle function, with weakness and exercise intolerance as key symptoms. Here we examine the time course of development of muscle dysfunction in a mouse model of premature ageing induced by defective proofreading function of mitochondrial DNA (mtDNA) polymerase (mtDNA mutator mouse). Isolated fast-twitch muscles and single muscle fibres from young (3-5 months) and end-stage (11 months) mtDNA mutator mice were compared to age-matched control mice. Force and free myoplasmic [Ca(2+)] ([Ca(2+)](i)) were measured under resting conditions and during fatigue induced by repeated tetani. Muscles of young mtDNA mutator mice displayed no weakness in the rested state, but had lower force and [Ca(2+)](i) than control mice during induction of fatigue. Muscles of young mtDNA mutator mice showed decreased activities of citrate synthase and β-hydroxyacyl-coenzyme A dehydrogenase, reduced expression of cytochrome c oxidase, and decreased expression of triggers of mitochondrial biogenesis (PGC-1α, PPARα, AMPK). Muscles from end-stage mtDNA mutator mice showed weakness under resting conditions with markedly decreased tetanic [Ca(2+)](i), force per cross-sectional area and protein expression of the sarcoplasmic reticulum Ca(2+) pump (SERCA1). In conclusion, fast-twitch muscles of prematurely ageing mtDNA mutator mice display a sequence of deleterious mitochondrial-to-nucleus signalling with an initial decrease in oxidative capacity, which was not counteracted by activation of signalling to increase mitochondrial biogenesis. This was followed by severe muscle weakness in the end stage. These results have implication for normal ageing and suggest that decreased mitochondrial oxidative capacity due to a sedentary lifestyle may predispose towards muscle weakness developing later in life.

  10. Late Mitochondrial Acquisition, Really?

    PubMed Central

    Degli Esposti, Mauro

    2016-01-01

    This article provides a timely critique of a recent Nature paper by Pittis and Gabaldón that has suggested a late origin of mitochondria in eukaryote evolution. It shows that the inferred ancestry of many mitochondrial proteins has been incorrectly assigned by Pittis and Gabaldón to bacteria other than the aerobic proteobacteria from which the ancestor of mitochondria originates, thereby questioning the validity of their suggestion that mitochondrial acquisition may be a late event in eukaryote evolution. The analysis and approach presented here may guide future studies to resolve the true ancestry of mitochondria. PMID:27289097

  11. Parkin loss leads to PARIS-dependent declines in mitochondrial mass and respiration

    PubMed Central

    Stevens, Daniel A.; Lee, Yunjong; Kang, Ho Chul; Lee, Byoung Dae; Lee, Yun-Il; Bower, Aaron; Jiang, Haisong; Kang, Sung-Ung; Andrabi, Shaida A.; Dawson, Valina L.; Shin, Joo-Ho; Dawson, Ted M.

    2015-01-01

    Mutations in parkin lead to early-onset autosomal recessive Parkinson’s disease (PD) and inactivation of parkin is thought to contribute to sporadic PD. Adult knockout of parkin in the ventral midbrain of mice leads to an age-dependent loss of dopamine neurons that is dependent on the accumulation of parkin interacting substrate (PARIS), zinc finger protein 746 (ZNF746), and its transcriptional repression of PGC-1α. Here we show that adult knockout of parkin in mouse ventral midbrain leads to decreases in mitochondrial size, number, and protein markers consistent with a defect in mitochondrial biogenesis. This decrease in mitochondrial mass is prevented by short hairpin RNA knockdown of PARIS. PARIS overexpression in mouse ventral midbrain leads to decreases in mitochondrial number and protein markers and PGC-1α–dependent deficits in mitochondrial respiration. Taken together, these results suggest that parkin loss impairs mitochondrial biogenesis, leading to declining function of the mitochondrial pool and cell death. PMID:26324925

  12. Pharmacologic Effects on Mitochondrial Function

    ERIC Educational Resources Information Center

    Cohen, Bruce H.

    2010-01-01

    The vast majority of energy necessary for cellular function is produced in mitochondria. Free-radical production and apoptosis are other critical mitochondrial functions. The complex structure, electrochemical properties of the inner mitochondrial membrane (IMM), and genetic control from both mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) are…

  13. Plasma membrane depolarization and Na,K-ATPase impairment induced by mitochondrial toxins augment leukemia cell apoptosis via a novel mitochondrial amplification mechanism.

    PubMed

    Yin, Wu; Li, Xiang; Feng, Su; Cheng, Wei; Tang, Bo; Shi, Yi-Lin; Hua, Zi-Chun

    2009-07-15

    Na,K-ATPase is a ubiquitous transmembrane protein that regulates and maintains the intracellular Na(+) and K(+) gradient necessary for cell homeostasis. Recently, the importance of this pump in external stimuli-induced leukemia cell apoptosis has been increasingly appreciated, however, the exact role of Na,K-ATPase in mitochondrial apoptotic pathway still remains little understood. In this study, we found mitochondrial toxin rotenone caused a rapid mitochondrial membrane potential (MMP) collapse in Jurkat cells followed by plasma membrane depolarization (PMP). Similar results were also obtained in human U937 cells and non-cancerous mouse primary T cells. Rotenone-induced PMP depolarization occurred before apoptosis and well correlated with Na,K-ATPase impairment. To understand the mechanisms, Jurkat cells with mtDNA depletion and catalase overexpression were used. The results demonstrated that both PMP depolarization and Na,K-ATPase impairment induced by rotenone were regulated by mitochondrial H(2)O(2) and Bcl-2. Finally, Na,K-ATPase suppression by ouabain greatly accelerated and enhanced mitochondrial toxins-induced cells apoptosis in Jurkat, U937 and primary T cells. In sum, by using leukemia cells and mouse primary T cells, we confirmed that mitochondria-to-Na,K-ATPase and PMP depolarization might represent a novel mechanism for mitochondria to amplify death signals in the initiation stage of cells apoptosis induced by mitochondrial toxins.

  14. Restriction enzyme analysis of the mitochondrial genome in mitochondrial myopathy.

    PubMed Central

    Poulton, J; Turnbull, D M; Mehta, A B; Wilson, J; Gardiner, R M

    1988-01-01

    The mitochondrial myopathies are a heterogeneous group of disorders some of which may be caused by mutations in the mitochondrial genome. Mitochondrial DNA from 10 patients with mitochondrial myopathy and their mothers was analysed using five restriction enzymes and 11 mitochondrial probes in bacteriophage M13. No abnormalities were found in seven out of the 10 patients. Polymorphisms which have not previously been reported were detected in three patients and two of their mothers. These results exclude the presence of deletions or insertions of greater than 60 bp in the region of the mitochondrial genome examined. Any causative mitochondrial DNA mutations in these disorders are therefore likely to be point mutations or small structural rearrangements. Images PMID:2903249

  15. All-Trans-Retinoic Acid Enhances Mitochondrial Function in Models of Human Liver.

    PubMed

    Tripathy, Sasmita; Chapman, John D; Han, Chang Y; Hogarth, Cathryn A; Arnold, Samuel L M; Onken, Jennifer; Kent, Travis; Goodlett, David R; Isoherranen, Nina

    2016-05-01

    All-trans-retinoic acid (atRA) is the active metabolite of vitamin A. The liver is the main storage organ of vitamin A, but activation of the retinoic acid receptors (RARs) in mouse liver and in human liver cell lines has also been shown. AlthoughatRA treatment improves mitochondrial function in skeletal muscle in rodents, its role in modulating mitochondrial function in the liver is controversial, and little data are available regarding the human liver. The aim of this study was to determine whetheratRA regulates hepatic mitochondrial activity.atRA treatment increased the mRNA and protein expression of multiple components of mitochondrialβ-oxidation, tricarboxylic acid (TCA) cycle, and respiratory chain. Additionally,atRA increased mitochondrial biogenesis in human hepatocytes and in HepG2 cells with and without lipid loading based on peroxisome proliferator activated receptor gamma coactivator 1αand 1βand nuclear respiratory factor 1 mRNA and mitochondrial DNA quantification.atRA also increasedβ-oxidation and ATP production in HepG2 cells and in human hepatocytes. Knockdown studies of RARα, RARβ, and PPARδrevealed that the enhancement of mitochondrial biogenesis andβ-oxidation byatRA requires peroxisome proliferator activated receptor delta. In vivo in mice,atRA treatment increased mitochondrial biogenesis markers after an overnight fast. Inhibition ofatRA metabolism by talarozole, a cytochrome P450 (CYP) 26 specific inhibitor, increased the effects ofatRA on mitochondrial biogenesis markers in HepG2 cells and in vivo in mice. These studies show thatatRA regulates mitochondrial function and lipid metabolism and that increasingatRA concentrations in human liver via CYP26 inhibition may increase mitochondrial biogenesis and fatty acidβ-oxidation and provide therapeutic benefit in diseases associated with mitochondrial dysfunction. PMID:26921399

  16. Abnormal Mitochondrial L-Arginine Transport Contributes to the Pathogenesis of Heart Failure and Rexoygenation Injury

    PubMed Central

    Byrne, Melissa; Joshi, Mandar; Horlock, Duncan; Lam, Nicholas T.; Gregorevic, Paul; McGee, Sean L.; Kaye, David M.

    2014-01-01

    Background Impaired mitochondrial function is fundamental feature of heart failure (HF) and myocardial ischemia. In addition to the effects of heightened oxidative stress, altered nitric oxide (NO) metabolism, generated by a mitochondrial NO synthase, has also been proposed to impact upon mitochondrial function. However, the mechanism responsible for arginine transport into mitochondria and the effect of HF on such a process is unknown. We therefore aimed to characterize mitochondrial L-arginine transport and to investigate the hypothesis that impaired mitochondrial L-arginine transport plays a key role in the pathogenesis of heart failure and myocardial injury. Methods and Results In mitochondria isolated from failing hearts (sheep rapid pacing model and mouse Mst1 transgenic model) we demonstrated a marked reduction in L-arginine uptake (p<0.05 and p<0.01 respectively) and expression of the principal L-arginine transporter, CAT-1 (p<0.001, p<0.01) compared to controls. This was accompanied by significantly lower NO production and higher 3-nitrotyrosine levels (both p<0.05). The role of mitochondrial L-arginine transport in modulating cardiac stress responses was examined in cardiomyocytes with mitochondrial specific overexpression of CAT-1 (mtCAT1) exposed to hypoxia-reoxygenation stress. mtCAT1 cardiomyocytes had significantly improved mitochondrial membrane potential, respiration and ATP turnover together with significantly decreased reactive oxygen species production and cell death following mitochondrial stress. Conclusion These data provide new insights into the role of L-arginine transport in mitochondrial biology and cardiovascular disease. Augmentation of mitochondrial L-arginine availability may be a novel therapeutic strategy for myocardial disorders involving mitochondrial stress such as heart failure and reperfusion injury. PMID:25111602

  17. Isomerically Pure Tetramethylrhodamine Voltage Reporters.

    PubMed

    Deal, Parker E; Kulkarni, Rishikesh U; Al-Abdullatif, Sarah H; Miller, Evan W

    2016-07-27

    We present the design, synthesis, and application of a new family of fluorescent voltage indicators based on isomerically pure tetramethylrhodamines. These new Rhodamine Voltage Reporters, or RhoVRs, use photoinduced electron transfer (PeT) as a trigger for voltage sensing, display excitation and emission profiles in the green to orange region of the visible spectrum, demonstrate high sensitivity to membrane potential changes (up to 47% ΔF/F per 100 mV), and employ a tertiary amide derived from sarcosine, which aids in membrane localization and simultaneously simplifies the synthetic route to the voltage sensors. The most sensitive of the RhoVR dyes, RhoVR 1, features a methoxy-substituted diethylaniline donor and phenylenevinylene molecular wire at the 5'-position of the rhodamine aryl ring, exhibits the highest voltage sensitivity to date for red-shifted PeT-based voltage sensors, and is compatible with simultaneous imaging alongside green fluorescent protein-based indicators. The discoveries that sarcosine-based tertiary amides in the context of molecular-wire voltage indicators prevent dye internalization and 5'-substituted voltage indicators exhibit improved voltage sensitivity should be broadly applicable to other types of PeT-based voltage-sensitive fluorophores. PMID:27428174

  18. Transistor voltage comparator performs own sensing

    NASA Technical Reports Server (NTRS)

    Cliff, R. A.

    1965-01-01

    Detection of the highest voltage input among a group of varying voltage inputs is accomplished by a transistorized voltage comparison circuit. The collector circuits of the transistors perform the sensing function. Input voltage levels are governed by the transistors.

  19. 'Mitochondrial energy imbalance and lipid peroxidation cause cell death in Friedreich's ataxia'

    PubMed Central

    Abeti, R; Parkinson, M H; Hargreaves, I P; Angelova, P R; Sandi, C; Pook, M A; Giunti, P; Abramov, A Y

    2016-01-01

    Friedreich's ataxia (FRDA) is an inherited neurodegenerative disease. The mutation consists of a GAA repeat expansion within the FXN gene, which downregulates frataxin, leading to abnormal mitochondrial iron accumulation, which may in turn cause changes in mitochondrial function. Although, many studies of FRDA patients and mouse models have been conducted in the past two decades, the role of frataxin in mitochondrial pathophysiology remains elusive. Are the mitochondrial abnormalities only a side effect of the increased accumulation of reactive iron, generating oxidative stress? Or does the progressive lack of iron-sulphur clusters (ISCs), induced by reduced frataxin, cause an inhibition of the electron transport chain complexes (CI, II and III) leading to reactive oxygen species escaping from oxidative phosphorylation reactions? To answer these crucial questions, we have characterised the mitochondrial pathophysiology of a group of disease-relevant and readily accessible neurons, cerebellar granule cells, from a validated FRDA mouse model. By using live cell imaging and biochemical techniques we were able to demonstrate that mitochondria are deregulated in neurons from the YG8R FRDA mouse model, causing a decrease in mitochondrial membrane potential (▵Ψm) due to an inhibition of Complex I, which is partially compensated by an overactivation of Complex II. This complex activity imbalance leads to ROS generation in both mitochondrial matrix and cytosol, which results in glutathione depletion and increased lipid peroxidation. Preventing this increase in lipid peroxidation, in neurons, protects against in cell death. This work describes the pathophysiological properties of the mitochondria in neurons from a FRDA mouse model and shows that lipid peroxidation could be an important target for novel therapeutic strategies in FRDA, which still lacks a cure. PMID:27228352

  20. Preventive mitochondrial replacement.

    PubMed

    Orgel, L E

    1997-03-01

    Techniques used recently to clone a sheep generate chimeras with the genome of the donor cell and mainly the mitochondria of the acceptor egg. The use of the same techniques should allow a mother carrying a mitochondrial defect to bear a normal child with normal mitochondria.

  1. Modeling mitochondrial function.

    PubMed

    Balaban, Robert S

    2006-12-01

    The mitochondrion represents a unique opportunity to apply mathematical modeling to a complex biological system. Understanding mitochondrial function and control is important since this organelle is critical in energy metabolism as well as playing key roles in biochemical synthesis, redox control/signaling, and apoptosis. A mathematical model, or hypothesis, provides several useful insights including a rigorous test of the consensus view of the operation of a biological process as well as providing methods of testing and creating new hypotheses. The advantages of the mitochondrial system for applying a mathematical model include the relative simplicity and understanding of the matrix reactions, the ability to study the mitochondria as a independent contained organelle, and, most importantly, one can dynamically measure many of the internal reaction intermediates, on line. The developing ability to internally monitor events within the metabolic network, rather than just the inflow and outflow, is extremely useful in creating critical bounds on complex mathematical models using the individual reaction mechanisms available. However, many serious problems remain in creating a working model of mitochondrial function including the incomplete definition of metabolic pathways, the uncertainty of using in vitro enzyme kinetics, as well as regulatory data in the intact system and the unknown chemical activities of relevant molecules in the matrix. Despite these formidable limitations, the advantages of the mitochondrial system make it one of the best defined mammalian metabolic networks that can be used as a model system for understanding the application and use of mathematical models to study biological systems.

  2. ENERGETICS, EPIGENETICS, MITOCHONDRIAL GENETICS

    PubMed Central

    Wallace, Douglas C.; Fan, Weiwei

    2011-01-01

    The epigenome has been hypothesized to provide the interface between the environment and the nuclear DNA (nDNA) genes. Key factors in the environment are the availability of calories and demands on the organism’s energetic capacity. Energy is funneled through glycolysis and mitochondrial oxidative phosphorylation (OXPHOS), the cellular bioenergetic systems. Since there are thousands of bioenergetic genes dispersed across the chromosomes and mitochondrial DNA (mtDNA), both cis and trans regulation of the nDNA genes is required. The bioenergetic systems convert environmental calories into ATP, acetyl-Coenzyme A (acetyl-CoA), S-adenosyl-methionine (SAM), and reduced NAD+. When calories are abundant, ATP and acetyl-CoA phosphorylate and acetylate chromatin, opening the nDNA for transcription and replication. When calories are limiting, chromatin phosphorylation and acetylation are lost and gene expression is suppressed. DNA methylaton via SAM can also be modulated by mitochondrial function. Phosphorylation and acetylation are also pivotal to regulating cellular signal transduction pathways. Therefore, bioenergetics provides the interface between the environment and the epigenome. Consistent with this conclusion, the clinical phenotypes of bioenergetic diseases are strikingly similar to those observed in epigenetic diseases (Angelman, Rett, Fragile X Syndromes, the laminopathies, cancer, etc.), and an increasing number of epigenetic diseases are being associated with mitochondrial dysfunction. This bioenergetic-epigenomic hypothesis has broad implications for the etiology, pathophysiology, and treatment of a wide range of common diseases. PMID:19796712

  3. Altered brain energetics induces mitochondrial fission arrest in Alzheimer's Disease.

    PubMed

    Zhang, Liang; Trushin, Sergey; Christensen, Trace A; Bachmeier, Benjamin V; Gateno, Benjamin; Schroeder, Andreas; Yao, Jia; Itoh, Kie; Sesaki, Hiromi; Poon, Wayne W; Gylys, Karen H; Patterson, Emily R; Parisi, Joseph E; Diaz Brinton, Roberta; Salisbury, Jeffrey L; Trushina, Eugenia

    2016-01-01

    Altered brain metabolism is associated with progression of Alzheimer's Disease (AD). Mitochondria respond to bioenergetic changes by continuous fission and fusion. To account for three dimensional architecture of the brain tissue and organelles, we applied 3-dimensional electron microscopy (3D EM) reconstruction to visualize mitochondrial structure in the brain tissue from patients and mouse models of AD. We identified a previously unknown mitochondrial fission arrest phenotype that results in elongated interconnected organelles, "mitochondria-on-a-string" (MOAS). Our data suggest that MOAS formation may occur at the final stages of fission process and was not associated with altered translocation of activated dynamin related protein 1 (Drp1) to mitochondria but with reduced GTPase activity. Since MOAS formation was also observed in the brain tissue of wild-type mice in response to hypoxia or during chronological aging, fission arrest may represent fundamental compensatory adaptation to bioenergetic stress providing protection against mitophagy that may preserve residual mitochondrial function. The discovery of novel mitochondrial phenotype that occurs in the brain tissue in response to energetic stress accurately detected only using 3D EM reconstruction argues for a major role of mitochondrial dynamics in regulating neuronal survival. PMID:26729583

  4. High voltage feedthrough bushing

    DOEpatents

    Brucker, John P.

    1993-01-01

    A feedthrough bushing for a high voltage diode provides for using compression sealing for all sealing surfaces. A diode assembly includes a central conductor extending through the bushing and a grading ring assembly circumferentially surrounding and coaxial with the central conductor. A flexible conductive plate extends between and compressively seals against the central conductor and the grading ring assembly, wherein the flexibility of the plate allows inner and outer portions of the plate to axially translate for compression sealing against the central conductor and the grading ring assembly, respectively. The inner portion of the plate is bolted to the central conductor for affecting sealing. A compression beam is also bolted to the central conductor and engages the outer portion of the plate to urge the outer portion toward the grading ring assembly to obtain compression sealing therebetween.

  5. High voltage isolation transformer

    NASA Technical Reports Server (NTRS)

    Clatterbuck, C. H.; Ruitberg, A. P. (Inventor)

    1985-01-01

    A high voltage isolation transformer is provided with primary and secondary coils separated by discrete electrostatic shields from the surfaces of insulating spools on which the coils are wound. The electrostatic shields are formed by coatings of a compound with a low electrical conductivity which completely encase the coils and adhere to the surfaces of the insulating spools adjacent to the coils. Coatings of the compound also line axial bores of the spools, thereby forming electrostatic shields separating the spools from legs of a ferromagnetic core extending through the bores. The transformer is able to isolate a high constant potential applied to one of its coils, without the occurrence of sparking or corona, by coupling the coatings, lining the axial bores to the ferromagnetic core and by coupling one terminal of each coil to the respective coating encasing the coil.

  6. Mitochondrial Atpif1 regulates heme synthesis in developing erythroblasts

    PubMed Central

    Shah, Dhvanit I.; Takahashi-Makise, Naoko; Cooney, Jeffrey D.; Li, Liangtao; Schultz, Iman J.; Pierce, Eric L.; Narla, Anupama; Seguin, Alexandra; Hattangadi, Shilpa M.; Medlock, Amy E.; Langer, Nathaniel B.; Dailey, Tamara A.; Hurst, Slater N.; Faccenda, Danilo; Wiwczar, Jessica M.; Heggers, Spencer K.; Vogin, Guillaume; Chen, Wen; Chen, Caiyong; Campagna, Dean R.; Brugnara, Carlo; Zhou, Yi; Ebert, Benjamin L.; Danial, Nika N.; Fleming, Mark D.; Ward, Diane M.; Campanella, Michelangelo; Dailey, Harry A.; Kaplan, Jerry; Paw, Barry H.

    2012-01-01

    SUMMARY Defects in the availability of heme substrates or the catalytic activity of the terminal enzyme in heme biosynthesis, ferrochelatase (Fech), impair heme synthesis, and thus cause human congenital anemias1,2. The inter-dependent functions of regulators of mitochondrial homeostasis and enzymes responsible for heme synthesis are largely unknown. To uncover this unmet need, we utilized zebrafish genetic screens and cloned mitochondrial ATPase inhibitory factor 1 (atpif1) from a zebrafish mutant with profound anemia, pinotage (pnt tq209). We now report a direct mechanism establishing that Atpif1 regulates the catalytic efficiency of vertebrate Fech to synthesize heme. The loss of Atpif1 impairs hemoglobin synthesis in zebrafish, mouse, and human hematopoietic models as a consequence of diminished Fech activity, and elevated mitochondrial pH. To understand the relationship among mitochondrial pH, redox potential, [2Fe-2S] clusters, and Fech activity, we used (1) genetic complementation studies of Fech constructs with or without [2Fe-2S] clusters in pnt, and (2) pharmacological agents modulating mitochondrial pH and redox potential. The presence of [2Fe-2S] cluster renders vertebrate Fech vulnerable to Atpif1-regulated mitochondrial pH and redox potential perturbations. Therefore, Atpif1 deficiency reduces the efficiency of vertebrate Fech to synthesize heme, resulting in anemia. The novel mechanism of Atpif1 as a regulator of heme synthesis advances the understanding of mitochondrial heme homeostasis and red blood cell development. A deficiency of Atpif1 may contribute to important human diseases, such as congenital sideroblastic anemias and mitochondriopathies. PMID:23135403

  7. [The mitochondrial genome of protists].

    PubMed

    Odintsova, M S; Iurina, N P

    2002-06-01

    The data on the structure and functions of the mitochondrial genomes of protists (Protozoa and unicellular red and green algae) are reviewed. It is emphasized that mitochondrial gene structure and composition, as well as organization of mitochondrial genomes in protists are more diverse than in multicellular eukaryotes. The gene content of mitochondrial genomes of protists are closer to those of plants than animals or fungi. In the protist mitochondrial DNA, both the universal (as in higher plants) and modified (as in animals and fungi) genetic codes are used. In the overwhelming majority of cases, protist mitochondrial genomes code for the major and minor rRNA components, some tRNAs, and about 30 proteins of the respiratory chain and ribosomes. Based on comparison of the mitochondrial genomes of various protists, the origin and evolution of mitochondria are briefly discussed.

  8. Apoptotic Signaling in Mouse Odontogenesis

    PubMed Central

    Svandova, Eva; Tucker, Abigail S.

    2012-01-01

    Abstract Apoptosis is an important morphogenetic event in embryogenesis as well as during postnatal life. In the last 2 decades, apoptosis in tooth development (odontogenesis) has been investigated with gradually increasing focus on the mechanisms and signaling pathways involved. The molecular machinery responsible for apoptosis exhibits a high degree of conservation but also organ and tissue specific patterns. This review aims to discuss recent knowledge about apoptotic signaling networks during odontogenesis, concentrating on the mouse, which is often used as a model organism for human dentistry. Apoptosis accompanies the entire development of the tooth and corresponding remodeling of the surrounding bony tissue. It is most evident in its role in the elimination of signaling centers within developing teeth, removal of vestigal tooth germs, and in odontoblast and ameloblast organization during tooth mineralization. Dental apoptosis is caspase dependent and proceeds via mitochondrial mediated cell death with possible amplification by Fas-FasL signaling modulated by Bcl-2 family members. PMID:22204278

  9. Apoptotic signaling in mouse odontogenesis.

    PubMed

    Matalova, Eva; Svandova, Eva; Tucker, Abigail S

    2012-01-01

    Apoptosis is an important morphogenetic event in embryogenesis as well as during postnatal life. In the last 2 decades, apoptosis in tooth development (odontogenesis) has been investigated with gradually increasing focus on the mechanisms and signaling pathways involved. The molecular machinery responsible for apoptosis exhibits a high degree of conservation but also organ and tissue specific patterns. This review aims to discuss recent knowledge about apoptotic signaling networks during odontogenesis, concentrating on the mouse, which is often used as a model organism for human dentistry. Apoptosis accompanies the entire development of the tooth and corresponding remodeling of the surrounding bony tissue. It is most evident in its role in the elimination of signaling centers within developing teeth, removal of vestigal tooth germs, and in odontoblast and ameloblast organization during tooth mineralization. Dental apoptosis is caspase dependent and proceeds via mitochondrial mediated cell death with possible amplification by Fas-FasL signaling modulated by Bcl-2 family members.

  10. Mitochondrial transcription factor A regulation of mitochondrial degeneration in experimental diabetic neuropathy

    PubMed Central

    Chandrasekaran, Krish; Anjaneyulu, Muragundla; Inoue, Tatsuya; Choi, Joungil; Sagi, Avinash Rao; Chen, Chen; Ide, Tamomi

    2015-01-01

    Oxidative stress-induced mitochondrial dysfunction and mitochondrial DNA (mtDNA) damage in peripheral neurons is considered to be important in the development of diabetic neuropathy. Mitochondrial transcription factor A (TFAM) wraps mtDNA and promotes mtDNA replication and transcription. We studied whether overexpression of TFAM reverses experimental peripheral diabetic neuropathy using TFAM transgenic mice (TFAM Tg) that express human TFAM (hTFAM). Levels of mouse mtDNA and the total TFAM (mouse TFAM + hTFAM) in the dorsal root ganglion (DRG) increased by approximately twofold in the TFAM Tg mice compared with control (WT) mice. WT and TFAM Tg mice were made diabetic by the administration of streptozotocin. Neuropathy end points were motor and sensory nerve conduction velocities, mechanical allodynia, thermal nociception, and intraepidermal nerve fiber density (IENFD). In the DRG neurons, mtDNA copy number and damage to mtDNA were quantified by qPCR, and TFAM levels were measured by Western blot. Mice with 16-wk duration of diabetes developed motor and sensory nerve conduction deficits, behavioral deficits, and intraepidermal nerve fiber loss. All of these changes were mostly prevented in diabetic TFAM Tg mice and were independent of changes in blood parameters. Mice with 16 wk of diabetes had a 40% decrease in mtDNA copy number compared with nondiabetic mice (P < 0.01). Importantly, the mtDNA copy number in diabetic TFAM Tg mice reached the same level as that of WT nondiabetic mice. In comparison, there was upregulation of mtDNA and TFAM in 6-wk diabetic mice, suggesting that TFAM activation could be a therapeutic strategy to treat peripheral neuropathy. PMID:25944881

  11. Mitochondrial transcription factor A regulation of mitochondrial degeneration in experimental diabetic neuropathy.

    PubMed

    Chandrasekaran, Krish; Anjaneyulu, Muragundla; Inoue, Tatsuya; Choi, Joungil; Sagi, Avinash Rao; Chen, Chen; Ide, Tamomi; Russell, James W

    2015-07-15

    Oxidative stress-induced mitochondrial dysfunction and mitochondrial DNA (mtDNA) damage in peripheral neurons is considered to be important in the development of diabetic neuropathy. Mitochondrial transcription factor A (TFAM) wraps mtDNA and promotes mtDNA replication and transcription. We studied whether overexpression of TFAM reverses experimental peripheral diabetic neuropathy using TFAM transgenic mice (TFAM Tg) that express human TFAM (hTFAM). Levels of mouse mtDNA and the total TFAM (mouse TFAM + hTFAM) in the dorsal root ganglion (DRG) increased by approximately twofold in the TFAM Tg mice compared with control (WT) mice. WT and TFAM Tg mice were made diabetic by the administration of streptozotocin. Neuropathy end points were motor and sensory nerve conduction velocities, mechanical allodynia, thermal nociception, and intraepidermal nerve fiber density (IENFD). In the DRG neurons, mtDNA copy number and damage to mtDNA were quantified by qPCR, and TFAM levels were measured by Western blot. Mice with 16-wk duration of diabetes developed motor and sensory nerve conduction deficits, behavioral deficits, and intraepidermal nerve fiber loss. All of these changes were mostly prevented in diabetic TFAM Tg mice and were independent of changes in blood parameters. Mice with 16 wk of diabetes had a 40% decrease in mtDNA copy number compared with nondiabetic mice (P < 0.01). Importantly, the mtDNA copy number in diabetic TFAM Tg mice reached the same level as that of WT nondiabetic mice. In comparison, there was upregulation of mtDNA and TFAM in 6-wk diabetic mice, suggesting that TFAM activation could be a therapeutic strategy to treat peripheral neuropathy.

  12. Targeting lipid peroxidation and mitochondrial imbalance in Friedreich's ataxia.

    PubMed

    Abeti, Rosella; Uzun, Ebru; Renganathan, Indhushri; Honda, Tadashi; Pook, Mark A; Giunti, Paola

    2015-09-01

    Friedreich's ataxia (FRDA) is an autosomal recessive disorder, caused by reduced levels of the protein frataxin. This protein is located in the mitochondria, where it functions in the biogenesis of iron-sulphur clusters (ISCs), which are important for the function of the mitochondrial respiratory chain complexes. Moreover, disruption in iron biogenesis may lead to oxidative stress. Oxidative stress can be the cause and/or the consequence of mitochondrial energy imbalance, leading to cell death. Fibroblasts from two FRDA mouse models, YG8R and KIKO, were used to analyse two different categories of protective compounds: deuterised poly-unsaturated fatty acids (dPUFAs) and Nrf2-inducers. The former have been shown to protect the cell from damage induced by lipid peroxidation and the latter trigger the well-known Nrf2 antioxidant pathway. Our results show that the sensitivity to oxidative stress of YG8R and KIKO mouse fibroblasts, resulting in cell death and lipid peroxidation, can be prevented by d4-PUFA and Nrf2-inducers (SFN and TBE-31). The mitochondrial membrane potential (ΔΨm) of YG8R and KIKO fibroblasts revealed a difference in their mitochondrial pathophysiology, which may be due to the different genetic basis of the two models. This suggests that variable levels of reduced frataxin may act differently on mitochondrial pathophysiology and that these two cell models could be useful in recapitulating the observed differences in the FRDA phenotype. This may reflect a different modulatory effect towards cell death that will need to be investigated further. PMID:26141703

  13. Modulation of mitochondrial function and morphology by interaction of Omi/HtrA2 with the mitochondrial fusion factor OPA1

    SciTech Connect

    Kieper, Nicole; Holmstroem, Kira M.; Ciceri, Dalila; Fiesel, Fabienne C.; Wolburg, Hartwig; Ziviani, Elena; Whitworth, Alexander J.; Martins, L. Miguel; Kahle, Philipp J.; Krueger, Rejko

    2010-04-15

    Loss of Omi/HtrA2 function leads to nerve cell loss in mouse models and has been linked to neurodegeneration in Parkinson's and Huntington's disease. Omi/HtrA2 is a serine protease released as a pro-apoptotic factor from the mitochondrial intermembrane space into the cytosol. Under physiological conditions, Omi/HtrA2 is thought to be involved in protection against cellular stress, but the cytological and molecular mechanisms are not clear. Omi/HtrA2 deficiency caused an accumulation of reactive oxygen species and reduced mitochondrial membrane potential. In Omi/HtrA2 knockout mouse embryonic fibroblasts, as well as in Omi/HtrA2 silenced human HeLa cells and Drosophila S2R+ cells, we found elongated mitochondria by live cell imaging. Electron microscopy confirmed the mitochondrial morphology alterations and showed abnormal cristae structure. Examining the levels of proteins involved in mitochondrial fusion, we found a selective up-regulation of more soluble OPA1 protein. Complementation of knockout cells with wild-type Omi/HtrA2 but not with the protease mutant [S306A]Omi/HtrA2 reversed the mitochondrial elongation phenotype and OPA1 alterations. Finally, co-immunoprecipitation showed direct interaction of Omi/HtrA2 with endogenous OPA1. Thus, we show for the first time a direct effect of loss of Omi/HtrA2 on mitochondrial morphology and demonstrate a novel role of this mitochondrial serine protease in the modulation of OPA1. Our results underscore a critical role of impaired mitochondrial dynamics in neurodegenerative disorders.

  14. Integrative Toxicoproteomics Implicates Impaired Mitochondrial Glutathione Import as an Off-Target Effect of Troglitazone

    PubMed Central

    2013-01-01

    Troglitazone, a first-generation thiazolidinedione of antihyperglycaemic properties, was withdrawn from the market due to unacceptable idiosyncratic hepatotoxicity. Despite intensive research, the underlying mechanism of troglitazone-induced liver toxicity remains unknown. Here we report the use of the Sod2+/– mouse model of silent mitochondrial oxidative-stress-based and quantitative mass spectrometry-based proteomics to track the mitochondrial proteome changes induced by physiologically relevant troglitazone doses. By quantitative untargeted proteomics, we first globally profiled the Sod2+/– hepatic mitochondria proteome and found perturbations including GSH metabolism that enhanced the toxicity of the normally nontoxic troglitazone. Short- and long-term troglitazone administration in Sod2+/– mouse led to a mitochondrial proteome shift from an early compensatory response to an eventual phase of intolerable oxidative stress, due to decreased mitochondrial glutathione (mGSH) import protein, decreased dicarboxylate ion carrier (DIC), and the specific activation of ASK1-JNK and FOXO3a with prolonged troglitazone exposure. Furthermore, mapping of the detected proteins onto mouse specific protein-centered networks revealed lipid-associated proteins as contributors to overt mitochondrial and liver injury when under prolonged exposure to the lipid-normalizing troglitazone. By integrative toxicoproteomics, we demonstrated a powerful systems approach in identifying the collapse of specific fragile nodes and activation of crucial proteome reconfiguration regulators when targeted by an exogenous toxicant. PMID:23659346

  15. FAST KINASE DOMAIN-CONTAINING PROTEIN 3 IS A MITOCHONDRIAL PROTEIN ESSENTIAL FOR CELLULAR RESPIRATION

    PubMed Central

    Simarro, Maria; Gimenez-Cassina, Alfredo; Kedersha, Nancy; Lazaro, Jean-Bernard; Adelmant, Guillaume O; Marto, Jarrod A; Rhee, Kirsten; Tisdale, Sarah; Danial, Nika; Benarafa, Charaf; Orduña, Anonio; Anderson, Paul

    2010-01-01

    Fas-activated serine/threonine phosphoprotein (FAST) is the founding member of the FAST kinase domain-containing protein (FASTKD) family that includes FASTKD1-5. FAST is a sensor of mitochondrial stress that modulates protein translation to promote the survival of cells exposed to adverse conditions. Mutations in FASTKD2 have been linked to a mitochondrial encephalomyopathy that is associated with reduced cytochrome c oxidase activity, an essential component of the mitochondrial electron transport chain. We have confirmed the mitochondrial localization of FASTKD2 and shown that all FASTKD family members are found in mitochondria. Although human and mouse FASTKD1-5 genes are expressed ubiquitously, some of them are most abundantly expressed in mitochondria-enriched tissues. We have found that RNA interference-mediated knockdown of FASTKD3 severely blunts basal and stress-induced mitochondrial oxygen consumption without disrupting the assembly of respiratory chain complexes. Tandem affinity purification reveals that FASTKD3 interacts with components of mitochondrial respiratory and translation machineries. Our results introduce FASTKD3 as an essential component of mitochondrial respiration that may modulate energy balance in cells exposed to adverse conditions by functionally coupling mitochondrial protein synthesis to respiration. PMID:20869947

  16. Geniposide attenuates mitochondrial dysfunction and memory deficits in APP/PS1 transgenic mice.

    PubMed

    Lv, Cui; Liu, Xiaoli; Liu, Hongjuan; Chen, Tong; Zhang, Wensheng

    2014-01-01

    Oxidative stress and mitochondrial dysfunction appear early and contribute to the disease progression in Alzheimer's disease (AD), which can be detected extensively in AD patients brains as well as in transgenic AD mice brains. Thus, treatments that result in attenuation of oxidative stress and mitochondrial dysfunction may hold potential for AD treatment. Geniposide, a pharmacologically active component purified from gardenia fruit, exhibits anti-oxidative, antiinflammatory and other important therapeutic properties. However, whether geniposide has any protective effect on oxidative stress and mitochondrial dysfunction in AD transgenic mouse model has not yet been reported. Here, we demonstrate that intragastric administration of geniposide significantly reduces oxidative stress and mitochondrial dysfunction in addition to improving learning and memory in APP/PS1 mice. Geniposide exerts protective effects on mitochondrial dysfunction in APP/PS1 mice through suppressing the mitochondrial oxidative damage and increasing the mitochondrial membrane potential and activity of cytochrome c oxidase. These studies indicate that geniposide may attenuate memory deficits through the suppression of mitochondrial oxidative stress. Thus, geniposide may be a potential therapeutic reagent for halting and preventing AD progress.

  17. The development of mitochondrial medicine.

    PubMed Central

    Luft, R

    1994-01-01

    Primary defects in mitochondrial function are implicated in over 100 diseases, and the list continues to grow. Yet the first mitochondrial defect--a myopathy--was demonstrated only 35 years ago. The field's dramatic expansion reflects growth of knowledge in three areas: (i) characterization of mitochondrial structure and function, (ii) elucidation of the steps involved in mitochondrial biosynthesis, and (iii) discovery of specific mitochondrial DNA. Many mitochondrial diseases are accompanied by mutations in this DNA. Inheritance is by maternal transmission. The metabolic defects encompass the electron transport complexes, intermediates of the tricarboxylic acid cycle, and substrate transport. The clinical manifestations are protean, most often involving skeletal muscle and the central nervous system. In addition to being a primary cause of disease, mitochondrial DNA mutations and impaired oxidation have now been found to occur as secondary phenomena in aging as well as in age-related degenerative diseases such as Parkinson, Alzheimer, and Huntington diseases, amyotrophic lateral sclerosis and cardiomyopathies, atherosclerosis, and diabetes mellitus. Manifestations of both the primary and secondary mitochondrial diseases are thought to result from the production of oxygen free radicals. With increased understanding of the mechanisms underlying the mitochondrial dysfunctions has come the beginnings of therapeutic strategies, based mostly on the administration of antioxidants, replacement of cofactors, and provision of nutrients. At the present accelerating pace of development of what may be called mitochondrial medicine, much more is likely to be achieved within the next few years. Images PMID:8090715

  18. Release of targeted p53 from the mitochondrion as an early signal during mitochondrial dysfunction

    EPA Science Inventory

    Increased accumulation of p53 tumor suppressor protein is an early response to low-level stressors. To investigate the fate of mitochondrial-sequestered p53, mouse embryonic fibroblast cells (MEFs) on a p53-deficient genetic background were transfected with p53-EGFP fusion protei...

  19. Platyzoan mitochondrial genomes.

    PubMed

    Wey-Fabrizius, Alexandra R; Podsiadlowski, Lars; Herlyn, Holger; Hankeln, Thomas

    2013-11-01

    Platyzoa is a putative lophotrochozoan (spiralian) subtaxon within the protostome clade of Metazoa, comprising a range of biologically diverse, mostly small worm-shaped animals. The monophyly of Platyzoa, the relationships between the putative subgroups Platyhelminthes, Gastrotricha and Gnathifera (the latter comprising at least Gnathostomulida, "Rotifera" and Acanthocephala) as well as some aspects of the internal phylogenies of these subgroups are highly debated. Here we review how complete mitochondrial (mt) genome data contribute to these debates. We highlight special features of the mt genomes and discuss problems in mtDNA phylogenies of the clade. Mitochondrial genome data seem to be insufficient to resolve the position of the platyzoan clade within the Spiralia but can help to address internal phylogenetic questions. The present review includes a tabular survey of all published platyzoan mt genomes. PMID:23274056

  20. Platyzoan mitochondrial genomes.

    PubMed

    Wey-Fabrizius, Alexandra R; Podsiadlowski, Lars; Herlyn, Holger; Hankeln, Thomas

    2013-11-01

    Platyzoa is a putative lophotrochozoan (spiralian) subtaxon within the protostome clade of Metazoa, comprising a range of biologically diverse, mostly small worm-shaped animals. The monophyly of Platyzoa, the relationships between the putative subgroups Platyhelminthes, Gastrotricha and Gnathifera (the latter comprising at least Gnathostomulida, "Rotifera" and Acanthocephala) as well as some aspects of the internal phylogenies of these subgroups are highly debated. Here we review how complete mitochondrial (mt) genome data contribute to these debates. We highlight special features of the mt genomes and discuss problems in mtDNA phylogenies of the clade. Mitochondrial genome data seem to be insufficient to resolve the position of the platyzoan clade within the Spiralia but can help to address internal phylogenetic questions. The present review includes a tabular survey of all published platyzoan mt genomes.

  1. Pharmacological NAD-Boosting Strategies Improve Mitochondrial Homeostasis in Human Complex I-Mutant Fibroblasts.

    PubMed

    Felici, Roberta; Lapucci, Andrea; Cavone, Leonardo; Pratesi, Sara; Berlinguer-Palmini, Rolando; Chiarugi, Alberto

    2015-06-01

    Mitochondrial disorders are devastating genetic diseases for which efficacious therapies are still an unmet need. Recent studies report that increased availability of intracellular NAD obtained by inhibition of the NAD-consuming enzyme poly(ADP-ribose) polymerase (PARP)-1 or supplementation with the NAD-precursor nicotinamide riboside (NR) ameliorates energetic derangement and symptoms in mouse models of mitochondrial disorders. Whether these pharmacological approaches also improve bioenergetics of human cells harboring mitochondrial defects is unknown. It is also unclear whether the same signaling cascade is prompted by PARP-1 inhibitors and NR supplementation to improve mitochondrial homeostasis. Here, we show that human fibroblasts mutant for the NADH dehydrogenase (ubiquinone) Fe-S protein 1 (NDUFS1) subunit of respiratory complex I have similar ATP, NAD, and mitochondrial content compared with control cells, but show reduced mitochondrial membrane potential. Interestingly, mutant cells also show increased transcript levels of mitochondrial DNA but not nuclear DNA respiratory complex subunits, suggesting activation of a compensatory response. At variance with prior work in mice, however, NR supplementation, but not PARP-1 inhibition, increased intracellular NAD content in NDUFS1 mutant human fibroblasts. Conversely, PARP-1 inhibitors, but not NR supplementation, increased transcription of mitochondrial transcription factor A and mitochondrial DNA-encoded respiratory complexes constitutively induced in mutant cells. Still, both NR and PARP-1 inhibitors restored mitochondrial membrane potential and increased organelle content as well as oxidative activity of NDUFS1-deficient fibroblasts. Overall, data provide the first evidence that in human cells harboring a mitochondrial respiratory defect exposure to NR or PARP-1, inhibitors activate different signaling pathways that are not invariantly prompted by NAD increases, but equally able to improve energetic

  2. Endosymbionts and mitochondrial origins

    NASA Technical Reports Server (NTRS)

    Woese, C. R.

    1977-01-01

    The possibility is put forth that the mitochondrion did not originate from an endosymbiosis 1-2 billion years ago involving an aerobic bacterium. Rather, it arose by endosymbiosis in a much earlier anaerobic period and was initially a photosynthetic organelle analogous to the modern chloroplast. This suggestion arises from a reconsideration of the nature of endosymbiosis. It explains the remarkable diversity in mitochondrial information storage and processing systems.

  3. Temperature controlled high voltage regulator

    DOEpatents

    Chiaro, Jr., Peter J.; Schulze, Gerald K.

    2004-04-20

    A temperature controlled high voltage regulator for automatically adjusting the high voltage applied to a radiation detector is described. The regulator is a solid state device that is independent of the attached radiation detector, enabling the regulator to be used by various models of radiation detectors, such as gas flow proportional radiation detectors.

  4. Voltage sensor and dielectric material

    DOEpatents

    Yakymyshyn, Christopher Paul; Yakymyshyn, Pamela Jane; Brubaker, Michael Allen

    2006-10-17

    A voltage sensor is described that consists of an arrangement of impedance elements. The sensor is optimized to provide an output ratio that is substantially immune to changes in voltage, temperature variations or aging. Also disclosed is a material with a large and stable dielectric constant. The dielectric constant can be tailored to vary with position or direction in the material.

  5. Mitochondrial dysfunction is an important cause of neurological deficits in an inflammatory model of multiple sclerosis.

    PubMed

    Sadeghian, Mona; Mastrolia, Vincenzo; Rezaei Haddad, Ali; Mosley, Angelina; Mullali, Gizem; Schiza, Dimitra; Sajic, Marija; Hargreaves, Iain; Heales, Simon; Duchen, Michael R; Smith, Kenneth J

    2016-01-01

    Neuroinflammation can cause major neurological dysfunction, without demyelination, in both multiple sclerosis (MS) and a mouse model of the disease (experimental autoimmune encephalomyelitis; EAE), but the mechanisms remain obscure. Confocal in vivo imaging of the mouse EAE spinal cord reveals that impaired neurological function correlates with the depolarisation of both the axonal mitochondria and the axons themselves. Indeed, the depolarisation parallels the expression of neurological deficit at the onset of disease, and during relapse, improving during remission in conjunction with the deficit. Mitochondrial dysfunction, fragmentation and impaired trafficking were most severe in regions of extravasated perivascular inflammatory cells. The dysfunction at disease onset was accompanied by increased expression of the rate-limiting glycolytic enzyme phosphofructokinase-2 in activated astrocytes, and by selective reduction in spinal mitochondrial complex I activity. The metabolic changes preceded any demyelination or axonal degeneration. We conclude that mitochondrial dysfunction is a major cause of reversible neurological deficits in neuroinflammatory disease, such as MS. PMID:27624721

  6. Mitochondrial dysfunction is an important cause of neurological deficits in an inflammatory model of multiple sclerosis

    PubMed Central

    Sadeghian, Mona; Mastrolia, Vincenzo; Rezaei Haddad, Ali; Mosley, Angelina; Mullali, Gizem; Schiza, Dimitra; Sajic, Marija; Hargreaves, Iain; Heales, Simon; Duchen, Michael R.; Smith, Kenneth J.

    2016-01-01

    Neuroinflammation can cause major neurological dysfunction, without demyelination, in both multiple sclerosis (MS) and a mouse model of the disease (experimental autoimmune encephalomyelitis; EAE), but the mechanisms remain obscure. Confocal in vivo imaging of the mouse EAE spinal cord reveals that impaired neurological function correlates with the depolarisation of both the axonal mitochondria and the axons themselves. Indeed, the depolarisation parallels the expression of neurological deficit at the onset of disease, and during relapse, improving during remission in conjunction with the deficit. Mitochondrial dysfunction, fragmentation and impaired trafficking were most severe in regions of extravasated perivascular inflammatory cells. The dysfunction at disease onset was accompanied by increased expression of the rate-limiting glycolytic enzyme phosphofructokinase-2 in activated astrocytes, and by selective reduction in spinal mitochondrial complex I activity. The metabolic changes preceded any demyelination or axonal degeneration. We conclude that mitochondrial dysfunction is a major cause of reversible neurological deficits in neuroinflammatory disease, such as MS. PMID:27624721

  7. Human Mitochondrial DNA Replication

    PubMed Central

    Holt, Ian J.; Reyes, Aurelio

    2012-01-01

    Elucidation of the process of DNA replication in mitochondria is in its infancy. For many years, maintenance of the mitochondrial genome was regarded as greatly simplified compared to the nucleus. Mammalian mitochondria were reported to lack all DNA repair systems, to eschew DNA recombination, and to possess but a single DNA polymerase, polymerase γ. Polγ was said to replicate mitochondrial DNA exclusively via one mechanism, involving only two priming events and a handful of proteins. In this “strand-displacement model,” leading strand DNA synthesis begins at a specific site and advances approximately two-thirds of the way around the molecule before DNA synthesis is initiated on the “lagging” strand. Although the displaced strand was long-held to be coated with protein, RNA has more recently been proposed in its place. Furthermore, mitochondrial DNA molecules with all the features of products of conventional bidirectional replication have been documented, suggesting that the process and regulation of replication in mitochondria is complex, as befits a genome that is a core factor in human health and longevity. PMID:23143808

  8. Unique quadruple immunofluorescence assay demonstrates mitochondrial respiratory chain dysfunction in osteoblasts of aged and PolgA(-/-) mice.

    PubMed

    Dobson, Philip F; Rocha, Mariana C; Grady, John P; Chrysostomou, Alexia; Hipps, Daniel; Watson, Sharon; Greaves, Laura C; Deehan, David J; Turnbull, Doug M

    2016-01-01

    Fragility fractures caused by osteoporosis affect millions of people worldwide every year with significant levels of associated morbidity, mortality and costs to the healthcare economy. The pathogenesis of declining bone mineral density is poorly understood but it is inherently related to increasing age. Growing evidence in recent years, especially that provided by mouse models, suggest that accumulating somatic mitochondrial DNA mutations may cause the phenotypic changes associated with the ageing process including osteoporosis. Methods to study mitochondrial abnormalities in individual osteoblasts, osteoclasts and osteocytes are limited and impair our ability to assess the changes seen with age and in animal models of ageing. To enable the assessment of mitochondrial protein levels, we have developed a quadruple immunofluorescence method to accurately quantify the presence of mitochondrial respiratory chain components within individual bone cells. We have applied this technique to a well-established mouse model of ageing and osteoporosis and show respiratory chain deficiency. PMID:27553587

  9. Unique quadruple immunofluorescence assay demonstrates mitochondrial respiratory chain dysfunction in osteoblasts of aged and PolgA−/− mice

    PubMed Central

    Dobson, Philip F.; Rocha, Mariana C.; Grady, John P.; Chrysostomou, Alexia; Hipps, Daniel; Watson, Sharon; Greaves, Laura C.; Deehan, David J.; Turnbull, Doug M.

    2016-01-01

    Fragility fractures caused by osteoporosis affect millions of people worldwide every year with significant levels of associated morbidity, mortality and costs to the healthcare economy. The pathogenesis of declining bone mineral density is poorly understood but it is inherently related to increasing age. Growing evidence in recent years, especially that provided by mouse models, suggest that accumulating somatic mitochondrial DNA mutations may cause the phenotypic changes associated with the ageing process including osteoporosis. Methods to study mitochondrial abnormalities in individual osteoblasts, osteoclasts and osteocytes are limited and impair our ability to assess the changes seen with age and in animal models of ageing. To enable the assessment of mitochondrial protein levels, we have developed a quadruple immunofluorescence method to accurately quantify the presence of mitochondrial respiratory chain components within individual bone cells. We have applied this technique to a well-established mouse model of ageing and osteoporosis and show respiratory chain deficiency. PMID:27553587

  10. Transient voltage oscillations in coils

    SciTech Connect

    Chowdhuri, P.

    1985-01-01

    Magnet coils may be excited into internal voltage oscillations by transient voltages. Such oscillations may electrically stress the magnet's dielectric components to many times its normal stress. This may precipitate a dielectric failure, and the attendant prolonged loss of service and costly repair work. Therefore, it is important to know the natural frequencies of oscillations of a magnet during the design stage, and to determine whether the expected switching transient voltages can excite the magnet into high-voltage internal oscillations. The series capacitance of a winding significantly affects its natural frequencies. However, the series capacitance is difficult to calculate, because it may comprise complex capacitance network, consisting of intra- and inter-coil turn-to-turn capacitances of the coil sections. A method of calculating the series capacitance of a winding is proposed. This method is rigorous but simple to execute. The time-varying transient voltages along the winding are also calculated.

  11. Quantitative mitochondrial redox imaging of breast cancer metastatic potential

    NASA Astrophysics Data System (ADS)

    Xu, He N.; Nioka, Shoko; Glickson, Jerry D.; Chance, Britton; Li, Lin Z.

    2010-05-01

    Predicting tumor metastatic potential remains a challenge in cancer research and clinical practice. Our goal was to identify novel biomarkers for differentiating human breast tumors with different metastatic potentials by imaging the in vivo mitochondrial redox states of tumor tissues. The more metastatic (aggressive) MDA-MB-231 and less metastatic (indolent) MCF-7 human breast cancer mouse xenografts were imaged with the low-temperature redox scanner to obtain multi-slice fluorescence images of reduced nicotinamide adenine dinucleotide (NADH) and oxidized flavoproteins (Fp). The nominal concentrations of NADH and Fp in tissue were measured using reference standards and used to calculate the Fp redox ratio, Fp/(NADH+Fp). We observed significant core-rim differences, with the core being more oxidized than the rim in all aggressive tumors but not in the indolent tumors. These results are consistent with our previous observations on human melanoma mouse xenografts, indicating that mitochondrial redox imaging potentially provides sensitive markers for distinguishing aggressive from indolent breast tumor xenografts. Mitochondrial redox imaging can be clinically implemented utilizing cryogenic biopsy specimens and is useful for drug development and for clinical diagnosis of breast cancer.

  12. Mitochondrial protein adducts formation and mitochondrial dysfunction during N-acetyl-m-aminophenol (AMAP)-induced hepatotoxicity in primary human hepatocytes.

    PubMed

    Xie, Yuchao; McGill, Mitchell R; Du, Kuo; Dorko, Kenneth; Kumer, Sean C; Schmitt, Timothy M; Ding, Wen-Xing; Jaeschke, Hartmut

    2015-12-01

    3'-Hydroxyacetanilide orN-acetyl-meta-aminophenol (AMAP) is generally regarded as a non-hepatotoxic analog of acetaminophen (APAP). Previous studies demonstrated the absence of toxicity after AMAP in mice, hamsters, primary mouse hepatocytes and several cell lines. In contrast, experiments with liver slices suggested that it may be toxic to human hepatocytes; however, the mechanism of toxicity is unclear. To explore this,we treated primary human hepatocytes (PHH) with AMAP or APAP for up to 48 h and measured several parameters to assess metabolism and injury. Although less toxic than APAP, AMAP dose-dependently triggered cell death in PHH as indicated by alanine aminotransferase (ALT) release and propidium iodide (PI) staining. Similar to APAP, AMAP also significantly depleted glutathione (GSH) in PHH and caused mitochondrial damage as indicated by glutamate dehydrogenase (GDH) release and the JC-1 assay. However, unlike APAP, AMAP treatment did not cause relevant c-jun-N-terminal kinase (JNK) activation in the cytosol or phospho-JNK translocation to mitochondria. To compare, AMAP toxicity was assessed in primary mouse hepatocytes (PMH). No cytotoxicity was observed as indicated by the lack of lactate dehydrogenase release and no PI staining. Furthermore, there was no GSH depletion or mitochondrial dysfunction after AMAP treatment in PMH. Immunoblotting for arylated proteins suggested that AMAP treatment caused extensive mitochondrial protein adduct formation in PHH but not in PMH. In conclusion, AMAP is hepatotoxic in PHH and the mechanism involves the formation of mitochondrial protein adducts and mitochondrial dysfunction. PMID:26431796

  13. Mitochondrial DNA damage and atherosclerosis.

    PubMed

    Yu, Emma P K; Bennett, Martin R

    2014-09-01

    Mitochondria are often regarded as the cellular powerhouses through their ability to generate ATP, the universal fuel for metabolic processes. However, in recent years mitochondria have been recognised as critical regulators of cell death, inflammation, metabolism, and the generation of reactive oxygen species (ROS). Thus, mitochondrial dysfunction directly promotes cell death, inflammation, and oxidative stress and alters metabolism. These are key processes in atherosclerosis and there is now evidence that mitochondrial DNA (mtDNA) damage leads to mitochondrial dysfunction and promotes atherosclerosis directly. In this review we discuss the recent evidence for and mechanisms linking mtDNA defects and atherosclerosis and suggest areas of mitochondrial biology that are potential therapeutic targets.

  14. Evolution meets disease: penetrance and functional epistasis of mitochondrial tRNA mutations.

    PubMed

    Moreno-Loshuertos, Raquel; Ferrín, Gustavo; Acín-Pérez, Rebeca; Gallardo, M Esther; Viscomi, Carlo; Pérez-Martos, Acisclo; Zeviani, Massimo; Fernández-Silva, Patricio; Enríquez, José Antonio

    2011-04-01

    About half of the mitochondrial DNA (mtDNA) mutations causing diseases in humans occur in tRNA genes. Particularly intriguing are those pathogenic tRNA mutations than can reach homoplasmy and yet show very different penetrance among patients. These mutations are scarce and, in addition to their obvious interest for understanding human pathology, they can be excellent experimental examples to model evolution and fixation of mitochondrial tRNA mutations. To date, the only source of this type of mutations is human patients. We report here the generation and characterization of the first mitochondrial tRNA pathological mutation in mouse cells, an m.3739G>A transition in the mitochondrial mt-Ti gene. This mutation recapitulates the molecular hallmarks of a disease-causing mutation described in humans, an m.4290T>C transition affecting also the human mt-Ti gene. We could determine that the pathogenic molecular mechanism, induced by both the mouse and the human mutations, is a high frequency of abnormal folding of the tRNA(Ile) that cannot be charged with isoleucine. We demonstrate that the cells harboring the mouse or human mutant tRNA have exacerbated mitochondrial biogenesis triggered by an increase in mitochondrial ROS production as a compensatory response. We propose that both the nature of the pathogenic mechanism combined with the existence of a compensatory mechanism can explain the penetrance pattern of this mutation. This particular behavior can allow a scenario for the evolution of mitochondrial tRNAs in which the fixation of two alleles that are individually deleterious can proceed in two steps and not require the simultaneous mutation of both. PMID:21533077

  15. Improved Programmable High-Voltage Power Supply

    NASA Technical Reports Server (NTRS)

    Castell, Karen; Rutberg, Arthur

    1994-01-01

    Improved dc-to-dc converter functions as programmable high-voltage power supply with low-power-dissipation voltage regulator on high-voltage side. Design of power supply overcomes deficiencies of older designs. Voltage regulation with low power dissipation provided on high-voltage side.

  16. Voltage Dependence of Conformational Dynamics and Subconducting States of VDAC-1.

    PubMed

    Briones, Rodolfo; Weichbrodt, Conrad; Paltrinieri, Licia; Mey, Ingo; Villinger, Saskia; Giller, Karin; Lange, Adam; Zweckstetter, Markus; Griesinger, Christian; Becker, Stefan; Steinem, Claudia; de Groot, Bert L

    2016-09-20

    The voltage-dependent anion channel 1 (VDAC-1) is an important protein of the outer mitochondrial membrane that transports energy metabolites and is involved in apoptosis. The available structures of VDAC proteins show a wide β-stranded barrel pore, with its N-terminal α-helix (N-α) bound to its interior. Electrophysiology experiments revealed that voltage, its polarity, and membrane composition modulate VDAC currents. Experiments with VDAC-1 mutants identified amino acids that regulate the gating process. However, the mechanisms for how these factors regulate VDAC-1, and which changes they trigger in the channel, are still unknown. In this study, molecular dynamics simulations and single-channel experiments of VDAC-1 show agreement for the current-voltage relationships of an "open" channel and they also show several subconducting transient states that are more cation selective in the simulations. We observed voltage-dependent asymmetric distortions of the VDAC-1 barrel and the displacement of particular charged amino acids. We constructed conformational models of the protein voltage response and the pore changes that consistently explain the protein conformations observed at opposite voltage polarities, either in phosphatidylethanolamine or phosphatidylcholine membranes. The submicrosecond VDAC-1 voltage response shows intrinsic structural changes that explain the role of key gating amino acids and support some of the current gating hypotheses. These voltage-dependent protein changes include asymmetric barrel distortion, its interaction with the membrane, and significant displacement of N-α amino acids. PMID:27653481

  17. Mitochondrial Ca(2+) mobilization is a key element in olfactory signaling.

    PubMed

    Fluegge, Daniela; Moeller, Lisa M; Cichy, Annika; Gorin, Monika; Weth, Agnes; Veitinger, Sophie; Cainarca, Silvia; Lohmer, Stefan; Corazza, Sabrina; Neuhaus, Eva M; Baumgartner, Werner; Spehr, Jennifer; Spehr, Marc

    2012-05-01

    In olfactory sensory neurons (OSNs), cytosolic Ca(2+) controls the gain and sensitivity of olfactory signaling. Important components of the molecular machinery that orchestrates OSN Ca(2+) dynamics have been described, but key details are still missing. Here, we demonstrate a critical physiological role of mitochondrial Ca(2+) mobilization in mouse OSNs. Combining a new mitochondrial Ca(2+) imaging approach with patch-clamp recordings, organelle mobility assays and ultrastructural analyses, our study identifies mitochondria as key determinants of olfactory signaling. We show that mitochondrial Ca(2+) mobilization during sensory stimulation shapes the cytosolic Ca(2+) response profile in OSNs, ensures a broad dynamic response range and maintains sensitivity of the spike generation machinery. When mitochondrial function is impaired, olfactory neurons function as simple stimulus detectors rather than as intensity encoders. Moreover, we describe activity-dependent recruitment of mitochondria to olfactory knobs, a mechanism that provides a context-dependent tool for OSNs to maintain cellular homeostasis and signaling integrity. PMID:22446879

  18. Low voltage nonprimary explosive detonator

    DOEpatents

    Dinegar, Robert H.; Kirkham, John

    1982-01-01

    A low voltage, electrically actuated, nonprimary explosive detonator is disclosed wherein said detonation is achieved by means of an explosive train in which a deflagration-to-detonation transition is made to occur. The explosive train is confined within a cylindrical body and positioned adjacent to low voltage ignition means have electrical leads extending outwardly from the cylindrical confining body. Application of a low voltage current to the electrical leads ignites a self-sustained deflagration in a donor portion of the explosive train which then is made to undergo a transition to detonation further down the train.

  19. Voltage Sensors Monitor Harmful Static

    NASA Technical Reports Server (NTRS)

    2009-01-01

    A tiny sensor, small enough to be worn on clothing, now monitors voltage changes near sensitive instruments after being created to alert Agency workers to dangerous static buildup near fuel operations and avionics. San Diego s Quasar Federal Systems received a Small Business Innovation Research (SBIR) contract from Kennedy Space Center to develop its remote voltage sensor (RVS), a dime-sized electrometer designed to measure triboelectric changes in the environment. One of the unique qualities of the RVS is that it can detect static at greater distances than previous devices, measuring voltage changes from a few centimeters to a few meters away, due to its much-improved sensitivity.

  20. Mitochondrial toxicity and HIV therapy

    PubMed Central

    White, A.

    2001-01-01

    Nucleoside reverse transcriptase inhibitors (NRTIs) remain the cornerstone of highly active antiretroviral therapy (HAART) combination regimens. However, it has been known for some time that these agents have the potential to cause varied side effects, many of which are thought to be due to their effects on mitochondria. Mitochondria, the key energy generating organelles in the cell, are unique in having their own DNA, a double stranded circular genome of about 16 000 bases. There is a separate enzyme present inside the cell that replicates mitochondrial DNA, polymerase gamma. NRTIs can affect the function of this enzyme and this may lead to depletion of mitochondrial DNA or qualitative changes. The study of inherited mitochondrial diseases has led to further understanding of the consequences of mutations or depletion in mitochondrial DNA. Key among these is the realisation that there may be substantial heteroplasmy among mitochondria within a given cell, and among cells in a particular tissue. The unpredictable nature of mitochondrial segregation during cellular replication makes it difficult to predict the likelihood of dysfunction in a given tissue. In addition, there is a threshold effect for the expression of mitochondrial dysfunction, both at the mitochondrial and cellular level. Various clinical and in vitro studies have suggested that NRTIs are associated with mitochondrial dysfunction in different tissues, although the weight of evidence is limited in many cases. The heterogeneity in the tissues affected by the different drugs raises interesting questions, and possible explanations include differential distribution or activation of these agents. This article reviews the major recognised toxicities associated with NRTI therapy and evidence for mitochondrial dysfunction in these complications. Data were identified through searching of online databases including Medline and Current Contents for relevant articles, along with abstracts and posters from recent

  1. NOTCH reprograms mitochondrial metabolism for proinflammatory macrophage activation

    PubMed Central

    Xu, Jun; Chi, Feng; Guo, Tongsheng; Punj, Vasu; Lee, W.N. Paul; French, Samuel W.; Tsukamoto, Hidekazu

    2015-01-01

    Metabolic reprogramming is implicated in macrophage activation, but the underlying mechanisms are poorly understood. Here, we demonstrate that the NOTCH1 pathway dictates activation of M1 phenotypes in isolated mouse hepatic macrophages (HMacs) and in a murine macrophage cell line by coupling transcriptional upregulation of M1 genes with metabolic upregulation of mitochondrial oxidative phosphorylation and ROS (mtROS) to augment induction of M1 genes. Enhanced mitochondrial glucose oxidation was achieved by increased recruitment of the NOTCH1 intracellular domain (NICD1) to nuclear and mitochondrial genes that encode respiratory chain components and by NOTCH-dependent induction of pyruvate dehydrogenase phosphatase 1 (Pdp1) expression, pyruvate dehydrogenase activity, and glucose flux to the TCA cycle. As such, inhibition of the NOTCH pathway or Pdp1 knockdown abrogated glucose oxidation, mtROS, and M1 gene expression. Conditional NOTCH1 deficiency in the myeloid lineage attenuated HMac M1 activation and inflammation in a murine model of alcoholic steatohepatitis and markedly reduced lethality following endotoxin-mediated fulminant hepatitis in mice. In vivo monocyte tracking further demonstrated the requirement of NOTCH1 for the migration of blood monocytes into the liver and subsequent M1 differentiation. Together, these results reveal that NOTCH1 promotes reprogramming of mitochondrial metabolism for M1 macrophage activation. PMID:25798621

  2. MCU encodes the pore conducting mitochondrial calcium currents.

    PubMed

    Chaudhuri, Dipayan; Sancak, Yasemin; Mootha, Vamsi K; Clapham, David E

    2013-06-04

    Mitochondrial calcium (Ca(2+)) import is a well-described phenomenon regulating cell survival and ATP production. Of multiple pathways allowing such entry, the mitochondrial Ca(2+) uniporter is a highly Ca(2+)-selective channel complex encoded by several recently-discovered genes. However, the identity of the pore-forming subunit remains to be established, since knockdown of all the candidate uniporter genes inhibit Ca(2+) uptake in imaging assays, and reconstitution experiments have been equivocal. To definitively identify the channel, we use whole-mitoplast voltage-clamping, the technique that originally established the uniporter as a Ca(2+) channel. We show that RNAi-mediated knockdown of the mitochondrial calcium uniporter (MCU) gene reduces mitochondrial Ca(2+) current (I MiCa ), whereas overexpression increases it. Additionally, a classic feature of I MiCa , its sensitivity to ruthenium red inhibition, can be abolished by a point mutation in the putative pore domain without altering current magnitude. These analyses establish that MCU encodes the pore-forming subunit of the uniporter channel. DOI:http://dx.doi.org/10.7554/eLife.00704.001.

  3. Sealing the mitochondrial respirasome.

    PubMed

    Winge, Dennis R

    2012-07-01

    The mitochondrial respiratory chain is organized within an array of supercomplexes that function to minimize the generation of reactive oxygen species (ROS) during electron transfer reactions. Structural models of supercomplexes are now known. Another recent advance is the discovery of non-OXPHOS complex proteins that appear to adhere to and seal the individual respiratory complexes to form stable assemblages that prevent electron leakage. This review highlights recent advances in our understanding of the structures of supercomplexes and the factors that mediate their stability.

  4. Mitochondrial form and function

    PubMed Central

    Friedman, Jonathan R.; Nunnari, Jodi

    2014-01-01

    Mitochondria are one of the major ancient endomembrane systems in eukaryotic cells. Owing to their ability to produce ATP through respiration, they became a driving force in evolution. As an essential step in the process of eukaryotic evolution, the size of the mitochondrial chromosome was drastically reduced, and the behaviour of mitochondria within eukaryotic cells radically changed. Recent advances have revealed how the organelle’s behaviour has evolved to allow the accurate transmission of its genome and to become responsive to the needs of the cell and its own dysfunction. PMID:24429632

  5. Mitochondrial Genome Instability and ROS Enhance Intestinal Tumorigenesis in APCMin/+ Mice

    PubMed Central

    Woo, Dong Kyun; Green, Paula D.; Santos, Janine H.; D'Souza, Anthony D.; Walther, Zenta; Martin, W. David; Christian, Brooke E.; Chandel, Navdeep S.; Shadel, Gerald S.

    2012-01-01

    Alterations in mitochondrial oxidative phosphorylation have long been documented in tumors. Other types of mitochondrial dysfunction, including altered reactive oxygen species (ROS) production and apoptosis, also can contribute to tumorigenesis and cancer phenotypes. Furthermore, mutation and altered amounts of mitochondrial DNA (mtDNA) have been observed in cancer cells. However, how mtDNA instability per se contributes to cancer remains largely undetermined. Mitochondrial transcription factor A (TFAM) is required for expression and maintenance of mtDNA. Tfam heterozygous knock-out (Tfam+/−) mice show mild mtDNA depletion, but have no overt phenotypes. We show that Tfam+/− mouse cells and tissues not only possess less mtDNA but also increased oxidative mtDNA damage. Crossing Tfam+/− mice to the adenomatous polyposis coli multiple intestinal neoplasia (APCMin/+) mouse cancer model revealed that mtDNA instability increases tumor number and growth in the small intestine. This was not a result of enhancement of Wnt/β-catenin signaling, but rather appears to involve a propensity for increased mitochondrial ROS production. Direct involvement of mitochondrial ROS in intestinal tumorigenesis was shown by crossing APCMin/+ mice to those that have catalase targeted to mitochondria, which resulted in a significant reduction in tumorigenesis in the colon. Thus, mitochondrial genome instability and ROS enhance intestinal tumorigenesis and Tfam+/− mice are a relevant model to address the role of mtDNA instability in disease states in which mitochondrial dysfunction is implicated, such as cancer, neurodegeneration, and aging. PMID:22056359

  6. Neuronal and astrocyte dysfunction diverges from embryonic fibroblasts in the Ndufs4fky/fky mouse

    PubMed Central

    Bird, Matthew J.; Wijeyeratne, Xiaonan W.; Komen, Jasper C.; Laskowski, Adrienne; Ryan, Michael T.; Thorburn, David R.; Frazier, Ann E.

    2014-01-01

    Mitochondrial dysfunction causes a range of early-onset neurological diseases and contributes to neurodegenerative conditions. The mechanisms of neurological damage however are poorly understood, as accessing relevant tissue from patients is difficult, and appropriate models are limited. Hence, we assessed mitochondrial function in neurologically relevant primary cell lines from a CI (complex I) deficient Ndufs4 KO (knockout) mouse (Ndufs4fky/fky) modelling aspects of the mitochondrial disease LS (Leigh syndrome), as well as MEFs (mouse embryonic fibroblasts). Although CI structure and function were compromised in all Ndufs4fky/fky cell types, the mitochondrial membrane potential was selectively impaired in the MEFs, correlating with decreased CI-dependent ATP synthesis. In addition, increased ROS (reactive oxygen species) generation and altered sensitivity to cell death were only observed in Ndufs4fky/fky primary MEFs. In contrast, Ndufs4fky/fky primary isocortical neurons and primary isocortical astrocytes displayed only impaired ATP generation without mitochondrial membrane potential changes. Therefore the neurological dysfunction in the Ndufs4fky/fky mouse may partly originate from a more severe ATP depletion in neurons and astrocytes, even at the expense of maintaining the mitochondrial membrane potential. This may provide protection from cell death, but would ultimately compromise cell functionality in neurons and astrocytes. Furthermore, RET (reverse electron transfer) from complex II to CI appears more prominent in neurons than MEFs or astrocytes, and is attenuated in Ndufs4fky/fky cells. PMID:25312000

  7. Genetically Engineered Fluorescent Voltage Reporters

    PubMed Central

    2012-01-01

    Fluorescent membrane voltage indicators that enable optical imaging of neuronal circuit operations in the living mammalian brain are powerful tools for biology and particularly neuroscience. Classical voltage-sensitive dyes, typically low molecular-weight organic compounds, have been in widespread use for decades but are limited by issues related to optical noise, the lack of generally applicable procedures that enable staining of specific cell populations, and difficulties in performing imaging experiments over days and weeks. Genetically encoded voltage indicators (GEVIs) represent a newer alternative that overcomes several of the limitations inherent to classical voltage-sensitive dyes. We critically review the fundamental concepts of this approach, the variety of available probes and their state of development. PMID:22896802

  8. High voltage solar array experiments

    NASA Technical Reports Server (NTRS)

    Kennerud, K. L.

    1974-01-01

    The interaction between the components of a high voltage solar array and a simulated space plasma is studied to obtain data for the design of a high voltage solar array capable of 15kW at 2 to 16kV. Testing was conducted in a vacuum chamber 1.5-m long by 1.5-m diameter having a plasma source which simulated the plasma conditions existing in earth orbit between 400 nautical miles and synchronous altitude. Test samples included solar array segments pinholes in insulation covering high voltage electrodes, and plain dielectric samples. Quantitative data are presented in the areas of plasma power losses, plasma and high voltage induced damage, and dielectric properties. Limitations of the investigation are described.

  9. Frontal cortical mitochondrial dysfunction and mitochondria-related β-amyloid accumulation by chronic sleep restriction in mice

    PubMed Central

    Zhao, Hongyi; He, Jialin; Zhuang, Jianhua; Liu, Zhenyu; Yang, Yang; Huang, Liuqing; Zhao, Zhongxin

    2016-01-01

    Mitochondrial dysfunction induced by mitochondria-related β-amyloid (Aβ) accumulation is increasingly being considered a novel risk factor for sporadic Alzheimer’s disease pathophysiology. The close relationship between chronic sleep restriction (CSR) and cortical Aβ elevation was confirmed recently. By assessing frontal cortical mitochondrial function (electron microscopy manifestation, cytochrome C oxidase concentration, ATP level, and mitochondrial membrane potential) and the levels of mitochondria-related Aβ in 9-month-old adult male C57BL/6J mice subjected to CSR and as an environmental control (CO) group, we aimed to evaluate the association of CSR with mitochondrial dysfunction and mitochondria-related Aβ accumulation. In this study, frontal cortical mitochondrial dysfunction was significantly more severe in CSR mice compared with CO animals. Furthermore, CSR mice showed higher mitochondria-associated Aβ, total Aβ, and mitochondria-related β-amyloid protein precursor (AβPP) levels compared with CO mice. In the CSR model, mouse frontal cortical mitochondrial dysfunction was correlated with mitochondria-associated Aβ and mitochondria-related AβPP levels. However, frontal cortical mitochondria-associated Aβ levels showed no significant association with cortical total Aβ and mitochondrial AβPP concentrations. These findings indicated that CSR-induced frontal cortical mitochondrial dysfunction and mitochondria-related Aβ accumulation, which was closely related to mitochondrial dysfunction under CSR. PMID:27341212

  10. Frontal cortical mitochondrial dysfunction and mitochondria-related β-amyloid accumulation by chronic sleep restriction in mice.

    PubMed

    Zhao, Hongyi; Wu, Huijuan; He, Jialin; Zhuang, Jianhua; Liu, Zhenyu; Yang, Yang; Huang, Liuqing; Zhao, Zhongxin

    2016-08-17

    Mitochondrial dysfunction induced by mitochondria-related β-amyloid (Aβ) accumulation is increasingly being considered a novel risk factor for sporadic Alzheimer's disease pathophysiology. The close relationship between chronic sleep restriction (CSR) and cortical Aβ elevation was confirmed recently. By assessing frontal cortical mitochondrial function (electron microscopy manifestation, cytochrome C oxidase concentration, ATP level, and mitochondrial membrane potential) and the levels of mitochondria-related Aβ in 9-month-old adult male C57BL/6J mice subjected to CSR and as an environmental control (CO) group, we aimed to evaluate the association of CSR with mitochondrial dysfunction and mitochondria-related Aβ accumulation. In this study, frontal cortical mitochondrial dysfunction was significantly more severe in CSR mice compared with CO animals. Furthermore, CSR mice showed higher mitochondria-associated Aβ, total Aβ, and mitochondria-related β-amyloid protein precursor (AβPP) levels compared with CO mice. In the CSR model, mouse frontal cortical mitochondrial dysfunction was correlated with mitochondria-associated Aβ and mitochondria-related AβPP levels. However, frontal cortical mitochondria-associated Aβ levels showed no significant association with cortical total Aβ and mitochondrial AβPP concentrations. These findings indicated that CSR-induced frontal cortical mitochondrial dysfunction and mitochondria-related Aβ accumulation, which was closely related to mitochondrial dysfunction under CSR.

  11. Synaptic dysfunction, memory deficits and hippocampal atrophy due to ablation of mitochondrial fission in adult forebrain neurons

    PubMed Central

    Oettinghaus, B; Schulz, J M; Restelli, L M; Licci, M; Savoia, C; Schmidt, A; Schmitt, K; Grimm, A; Morè, L; Hench, J; Tolnay, M; Eckert, A; D'Adamo, P; Franken, P; Ishihara, N; Mihara, K; Bischofberger, J; Scorrano, L; Frank, S

    2016-01-01

    Well-balanced mitochondrial fission and fusion processes are essential for nervous system development. Loss of function of the main mitochondrial fission mediator, dynamin-related protein 1 (Drp1), is lethal early during embryonic development or around birth, but the role of mitochondrial fission in adult neurons remains unclear. Here we show that inducible Drp1 ablation in neurons of the adult mouse forebrain results in progressive, neuronal subtype-specific alterations of mitochondrial morphology in the hippocampus that are marginally responsive to antioxidant treatment. Furthermore, DRP1 loss affects synaptic transmission and memory function. Although these changes culminate in hippocampal atrophy, they are not sufficient to cause neuronal cell death within 10 weeks of genetic Drp1 ablation. Collectively, our in vivo observations clarify the role of mitochondrial fission in neurons, demonstrating that Drp1 ablation in adult forebrain neurons compromises critical neuronal functions without causing overt neurodegeneration. PMID:25909888

  12. Methods for Efficient Elimination of Mitochondrial DNA from Cultured Cells

    PubMed Central

    Spadafora, Domenico; Kozhukhar, Nataliya; Chouljenko, Vladimir N.; Kousoulas, Konstantin G.; Alexeyev, Mikhail F.

    2016-01-01

    Here, we document that persistent mitochondria DNA (mtDNA) damage due to mitochondrial overexpression of the Y147A mutant uracil-N-glycosylase as well as mitochondrial overexpression of bacterial Exonuclease III or Herpes Simplex Virus protein UL12.5M185 can induce a complete loss of mtDNA (ρ0 phenotype) without compromising the viability of cells cultured in media supplemented with uridine and pyruvate. Furthermore, we use these observations to develop rapid, sequence-independent methods for the elimination of mtDNA, and demonstrate utility of these methods for generating ρ0 cells of human, mouse and rat origin. We also demonstrate that ρ0 cells generated by each of these three methods can serve as recipients of mtDNA in fusions with enucleated cells. PMID:27136098

  13. Molecular Genetics of Mitochondrial Disorders

    ERIC Educational Resources Information Center

    Wong, Lee-Jun C.

    2010-01-01

    Mitochondrial respiratory chain (RC) disorders (RCDs) are a group of genetically and clinically heterogeneous diseases because of the fact that protein components of the RC are encoded by both mitochondrial and nuclear genomes and are essential in all cells. In addition, the biogenesis, structure, and function of mitochondria, including DNA…

  14. CCN6 regulates mitochondrial function.

    PubMed

    Patra, Milan; Mahata, Sushil K; Padhan, Deepesh K; Sen, Malini

    2016-07-15

    Despite established links of CCN6, or Wnt induced signaling protein-3 (WISP3), with progressive pseudo rheumatoid dysplasia, functional characterization of CCN6 remains incomplete. In light of the documented negative correlation between accumulation of reactive oxygen species (ROS) and CCN6 expression, we investigated whether CCN6 regulates ROS accumulation through its influence on mitochondrial function. We found that CCN6 localizes to mitochondria, and depletion of CCN6 in the chondrocyte cell line C-28/I2 by using siRNA results in altered mitochondrial electron transport and respiration. Enhanced electron transport chain (ETC) activity of CCN6-depleted cells was reflected by increased mitochondrial ROS levels in association with augmented mitochondrial ATP synthesis, mitochondrial membrane potential and Ca(2+) Additionally, CCN6-depleted cells display ROS-dependent PGC1α (also known as PPARGC1A) induction, which correlates with increased mitochondrial mass and volume density, together with altered mitochondrial morphology. Interestingly, transcription factor Nrf2 (also known as NFE2L2) repressed CCN6 expression. Taken together, our results suggest that CCN6 acts as a molecular brake, which is appropriately balanced by Nrf2, in regulating mitochondrial function. PMID:27252383

  15. Mitochondrial genomes as living 'fossils'.

    PubMed

    Small, Ian

    2013-04-15

    The huge variation between mitochondrial genomes makes untangling their evolutionary histories difficult. Richardson et al. report on the remarkably unaltered 'fossil' genome of the tulip tree, giving us many clues as to how the mitochondrial genomes of flowering plants have evolved over the last 150 million years, and raising questions about how such extraordinary sequence conservation can be maintained.

  16. Exercise-Induced Neuroprotection of Hippocampus in APP/PS1 Transgenic Mice via Upregulation of Mitochondrial 8-Oxoguanine DNA Glycosylase

    PubMed Central

    Kang, Weimin; Jiang, Ning; Wang, Xun; Zhang, Yong; Ji, Li Li

    2014-01-01

    Improving mitochondrial function has been proposed as a reasonable therapeutic strategy to reduce amyloid-β (Aβ) load and to modify the progression of Alzheimer's disease (AD). However, the relationship between mitochondrial adaptation and brain neuroprotection caused by physical exercise in AD is poorly understood. This study was undertaken to investigate the effects of long-term treadmill exercise on mitochondrial 8-oxoguanine DNA glycosylase-1 (OGG1) level, mtDNA oxidative damage, and mitochondrial function in the hippocampus of APP/PS1 transgenic mouse model of AD. In the present study, twenty weeks of treadmill training significantly improved the cognitive function and reduced the expression of Aβ-42 in APP/PS1 transgenic (Tg) mice. Training also ameliorated mitochondrial respiratory function by increasing the complexes I, and IV and ATP synthase activities, whereas it attenuated ROS generation and mtDNA oxidative damage in Tg mice. Furthermore, the impaired mitochondrial antioxidant enzymes and mitochondrial OGG1 activities seen in Tg mice were restored with training. Acetylation level of mitochondrial OGG1 and MnSOD was markedly suppressed in Tg mice after exercise training, in parallel with increased level of SIRT3. These findings suggest that exercise training could increase mtDNA repair capacity in the mouse hippocampus, which in turn would result in protection against AD-related mitochondrial dysfunction and phenotypic deterioration. PMID:25538817

  17. Low-Voltage Bypass Device

    NASA Technical Reports Server (NTRS)

    Wilson, J. P.

    1994-01-01

    Improved bypass device provides low-resistance current shunt around low-voltage power cell when cell fails in open-circuit condition during operation. In comparison with older bypass devices for same application, this one weighs less, generates less heat, and has lower voltage drop (less resistance). Bypass device connected in parallel with power cell. Draws very little current during normal operation of cell.

  18. Switched-Capacitor Voltage Multiplier

    NASA Technical Reports Server (NTRS)

    Sridharan, Govind

    1991-01-01

    Dc-to-dc power converter multiplies input supply potential by factor of nearly 40. Design does not make use of transformers or inductors but effects voltage boost-up by capacitive energy transfer. Circuit primarily made up of banks of capacitors, connected by network of integrated-circuit relays. Converter functionally linear voltage amplifier with fixed gain figure. Bipolar in operation. Output fully floating, and excellent dc isolation between input and output terminals.

  19. High voltage power transistor development

    NASA Technical Reports Server (NTRS)

    Hower, P. L.

    1981-01-01

    Design considerations, fabrication procedures, and methods of evaluation for high-voltage power-transistor development are discussed. Technique improvements such as controlling the electric field at the surface and perserving lifetimes in the collector region which have advanced the state of the art in high-voltage transistors are discussed. These improvements can be applied directly to the development of 1200 volt, 200 ampere transistors.

  20. Depletion of PINK1 affects mitochondrial metabolism, calcium homeostasis and energy maintenance.

    PubMed

    Heeman, Bavo; Van den Haute, Chris; Aelvoet, Sarah-Ann; Valsecchi, Federica; Rodenburg, Richard J; Reumers, Veerle; Debyser, Zeger; Callewaert, Geert; Koopman, Werner J H; Willems, Peter H G M; Baekelandt, Veerle

    2011-04-01

    Loss-of-function mutations in the gene encoding the mitochondrial PTEN-induced putative kinase 1 (PINK1) are a major cause of early-onset familial Parkinson's disease (PD). Recent studies have highlighted an important function for PINK1 in clearing depolarized mitochondria by mitophagy. However, the role of PINK1 in mitochondrial and cellular functioning in physiological conditions is still incompletely understood. Here, we investigate mitochondrial and cellular calcium (Ca(2+)) homeostasis in PINK1-knockdown and PINK1-knockout mouse cells, both in basal metabolic conditions and after physiological stimulation, using unbiased automated live single-cell imaging in combination with organelle-specific fluorescent probes. Our data reveal that depletion of PINK1 induces moderate fragmentation of the mitochondrial network, mitochondrial membrane depolarization and increased production of reactive oxygen species. This results in reduced uptake of Ca(2+) by mitochondria after physiological stimulation. As a consequence, cells with knockdown or knockout of PINK1 display impaired mitochondrial ATP synthesis, which is exacerbated under conditions of increased ATP demand, thereby affecting cytosolic Ca(2+) extrusion. The impairment in energy maintenance was confirmed in the brain of PINK1-knockout mice by in vivo bioluminescence imaging. Our findings demonstrate a key role for PINK1 in the regulation of mitochondrial homeostasis and energy metabolism under physiological conditions. PMID:21385841

  1. PGC-1α regulation of mitochondrial degeneration in experimental diabetic neuropathy.

    PubMed

    Choi, Joungil; Chandrasekaran, Krish; Inoue, Tatsuya; Muragundla, Anjaneyulu; Russell, James W

    2014-04-01

    Mitochondrial degeneration is considered to play an important role in the development of diabetic peripheral neuropathy in humans. Mitochondrial degeneration and the corresponding protein regulation associated with the degeneration were studied in an animal model of diabetic neuropathy. PGC-1α and its-regulated transcription factors including TFAM and NRF1, which are master regulators of mitochondrial biogenesis, are significantly downregulated in streptozotocin diabetic dorsal root ganglion (DRG) neurons. Diabetic mice develop peripheral neuropathy, loss of mitochondria, decreased mitochondrial DNA content and increased protein oxidation. Importantly, this phenotype is exacerbated in PGC-1α (-/-) diabetic mice, which develop a more severe neuropathy with reduced mitochondrial DNA and a further increase in protein oxidation. PGC-1α (-/-) diabetic mice develop an increase in total cholesterol and triglycerides, and a decrease in TFAM and NRF1 protein levels. Loss of PGC-1α causes severe mitochondrial degeneration with vacuolization in DRG neurons, coupled with reduced state 3 and 4 respiration, reduced expression of oxidative stress response genes and an increase in protein oxidation. In contrast, overexpression of PGC-1α in cultured adult mouse neurons prevents oxidative stress associated with increased glucose levels. The study provides new insights into the role of PGC-1α in mitochondrial regeneration in peripheral neurons and suggests that therapeutic modulation of PGC-1α function may be an attractive approach for treatment of diabetic neuropathy.

  2. Mitochondrial Biogenesis in the Pulmonary Vasculature During Inhalational Lung Injury and Fibrosis

    PubMed Central

    CARRAWAY, MARTHA S.; SULIMAN, HAGIR B.; KLIMENT, CORRINE; WELTY-WOLF, KAREN E.; OURY, TIM D.; PIANTADOSI, CLAUDE A.

    2008-01-01

    Cell survival and injury repair is facilitated by mitochondrial biogenesis; however, the role of this process in lung repair is unknown. We evaluated mitochondrial biogenesis in the mouse lung in two injuries that cause acute inflammation and in two that cause chronic inflammation and pulmonary fibrosis. By using reporter mice that express green fluorescent protein (GFP) exclusively in mitochondria, we tracked mitochondrial biogenesis and correlated it with histologic lung injury, proliferation, and fibrosis. At 72 hours after acute LPS or continuous exposure to hyperoxia (Fio2, 1.0), the lungs showed diffuse infiltration by inflammatory cells in the alveolar region. In reporter mice, patchy new mitochondrial fluorescence was found in the alveolar region but was most prominent and unexpected in perivascular regions. At 14 days after instillation of asbestos or bleomycin, diffuse chronic inflammation had developed, and green fluorescence appeared in inflammatory cells in the expanded interstitium and was most intense in smooth muscle cells of pulmonary vessels. In all four lung injuries, mitochondrial fluorescence colocalized with mitochondrial superoxide dismutase, but not with proliferating cell nuclear antigen. These data indicate that vascular mitochondrial biogenesis is activated in diverse inhalational lung injuries along with oxidative stress. This finding indicates a unique and unexpected mechanism of metabolic adaptation to pulmonary fibrotic injuries. PMID:17999632

  3. Parkinson's disease-associated mutant VPS35 causes mitochondrial dysfunction by recycling DLP1 complexes.

    PubMed

    Wang, Wenzhang; Wang, Xinglong; Fujioka, Hisashi; Hoppel, Charles; Whone, Alan L; Caldwell, Maeve A; Cullen, Peter J; Liu, Jun; Zhu, Xiongwei

    2016-01-01

    Mitochondrial dysfunction represents a critical step during the pathogenesis of Parkinson's disease (PD), and increasing evidence suggests abnormal mitochondrial dynamics and quality control as important underlying mechanisms. The VPS35 gene, which encodes a key component of the membrane protein-recycling retromer complex, is the third autosomal-dominant gene associated with PD. However, how VPS35 mutations lead to neurodegeneration remains unclear. Here we demonstrate that PD-associated VPS35 mutations caused mitochondrial fragmentation and cell death in cultured neurons in vitro, in mouse substantia nigra neurons in vivo and in human fibroblasts from an individual with PD who has the VPS35(D620N) mutation. VPS35-induced mitochondrial deficits and neuronal dysfunction could be prevented by inhibition of mitochondrial fission. VPS35 mutants showed increased interaction with dynamin-like protein (DLP) 1, which enhanced turnover of the mitochondrial DLP1 complexes via the mitochondria-derived vesicle-dependent trafficking of the complexes to lysosomes for degradation. Notably, oxidative stress increased the VPS35-DLP1 interaction, which we also found to be increased in the brains of sporadic PD cases. These results revealed a novel cellular mechanism for the involvement of VPS35 in mitochondrial fission, dysregulation of which is probably involved in the pathogenesis of familial, and possibly sporadic, PD. PMID:26618722

  4. Requirement for the Mitochondrial Pyruvate Carrier in Mammalian Development Revealed by a Hypomorphic Allelic Series.

    PubMed

    Bowman, Caitlyn E; Zhao, Liang; Hartung, Thomas; Wolfgang, Michael J

    2016-08-01

    Glucose and oxygen are two of the most important molecules transferred from mother to fetus during eutherian pregnancy, and the metabolic fates of these nutrients converge at the transport and metabolism of pyruvate in mitochondria. Pyruvate enters the mitochondrial matrix through the mitochondrial pyruvate carrier (MPC), a complex in the inner mitochondrial membrane that consists of two essential components, MPC1 and MPC2. Here, we define the requirement for mitochondrial pyruvate metabolism during development with a progressive allelic series of Mpc1 deficiency in mouse. Mpc1 deletion was homozygous lethal in midgestation, but Mpc1 hypomorphs and tissue-specific deletion of Mpc1 presented as early perinatal lethality. The allelic series demonstrated that graded suppression of MPC resulted in dose-dependent metabolic and transcriptional changes. Steady-state metabolomics analysis of brain and liver from Mpc1 hypomorphic embryos identified compensatory changes in amino acid and lipid metabolism. Flux assays in Mpc1-deficient embryonic fibroblasts also reflected these changes, including a dramatic increase in mitochondrial alanine utilization. The mitochondrial alanine transaminase GPT2 was found to be necessary and sufficient for increased alanine flux upon MPC inhibition. These data show that impaired mitochondrial pyruvate transport results in biosynthetic deficiencies that can be mitigated in part by alternative anaplerotic substrates in utero. PMID:27215380

  5. Loss of mitochondrial fission depletes axonal mitochondria in midbrain dopamine neurons.

    PubMed

    Berthet, Amandine; Margolis, Elyssa B; Zhang, Jue; Hsieh, Ivy; Zhang, Jiasheng; Hnasko, Thomas S; Ahmad, Jawad; Edwards, Robert H; Sesaki, Hiromi; Huang, Eric J; Nakamura, Ken

    2014-10-22

    Disruptions in mitochondrial dynamics may contribute to the selective degeneration of dopamine (DA) neurons in Parkinson's disease (PD). However, little is known about the normal functions of mitochondrial dynamics in these neurons, especially in axons where degeneration begins, and this makes it difficult to understand the disease process. To study one aspect of mitochondrial dynamics-mitochondrial fission-in mouse DA neurons, we deleted the central fission protein dynamin-related protein 1 (Drp1). Drp1 loss rapidly eliminates the DA terminals in the caudate-putamen and causes cell bodies in the midbrain to degenerate and lose α-synuclein. Without Drp1, mitochondrial mass dramatically decreases, especially in axons, where the mitochondrial movement becomes uncoordinated. However, in the ventral tegmental area (VTA), a subset of midbrain DA neurons characterized by small hyperpolarization-activated cation currents (Ih) is spared, despite near complete loss of their axonal mitochondria. Drp1 is thus critical for targeting mitochondria to the nerve terminal, and a disruption in mitochondrial fission can contribute to the preferential death of nigrostriatal DA neurons.

  6. Peroxynitrite induced mitochondrial biogenesis following MnSOD knockdown in normal rat kidney (NRK) cells.

    PubMed

    Marine, Akira; Krager, Kimberly J; Aykin-Burns, Nukhet; Macmillan-Crow, Lee Ann

    2014-01-01

    Superoxide is widely regarded as the primary reactive oxygen species (ROS) which initiates downstream oxidative stress. Increased oxidative stress contributes, in part, to many disease conditions such as cancer, atherosclerosis, ischemia/reperfusion, diabetes, aging, and neurodegeneration. Manganese superoxide dismutase (MnSOD) catalyzes the dismutation of superoxide into hydrogen peroxide which can then be further detoxified by other antioxidant enzymes. MnSOD is critical in maintaining the normal function of mitochondria, thus its inactivation is thought to lead to compromised mitochondria. Previously, our laboratory observed increased mitochondrial biogenesis in a novel kidney-specific MnSOD knockout mouse. The current study used transient siRNA mediated MnSOD knockdown of normal rat kidney (NRK) cells as the in vitro model, and confirmed functional mitochondrial biogenesis evidenced by increased PGC1α expression, mitochondrial DNA copy numbers and integrity, electron transport chain protein CORE II, mitochondrial mass, oxygen consumption rate, and overall ATP production. Further mechanistic studies using mitoquinone (MitoQ), a mitochondria-targeted antioxidant and L-NAME, a nitric oxide synthase (NOS) inhibitor demonstrated that peroxynitrite (at low micromolar levels) induced mitochondrial biogenesis. These findings provide the first evidence that low levels of peroxynitrite can initiate a protective signaling cascade involving mitochondrial biogenesis which may help to restore mitochondrial function following transient MnSOD inactivation. PMID:24563852

  7. A matter of quantum voltages.

    PubMed

    Sellner, Bernhard; Kathmann, Shawn M

    2014-11-14

    Voltages inside matter are relevant to crystallization, materials science, biology, catalysis, and aqueous chemistry. The variation of voltages in matter can be measured by experiment, however, modern supercomputers allow the calculation of accurate quantum voltages with spatial resolutions of bulk systems well beyond what can currently be measured provided a sufficient level of theory is employed. Of particular interest is the Mean Inner Potential (V(o))--the spatial average of these quantum voltages referenced to the vacuum. Here we establish a protocol to reliably evaluate V(o) from quantum calculations. Voltages are very sensitive to the distribution of electrons and provide metrics to understand interactions in condensed phases. In the present study, we find excellent agreement with measurements of V(o) for vitrified water and salt crystals and demonstrate the impact of covalent and ionic bonding as well as intermolecular/atomic interactions. Certain aspects in this regard are highlighted making use of simple model systems/approximations. Furthermore, we predict V(o) as well as the fluctuations of these voltages in aqueous NaCl electrolytes and characterize the changes in their behavior as the resolution increases below the size of atoms. PMID:25399199

  8. A matter of quantum voltages.

    PubMed

    Sellner, Bernhard; Kathmann, Shawn M

    2014-11-14

    Voltages inside matter are relevant to crystallization, materials science, biology, catalysis, and aqueous chemistry. The variation of voltages in matter can be measured by experiment, however, modern supercomputers allow the calculation of accurate quantum voltages with spatial resolutions of bulk systems well beyond what can currently be measured provided a sufficient level of theory is employed. Of particular interest is the Mean Inner Potential (V(o))--the spatial average of these quantum voltages referenced to the vacuum. Here we establish a protocol to reliably evaluate V(o) from quantum calculations. Voltages are very sensitive to the distribution of electrons and provide metrics to understand interactions in condensed phases. In the present study, we find excellent agreement with measurements of V(o) for vitrified water and salt crystals and demonstrate the impact of covalent and ionic bonding as well as intermolecular/atomic interactions. Certain aspects in this regard are highlighted making use of simple model systems/approximations. Furthermore, we predict V(o) as well as the fluctuations of these voltages in aqueous NaCl electrolytes and characterize the changes in their behavior as the resolution increases below the size of atoms.

  9. A Matter of Quantum Voltages

    SciTech Connect

    Sellner, Bernhard; Kathmann, Shawn M.

    2014-11-14

    Voltages inside matter are relevant to crystallization, materials science, biology, catalysis, and aqueous chemistry. Electron holography is able to measure the variation of voltages in matter and modern supercomputers allow the calculation of quantum voltages with practically unlimited spatial and temporal resolution of bulk systems. Of particular interest is the Mean Inner Potential (Vo) - the spatial average of these voltages. Voltages are very sensitive to the distribution of electrons and provide metrics to understand interactions in condensed phases. In the present study, we find excellent agreement with measurements of Vo for vitrified water and salt crystals and demonstrate the impact of covalent and ionic bonding as well as intermolecular/atomic interactions. Furthermore, we predict Vo as well as the fluctuations of these voltages in aqueous NaCl electrolytes and characterize the changes in their behavior as the resolution increases below the size of atoms. This work was supported by the U.S. Department of Energy, Office of Basic Energy Sciences, Division of Chemical Sciences, Geosciences & Biosciences. Pacific Northwest National Laboratory (PNNL) is a multiprogram national laboratory operated for DOE by Battelle. This research used resources of the National Energy Research Scientific Computing Center, which is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231.

  10. A matter of quantum voltages

    SciTech Connect

    Sellner, Bernhard; Kathmann, Shawn M.

    2014-11-14

    Voltages inside matter are relevant to crystallization, materials science, biology, catalysis, and aqueous chemistry. The variation of voltages in matter can be measured by experiment, however, modern supercomputers allow the calculation of accurate quantum voltages with spatial resolutions of bulk systems well beyond what can currently be measured provided a sufficient level of theory is employed. Of particular interest is the Mean Inner Potential (V{sub o}) – the spatial average of these quantum voltages referenced to the vacuum. Here we establish a protocol to reliably evaluate V{sub o} from quantum calculations. Voltages are very sensitive to the distribution of electrons and provide metrics to understand interactions in condensed phases. In the present study, we find excellent agreement with measurements of V{sub o} for vitrified water and salt crystals and demonstrate the impact of covalent and ionic bonding as well as intermolecular/atomic interactions. Certain aspects in this regard are highlighted making use of simple model systems/approximations. Furthermore, we predict V{sub o} as well as the fluctuations of these voltages in aqueous NaCl electrolytes and characterize the changes in their behavior as the resolution increases below the size of atoms.

  11. Voltage-Boosting Driver For Switching Regulator

    NASA Technical Reports Server (NTRS)

    Trump, Ronald C.

    1990-01-01

    Driver circuit assures availability of 10- to 15-V gate-to-source voltage needed to turn on n-channel metal oxide/semiconductor field-effect transistor (MOSFET) acting as switch in switching voltage regulator. Includes voltage-boosting circuit efficiently providing gate voltage 10 to 15 V above supply voltage. Contains no exotic parts and does not require additional power supply. Consists of NAND gate and dual voltage booster operating in conjunction with pulse-width modulator part of regulator.

  12. [Mitochondrial neurogastrointestinal encephalopathy disease].

    PubMed

    Benureau, A; Meyer, P; Maillet, O; Leboucq, N; Legras, S; Jeziorski, E; Fournier-Favre, S; Jeandel, C; Gaignard, P; Slama, A; Rivier, F; Roubertie, A; Carneiro, M

    2014-12-01

    Mitochondrial neurogastrointestinal encephalopathy disease (MNGIE) is a rare autosomal-recessive syndrome, resulting from mutations in the TYMP gene, located at 22q13. The mutation induces a thymidine phosphorylase (TP) deficit, which leads to a nucleotide pool imbalance and to instability of the mitochondrial DNA. The clinical picture regroups gastrointestinal dysmotility, cachexia, ptosis, ophthalmoplegia, peripheral neuropathy, and asymptomatic leukoencephalopathy. The prognosis is unfavorable. We present the case of a 14-year-old Caucasian female whose symptoms started in early childhood. The diagnosis was suspected after magnetic resonance imaging (MRI), performed given the atypical features of mental anorexia, which revealed white matter abnormalities. She presented chronic vomiting, postprandial abdominal pain, and problems gaining weight accompanied by cachexia. This diagnosis led to establishing proper care, in particular an enteral and parenteral nutrition program. There is no known specific effective treatment, but numerous studies are in progress. In this article, after reviewing the existing studies, we discuss the main diagnostic and therapeutic aspects of the disease. We argue for the necessity of performing a cerebral MRI given the atypical features of a patient with suspected mental anorexia (or when the clinical pattern of a patient with mental anorexia seems atypical), so that MNGIE can be ruled out. PMID:25282463

  13. Ubiquitylation of voltage-gated sodium channels.

    PubMed

    Laedermann, Cédric J; Decosterd, Isabelle; Abriel, Hugues

    2014-01-01

    Ion channel proteins are regulated by different types of posttranslational modifications. The focus of this review is the regulation of voltage-gated sodium channels (Navs) upon their ubiquitylation. The amiloride-sensitive epithelial sodium channel (ENaC) was the first ion channel shown to be regulated upon ubiquitylation. This modification results from the binding of ubiquitin ligase from the Nedd4 family to a protein-protein interaction domain, known as the PY motif, in the ENaC subunits. Many of the Navs have similar PY motifs, which have been demonstrated to be targets of Nedd4-dependent ubiquitylation, tagging them for internalization from the cell surface. The role of Nedd4-dependent regulation of the Nav membrane density in physiology and disease remains poorly understood. Two recent studies have provided evidence that Nedd4-2 is downregulated in dorsal root ganglion (DRG) neurons in both rat and mouse models of nerve injury-induced neuropathic pain. Using two different mouse models, one with a specific knockout of Nedd4-2 in sensory neurons and another where Nedd4-2 was overexpressed with the use of viral vectors, it was demonstrated that the neuropathy-linked neuronal hyperexcitability was the result of Nav1.7 and Nav1.8 overexpression due to Nedd4-2 downregulation. These studies provided the first in vivo evidence of the role of Nedd4-2-dependent regulation of Nav channels in a disease state. This ubiquitylation pathway may be involved in the development of symptoms and diseases linked to Nav-dependent hyperexcitability, such as pain, cardiac arrhythmias, epilepsy, migraine, and myotonias. PMID:24737239

  14. On voltage collapse in electric power systems

    SciTech Connect

    Chiang, H.D.; Dobson, I.; Thomas, R.J.; Thorp, J.S.; Fekih-Ahmed, L. . School of Electrical Engineering)

    1990-05-01

    Several voltage collapses have had a period of slowly decreasing voltage followed by an accelerating collapse in voltage. This paper analyzes this type of voltage collapse based on a center manifold voltage collapse model. The essence of this model is that the system dynamics after bifurcation are captured by the center manifold trajectory and it is a computable model that allows prediction of voltage collapse. Both physical explanations and computational considerations of this model are presented. The authors clarify the use of static and dynamic models to explain voltage collapse. Voltage collapse dynamics are demonstrated on a simple power system model.

  15. High resolution respirometry analysis of polyethylenimine-mediated mitochondrial energy crisis and cellular stress: Mitochondrial proton leak and inhibition of the electron transport system.

    PubMed

    Hall, Arnaldur; Larsen, Anna K; Parhamifar, Ladan; Meyle, Kathrine D; Wu, Lin-Ping; Moghimi, S Moein

    2013-10-01

    Polyethylenimines (PEIs) are highly efficient non-viral transfectants, but can induce cell death through poorly understood necrotic and apoptotic processes as well as autophagy. Through high resolution respirometry studies in H1299 cells we demonstrate that the 25kDa branched polyethylenimine (25k-PEI-B), in a concentration and time-dependent manner, facilitates mitochondrial proton leak and inhibits the electron transport system. These events were associated with gradual reduction of the mitochondrial membrane potential and mitochondrial ATP synthesis. The intracellular ATP levels further declined as a consequence of PEI-mediated plasma membrane damage and subsequent ATP leakage to the extracellular medium. Studies with freshly isolated mouse liver mitochondria corroborated with bioenergetic findings and demonstrated parallel polycation concentration- and time-dependent changes in state 2 and state 4o oxygen flux as well as lowered ADP phosphorylation (state 3) and mitochondrial ATP synthesis. Polycation-mediated reduction of electron transport system activity was further demonstrated in 'broken mitochondria' (freeze-thawed mitochondrial preparations). Moreover, by using both high-resolution respirometry and spectrophotometry analysis of cytochrome c oxidase activity we were able to identify complex IV (cytochrome c oxidase) as a likely specific site of PEI mediated inhibition within the electron transport system. Unraveling the mechanisms of PEI-mediated mitochondrial energy crisis is central for combinatorial design of safer polymeric non-viral gene delivery systems. PMID:23850549

  16. Mouse Curve Biometrics

    SciTech Connect

    Schulz, Douglas A.

    2007-10-08

    A biometric system suitable for validating user identity using only mouse movements and no specialized equipment is presented. Mouse curves (mouse movements with little or no pause between them) are individually classied and used to develop classication histograms, which are representative of an individual's typical mouse use. These classication histograms can then be compared to validate identity. This classication approach is suitable for providing continuous identity validation during an entire user session.

  17. Identification of two voltage-dependent anion channel-like protein sequences conserved in Kinetoplastida

    PubMed Central

    Flinner, Nadine; Schleiff, Enrico; Mirus, Oliver

    2012-01-01

    The eukaryotic porin superfamily consists of two families, voltage-dependent anion channel (VDAC) and Tom40, which are both located in the mitochondrial outer membrane. In Trypanosoma brucei, only a single member of the VDAC family has been described. We report the detection of two additional eukaryotic porin-like sequences in T. brucei. By bioinformatic means, we classify both as putative VDAC isoforms. PMID:22219392

  18. Electrophysiological and metabolic effects of a convulsant barbiturate on dissociated mouse primary sensory neurons.

    PubMed Central

    Pearce, R J; Duchen, M R

    1995-01-01

    1. The convulsant barbiturate 5-(2-cyclohexylidene-ethyl)-5-ethyl barbituric acid (CHEB) depolarizes dorsal root ganglion (DRG) neurons. We have applied microfluorimetric and whole-cell patch clamp techniques to investigate the mechanisms underlying this response in freshly dissociated mouse DRG cells. 2. Application of CHEB (2-200 microM) raised cytosolic calcium concentration ([Ca2+]i) rapidly and reversibly in 55% of eighty-three neurons tested. This population did not correlate with other classifications of sensory neurons based on either cell size or the expression of membrane currents. 3. The response was dependent on external calcium and was reduced by 81 +/- 22% by Ruthenium Red. A rise in [Ca2+]i was still seen with the membrane potential clamped at -70 mV, excluding membrane depolarization and activation of voltage-dependent Ca2+ channels as the principal mechanism for the response. 4. The rise in [Ca2+]i was associated with an increase in membrane conductance and a current, ICHEB, which was inward at -70 mV. Both the rise in [Ca2+]i and the current showed 'run-down' under whole-cell recording conditions. When K+ conductances were blocked, the reversal potential of ICHEB was close to 0 mV. This was independent of the Cl- reversal potential, suggesting that ICHEB is carried as a non-specific cation current. 5. In contrast to the change in [Ca2+]i, ICHEB was not dependent on external Ca2+ and the current was still seen when [Ca2+]i as strongly buffered by the pipette filling solution. These data suggest that CHEB opens a non-selective cation channel permeant to Ca2+, raising [Ca2+]i and further depolarizing the cell membrane potential. The exact nature of this conductance remains unknown. These actions could readily account for the convulsant actions of the drug, depolarizing neurons and increasing transmitter release. 6. It was also noted that CHEB increases autofluorescence derived from mitochondrial NAD(P)H. Further examination of this phenomenon using

  19. Building a Brainier Mouse.

    ERIC Educational Resources Information Center

    Tsien, Joe Z.

    2000-01-01

    Describes a genetic engineering project to build an intelligent mouse. Cites understanding the molecular basis of learning and memory as a very important step. Concludes that while science will never create a genius mouse that plays the stock market, it can turn a mouse into a quick learner with a better memory. (YDS)

  20. PRKAA1/AMPKα1 is required for autophagy-dependent mitochondrial clearance during erythrocyte maturation.

    PubMed

    Zhu, Huaiping; Foretz, Marc; Xie, Zhonglin; Zhang, Miao; Zhu, Zhiren; Xing, Junjie; Leclerc, Jocelyne; Gaudry, Murielle; Viollet, Benoit; Zou, Ming-Hui

    2014-09-01

    AMP-activated protein kinase α1 knockout (prkaa1(-/-)) mice manifest splenomegaly and anemia. The underlying molecular mechanisms, however, remain to be established. In this study, we tested the hypothesis that defective autophagy-dependent mitochondrial clearance in prkaa1(-/-) mice exacerbates oxidative stress, thereby enhancing erythrocyte destruction. The levels of ULK1 phosphorylation, autophagical flux, mitochondrial contents, and reactive oxygen species (ROS) were examined in human erythroleukemia cell line, K562 cells, as well as prkaa1(-/-) mouse embryonic fibroblasts and erythrocytes. Deletion of Prkaa1 resulted in the inhibition of ULK1 phosphorylation at Ser555, prevented the formation of ULK1 and BECN1- PtdIns3K complexes, and reduced autophagy capacity. The suppression of autophagy was associated with enhanced damaged mitochondrial accumulation and ROS production. Compared with wild-type (WT) mice, prkaa1(-/-) mice exhibited a shortened erythrocyte life span, hemolytic destruction of erythrocytes, splenomegaly, and anemia, all of which were alleviated by the administration of either rapamycin to activate autophagy or Mito-tempol, a mitochondria-targeted antioxidant, to scavenge mitochondrial ROS. Furthermore, transplantation of WT bone marrow into prkaa1(-/-) mice restored mitochondrial removal, reduced intracellular ROS levels, and normalized hematologic parameters and spleen size. Conversely, transplantation of prkaa1 (-/-) bone marrow into WT mice recapitulated the prkaa1(-/-) mouse phenotypes. We conclude that PRKAA1-dependent autophagy-mediated clearance of damaged mitochondria is required for erythrocyte maturation and homeostasis.

  1. Electrode voltage fall and total voltage of a transient arc

    NASA Astrophysics Data System (ADS)

    Valensi, F.; Ratovoson, L.; Razafinimanana, M.; Masquère, M.; Freton, P.; Gleizes, A.

    2016-06-01

    This paper deals with an experimental study of the components of a transient arc total voltage with duration of a few tens of ms and a current peak close to 1000 A. The cathode tip is made of graphite whereas the flat anode is made either of copper or of graphite; the electrodes gap is a few mm. The analysis of the electrical parameters is supported and validated by fast imaging and by two models: the first one is a 2D physical model of the arc allowing to calculate both the plasma temperature field and the arc voltage; the second model is able to estimate the transient heating of the graphite electrode. The main aim of the study was to detect the possible change of the cathode voltage fall (CVF) during the first instants of the arc. Indeed it is expected that during the first ms the graphite cathode is rather cool and the main mechanism of the electron emission should be the field effect emission, whereas after several tens of ms the cathode is strongly heated and thermionic emission should be predominant. We have observed some change in the apparent CVF but we have shown that this apparent change can be attributed to the variation of the solid cathode resistance. On the other hand, the possible change of CVF corresponding to the transition between a ‘cold’ and a ‘hot’ cathode should be weak and could not be characterized considering our measurement uncertainty of about 2 V. The arc column voltage (ACV) was estimated by subtracting the electrode voltage fall from the total arc voltage. The experimental transient evolution of the ACV is in very good agreement with the theoretical variation predicted by the model, showing the good ability of the model to study this kind of transient arc.

  2. Mitochondrial dysfunction in heart failure.

    PubMed

    Rosca, Mariana G; Hoppel, Charles L

    2013-09-01

    Heart failure (HF) is a complex chronic clinical syndrome. Energy deficit is considered to be a key contributor to the development of both cardiac and skeletal myopathy. In HF, several components of cardiac and skeletal muscle bioenergetics are altered, such as oxygen availability, substrate oxidation, mitochondrial ATP production, and ATP transfer to the contractile apparatus via the creatine kinase shuttle. This review focuses on alterations in mitochondrial biogenesis and respirasome organization, substrate oxidation coupled with ATP synthesis in the context of their contribution to the chronic energy deficit, and mechanical dysfunction of the cardiac and skeletal muscle in HF. We conclude that HF is associated with decreased mitochondrial biogenesis and function in both heart and skeletal muscle, supporting the concept of a systemic mitochondrial cytopathy. The sites of mitochondrial defects are located within the electron transport and phosphorylation apparatus and differ with the etiology and progression of HF in the two mitochondrial populations (subsarcolemmal and interfibrillar) of cardiac and skeletal muscle. The roles of adrenergic stimulation, the renin-angiotensin system, and cytokines are evaluated as factors responsible for the systemic energy deficit. We propose a cyclic AMP-mediated mechanism by which increased adrenergic stimulation contributes to the mitochondrial dysfunction.

  3. Mitochondrial dysfunction in heart failure

    PubMed Central

    Rosca, Mariana G.; Hoppel, Charles L.

    2013-01-01

    Heart failure (HF) is a complex chronic clinical syndrome. Energy deficit is considered to be a key contributor to the development of both cardiac and skeletal myopathy. In HF several components of cardiac and skeletal muscle bioenergetics are altered, such as oxygen availability, substrate oxidation, mitochondrial ATP production, and ATP transfer to the contractile apparatus via the creatine kinase shuttle. This review focuses on alterations in mitochondrial biogenesis and respirasome organization, substrate oxidation coupled with ATP synthesis in the context of their contribution to the chronic energy deficit, and mechanical dysfunction of the cardiac and skeletal muscle in HF. We conclude that HF is associated with decreased mitochondrial biogenesis and function in both heart and skeletal muscle, supporting the concept of a systemic mitochondrial cytopathy. The sites of mitochondrial defects are located within the electron transport and phosphorylation apparatus, and differ with the etiology and progression of HF in the two mitochondrial populations (subsarcolemmal and interfibrillar) of cardiac and skeletal muscle. The roles of adrenergic stimulation, the renin-angiotensin system, and cytokines are evaluated as factors responsible for the systemic energy deficit. We propose a cylic AMP-mediated mechanism by which increased adrenergic stimulation contributes to the mitochondrial dysfunction. PMID:22948484

  4. Role and Treatment of Mitochondrial DNA-Related Mitochondrial Dysfunction in Sporadic Neurodegenerative Diseases

    PubMed Central

    Swerdlow, Russell H.

    2012-01-01

    Several sporadic neurodegenerative diseases display phenomena that directly or indirectly relate to mitochondrial function. Data suggesting altered mitochondrial function in these diseases could arise from mitochondrial DNA (mtDNA) are reviewed. Approaches for manipulating mitochondrial function and minimizing the downstream consequences of mitochondrial dysfunction are discussed. PMID:21902672

  5. Mitochondrial Quality Control in Cardiac Diseases

    PubMed Central

    Campos, Juliane C.; Bozi, Luiz H. M.; Bechara, Luiz R. G.; Lima, Vanessa M.; Ferreira, Julio C. B.

    2016-01-01

    Disruption of mitochondrial homeostasis is a hallmark of cardiac diseases. Therefore, maintenance of mitochondrial integrity through different surveillance mechanisms is critical for cardiomyocyte survival. In this review, we discuss the most recent findings on the central role of mitochondrial quality control processes including regulation of mitochondrial redox balance, aldehyde metabolism, proteostasis, dynamics, and clearance in cardiac diseases, highlighting their potential as therapeutic targets.

  6. Respiratory active mitochondrial supercomplexes.

    PubMed

    Acín-Pérez, Rebeca; Fernández-Silva, Patricio; Peleato, Maria Luisa; Pérez-Martos, Acisclo; Enriquez, Jose Antonio

    2008-11-21

    The structural organization of the mitochondrial respiratory complexes as four big independently moving entities connected by the mobile carriers CoQ and cytochrome c has been challenged recently. Blue native gel electrophoresis reveals the presence of high-molecular-weight bands containing several respiratory complexes and suggesting an in vivo assembly status of these structures (respirasomes). However, no functional evidence of the activity of supercomplexes as true respirasomes has been provided yet. We have observed that (1) supercomplexes are not formed when one of their component complexes is absent; (2) there is a temporal gap between the formation of the individual complexes and that of the supercomplexes; (3) some putative respirasomes contain CoQ and cytochrome c; (4) isolated respirasomes can transfer electrons from NADH to O(2), that is, they respire. Therefore, we have demonstrated the existence of a functional respirasome and propose a structural organization model that accommodates these findings.

  7. Protecting the mitochondrial powerhouse.

    PubMed

    Scheibye-Knudsen, Morten; Fang, Evandro F; Croteau, Deborah L; Wilson, David M; Bohr, Vilhelm A

    2015-03-01

    Mitochondria are the oxygen-consuming power plants of cells. They provide a critical milieu for the synthesis of many essential molecules and allow for highly efficient energy production through oxidative phosphorylation. The use of oxygen is, however, a double-edged sword that on the one hand supplies ATP for cellular survival, and on the other leads to the formation of damaging reactive oxygen species (ROS). Different quality control pathways maintain mitochondria function including mitochondrial DNA (mtDNA) replication and repair, fusion-fission dynamics, free radical scavenging, and mitophagy. Further, failure of these pathways may lead to human disease. We review these pathways and propose a strategy towards a treatment for these often untreatable disorders.

  8. Lophotrochozoan mitochondrial genomes

    SciTech Connect

    Valles, Yvonne; Boore, Jeffrey L.

    2005-10-01

    Progress in both molecular techniques and phylogeneticmethods has challenged many of the interpretations of traditionaltaxonomy. One example is in the recognition of the animal superphylumLophotrochozoa (annelids, mollusks, echiurans, platyhelminthes,brachiopods, and other phyla), although the relationships within thisgroup and the inclusion of some phyla remain uncertain. While much ofthis progress in phylogenetic reconstruction has been based on comparingsingle gene sequences, we are beginning to see the potential of comparinglarge-scale features of genomes, such as the relative order of genes.Even though tremendous progress is being made on the sequencedetermination of whole nuclear genomes, the dataset of choice forgenome-level characters for many animals across a broad taxonomic rangeremains mitochondrial genomes. We review here what is known aboutmitochondrial genomes of the lophotrochozoans and discuss the promisethat this dataset will enable insight into theirrelationships.

  9. Lysine Acetylation Activates Mitochondrial Aconitase in the Heart

    PubMed Central

    Fernandes, Jolyn; Weddle, Alexis; Kinter, Caroline S.; Humphries, Kenneth M.; Mather, Timothy; Szweda, Luke I.; Kinter, Michael

    2015-01-01

    High throughput proteomics studies have identified several thousand acetylation sites on over one thousand proteins. Mitochondrial aconitase, the Krebs cycle enzyme that converts citrate to isocitrate, has been identified in many of these reports. Acetylated mitochondrial aconitase has also been identified as a target for sirtuin 3 (SIRT3) catalyzed deacetylation. However, the functional significance of mitochondrial aconitase acetylation has not been determined. Using in vitro strategies, mass spectrometric analyses, and an in vivo mouse model of obesity, we found a significant acetylation-dependent activation of aconitase. Isolated heart mitochondria subjected to in vitro chemical acetylation with either acetic anhydride or acetyl-CoA resulted in increased aconitase activity that was reversed with SIRT3 treatment. Quantitative mass spectrometry was used to measure acetylation at 21 lysine residues and found significant increases with both in vitro treatments. A high fat diet (60% kcal from fat) was used as an in vivo model and also showed significantly increased mitochondrial aconitase activity without changes in protein level. The high fat diet also produced increased aconitase acetylation at multiple sites as measured by the quantitative mass spectrometry assays. Treatment of isolated mitochondria from these mice with SIRT3 abolished the high fat diet-induced activation of aconitase and reduced acetylation. Finally, kinetic analyses found that the increase in activity was a result of increased maximal velocity and molecular modeling suggests the potential for acetylation at K144 to perturb the tertiary structure of the enzyme. The results of this study reveal a novel activation of mitochondrial aconitase by acetylation. PMID:26061789

  10. Early effects of the antineoplastic agent salinomycin on mitochondrial function.

    PubMed

    Managò, A; Leanza, L; Carraretto, L; Sassi, N; Grancara, S; Quintana-Cabrera, R; Trimarco, V; Toninello, A; Scorrano, L; Trentin, L; Semenzato, G; Gulbins, E; Zoratti, M; Szabò, I

    2015-01-01

    Salinomycin, isolated from Streptomyces albus, displays antimicrobial activity. Recently, a large-scale screening approach identified salinomycin and nigericin as selective apoptosis inducers of cancer stem cells. Growing evidence suggests that salinomycin is able to kill different types of non-stem tumor cells that usually display resistance to common therapeutic approaches, but the mechanism of action of this molecule is still poorly understood. Since salinomycin has been suggested to act as a K(+) ionophore, we explored its impact on mitochondrial bioenergetic performance at an early time point following drug application. In contrast to the K(+) ionophore valinomycin, salinomycin induced a rapid hyperpolarization. In addition, mitochondrial matrix acidification and a significant decrease of respiration were observed in intact mouse embryonic fibroblasts (MEFs) and in cancer stem cell-like HMLE cells within tens of minutes, while increased production of reactive oxygen species was not detected. By comparing the chemical structures and cellular effects of this drug with those of valinomycin (K(+) ionophore) and nigericin (K(+)/H(+) exchanger), we conclude that salinomycin mediates K(+)/H(+) exchange across the inner mitochondrial membrane. Compatible with its direct modulation of mitochondrial function, salinomycin was able to induce cell death also in Bax/Bak-less double-knockout MEF cells. Since at the concentration range used in most studies (around 10 μM) salinomycin exerts its effect at the level of mitochondria and alters bioenergetic performance, the specificity of its action on pathologic B cells isolated from patients with chronic lymphocytic leukemia (CLL) versus B cells from healthy subjects was investigated. Mesenchymal stromal cells (MSCs), proposed to mimic the tumor environment, attenuated the apoptotic effect of salinomycin on B-CLL cells. Apoptosis occurred to a significant extent in healthy B cells as well as in MSCs and human primary

  11. Mitochondrial Ceramide-Rich Macrodomains Functionalize Bax upon Irradiation

    PubMed Central

    Lee, Hyunmi; Rotolo, Jimmy A.; Mesicek, Judith; Penate-Medina, Tuula; Rimner, Andreas; Liao, Wen-Chieh; Yin, Xianglei; Ragupathi, Govind; Ehleiter, Desiree; Gulbins, Erich; Zhai, Dayong; Reed, John C.; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

    2011-01-01

    Background Evidence indicates that Bax functions as a “lipidic” pore to regulate mitochondrial outer membrane permeabilization (MOMP), the apoptosis commitment step, through unknown membrane elements. Here we show mitochondrial ceramide elevation facilitates MOMP-mediated cytochrome c release in HeLa cells by generating a previously-unrecognized mitochondrial ceramide-rich macrodomain (MCRM), which we visualize and isolate, into which Bax integrates. Methodology/Principal Findings MCRMs, virtually non-existent in resting cells, form upon irradiation coupled to ceramide synthase-mediated ceramide elevation, optimizing Bax insertion/oligomerization and MOMP. MCRMs are detected by confocal microscopy in intact HeLa cells and isolated biophysically as a light membrane fraction from HeLa cell lysates. Inhibiting ceramide generation using a well-defined natural ceramide synthase inhibitor, Fumonisin B1, prevented radiation-induced Bax insertion, oligomerization and MOMP. MCRM deconstruction using purified mouse hepatic mitochondria revealed ceramide alone is non-apoptogenic. Rather Bax integrates into MCRMs, oligomerizing therein, conferring 1–2 log enhanced cytochrome c release. Consistent with this mechanism, MCRM Bax isolates as high molecular weight “pore-forming” oligomers, while non-MCRM membrane contains exclusively MOMP-incompatible monomeric Bax. Conclusions/Significance Our recent studies in the C. elegans germline indicate that mitochondrial ceramide generation is obligate for radiation-induced apoptosis, although a mechanism for ceramide action was not delineated. Here we demonstrate that ceramide, generated in the mitochondrial outer membrane of mammalian cells upon irradiation, forms a platform into which Bax inserts, oligomerizes and functionalizes as a pore. We posit conceptualization of ceramide as a membrane-based stress calibrator, driving membrane macrodomain organization, which in mitochondria regulates intensity of Bax-induced MOMP, and is

  12. Integrating mitochondrial translation into the cellular context.

    PubMed

    Richter-Dennerlein, Ricarda; Dennerlein, Sven; Rehling, Peter

    2015-10-01

    Mitochondrial-encoded subunits of the oxidative phosphorylation system assemble with nuclear-encoded subunits into enzymatic complexes. Recent findings showed that mitochondrial translation is linked to other mitochondrial functions, as well as to cellular processes. The supply of mitochondrial-encoded proteins is coordinated by the coupling of mitochondrial protein synthesis with assembly of respiratory chain complexes. MicroRNAs imported from the cytoplasm into mitochondria were, surprisingly, found to act as regulators of mitochondrial translation. In turn, translation in mitochondria controls cellular proliferation, and mitochondrial ribosomal subunits contribute to the cytoplasmic stress response. Thus, translation in mitochondria is apparently integrated into cellular processes. PMID:26535422

  13. [Is glaucoma a mitochondrial neurodegenerative disease].

    PubMed

    Zhang, Z; Ma, J M; Wang, N L

    2016-09-11

    The retinal ganglion cell, due to peculiar structural and energetic constraints, appears acutely susceptible to mitochondrial dysfunction. Emerging evidence suggests that changes in the mitochondrial DNA(mtDNA)and in nuclear DNA genes that encode mitochondrial proteins may influence mitochondrial structure and function and, therefore, contribute to the pathogenesis of primary open angle glaucoma. As the main glaucoma risk factors are elevated intraocular pressure and older age, we discuss their relationship with mitochondrial dysfunction. If the contribution of mitochondrial dysfunction to glaucoma pathogenesis is further established, emerging therapies aiming to optimize mitochondrial function represent potential clinical treatments. (Chin J Ophthalmol, 2016, 52: 714-717). PMID:27647253

  14. The mitochondrial nucleoid: integrating mitochondrial DNA into cellular homeostasis.

    PubMed

    Gilkerson, Robert; Bravo, Liliana; Garcia, Iraselia; Gaytan, Norma; Herrera, Alan; Maldonado, Alicia; Quintanilla, Brandi

    2013-05-01

    The packaging of mitochondrial DNA (mtDNA) into DNA-protein assemblies called nucleoids provides an efficient segregating unit of mtDNA, coordinating mtDNA's involvement in cellular metabolism. From the early discovery of mtDNA as "extranuclear" genetic material, its organization into nucleoids and integration into both the mitochondrial organellar network and the cell at large via a variety of signal transduction pathways, mtDNA is a crucial component of the cell's homeostatic network. The mitochondrial nucleoid is composed of a set of DNA-binding core proteins involved in mtDNA maintenance and transcription, and a range of peripheral factors, which are components of signaling pathways controlling mitochondrial biogenesis, metabolism, apoptosis, and retrograde mitochondria-to-nucleus signaling. The molecular interactions of nucleoid components with the organellar network and cellular signaling pathways provide exciting clues to the dynamic integration of mtDNA into cellular metabolic homeostasis.

  15. Modulated voltage metastable ionization detector

    NASA Technical Reports Server (NTRS)

    Carle, G. C.; Kojiro, D. R.; Humphrey, D. E. (Inventor)

    1985-01-01

    The output current from a metastable ionization detector (MID) is applied to a modulation voltage circuit. An adjustment is made to balance out the background current, and an output current, above background, is applied to an input of a strip chart recorder. For low level concentrations, i.e., low detected output current, the ionization potential will be at a maximum and the metastable ionization detector will operate at its most sensitive level. When the detected current from the metastable ionization detector increases above a predetermined threshold level, a voltage control circuit is activated which turns on a high voltage transistor which acts to reduce the ionization potential. The ionization potential applied to the metastable ionization detector is then varied so as to maintain the detected signal level constant. The variation in ionization potential is now related to the concentration of the constituent and a representative amplitude is applied to another input of said strip chart recorder.

  16. Voltage-Gated Hydrophobic Nanopores

    SciTech Connect

    Lavrik, Nickolay V

    2011-01-01

    Hydrophobicity is a fundamental property that is responsible for numerous physical and biophysical aspects of molecular interactions in water. Peculiar behavior is expected for water in the vicinity of hydrophobic structures, such as nanopores. Indeed, hydrophobic nanopores can be found in two distinct states, dry and wet, even though the latter is thermodynamically unstable. Transitions between these two states are kinetically hindered in long pores but can be much faster in shorter pores. As it is demonstrated for the first time in this paper, these transitions can be induced by applying a voltage across a membrane with a single hydrophobic nanopore. Such voltage-induced gating in single nanopores can be realized in a reversible manner through electrowetting of inner walls of the nanopores. The resulting I-V curves of such artificial hydrophobic nanopores mimic biological voltage-gated channels.

  17. Loss of Prohibitin Membrane Scaffolds Impairs Mitochondrial Architecture and Leads to Tau Hyperphosphorylation and Neurodegeneration

    PubMed Central

    Merkwirth, Carsten; Morbin, Michela; Brönneke, Hella S.; Jordan, Sabine D.; Rugarli, Elena I.; Langer, Thomas

    2012-01-01

    Fusion and fission of mitochondria maintain the functional integrity of mitochondria and protect against neurodegeneration, but how mitochondrial dysfunctions trigger neuronal loss remains ill-defined. Prohibitins form large ring complexes in the inner membrane that are composed of PHB1 and PHB2 subunits and are thought to function as membrane scaffolds. In Caenorhabditis elegans, prohibitin genes affect aging by moderating fat metabolism and energy production. Knockdown experiments in mammalian cells link the function of prohibitins to membrane fusion, as they were found to stabilize the dynamin-like GTPase OPA1 (optic atrophy 1), which mediates mitochondrial inner membrane fusion and cristae morphogenesis. Mutations in OPA1 are associated with dominant optic atrophy characterized by the progressive loss of retinal ganglion cells, highlighting the importance of OPA1 function in neurons. Here, we show that neuron-specific inactivation of Phb2 in the mouse forebrain causes extensive neurodegeneration associated with behavioral impairments and cognitive deficiencies. We observe early onset tau hyperphosphorylation and filament formation in the hippocampus, demonstrating a direct link between mitochondrial defects and tau pathology. Loss of PHB2 impairs the stability of OPA1, affects mitochondrial ultrastructure, and induces the perinuclear clustering of mitochondria in hippocampal neurons. A destabilization of the mitochondrial genome and respiratory deficiencies manifest in aged neurons only, while the appearance of mitochondrial morphology defects correlates with tau hyperphosphorylation in the absence of PHB2. These results establish an essential role of prohibitin complexes for neuronal survival in vivo and demonstrate that OPA1 stability, mitochondrial fusion, and the maintenance of the mitochondrial genome in neurons depend on these scaffolding proteins. Moreover, our findings establish prohibitin-deficient mice as a novel genetic model for tau pathologies

  18. Ancillary service details: Voltage control

    SciTech Connect

    Kirby, B.; Hirst, E.

    1997-12-01

    Voltage control is accomplished by managing reactive power on an alternating-current power system. Reactive power can be produced and absorbed by both generation and transmission equipment. Reactive-power devices differ substantially in the magnitude and speed of response and in their capital costs. System operators, transmission owners, generators, customers, power marketers, and government regulators need to pay close attention to voltage control as they restructure the U.S. electricity industry. Voltage control can affect reliability and commerce in three ways: (1) Voltages must be maintained within an acceptable range for both customer and power-system equipment to function properly. (2) The movement of reactive power consumes transmission resources, which limits the ability to move real power and worsens congestion. (3) The movement of reactive power results in real-power losses. When generators are required to supply excessive amounts of reactive power, their real-power production must be curtailed. These opportunity costs are not currently compensated for in most regions. Current tariffs are based on embedded costs. These embedded-cost tariffs average about $0.51/MWh, equivalent to $1.5 billion annually for the United States as a whole. Although this cost is low when compared with the cost of energy, it still aggregates to a significant amount of money. This report takes a basic look at why the power system requires reactive power (an appendix explains the fundamentals of real and reactive power). The report then examines the various types of generation and transmission resources used to supply reactive power and to control voltage. Finally it discusses how these resources are deployed and paid for in several reliability regions around the country. As the U.S. electricity industry is restructured, the generation, transmission, and system-control equipment and functions that maintain voltages within the appropriate ranges are being deintegrated.

  19. A low voltage CMOS low drop-out voltage regulator

    NASA Astrophysics Data System (ADS)

    Bakr, Salma Ali; Abbasi, Tanvir Ahmad; Abbasi, Mohammas Suhaib; Aldessouky, Mohamed Samir; Abbasi, Mohammad Usaid

    2009-05-01

    A low voltage implementation of a CMOS Low Drop-Out voltage regulator (LDO) is presented. The requirement of low voltage devices is crucial for portable devices that require extensive computations in a low power environment. The LDO is implemented in 90nm generic CMOS technology. It generates a fixed 0.8V from a 2.5V supply which on discharging goes to 1V. The buffer stage used is unity gain configured unbuffered OpAmp with rail-to-rail swing input stage. The simulation result shows that the implemented circuit provides load regulation of 0.004%/mA and line regulation of -11.09mV/V. The LDO provides full load transient response with a settling time of 5.2μs. Further, the dropout voltage is 200mV and the quiescent current through the pass transistor (Iload=0) is 20μA. The total power consumption of this LDO (excluding bandgap reference) is only 80μW.

  20. Caffeine increases mitochondrial function and blocks melatonin signaling to mitochondria in Alzheimer's mice and cells.

    PubMed

    Dragicevic, Natasa; Delic, Vedad; Cao, Chuanhai; Copes, Neil; Lin, Xiaoyang; Mamcarz, Maggie; Wang, Li; Arendash, Gary W; Bradshaw, Patrick C

    2012-12-01

    Caffeine and melatonin have been shown to protect the Swedish mutant amyloid precursor protein (APP(sw)) transgenic mouse model of Alzheimer's disease from cognitive dysfunction. But their mechanisms of action remain incompletely understood. These Alzheimer's mice have extensive mitochondrial dysfunction, which likely contributes to their cognitive decline. To further explore the mechanism through which caffeine and melatonin protect cognitive function in these mice, we monitored the function of isolated mitochondria from APP(sw) mice treated with caffeine, melatonin, or both in their drinking water for one month. Melatonin treatment yielded a near complete restoration of mitochondrial function in assays of respiratory rate, membrane potential, reactive oxygen species production, and ATP levels. Caffeine treatment by itself yielded a small increase in mitochondrial function. However, caffeine largely blocked the large enhancement of mitochondrial function provided by melatonin. Studies with N2a neuroblastoma cells stably expressing APP(sw) showed that specific inhibition of cAMP-dependent phosphodiesterase (PDE) 4 or cGMP-dependent PDE5 also blocked melatonin protection of mitochondrial function, but A(2a) and A₁ adenosine receptor antagonists were without effect. Melatonin or caffeine at the concentrations used to modulate mitochondrial function in the cells had no effect on cAMP-dependent PDE activity or cellular cAMP or cGMP levels. Therefore, caffeine and increased cyclic nucleotide levels likely block melatonin signaling to mitochondria by independent mechanisms that do not involve adenosine receptor antagonism. The results of this study indicate that melatonin restores mitochondrial function much more potently than caffeine in APP(sw) transgenic mouse and cell models of Alzheimer's disease.

  1. Low Voltage Spatial Light Modulator

    SciTech Connect

    Papavasiliou, A

    2003-02-19

    This project studied the feasibility of a Low-Voltage actuator technology that promises to reduce the switched voltage requirements and linearize the response of spatial light modulators. We created computer models that demonstrate substantial advantages offered by this technology, and fabricated and tested those devices. SLMs are electro-optic devices for modulating the phase, amplitude or angle of light beams, laser or other. Applications for arrays of SLMs include turbulence correction for high-speed optical communications, imaging through distorting media, input devices for holographic memories, optical manipulation of DNA molecules, and optical computers. Devices based on micro electro-mechanical systems (MEMS) technology have recently become of special interest because of their potential for greatly improved performance at a much lower cost than piezoelectric or liquid crystal based devices. The new MEMS-based SLM devices could have important applications in high-speed optical communication and remote optical sensing, in support of DoD and DOE missions. Virtually all previously demonstrated MEMS SLMs are based on parallel-plate capacitors where an applied voltage causes a mirror attached to a suspended electrode to move towards a fixed electrode. They require relatively high voltages, typically on the order of 100 V, resulting in (1) large transistor sizes, available only from specialized foundries at significant cost and limiting the amount/sophistication of electronics under each SLM pixel, and (2) large power dissipation/area, resulting in a heat removal issue because of the optical precision required ({approx} 1/50-th of a wavelength). The actuator described in this process uses an advanced geometry that was invented at LLNL and is currently still proprietary. The new geometry allows the application of a bias voltage. This applied bias voltage results in a reduction of the required switched voltage and a linearization of the response curve. When this

  2. Mitochondrial DNA-deficient models and aging.

    PubMed

    Olgun, Abdullah; Akman, Serif

    2007-04-01

    Human mitochondrial DNA (mtDNA) encodes 13 subunits of oxidative phosphorylation (OXPHOS) enzyme complexes I, III, IV, and V except complex II. MtDNA is more sensitive to oxidative damage than nuclear DNA. MtDNA defects are involved in many pathologies including aging. Several mtDNA-deficient cell culture, yeast, and animal models were generated to study the role of mtDNA in many physiological processes. Ethidium bromide (EB), an agent that is known to inhibit mtDNA replication with a negligible effect on nuclear DNA, is generally used to generate mtDNA-deficient models. The antibiotics chloramphenicol and doxycycline, which were known to inhibit mitochondrial translation, were also used to generate the same phenotype. Cultured mtDNA-deficient cells need uridine and pyruvate to survive. At the organismal level, uridine can be supplemented, but pyruvate supplementation can cause a worser phenotype because of lactic acidosis. In C. elegans, EB, when used during larval development, increases life span, but decreases, when used after the beginning of adult stage. This should be kept in mind since mitochondria-related genes are generally detected in genome-wide screening studies for longevity. We believe that conditional knockout studies need to be carried out for these genes after reaching adulthood. MtDNA mutator mouse did not show an increase of free radical production. Therefore, the downstream phenomena to mtDNA defects are likely ineffective pyrimidine synthesis (dihydroorotate dehydrogenase, DHODH, needs a functional respiratory chain) and excess NADH (decreased NAD pool) in addition to free radicals. PMID:17460185

  3. The clinical maze of mitochondrial neurology

    PubMed Central

    DiMauro, Salvatore; Schon, Eric A.; Carelli, Valerio; Hirano, Michio

    2014-01-01

    Mitochondrial diseases involve the respiratory chain, which is under the dual control of nuclear and mitochondrial DNA (mtDNA). The complexity of mitochondrial genetics provides one explanation for the clinical heterogeneity of mitochondrial diseases, but our understanding of disease pathogenesis remains limited. Classification of Mendelian mitochondrial encephalomyopathies has been laborious, but whole-exome sequencing studies have revealed unexpected molecular aetiologies for both typical and atypical mitochondrial disease phenotypes. Mendelian mitochondrial defects can affect five components of mitochondrial biology: subunits of respiratory chain complexes (direct hits); mitochondrial assembly proteins; mtDNA translation; phospholipid composition of the inner mitochondrial membrane; or mitochondrial dynamics. A sixth category—defects of mtDNA maintenance—combines features of Mendelian and mitochondrial genetics. Genetic defects in mitochondrial dynamics are especially important in neurology as they cause optic atrophy, hereditary spastic paraplegia, and Charcot–Marie–Tooth disease. Therapy is inadequate and mostly palliative, but promising new avenues are being identified. Here, we review current knowledge on the genetics and pathogenesis of the six categories of mitochondrial disorders outlined above, focusing on their salient clinical manifestations and highlighting novel clinical entities. An outline of diagnostic clues for the various forms of mitochondrial disease, as well as potential therapeutic strategies, is also discussed. PMID:23835535

  4. Bioenergetic roles of mitochondrial fusion.

    PubMed

    Silva Ramos, Eduardo; Larsson, Nils-Göran; Mourier, Arnaud

    2016-08-01

    Mitochondria are bioenergetic hotspots, producing the bulk of ATP by the oxidative phosphorylation process. Mitochondria are also structurally dynamic and undergo coordinated fusion and fission to maintain their function. Recent studies of the mitochondrial fusion machinery have provided new evidence in detailing their role in mitochondrial metabolism. Remarkably, mitofusin 2, in addition to its role in fusion, is important for maintaining coenzyme Q levels and may be an integral player in the mevalonate synthesis pathway. Here, we review the bioenergetic roles of mitochondrial dynamics and emphasize the importance of the in vitro growth conditions when evaluating mitochondrial respiration. This article is part of a Special Issue entitled 'EBEC 2016: 19th European Bioenergetics Conference, Riva del Garda, Italy, July 2-6, 2016,' edited by Prof. Paolo Bernardi. PMID:27060252

  5. Pathological Significance of Mitochondrial Glycation

    PubMed Central

    Pun, Pamela Boon Li; Murphy, Michael P.

    2012-01-01

    Glycation, the nonenzymatic glycosylation of biomolecules, is commonly observed in diabetes and ageing. Reactive dicarbonyl species such as methylglyoxal and glyoxal are thought to be major physiological precursors of glycation. Because these dicarbonyls tend to be formed intracellularly, the levels of advanced glycation end products on cellular proteins are higher than on extracellular ones. The formation of glycation adducts within cells can have severe functional consequences such as inhibition of protein activity and promotion of DNA mutations. Although several lines of evidence suggest that there are specific mitochondrial targets of glycation, and mitochondrial dysfunction itself has been implicated in disease and ageing, it is unclear if glycation of biomolecules specifically within mitochondria induces dysfunction and contributes to disease pathology. We discuss here the possibility that mitochondrial glycation contributes to disease, focussing on diabetes, ageing, cancer, and neurodegeneration, and highlight the current limitations in our understanding of the pathological significance of mitochondrial glycation. PMID:22778743

  6. Incrementally Variable High-Voltage Supply

    NASA Technical Reports Server (NTRS)

    Potter, D. W.; Chin, J.; Anderson, H. R.; Loveless, R. L.

    1985-01-01

    Programable power supply provides regulated output ranging from 2.5 to 2,500 volts. Exponential digital-to-analog converter provides low-voltage analog signal to power converter and to negative and positive high-voltage regulators. In response, converter furnishes voltage of approximate magnitude represented by analog signal, and regulators adjust voltage to precise magnitude. Entire voltage range covered in 169 steps. Total power consumption expected to be less than 2 watts.

  7. To unite or divide: mitochondrial dynamics in the murine outer retina that preceded age related photoreceptor loss

    PubMed Central

    Kam, Jaimie Hoh; Jeffery, Glen

    2015-01-01

    Mitochondrial function declines with age and is associated with age-related disorders and cell death. In the retina this is critical as photoreceptor energy demands are the greatest in the body and aged cell loss large (~30%). But mitochondria can fuse or divide to accommodate changing demands. We explore ageing mitochondrial dynamics in young (1 month) and old (12 months) mouse retina, investigating changes in mitochondrial fission (Fis1) and fusion (Opa1) proteins, cytochrome C oxidase (COX III), which reflects mitochondrial metabolic status, and heat shock protein 60 (Hsp60) that is a mitochondrial chaperon for protein folding. Western blots showed each protein declined with age. However, within this, immunostaining revealed increases of around 50% in Fis1 and Opa1 in photoreceptor inner segments (IS). Electron microscope analysis revealed mitochondrial fragmentation with age and marked changes in morphology in IS, consistent with elevated dynamics. COX III declined by approximately 30% in IS, but Hsp60 reductions were around 80% in the outer plexiform layer. Our results are consistent with declining mitochondrial metabolism. But also with increased photoreceptor mitochondrial dynamics that differ from other retinal regions, perhaps reflecting attempts to maintain function. These changes are the platform for age related photoreceptor loss initiated after 12 months. PMID:26393878

  8. Slow mitochondrial repair of 5'-AMP renders mtDNA susceptible to damage in APTX deficient cells.

    PubMed

    Akbari, Mansour; Sykora, Peter; Bohr, Vilhelm A

    2015-08-10

    Aborted DNA ligation events in eukaryotic cells can generate 5'-adenylated (5'-AMP) DNA termini that can be removed from DNA by aprataxin (APTX). Mutations in APTX cause an inherited human disease syndrome characterized by early-onset progressive ataxia with ocular motor apraxia (AOA1). APTX is found in the nuclei and mitochondria of eukaryotic cells. Depletion of APTX causes mitochondrial dysfunction and renders the mitochondrial genome, but not the nuclear genome susceptible to damage. The biochemical processes that link APTX deficiency to mitochondrial dysfunction have not been well elucidated. Here, we monitored the repair of 5'-AMP DNA damage in nuclear and mitochondrial extracts from human APTX(+/+) and APTX(-/-) cells. The efficiency of repair of 5'-AMP DNA was much lower in mitochondrial than in nuclear protein extracts, and resulted in persistent DNA repair intermediates in APTX deficient cells. Moreover, the removal of 5'-AMP from DNA was significantly slower in the mitochondrial extracts from human cell lines and mouse tissues compared with their corresponding nuclear extracts. These results suggest that, contrary to nuclear DNA repair, mitochondrial DNA repair is not able to compensate for APTX deficiency resulting in the accumulation of mitochondrial DNA damage.

  9. Slow mitochondrial repair of 5'-AMP renders mtDNA susceptible to damage in APTX deficient cells.

    PubMed

    Akbari, Mansour; Sykora, Peter; Bohr, Vilhelm A

    2015-01-01

    Aborted DNA ligation events in eukaryotic cells can generate 5'-adenylated (5'-AMP) DNA termini that can be removed from DNA by aprataxin (APTX). Mutations in APTX cause an inherited human disease syndrome characterized by early-onset progressive ataxia with ocular motor apraxia (AOA1). APTX is found in the nuclei and mitochondria of eukaryotic cells. Depletion of APTX causes mitochondrial dysfunction and renders the mitochondrial genome, but not the nuclear genome susceptible to damage. The biochemical processes that link APTX deficiency to mitochondrial dysfunction have not been well elucidated. Here, we monitored the repair of 5'-AMP DNA damage in nuclear and mitochondrial extracts from human APTX(+/+) and APTX(-/-) cells. The efficiency of repair of 5'-AMP DNA was much lower in mitochondrial than in nuclear protein extracts, and resulted in persistent DNA repair intermediates in APTX deficient cells. Moreover, the removal of 5'-AMP from DNA was significantly slower in the mitochondrial extracts from human cell lines and mouse tissues compared with their corresponding nuclear extracts. These results suggest that, contrary to nuclear DNA repair, mitochondrial DNA repair is not able to compensate for APTX deficiency resulting in the accumulation of mitochondrial DNA damage. PMID:26256098

  10. Mitochondrial control of nuclear apoptosis

    PubMed Central

    1996-01-01

    Anucleate cells can be induced to undergo programmed cell death (PCD), indicating the existence of a cytoplasmic PCD pathway that functions independently from the nucleus. Cytoplasmic structures including mitochondria have been shown to participate in the control of apoptotic nuclear disintegration. Before cells exhibit common signs of nuclear apoptosis (chromatin condensation and endonuclease-mediated DNA fragmentation), they undergo a reduction of the mitochondrial transmembrane potential (delta psi m) that may be due to the opening of mitochondrial permeability transition (PT) pores. Here, we present direct evidence indicating that mitochondrial PT constitutes a critical early event of the apoptotic process. In a cell-free system combining purified mitochondria and nuclei, mitochondria undergoing PT suffice to induce chromatin condensation and DNA fragmentation. Induction of PT by pharmacological agents augments the apoptosis-inducing potential of mitochondria. In contrast, prevention of PT by pharmacological agents impedes nuclear apoptosis, both in vitro and in vivo. Mitochondria from hepatocytes or lymphoid cells undergoing apoptosis, but not those from normal cells, induce disintegration of isolated Hela nuclei. A specific ligand of the mitochondrial adenine nucleotide translocator (ANT), bongkreik acid, inhibits PT and reduces apoptosis induction by mitochondria in a cell-free system. Moreover, it inhibits the induction of apoptosis in intact cells. Several pieces of evidence suggest that the proto-oncogene product Bcl-2 inhibits apoptosis by preventing mitochondrial PT. First, to inhibit nuclear apoptosis, Bcl-2 must be localized in mitochondrial but not nuclear membranes. Second, transfection-enforced hyperexpression of Bcl-2 directly abolishes the induction of mitochondrial PT in response to a protonophore, a pro- oxidant, as well as to the ANT ligand atractyloside, correlating with its apoptosis-inhibitory effect. In conclusion, mitochondrial PT appears

  11. Mitochondrial division in Caenorhabditis elegans.

    PubMed

    Gandre, Shilpa; van der Bliek, Alexander M

    2007-01-01

    The study of mitochondrial division proteins has largely focused on yeast and mammalian cells. We describe methods to use Caenorhabditis elegans as an alternative model for studying mitochondrial division, taking advantage of the many wonderful resources provided by the C. elegans community. Our methods are largely based on manipulation of gene expression using classic and molecular genetic techniques combined with fluorescence microscopy. Some biochemical methods are also included. As antibodies become available, these biochemical methods are likely to become more sophisticated. PMID:18314747

  12. Mitochondrial dysfunction and organophosphorus compounds

    SciTech Connect

    Karami-Mohajeri, Somayyeh; Abdollahi, Mohammad

    2013-07-01

    Organophosphorous (OPs) pesticides are the most widely used pesticides in the agriculture and home. However, many acute or chronic poisoning reports about OPs have been published in the recent years. Mitochondria as a site of cellular oxygen consumption and energy production can be a target for OPs poisoning as a non-cholinergic mechanism of toxicity of OPs. In the present review, we have reviewed and criticized all the evidences about the mitochondrial dysfunctions as a mechanism of toxicity of OPs. For this purpose, all biochemical, molecular, and morphological data were retrieved from various studies. Some toxicities of OPs are arisen from dysfunction of mitochondrial oxidative phosphorylation through alteration of complexes I, II, III, IV and V activities and disruption of mitochondrial membrane. Reductions of adenosine triphosphate (ATP) synthesis or induction of its hydrolysis can impair the cellular energy. The OPs disrupt cellular and mitochondrial antioxidant defense, reactive oxygen species generation, and calcium uptake and promote oxidative and genotoxic damage triggering cell death via cytochrome C released from mitochondria and consequent activation of caspases. The mitochondrial dysfunction induced by OPs can be restored by use of antioxidants such as vitamin E and C, alpha-tocopherol, electron donors, and through increasing the cytosolic ATP level. However, to elucidate many aspect of mitochondrial toxicity of Ops, further studies should be performed. - Highlights: • As a non-cholinergic mechanism of toxicity, mitochondria is a target for OPs. • OPs affect action of complexes I, II, III, IV and V in the mitochondria. • OPs reduce mitochondrial ATP. • OPs promote oxidative and genotoxic damage via release of cytochrome C from mitochondria. • OP-induced mitochondrial dysfunction can be restored by increasing the cytosolic ATP.

  13. Mitochondrial efficiency and insulin resistance.

    PubMed

    Crescenzo, Raffaella; Bianco, Francesca; Mazzoli, Arianna; Giacco, Antonia; Liverini, Giovanna; Iossa, Susanna

    2014-01-01

    Insulin resistance, "a relative impairment in the ability of insulin to exert its effects on glucose, protein and lipid metabolism in target tissues," has many detrimental effects on metabolism and is strongly correlated to deposition of lipids in non-adipose tissues. Mitochondria are the main cellular sites devoted to ATP production and fatty acid oxidation. Therefore, a role for mitochondrial dysfunction in the onset of skeletal muscle insulin resistance has been proposed and many studies have dealt with possible alteration in mitochondrial function in obesity and diabetes, both in humans and animal models. Data reporting evidence of mitochondrial dysfunction in type two diabetes mellitus are numerous, even though the issue that this reduced mitochondrial function is causal in the development of the disease is not yet solved, also because a variety of parameters have been used in the studies carried out on this subject. By assessing the alterations in mitochondrial efficiency as well as the impact of this parameter on metabolic homeostasis of skeletal muscle cells, we have obtained results that allow us to suggest that an increase in mitochondrial efficiency precedes and therefore can contribute to the development of high-fat-induced insulin resistance in skeletal muscle. PMID:25601841

  14. Mitochondrial Epigenetics and Environmental Exposure.

    PubMed

    Lambertini, Luca; Byun, Hyang-Min

    2016-09-01

    The rising toll of chronic and debilitating diseases brought about by the exposure to an ever expanding number of environmental pollutants and socio-economic factors is calling for action. The understanding of the molecular mechanisms behind the effects of environmental exposures can lead to the development of biomarkers that can support the public health fields of both early diagnosis and intervention to limit the burden of environmental diseases. The study of mitochondrial epigenetics carries high hopes to provide important biomarkers of exposure and disease. Mitochondria are in fact on the frontline of the cellular response to the environment. Modifications of the epigenetic factors regulating the mitochondrial activity are emerging as informative tools that can effectively report on the effects of the environment on the phenotype. Here, we will discuss the emerging field of mitochondrial epigenetics. This review describes the main epigenetic phenomena that modify the activity of the mitochondrial DNA including DNA methylation, long and short non-coding RNAs. We will discuss the unique pattern of mitochondrial DNA methylation, describe the challenges of correctly measuring it, and report on the existing studies that have analysed the correlation between environmental exposures and mitochondrial DNA methylation. Finally, we provide a brief account of the therapeutic approaches targeting mitochondria currently under consideration.

  15. Mitochondrial dysfunction in Parkinson's disease.

    PubMed

    Hu, Qingsong; Wang, Guanghui

    2016-01-01

    Parkinson's disease (PD) is the second most common neurodegenerative disease, which is characterized by loss of dopaminergic (DA) neurons in the substantia nigra pars compacta and the formation of Lewy bodies and Lewy neurites in surviving DA neurons in most cases. Although the cause of PD is still unclear, the remarkable advances have been made in understanding the possible causative mechanisms of PD pathogenesis. Numerous studies showed that dysfunction of mitochondria may play key roles in DA neuronal loss. Both genetic and environmental factors that are associated with PD contribute to mitochondrial dysfunction and PD pathogenesis. The induction of PD by neurotoxins that inhibit mitochondrial complex I provides direct evidence linking mitochondrial dysfunction to PD. Decrease of mitochondrial complex I activity is present in PD brain and in neurotoxin- or genetic factor-induced PD cellular and animal models. Moreover, PINK1 and parkin, two autosomal recessive PD gene products, have important roles in mitophagy, a cellular process to clear damaged mitochondria. PINK1 activates parkin to ubiquitinate outer mitochondrial membrane proteins to induce a selective degradation of damaged mitochondria by autophagy. In this review, we summarize the factors associated with PD and recent advances in understanding mitochondrial dysfunction in PD. PMID:27453777

  16. CFTR activity and mitochondrial function☆

    PubMed Central

    Valdivieso, Angel Gabriel; Santa-Coloma, Tomás A.

    2013-01-01

    Cystic Fibrosis (CF) is a frequent and lethal autosomal recessive disease, caused by mutations in the gene encoding the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR). Before the discovery of the CFTR gene, several hypotheses attempted to explain the etiology of this disease, including the possible role of a chloride channel, diverse alterations in mitochondrial functions, the overexpression of the lysosomal enzyme α-glucosidase and a deficiency in the cytosolic enzyme glucose 6-phosphate dehydrogenase. Because of the diverse mitochondrial changes found, some authors proposed that the affected gene should codify for a mitochondrial protein. Later, the CFTR cloning and the demonstration of its chloride channel activity turned the mitochondrial, lysosomal and cytosolic hypotheses obsolete. However, in recent years, using new approaches, several investigators reported similar or new alterations of mitochondrial functions in Cystic Fibrosis, thus rediscovering a possible role of mitochondria in this disease. Here, we review these CFTR-driven mitochondrial defects, including differential gene expression, alterations in oxidative phosphorylation, calcium homeostasis, oxidative stress, apoptosis and innate immune response, which might explain some characteristics of the complex CF phenotype and reveals potential new targets for therapy. PMID:24024153

  17. Mitochondrial Metabolism in Aging Heart.

    PubMed

    Lesnefsky, Edward J; Chen, Qun; Hoppel, Charles L

    2016-05-13

    Altered mitochondrial metabolism is the underlying basis for the increased sensitivity in the aged heart to stress. The aged heart exhibits impaired metabolic flexibility, with a decreased capacity to oxidize fatty acids and enhanced dependence on glucose metabolism. Aging impairs mitochondrial oxidative phosphorylation, with a greater role played by the mitochondria located between the myofibrils, the interfibrillar mitochondria. With aging, there is a decrease in activity of complexes III and IV, which account for the decrease in respiration. Furthermore, aging decreases mitochondrial content among the myofibrils. The end result is that in the interfibrillar area, there is ≈50% decrease in mitochondrial function, affecting all substrates. The defective mitochondria persist in the aged heart, leading to enhanced oxidant production and oxidative injury and the activation of oxidant signaling for cell death. Aging defects in mitochondria represent new therapeutic targets, whether by manipulation of the mitochondrial proteome, modulation of electron transport, activation of biogenesis or mitophagy, or the regulation of mitochondrial fission and fusion. These mechanisms provide new ways to attenuate cardiac disease in elders by preemptive treatment of age-related defects, in contrast to the treatment of disease-induced dysfunction. PMID:27174952

  18. Voltage control of ferromagnetic resonance

    NASA Astrophysics Data System (ADS)

    Zhou, Ziyao; Peng, Bin; Zhu, Mingmin; Liu, Ming

    2016-05-01

    Voltage control of magnetism in multiferroics, where the ferromagnetism and ferroelectricity are simultaneously exhibiting, is of great importance to achieve compact, fast and energy efficient voltage controllable magnetic/microwave devices. Particularly, these devices are widely used in radar, aircraft, cell phones and satellites, where volume, response time and energy consumption is critical. Researchers realized electric field tuning of magnetic properties like magnetization, magnetic anisotropy and permeability in varied multiferroic heterostructures such as bulk, thin films and nanostructure by different magnetoelectric (ME) coupling mechanism: strain/stress, interfacial charge, spin-electromagnetic (EM) coupling and exchange coupling, etc. In this review, we focus on voltage control of ferromagnetic resonance (FMR) in multiferroics. ME coupling-induced FMR change is critical in microwave devices, where the electric field tuning of magnetic effective anisotropic field determines the tunability of the performance of microwave devices. Experimentally, FMR measurement technique is also an important method to determine the small effective magnetic field change in small amount of magnetic material precisely due to its high sensitivity and to reveal the deep science of multiferroics, especially, voltage control of magnetism in novel mechanisms like interfacial charge, spin-EM coupling and exchange coupling.

  19. High voltage MOSFET switching circuit

    DOEpatents

    McEwan, T.E.

    1994-07-26

    The problem of source lead inductance in a MOSFET switching circuit is compensated for by adding an inductor to the gate circuit. The gate circuit inductor produces an inductive spike which counters the source lead inductive drop to produce a rectangular drive voltage waveform at the internal gate-source terminals of the MOSFET. 2 figs.

  20. High Voltage Space Solar Arrays

    NASA Technical Reports Server (NTRS)

    Ferguson, D. C.; Hillard, G. B.; Vayner, B. V.; Galofaro, J. T.; Lyons, Valerie (Technical Monitor)

    2002-01-01

    Recent tests performed at the NASA Glenn Research Center and elsewhere have shown promise in the design and construction of high voltage (300-1000 V) solar arrays for space applications. Preliminary results and implications for solar array design will be discussed, with application to direct-drive electric propulsion and space solar power.

  1. High voltage MOSFET switching circuit

    DOEpatents

    McEwan, Thomas E.

    1994-01-01

    The problem of source lead inductance in a MOSFET switching circuit is compensated for by adding an inductor to the gate circuit. The gate circuit inductor produces an inductive spike which counters the source lead inductive drop to produce a rectangular drive voltage waveform at the internal gate-source terminals of the MOSFET.

  2. LHCb calorimeters high voltage system

    NASA Astrophysics Data System (ADS)

    Gilitsky, Yu.; Golutvin, A.; Konoplyannikov, A.; Lefrancois, J.; Perret, P.; Schopper, A.; Soldatov, M.; Yakimchuk, V.

    2007-02-01

    The calorimeter system in LHCb aims to identify electrons, photons and hadrons. All calorimeters are equipped with Hamamatsu photo tubes as devices for light to signal conversion. Eight thousand R7899-20 tubes are used for electromagnetic and hadronic calorimeters and two hundred 64 channels multi-anode R7600-00-M64 for Scintillator-Pad/Preshower detectors. The calorimeter high voltage (HV) system is based on a Cockroft Walton (CW) voltage converter and a control board connected to the Experiment Control System (ECS) by serial bus. The base of each photomultiplier tube (PMT) is built with a high voltage converter and constructed on an individual printed circuit board, using compact surface mount components. The base is attached directly to the PMT. There are no HV cables in the system. A Field Programmable Gate Array (FPGA) is used on the control board as an interface between the ECS and the 200 control channels. The FPGA includes also additional functionalities allowing automated monitoring and ramp up of the high voltage values. This paper describes the HV system architecture, some technical details of the electronics implementation and summarizes the system performance. This safe and low power consumption HV electronic system for the photomultiplier tubes can be used for various biomedical apparatus too.

  3. High-Voltage Droplet Dispenser

    NASA Technical Reports Server (NTRS)

    Eichenberg, Dennis J.

    2003-01-01

    An apparatus that is extremely effective in dispensing a wide range of droplets has been developed. This droplet dispenser is unique in that it utilizes a droplet bias voltage, as well as an ionization pulse, to release a droplet. Apparatuses that deploy individual droplets have been used in many applications, including, notably, study of combustion of liquid fuels. Experiments on isolated droplets are useful in that they enable the study of droplet phenomena under well-controlled and simplified conditions. In this apparatus, a syringe dispenses a known value of liquid, which emerges from, and hangs onto, the outer end of a flat-tipped, stainless steel needle. Somewhat below the needle tip and droplet is a ring electrode. A bias high voltage, followed by a high-voltage pulse, is applied so as to attract the droplet sufficiently to pull it off the needle. The voltages are such that the droplet and needle are negatively charged and the ring electrode is positively charged.

  4. Stimulatory effect of CSE-generated H2S on hepatic mitochondrial biogenesis and the underlying mechanisms.

    PubMed

    Untereiner, Ashley A; Fu, Ming; Módis, Katalin; Wang, Rui; Ju, YoungJun; Wu, Lingyun

    2016-08-31

    We previously showed that hydrogen sulfide (H2S) upregulates peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α in primary hepatocytes. PGC-1α is a crucial regulator of mitochondrial biogenesis, a process required to maintain cellular energy homeostasis. We investigated the regulation of hepatic mitochondrial biogenesis by cystathionine γ-lyase (CSE)-generated H2S under physiological conditions. Primary hepatocytes isolated from CSE knockout (KO) and wild-type (WT) mice were used in all experiments. Mitochondrial DNA (mtDNA) and mRNA levels were measured via real-time PCR. Protein S-sulfhydration was determined via a modified biotin switch assay. MitoTracker Green was used to quantify mitochondrial content and distribution. CSE-KO hepatocytes produced less mtDNA compared to WT hepatocytes. Mitochondrial content was reduced in CSE-KO hepatocytes compared to WT hepatocytes, which was restored with NaHS (an H2S donor) treatment. CSE-KO hepatocytes exhibited lower levels of mitochondrial transcription factors and the mitochondrial transcription coactivator, peroxisome proliferator-activated receptor-γ coactivator-related protein (PPRC) compared to WT hepatocytes. NaHS administration upregulated PPRC, yet downregulated PGC-1β protein level in mouse hepatocytes. Exogenous H2S induced the S-sulfhydration of PPRC, which was lower in untreated CSE-KO hepatocytes, but not that of PGC-1β. Finally, knockdown of either PGC-1α or PPRC significantly decreased NaHS-stimulated mitochondrial biogenesis in hepatocytes, where knockdown of both genes were required to abolish NaHS-induced mitochondrial biogenesis. Endogenous H2S-induced liver mitochondrial biogenesis is dependent upon PGC-1α and PPRC signaling in primary hepatocytes. This study may offer clues to the regulation of energy homeostasis under physiological conditions as well as mitochondrial dysregulation. PMID:27364855

  5. Krüppel-like factor 6 regulates mitochondrial function in the kidney

    PubMed Central

    Mallipattu, Sandeep K.; Horne, Sylvia J.; D’Agati, Vivette; Narla, Goutham; Liu, Ruijie; Frohman, Michael A.; Dickman, Kathleen; Chen, Edward Y.; Ma’ayan, Avi; Bialkowska, Agnieszka B.; Ghaleb, Amr M.; Nandan, Mandayam O.; Jain, Mukesh K.; Daehn, Ilse; Chuang, Peter Y.; Yang, Vincent W.; He, John C.

    2015-01-01

    Maintenance of mitochondrial structure and function is critical for preventing podocyte apoptosis and eventual glomerulosclerosis in the kidney; however, the transcription factors that regulate mitochondrial function in podocyte injury remain to be identified. Here, we identified Krüppel-like factor 6 (KLF6), a zinc finger domain transcription factor, as an essential regulator of mitochondrial function in podocyte apoptosis. We observed that podocyte-specific deletion of Klf6 increased the susceptibility of a resistant mouse strain to adriamycin-induced (ADR-induced) focal segmental glomerulosclerosis (FSGS). KLF6 expression was induced early in response to ADR in mice and cultured human podocytes, and prevented mitochondrial dysfunction and activation of intrinsic apoptotic pathways in these podocytes. Promoter analysis and chromatin immunoprecipitation studies revealed that putative KLF6 transcriptional binding sites are present in the promoter of the mitochondrial cytochrome c oxidase assembly gene (SCO2), which is critical for preventing cytochrome c release and activation of the intrinsic apoptotic pathway. Additionally, KLF6 expression was reduced in podocytes from HIV-1 transgenic mice as well as in renal biopsies from patients with HIV-associated nephropathy (HIVAN) and FSGS. Together, these findings indicate that KLF6-dependent regulation of the cytochrome c oxidase assembly gene is critical for maintaining mitochondrial function and preventing podocyte apoptosis. PMID:25689250

  6. The Mitochondrial Fission Regulator DRP3B Does Not Regulate Cell Death in Plants

    PubMed Central

    YOSHINAGA, KEIKO; FUJIMOTO, MASARU; ARIMURA, SHIN-ICHI; TSUTSUMI, NOBUHIRO; UCHIMIYA, HIROFUMI; KAWAI-YAMADA, MAKI

    2006-01-01

    • Background and Aims Recent reports have described dramatic alterations in mitochondrial morphology during metazoan apoptosis. A dynamin-related protein (DRP) associated with mitochondrial outer membrane fission is known to be involved in the regulation of apoptosis. This study analysed the relationship between mitochondrial fission and regulation of plant cell death. • Methods Transgenic plants were generated possessing Arabidopsis DRP3B (K56A), the dominant-negative form of Arabidopsis DRP, mitochondrial-targeted green fluorescent protein and mouse Bax. • Key Results Arabidopsis plants over-expressing DRP3B (K56A) exhibited long tubular mitochondria. In these plants, mitochondria appeared as a string-of-beads during cell death. This indicates that DRP3B (K56A) prevented mitochondrial fission during plant cell death. However, in contrast to results for mammalian cells and yeast, Bax-induced cell death was not inhibited in DRP3B (K56A)-expressing plant cells. Similarly, hydrogen peroxide-, menadione-, darkness- and salicylic acid-induced cell death was not inhibited by DRP3B (K56A) expression. • Conclusions These results indicate that the systems controlling cell death in animals and plants are not common in terms of mitochondrial fission. PMID:16533833

  7. Mitochondrial stress engages E2F1 apoptotic signaling to cause deafness

    PubMed Central

    Raimundo, Nuno; Song, Lei; Shutt, Timothy E.; McKay, Sharen E.; Cotney, Justin; Guan, Min-Xin; Gilliland, Thomas C.; Hohuan, David; Santos-Sacchi, Joseph; Shadel, Gerald S.

    2012-01-01

    SUMMARY Mitochondrial dysfunction causes poorly understood tissue-specific pathology stemming from primary defects in respiration, coupled with altered reactive oxygen species (ROS), metabolic signaling and apoptosis. The A1555G mtDNA mutation that causes maternally inherited deafness disrupts mitochondrial ribosome function, in part, via increased methylation of the mitochondrial 12S rRNA by the methyltransferase mtTFB1. In patient-derived A1555G cells, we show that 12S rRNA hyper-methylation causes ROS-dependent activation of AMP kinase and the pro-apoptotic nuclear transcription factor E2F1. This retrograde mitochondrial-stress relay is operative in vivo as transgenic-mtTFB1 mice exhibit enhanced 12S rRNA methylation in multiple tissues, increased E2F1 and apoptosis in the stria vascularis and spiral ganglion neurons of the inner ear, and progressive E2F1-dependent hearing loss. This transgenic-mtTFB1 mouse mitochondrial disease model provides a robust platform for deciphering the complex tissue-specificity of human mitochondrial-based disorders, as well as the precise pathogenic mechanism of maternally inherited deafness and its exacerbation by environmental factors. PMID:22341444

  8. Proteomic analysis of pRb loss highlights a signature of decreased mitochondrial oxidative phosphorylation.

    PubMed

    Nicolay, Brandon N; Danielian, Paul S; Kottakis, Filippos; Lapek, John D; Sanidas, Ioannis; Miles, Wayne O; Dehnad, Mantre; Tschöp, Katrin; Gierut, Jessica J; Manning, Amity L; Morris, Robert; Haigis, Kevin; Bardeesy, Nabeel; Lees, Jacqueline A; Haas, Wilhelm; Dyson, Nicholas J

    2015-09-01

    The retinoblastoma tumor suppressor (pRb) protein associates with chromatin and regulates gene expression. Numerous studies have identified Rb-dependent RNA signatures, but the proteomic effects of Rb loss are largely unexplored. We acutely ablated Rb in adult mice and conducted a quantitative analysis of RNA and proteomic changes in the colon and lungs, where Rb(KO) was sufficient or insufficient to induce ectopic proliferation, respectively. As expected, Rb(KO) caused similar increases in classic pRb/E2F-regulated transcripts in both tissues, but, unexpectedly, their protein products increased only in the colon, consistent with its increased proliferative index. Thus, these protein changes induced by Rb loss are coupled with proliferation but uncoupled from transcription. The proteomic changes in common between Rb(KO) tissues showed a striking decrease in proteins with mitochondrial functions. Accordingly, RB1 inactivation in human cells decreased both mitochondrial mass and oxidative phosphorylation (OXPHOS) function. RB(KO) cells showed decreased mitochondrial respiratory capacity and the accumulation of hypopolarized mitochondria. Additionally, RB/Rb loss altered mitochondrial pyruvate oxidation from (13)C-glucose through the TCA cycle in mouse tissues and cultured cells. Consequently, RB(KO) cells have an enhanced sensitivity to mitochondrial stress conditions. In summary, proteomic analyses provide a new perspective on Rb/RB1 mutation, highlighting the importance of pRb for mitochondrial function and suggesting vulnerabilities for treatment.

  9. Cardiolipin metabolism and the role it plays in heart failure and mitochondrial supercomplex formation.

    PubMed

    Mejia, Edgard M; Cole, Laura K; Hatch, Grant M

    2014-01-01

    Cardiolipin is a major membrane phospholipid in the mitochondria and is essential for cellular energy metabolism mediated through mitochondrial oxidative phosphorylation. Recent studies indicate that it plays a diverse role in cellular metabolism. Eukaryotic cardiolipin is synthesized de novo from phosphatidic acid via the cytidine-5'-diphosphate-1,2-diacyl-sn-glycerol pathway and is deacylated to monolysocardiolipin in order for it to be remodelled into the form that is observed in mitochondrial membranes. This resynthesis of deacylated cardiolipin from monolysocardiolipin occurs via the Barth Syndrome gene product tafazzin and acyllysocardiolipin acyltransferase-1, monolysocardiolipin acyltransferase-1 and the alpha subunit of trifunctional protein. Heart failure is a disease condition in which the amount and type of cardiolipin is altered. Several animal models have been generated to study the role of altered cardiolipin in heart failure. In many of these models loss of the tetralinoleoyl-cardiolipin species is observed during the development of the heart failure. In the doxycycline inducible short hairpin RNA tafazzin knock down mouse, loss of tetralinoleoyl-cardiolipin is associated with a mitochondrial bioenergetic disruption. Reduction in mitochondrial supercomplex formation and NADH dehydrogenase activity within these supercomplexes is observed. Modulation of CL fatty acyl composition may serve as a therapeutic strategy for the treatment of several pathologies including cardiac dysfunction.We propose that increasing cardiolipin may improve mitochondrial function and potentially serve as a therapy for diseases which exhibit mitochondrial dysfunction involving reduced cardiolipin. PMID:24801725

  10. Release of targeted p53 from the mitochondrion as an early signal during mitochondrial dysfunction.

    PubMed

    Green, M L; Pisano, M M; Prough, R A; Knudsen, T B

    2013-12-01

    Increased accumulation of p53 tumor suppressor protein is an early response to low-level stressors. To investigate the fate of mitochondrial-sequestered p53, mouse embryonic fibroblast cells (MEFs) on a p53-deficient genetic background were transfected with p53-EGFP fusion protein led by a sense (m53-EGFP) or antisense (c53-EGFP) mitochondrial import signal. Rotenone exposure (100nM, 1h) triggered the translocation of m53-EGFP from the mitochondrion to the nucleus, thus shifting the transfected cells from a mitochondrial p53 to a nuclear p53 state. Antibodies for p53 serine phosphorylation or lysine acetylation indicated a different post-translational status of recombinant p53 in the nucleus and mitochondrion, respectively. These data suggest that cycling of p53 through the mitochondria may establish a direct pathway for p53 signaling from the mitochondria to the nucleus during mitochondrial dysfunction. PK11195, a pharmacological ligand of mitochondrial TSPO (formerly known as the peripheral-type benzodiazepine receptor), partially suppressed the release of mitochondria-sequestered p53. These findings support the notion that p53 function mediates a direct signaling pathway from the mitochondria to nucleus during mitochondrial dysfunction.

  11. Monoamine oxidase inhibition prevents mitochondrial dysfunction and apoptosis in myoblasts from patients with collagen VI myopathies.

    PubMed

    Sorato, E; Menazza, S; Zulian, A; Sabatelli, P; Gualandi, F; Merlini, L; Bonaldo, P; Canton, M; Bernardi, P; Di Lisa, F

    2014-10-01

    Although mitochondrial dysfunction and oxidative stress have been proposed to play a crucial role in several types of muscular dystrophy (MD), whether a causal link between these two alterations exists remains an open question. We have documented that mitochondrial dysfunction through opening of the permeability transition pore plays a key role in myoblasts from patients as well as in mouse models of MD, and that oxidative stress caused by monoamine oxidases (MAO) is involved in myofiber damage. In the present study we have tested whether MAO-dependent oxidative stress is a causal determinant of mitochondrial dysfunction and apoptosis in myoblasts from patients affected by collagen VI myopathies. We find that upon incubation with hydrogen peroxide or the MAO substrate tyramine myoblasts from patients upregulate MAO-B expression and display a significant rise in reactive oxygen species (ROS) levels, with concomitant mitochondrial depolarization. MAO inhibition by pargyline significantly reduced both ROS accumulation and mitochondrial dysfunction, and normalized the increased incidence of apoptosis in myoblasts from patients. Thus, MAO-dependent oxidative stress is causally related to mitochondrial dysfunction and cell death in myoblasts from patients affected by collagen VI myopathies, and inhibition of MAO should be explored as a potential treatment for these diseases.

  12. TOM40 Mediates Mitochondrial Dysfunction Induced by α-Synuclein Accumulation in Parkinson’s Disease

    PubMed Central

    Rockenstein, Edward; Adame, Anthony; Elstner, Matthias; Laub, Christoph; Mueller, Sarina; Koob, Andrew O.; Mante, Michael; Pham, Emily; Klopstock, Thomas; Masliah, Eliezer

    2013-01-01

    Alpha-synuclein (α-Syn) accumulation/aggregation and mitochondrial dysfunction play prominent roles in the pathology of Parkinson’s disease. We have previously shown that postmortem human dopaminergic neurons from PD brains accumulate high levels of mitochondrial DNA (mtDNA) deletions. We now addressed the question, whether alterations in a component of the mitochondrial import machinery -TOM40- might contribute to the mitochondrial dysfunction and damage in PD. For this purpose, we studied levels of TOM40, mtDNA deletions, oxidative damage, energy production, and complexes of the respiratory chain in brain homogenates as well as in single neurons, using laser-capture-microdissection in transgenic mice overexpressing human wildtype α-Syn. Additionally, we used lentivirus-mediated stereotactic delivery of a component of this import machinery into mouse brain as a novel therapeutic strategy. We report here that TOM40 is significantly reduced in the brain of PD patients and in α-Syn transgenic mice. TOM40 deficits were associated with increased mtDNA deletions and oxidative DNA damage, and with decreased energy production and altered levels of complex I proteins in α-Syn transgenic mice. Lentiviral-mediated overexpression of Tom40 in α-Syn-transgenic mice brains ameliorated energy deficits as well as oxidative burden. Our results suggest that alterations in the mitochondrial protein transport machinery might contribute to mitochondrial impairment in α-Synucleinopathies. PMID:23626796

  13. Miro1 regulates intercellular mitochondrial transport & enhances mesenchymal stem cell rescue efficacy

    PubMed Central

    Ahmad, Tanveer; Mukherjee, Shravani; Pattnaik, Bijay; Kumar, Manish; Singh, Suchita; Kumar, Manish; Rehman, Rakhshinda; Tiwari, Brijendra K; Jha, Kumar A; Barhanpurkar, Amruta P; Wani, Mohan R; Roy, Soumya S; Mabalirajan, Ulaganathan; Ghosh, Balaram; Agrawal, Anurag

    2014-01-01

    There is emerging evidence that stem cells can rejuvenate damaged cells by mitochondrial transfer. Earlier studies show that epithelial mitochondrial dysfunction is critical in asthma pathogenesis. Here we show for the first time that Miro1, a mitochondrial Rho-GTPase, regulates intercellular mitochondrial movement from mesenchymal stem cells (MSC) to epithelial cells (EC). We demonstrate that overexpression of Miro1 in MSC (MSCmiroHi) leads to enhanced mitochondrial transfer and rescue of epithelial injury, while Miro1 knockdown (MSCmiroLo) leads to loss of efficacy. Treatment with MSCmiroHi was associated with greater therapeutic efficacy, when compared to control MSC, in mouse models of rotenone (Rot) induced airway injury and allergic airway inflammation (AAI). Notably, airway hyperresponsiveness and remodeling were reversed by MSCmiroHi in three separate allergen-induced asthma models. In a human in vitro system, MSCmiroHi reversed mitochondrial dysfunction in bronchial epithelial cells treated with pro-inflammatory supernatant of IL-13-induced macrophages. Anti-inflammatory MSC products like NO, TGF-β, IL-10 and PGE2, were unchanged by Miro1 overexpression, excluding non-specific paracrine effects. In summary, Miro1 overexpression leads to increased stem cell repair. PMID:24431222

  14. Proteomic analysis of pRb loss highlights a signature of decreased mitochondrial oxidative phosphorylation

    PubMed Central

    Nicolay, Brandon N.; Danielian, Paul S.; Kottakis, Filippos; Lapek, John D.; Sanidas, Ioannis; Miles, Wayne O.; Dehnad, Mantre; Tschöp, Katrin; Gierut, Jessica J.; Manning, Amity L.; Morris, Robert; Haigis, Kevin; Bardeesy, Nabeel; Lees, Jacqueline A.; Haas, Wilhelm; Dyson, Nicholas J.

    2015-01-01

    The retinoblastoma tumor suppressor (pRb) protein associates with chromatin and regulates gene expression. Numerous studies have identified Rb-dependent RNA signatures, but the proteomic effects of Rb loss are largely unexplored. We acutely ablated Rb in adult mice and conducted a quantitative analysis of RNA and proteomic changes in the colon and lungs, where RbKO was sufficient or insufficient to induce ectopic proliferation, respectively. As expected, RbKO caused similar increases in classic pRb/E2F-regulated transcripts in both tissues, but, unexpectedly, their protein products increased only in the colon, consistent with its increased proliferative index. Thus, these protein changes induced by Rb loss are coupled with proliferation but uncoupled from transcription. The proteomic changes in common between RbKO tissues showed a striking decrease in proteins with mitochondrial functions. Accordingly, RB1 inactivation in human cells decreased both mitochondrial mass and oxidative phosphorylation (OXPHOS) function. RBKO cells showed decreased mitochondrial respiratory capacity and the accumulation of hypopolarized mitochondria. Additionally, RB/Rb loss altered mitochondrial pyruvate oxidation from 13C-glucose through the TCA cycle in mouse tissues and cultured cells. Consequently, RBKO cells have an enhanced sensitivity to mitochondrial stress conditions. In summary, proteomic analyses provide a new perspective on Rb/RB1 mutation, highlighting the importance of pRb for mitochondrial function and suggesting vulnerabilities for treatment. PMID:26314710

  15. A PPARγ-Bnip3 Axis Couples Adipose Mitochondrial Fusion-Fission Balance to Systemic Insulin Sensitivity.

    PubMed

    Tol, Marc J; Ottenhoff, Roelof; van Eijk, Marco; Zelcer, Noam; Aten, Jan; Houten, Sander M; Geerts, Dirk; van Roomen, Cindy; Bierlaagh, Marlou C; Scheij, Saskia; Hoeksema, Marten A; Aerts, Johannes M; Bogan, Jonathan S; Dorn, Gerald W; Argmann, Carmen A; Verhoeven, Arthur J

    2016-09-01

    Aberrant mitochondrial fission plays a pivotal role in the pathogenesis of skeletal muscle insulin resistance. However, fusion-fission dynamics are physiologically regulated by inherent tissue-specific and nutrient-sensitive processes that may have distinct or even opposing effects with respect to insulin sensitivity. Based on a combination of mouse population genetics and functional in vitro assays, we describe here a regulatory circuit in which peroxisome proliferator-activated receptor γ (PPARγ), the adipocyte master regulator and receptor for the thiazolidinedione class of antidiabetic drugs, controls mitochondrial network fragmentation through transcriptional induction of Bnip3. Short hairpin RNA-mediated knockdown of Bnip3 in cultured adipocytes shifts the balance toward mitochondrial elongation, leading to compromised respiratory capacity, heightened fatty acid β-oxidation-associated mitochondrial reactive oxygen species generation, insulin resistance, and reduced triacylglycerol storage. Notably, the selective fission/Drp1 inhibitor Mdivi-1 mimics the effects of Bnip3 knockdown on adipose mitochondrial bioenergetics and glucose disposal. We further show that Bnip3 is reciprocally regulated in white and brown fat depots of diet-induced obesity and leptin-deficient ob/ob mouse models. Finally, Bnip3(-/-) mice trade reduced adiposity for increased liver steatosis and develop aggravated systemic insulin resistance in response to high-fat feeding. Together, our data outline Bnip3 as a key effector of PPARγ-mediated adipose mitochondrial network fragmentation, improving insulin sensitivity and limiting oxidative stress. PMID:27325287

  16. Mitochondrial oxidative stress caused by Sod2 deficiency promotes cellular senescence and aging phenotypes in