Publications by authors named "Ivano Di Meo"

23 Publications

  • Page 1 of 1

A novel homozygous MSTO1 mutation in Ashkenazi Jewish siblings with ataxia and myopathy.

J Hum Genet 2021 Feb 22. Epub 2021 Feb 22.

Pediatric Neurology Unit, Hadassah Hebrew University of Jerusalem, 91240, Jerusalem, Israel.

MSTO1 is a cytoplasmic protein that modulates mitochondrial dynamics by promoting mitochondrial fusion. Mutations in the MSTO1 gene are responsible for an extremely rare condition characterized by early-onset myopathy and cerebellar ataxia. We report here two siblings from a large Ashkenazi Jewish family, presenting with a progressive neuromuscular disease characterized by ataxia and myopathy. By whole exome sequencing, we found a novel homozygous missense mutation (c.1403T>A, p.Leu468Gln) in MSTO1. Studies performed on fibroblasts from the index patient demonstrated the pathogenic role of the identified variant; we found that MSTO1 protein level was reduced and that mitochondrial network was fragmented or formed enlarged structures. Moreover, patient's cells showed reduced mitochondrial DNA amount. Our report confirms that MSTO1 mutations are typically recessive, and associated with clinical phenotypes characterized by early-onset muscle impairment and ataxia, often with upper motor neuron signs and varied cognitive impairment.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s10038-020-00897-4DOI Listing
February 2021

Neuronal Ablation of CoA Synthase Causes Motor Deficits, Iron Dyshomeostasis, and Mitochondrial Dysfunctions in a CoPAN Mouse Model.

Int J Mol Sci 2020 Dec 19;21(24). Epub 2020 Dec 19.

Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, 20126 Milan, Italy.

COASY protein-associated neurodegeneration (CoPAN) is a rare but devastating genetic autosomal recessive disorder of inborn error of CoA metabolism, which shares with pantothenate kinase-associated neurodegeneration (PKAN) similar features, such as dystonia, parkinsonian traits, cognitive impairment, axonal neuropathy, and brain iron accumulation. These two disorders are part of the big group of neurodegenerations with brain iron accumulation (NBIA) for which no effective treatment is available at the moment. To date, the lack of a mammalian model, fully recapitulating the human disorder, has prevented the elucidation of pathogenesis and the development of therapeutic approaches. To gain new insights into the mechanisms linking CoA metabolism, iron dyshomeostasis, and neurodegeneration, we generated and characterized the first CoPAN disease mammalian model. Since CoA is a crucial metabolite, constitutive ablation of the gene is incompatible with life. On the contrary, a conditional neuronal-specific knock-out mouse model consistently developed a severe early onset neurological phenotype characterized by sensorimotor defects and dystonia-like movements, leading to premature death. For the first time, we highlighted defective brain iron homeostasis, elevation of iron, calcium, and magnesium, together with mitochondrial dysfunction. Surprisingly, total brain CoA levels were unchanged, and no signs of neurodegeneration were present.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms21249707DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7766928PMC
December 2020

Homozygous mutations in C1QBP as cause of progressive external ophthalmoplegia (PEO) and mitochondrial myopathy with multiple mtDNA deletions.

Hum Mutat 2020 Jul 11. Epub 2020 Jul 11.

Unit of Medical Genetics and Neurogenetics, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milano, Italy.

Biallelic mutations in the C1QBP gene have been associated with mitochondrial cardiomyopathy and combined respiratory-chain deficiencies, with variable onset (including intrauterine or neonatal forms), phenotypes, and severity. We studied two unrelated adult patients from consanguineous families, presenting with progressive external ophthalmoplegia (PEO), mitochondrial myopathy, and without any heart involvement. Muscle biopsies from both patients showed typical mitochondrial alterations and the presence of multiple mitochondrial DNA deletions, whereas biochemical defects of the respiratory chain were present only in one subject. Using next-generation sequencing approaches, we identified homozygous mutations in C1QBP. Immunoblot analyses in patients' muscle samples revealed a strong reduction in the amount of the C1QBP protein and varied impairment of respiratory chain complexes, correlating with disease severity. Despite the original study indicated C1QBP mutations as causative for mitochondrial cardiomyopathy, our data indicate that mutations in C1QBP have to be considered in subjects with PEO phenotype or primary mitochondrial myopathy and without cardiomyopathy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/humu.24081DOI Listing
July 2020

Harmful Iron-Calcium Relationship in Pantothenate kinase Associated Neurodegeneration.

Int J Mol Sci 2020 May 22;21(10). Epub 2020 May 22.

IRCCS San Raffaele Scientific Institute, 20132 Milan, Italy.

Pantothenate Kinase-associated Neurodegeneration (PKAN) belongs to a wide spectrum of diseases characterized by brain iron accumulation and extrapyramidal motor signs. PKAN is caused by mutations in PANK2, encoding the mitochondrial pantothenate kinase 2, which is the first enzyme of the biosynthesis of Coenzyme A. We established and characterized glutamatergic neurons starting from previously developed PKAN Induced Pluripotent Stem Cells (iPSCs). Results obtained by inductively coupled plasma mass spectrometry indicated a higher amount of total cellular iron in PKAN glutamatergic neurons with respect to controls. PKAN glutamatergic neurons, analyzed by electron microscopy, exhibited electron dense aggregates in mitochondria that were identified as granules containing calcium phosphate. Calcium homeostasis resulted compromised in neurons, as verified by monitoring the activity of calcium-dependent enzyme calpain1, calcium imaging and voltage dependent calcium currents. Notably, the presence of calcification in the internal was confirmed in seven out of 15 genetically defined PKAN patients for whom brain CT scan was available. Moreover, we observed a higher prevalence of brain calcification in females. Our data prove that high amount of iron coexists with an impairment of cytosolic calcium in PKAN glutamatergic neurons, indicating both, iron and calcium dys-homeostasis, as actors in pathogenesis of the disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/ijms21103664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7279353PMC
May 2020

SSBP1 mutations cause mtDNA depletion underlying a complex optic atrophy disorder.

J Clin Invest 2020 01;130(1):108-125

Unit of Neurology, Department of Biomedical and NeuroMotor Sciences (DIBINEM), University of Bologna, Bologna, Italy.

Inherited optic neuropathies include complex phenotypes, mostly driven by mitochondrial dysfunction. We report an optic atrophy spectrum disorder, including retinal macular dystrophy and kidney insufficiency leading to transplantation, associated with mitochondrial DNA (mtDNA) depletion without accumulation of multiple deletions. By whole-exome sequencing, we identified mutations affecting the mitochondrial single-strand binding protein (SSBP1) in 4 families with dominant and 1 with recessive inheritance. We show that SSBP1 mutations in patient-derived fibroblasts variably affect the amount of SSBP1 protein and alter multimer formation, but not the binding to ssDNA. SSBP1 mutations impaired mtDNA, nucleoids, and 7S-DNA amounts as well as mtDNA replication, affecting replisome machinery. The variable mtDNA depletion in cells was reflected in severity of mitochondrial dysfunction, including respiratory efficiency, OXPHOS subunits, and complex amount and assembly. mtDNA depletion and cytochrome c oxidase-negative cells were found ex vivo in biopsies of affected tissues, such as kidney and skeletal muscle. Reduced efficiency of mtDNA replication was also reproduced in vitro, confirming the pathogenic mechanism. Furthermore, ssbp1 suppression in zebrafish induced signs of nephropathy and reduced optic nerve size, the latter phenotype complemented by WT mRNA but not by SSBP1 mutant transcripts. This previously unrecognized disease of mtDNA maintenance implicates SSBP1 mutations as a cause of human pathology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/JCI128514DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6934201PMC
January 2020

Inborn errors of coenzyme A metabolism and neurodegeneration.

J Inherit Metab Dis 2019 01;42(1):49-56

Unit of Molecular Neurogenetics - Pierfranco and Luisa Mariani Centre for the Study of Mitochondrial Disorders in Children, Foundation IRCCS Neurological Institute C. Besta, Via Temolo 4, Milan 20126, Italy.

Two inborn errors of coenzyme A (CoA) metabolism are responsible for distinct forms of neurodegeneration with brain iron accumulation (NBIA), a heterogeneous group of neurodegenerative diseases having as a common denominator iron accumulation mainly in the inner portion of globus pallidus. Pantothenate kinase-associated neurodegeneration (PKAN), an autosomal recessive disorder with progressive impairment of movement, vision and cognition, is the most common form of NBIA and is caused by mutations in the pantothenate kinase 2 gene (PANK2), coding for a mitochondrial enzyme, which phosphorylates vitamin B5 in the first reaction of the CoA biosynthetic pathway. Another very rare but similar disorder, denominated CoPAN, is caused by mutations in coenzyme A synthase gene (COASY) coding for a bi-functional mitochondrial enzyme, which catalyzes the final steps of CoA biosynthesis. It still remains a mystery why dysfunctions in CoA synthesis lead to neurodegeneration and iron accumulation in specific brain regions, but it is now evident that CoA metabolism plays a crucial role in the normal functioning and metabolism of the nervous system.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jimd.12026DOI Listing
January 2019

Dissection of metabolic reprogramming in polycystic kidney disease reveals coordinated rewiring of bioenergetic pathways.

Commun Biol 2018 16;1:194. Epub 2018 Nov 16.

Division of Genetics and Cell Biology, San Raffaele Scientific Institute, Via Olgettina, 60, Milan, 20132, Italy.

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is a genetic disorder caused by loss-of-function mutations in or . Increased glycolysis is a prominent feature of the disease, but how it impacts on other metabolic pathways is unknown. Here, we present an analysis of mouse mutant cells and kidneys to investigate the metabolic reprogramming of this pathology. We show that loss of leads to profound metabolic changes that affect glycolysis, mitochondrial metabolism, and fatty acid synthesis (FAS). We find that -mutant cells preferentially use glutamine to fuel the TCA cycle and to sustain FAS. Interfering with either glutamine uptake or FAS retards cell growth and survival. We also find that glutamine is diverted to asparagine via asparagine synthetase (ASNS). Transcriptional profiling of -mutant human kidneys confirmed these alterations. We find that silencing of is lethal in -mutant cells when combined with glucose deprivation, suggesting therapeutic approaches for ADPKD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s42003-018-0200-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6240072PMC
November 2018

Classification and molecular pathogenesis of NBIA syndromes.

Eur J Paediatr Neurol 2018 Mar 17;22(2):272-284. Epub 2018 Jan 17.

Unit of Molecular Neurogenetics, Pierfranco and Luisa Mariani Centre for the Study of Mitochondrial Disorders in Children, Foundation IRCCS Neurological Institute C. Besta, Via Temolo 4, 20126, Milan, Italy. Electronic address:

Brain iron accumulation is the hallmark of a group of seriously invalidating and progressive rare diseases collectively denominated Neurodegeneration with Brain Iron Accumulation (NBIA), characterized by movement disorder, painful dystonia, parkinsonism, mental disability and early death. Currently there is no established therapy available to slow down or reverse the progression of these conditions. Several genes have been identified as responsible for NBIA but only two encode for proteins playing a direct role in iron metabolism. The other genes encode for proteins either with various functions in lipid metabolism, lysosomal activity and autophagic processes or with still unknown roles. The different NBIA subtypes have been classified and denominated on the basis of the mutated genes and, despite genetic heterogeneity, some of them code for proteins, which share or converge on common metabolic pathways. In the last ten years, the implementation of genetic screening based on Whole Exome Sequencing has greatly accelerated gene discovery, nevertheless our knowledge of the pathogenic mechanisms underlying the NBIA syndromes is still largely incomplete.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.ejpn.2018.01.008DOI Listing
March 2018

Clinical, biochemical, and genetic features associated with VARS2-related mitochondrial disease.

Hum Mutat 2018 04 7;39(4):563-578. Epub 2018 Feb 7.

Unit of Neuromuscular and Neurodegenerative Disorders, Laboratory of Molecular Medicine, 'Bambino Ges.' Children's Research Hospital, Rome, Italy.

In recent years, an increasing number of mitochondrial disorders have been associated with mutations in mitochondrial aminoacyl-tRNA synthetases (mt-aaRSs), which are key enzymes of mitochondrial protein synthesis. Bi-allelic functional variants in VARS2, encoding the mitochondrial valyl tRNA-synthetase, were first reported in a patient with psychomotor delay and epilepsia partialis continua associated with an oxidative phosphorylation (OXPHOS) Complex I defect, before being described in a patient with a neonatal form of encephalocardiomyopathy. Here we provide a detailed genetic, clinical, and biochemical description of 13 patients, from nine unrelated families, harboring VARS2 mutations. All patients except one, who manifested with a less severe disease course, presented at birth exhibiting severe encephalomyopathy and cardiomyopathy. Features included hypotonia, psychomotor delay, seizures, feeding difficulty, abnormal cranial MRI, and elevated lactate. The biochemical phenotype comprised a combined Complex I and Complex IV OXPHOS defect in muscle, with patient fibroblasts displaying normal OXPHOS activity. Homology modeling supported the pathogenicity of VARS2 missense variants. The detailed description of this cohort further delineates our understanding of the clinical presentation associated with pathogenic VARS2 variants and we recommend that this gene should be considered in early-onset mitochondrial encephalomyopathies or encephalocardiomyopathies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/humu.23398DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5873438PMC
April 2018

Acetyl-4'-phosphopantetheine is stable in serum and prevents phenotypes induced by pantothenate kinase deficiency.

Sci Rep 2017 09 12;7(1):11260. Epub 2017 Sep 12.

Department of Cell Biology, University Medical Center Groningen, University of Groningen, Ant. Deusinglaan 1, 9713 AV, Groningen, The Netherlands.

Coenzyme A is an essential metabolite known for its central role in over one hundred cellular metabolic reactions. In cells, Coenzyme A is synthesized de novo in five enzymatic steps with vitamin B5 as the starting metabolite, phosphorylated by pantothenate kinase. Mutations in the pantothenate kinase 2 gene cause a severe form of neurodegeneration for which no treatment is available. One therapeutic strategy is to generate Coenzyme A precursors downstream of the defective step in the pathway. Here we describe the synthesis, characteristics and in vivo rescue potential of the acetyl-Coenzyme A precursor S-acetyl-4'-phosphopantetheine as a possible treatment for neurodegeneration associated with pantothenate kinase deficiency.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-017-11564-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5595861PMC
September 2017

Transcription Factor EB Controls Metabolic Flexibility during Exercise.

Cell Metab 2017 01 20;25(1):182-196. Epub 2016 Dec 20.

Telethon Institute of Genetics and Medicine (TIGEM), Via Campi Flegrei 34, 80078 Pozzuoli, Naples, Italy; Medical Genetics, Department of Pediatrics, Federico II University, Via Pansini 5, 80131 Naples, Italy; Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA; Jan and Dan Duncan Neurological Research Institute, Texas Children's Hospital, Houston, TX 77030, USA. Electronic address:

The transcription factor EB (TFEB) is an essential component of lysosomal biogenesis and autophagy for the adaptive response to food deprivation. To address the physiological function of TFEB in skeletal muscle, we have used muscle-specific gain- and loss-of-function approaches. Here, we show that TFEB controls metabolic flexibility in muscle during exercise and that this action is independent of peroxisome proliferator-activated receptor-γ coactivator1α (PGC1α). Indeed, TFEB translocates into the myonuclei during physical activity and regulates glucose uptake and glycogen content by controlling expression of glucose transporters, glycolytic enzymes, and pathways related to glucose homeostasis. In addition, TFEB induces the expression of genes involved in mitochondrial biogenesis, fatty acid oxidation, and oxidative phosphorylation. This coordinated action optimizes mitochondrial substrate utilization, thus enhancing ATP production and exercise capacity. These findings identify TFEB as a critical mediator of the beneficial effects of exercise on metabolism.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cmet.2016.11.003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5241227PMC
January 2017

Coenzyme Q deficiency causes impairment of the sulfide oxidation pathway.

EMBO Mol Med 2017 01;9(1):96-111

Department of Neurology, Columbia University Medical Center, New York, NY, USA

Coenzyme Q (CoQ) is an electron acceptor for sulfide-quinone reductase (SQR), the first enzyme of the hydrogen sulfide oxidation pathway. Here, we show that lack of CoQ in human skin fibroblasts causes impairment of hydrogen sulfide oxidation, proportional to the residual levels of CoQ. Biochemical and molecular abnormalities are rescued by CoQ supplementation in vitro and recapitulated by pharmacological inhibition of CoQ biosynthesis in skin fibroblasts and ADCK3 depletion in HeLa cells. Kidneys of Pdss2 mice, which only have ~15% residual CoQ concentrations and are clinically affected, showed (i) reduced protein levels of SQR and downstream enzymes, (ii) accumulation of hydrogen sulfides, and (iii) glutathione depletion. These abnormalities were not present in brain, which maintains ~30% residual CoQ and is clinically unaffected. In Pdss2 mice, we also observed low levels of plasma and urine thiosulfate and increased blood C4-C6 acylcarnitines. We propose that impairment of the sulfide oxidation pathway induced by decreased levels of CoQ causes accumulation of sulfides and consequent inhibition of short-chain acyl-CoA dehydrogenase and glutathione depletion, which contributes to increased oxidative stress and kidney failure.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.15252/emmm.201606356DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5210092PMC
January 2017

C4.4A gene ablation is compatible with normal epidermal development and causes modest overt phenotypes.

Sci Rep 2016 05 12;6:25833. Epub 2016 May 12.

The Finsen Laboratory, Rigshospitalet, Copenhagen, Denmark.

C4.4A is a modular glycolipid-anchored Ly6/uPAR/α-neurotoxin multidomain protein that exhibits a prominent membrane-associated expression in stratified squamous epithelia. C4.4A is also expressed in various solid cancer lesions, where high expression levels often are correlated to poor prognosis. Circumstantial evidence suggests a role for C4.4A in cell adhesion, migration, and invasion, but a well-defined biological function is currently unknown. In the present study, we have generated and characterized the first C4.4A-deficient mouse line to gain insight into the functional significance of C4.4A in normal physiology and cancer progression. The unchallenged C4.4A-deficient mice were viable, fertile, born in a normal Mendelian distribution and, surprisingly, displayed normal development of squamous epithelia. The C4.4A-deficient mice were, nonetheless, significantly lighter than littermate controls predominantly due to differences in fat mass. Congenital C4.4A deficiency delayed migration of keratinocytes enclosing incisional skin wounds in male mice. In chemically induced bladder carcinomas, C4.4A deficiency attenuated the incidence of invasive lesions despite having no effect on total tumour burden. This new C4.4A-deficient mouse line provides a useful platform for future studies on functional aspects of C4.4A in tumour cell invasion in vivo.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/srep25833DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4864438PMC
May 2016

Liver transplant in ethylmalonic encephalopathy: a new treatment for an otherwise fatal disease.

Brain 2016 Apr 25;139(Pt 4):1045-51. Epub 2016 Feb 25.

Department of Surgery and Transplantation, Bambino Gesù Children's Research Hospital IRCCS, Rome, Italy.

Ethylmalonic encephalopathy is a fatal, rapidly progressive mitochondrial disorder caused by ETHE1 mutations, whose peculiar clinical and biochemical features are due to the toxic accumulation of hydrogen sulphide and of its metabolites, including thiosulphate. In mice with ethylmalonic encephalopathy, liver-targeted adeno-associated virus-mediated ETHE1 gene transfer dramatically improved both clinical course and metabolic abnormalities. Reasoning that the same achievement could be accomplished by liver transplantation, we performed living donor-liver transplantation in an infant with ethylmalonic encephalopathy. Unlike the invariably progressive deterioration of the disease, 8 months after liver transplantation, we observed striking neurological improvement with remarkable achievements in psychomotor development, along with dramatic reversion of biochemical abnormalities. These results clearly indicate that liver transplantation is a viable therapeutic option for ETHE1 disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/brain/aww013DOI Listing
April 2016

Mitochondrial diseases caused by toxic compound accumulation: from etiopathology to therapeutic approaches.

EMBO Mol Med 2015 Oct;7(10):1257-66

Unit of Molecular Neurogenetics, Foundation IRCCS Neurological Institute C. Besta, Milan, Italy

Mitochondrial disorders are a group of highly invalidating human conditions for which effective treatment is currently unavailable and characterized by faulty energy supply due to defective oxidative phosphorylation (OXPHOS). Given the complexity of mitochondrial genetics and biochemistry, mitochondrial inherited diseases may present with a vast range of symptoms, organ involvement, severity, age of onset, and outcome. Despite the wide spectrum of clinical signs and biochemical underpinnings of this group of dis-orders, some common traits can be identified, based on both pathogenic mechanisms and potential therapeutic approaches. Here, we will review two peculiar mitochondrial disorders, ethylmalonic encephalopathy (EE) and mitochondrial neurogastrointestinal encephalomyopathy (MNGIE), caused by mutations in the ETHE1 and TYMP nuclear genes, respectively. ETHE1 encodes for a mitochondrial enzyme involved in sulfide detoxification and TYMP for a cytosolic enzyme involved in the thymidine/deoxyuridine catabolic pathway. We will discuss these two clinical entities as a paradigm of mitochondrial diseases caused by the accumulation of compounds normally present in traces, which exerts a toxic and inhibitory effect on the OXPHOS system.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.15252/emmm.201505040DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4604682PMC
October 2015

Gene therapy using a liver-targeted AAV vector restores nucleoside and nucleotide homeostasis in a murine model of MNGIE.

Mol Ther 2014 May 22;22(5):901-7. Epub 2014 Jan 22.

1] Mitochondrial Disorders Unit, Vall d'Hebron Institut de Recerca, Universitat Autònoma de Barcelona, Barcelona, Spain; [2] Biomedical Network Research Centre on Rare Diseases, Instituto de Salud Carlos III, Madrid, Spain;

Mitochondrial neurogastrointestinal encephalomyopathy (MNGIE) is an autosomal recessive disorder caused by mutations in TYMP, enconding thymidine phosphorylase (TP). TP deficiency results in systemic accumulation of thymidine and deoxyuridine, which interferes with mitochondrial DNA (mtDNA) replication and leads to mitochondrial dysfunction. To date, the only treatment available for MNGIE patients is allogeneic hematopoietic stem cell transplantation, which is associated with high morbidity and mortality. Here, we report that AAV2/8-mediated transfer of the human TYMP coding sequence (hcTYMP) under the control of a liver-specific promoter prevents the biochemical imbalances in a murine model of MNGIE. hcTYMP expression was restricted to liver, and a dose as low as 2 × 10(11) genome copies/kg led to a permanent reduction in systemic nucleoside levels to normal values in about 50% of treated mice. Higher doses resulted in reductions to normal or slightly below normal levels in virtually all mice treated. The nucleoside reduction achieved by this treatment prevented deoxycytidine triphosphate (dCTP) depletion, which is the limiting factor affecting mtDNA replication in this disease. These results demonstrate that the use of AAV to direct TYMP expression in liver is feasible as a potentially safe gene therapy strategy for MNGIE.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/mt.2014.6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4015233PMC
May 2014

AAV-mediated liver-specific MPV17 expression restores mtDNA levels and prevents diet-induced liver failure.

Mol Ther 2014 Jan 3;22(1):10-7. Epub 2013 Jul 3.

1] Unit of Molecular Neurogenetics, The Foundation "Carlo Besta" Institute of Neurology IRCCS, Milan, Italy [2] MRC-Mitochondrial Biology Unit, Cambridge, UK.

Mutations in human MPV17 cause a hepatocerebral form of mitochondrial DNA depletion syndrome (MDS) hallmarked by early-onset liver failure, leading to premature death. Liver transplantation and frequent feeding using slow-release carbohydrates are the only available therapies, although surviving patients eventually develop slowly progressive peripheral and central neuropathy. The physiological role of Mpv17, including its functional link to mitochondrial DNA (mtDNA) maintenance, is still unclear. We show here that Mpv17 is part of a high molecular weight complex of unknown composition, which is essential for mtDNA maintenance in critical tissues, i.e. liver, of a Mpv17 knockout mouse model. On a standard diet, Mpv17-/- mouse shows hardly any symptom of liver dysfunction, but a ketogenic diet (KD) leads these animals to liver cirrhosis and failure. However, when expression of human MPV17 is carried out by adeno-associated virus (AAV)-mediated gene replacement, the Mpv17 knockout mice are able to reconstitute the Mpv17-containing supramolecular complex, restore liver mtDNA copy number and oxidative phosphorylation (OXPHOS) proficiency, and prevent liver failure induced by the KD. These results open new therapeutic perspectives for the treatment of MPV17-related liver-specific MDS.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/mt.2013.230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3880585PMC
January 2014

Proteome adaptations in Ethe1-deficient mice indicate a role in lipid catabolism and cytoskeleton organization via post-translational protein modifications.

Biosci Rep 2013 Jul 25;33(4). Epub 2013 Jul 25.

Institut für Pflanzengenetik, Leibniz Universität Hannover, Germany.

Hydrogen sulfide is a physiologically relevant signalling molecule. However, circulating levels of this highly biologically active substance have to be maintained within tightly controlled limits in order to avoid toxic side effects. In patients suffering from EE (ethylmalonic encephalopathy), a block in sulfide oxidation at the level of the SDO (sulfur dioxygenase) ETHE1 leads to severe dysfunctions in microcirculation and cellular energy metabolism. We used an Ethe1-deficient mouse model to investigate the effect of increased sulfide and persulfide concentrations on liver, kidney, muscle and brain proteomes. Major disturbances in post-translational protein modifications indicate that the mitochondrial sulfide oxidation pathway could have a crucial function during sulfide signalling most probably via the regulation of cysteine S-modifications. Our results confirm the involvement of sulfide in redox regulation and cytoskeleton dynamics. In addition, they suggest that sulfide signalling specifically regulates mitochondrial catabolism of FAs (fatty acids) and BCAAs (branched-chain amino acids). These findings are particularly relevant in the context of EE since they may explain major symptoms of the disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1042/BSR20130051DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3827611PMC
July 2013

Effective AAV-mediated gene therapy in a mouse model of ethylmalonic encephalopathy.

EMBO Mol Med 2012 Sep 20;4(9):1008-14. Epub 2012 Aug 20.

Unit of Molecular Neurogenetics, The Foundation "Carlo Besta" Institute of Neurology IRCCS, Milan, Italy.

Ethylmalonic encephalopathy (EE) is an invariably fatal disease, characterized by the accumulation of hydrogen sulfide (H(2)S), a highly toxic compound. ETHE1, encoding sulfur dioxygenase (SDO), which takes part in the mitochondrial pathway that converts sulfide into harmless sulfate, is mutated in EE. The main source of H(2)S is the anaerobic bacterial flora of the colon, although in trace amount it is also produced by tissues, where it acts as a 'gasotransmitter'. Here, we show that AAV2/8-mediated, ETHE1-gene transfer to the liver of a genetically, metabolically and clinically faithful EE mouse model resulted in full restoration of SDO activity, correction of plasma thiosulfate, a biomarker reflecting the accumulation of H(2)S, and spectacular clinical improvement. Most of treated animals were alive and well >6-8 months after birth, whereas untreated individuals live 26 ± 7 days. Our results provide proof of concept on the efficacy and safety of AAV2/8-mediated livergene therapy for EE, and alike conditions caused by the accumulation of harmful compounds in body fluids and tissues, which can directly be transferred to the clinic.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/emmm.201201433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3491831PMC
September 2012

Morphologic evidence of diffuse vascular damage in human and in the experimental model of ethylmalonic encephalopathy.

J Inherit Metab Dis 2012 May 22;35(3):451-8. Epub 2011 Oct 22.

Department of Radiological, Oncological and Pathological Sciences, Sapienza University, Policlinico Umberto I, Viale Regina Elena 324, 00161 Rome, Italy.

Ethylmalonic encephalopathy (EE) is a rare autosomal recessive disorder characterized by early onset encephalopathy, chronic diarrhoea, petechiae, orthostatic acrocyanosis and defective cytochrome c oxidase (COX) in muscle and brain. High levels of lactic, ethylmalonic and methylsuccinic acids are detected in body fluids. EE is caused by mutations in ETHE1, a mitochondrial sulphur dioxygenase. By studying a suitable mouse model, we found that loss of ETHE1 leads to accumulation of sulphide, which is a poison for COX and other enzymatic activities thus accounting for the main features of EE. We report here the first autopsy case of a child with a genetically confirmed diagnosis of EE, and compare the histological, histochemical and immunohistochemical findings with those of the constitutive Ethe1 (-/-) mice. In addition to COX depleted cells, widespread endothelial lesions of arterioles and capillaries of the brain and gastrointestinal tract were the pathologic hallmarks in both organisms. Our findings of diffuse vascular damage of target critical organs are in keeping with the hypothesis that the pathologic effects of ETHE1 deficiency may stem from high levels of circulating hydrogen sulphide rather than the inability of specific organs to detoxify its endogenous production.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s10545-011-9408-3DOI Listing
May 2012

Chronic exposure to sulfide causes accelerated degradation of cytochrome c oxidase in ethylmalonic encephalopathy.

Antioxid Redox Signal 2011 Jul 25;15(2):353-62. Epub 2011 Feb 25.

Unit of Molecular Neurogenetics, Pierfranco and Luisa Mariani Center for Research on Children's Mitochondrial Disorders, Institute of Neurology Carlo Besta-IRCCS Foundation, Milan, Italy.

Ethylmalonic encephalopathy (EE) is an autosomal recessive, invariably fatal disorder associated with mutations in ETHE1, a gene encoding a mitochondrial sulfur dioxygenase (SDO). The main consequence of the absence of Ethe1-SDO is the accumulation of sulfide (H(2)S) in critical tissues, including colonic mucosa, liver, muscle, and brain. To make progress in the elucidation of the biochemical mechanisms leading to cytochrome c oxidase (COX) deficiency, we (i) generated tissue-specific conditional Ethe1 knockout mice to clarify the different contributions of endogenous and exogenous H(2)S production, and (ii) studied the development of H(2)S-driven COX deficiency in Ethe1(-/-) mouse tissues and human cells. Ethe1(-/-) conditional animals displayed COX deficiency limited to the specific targeted tissue. The accumulation of H(2)S over time causes progressive COX deficiency in animal tissues and human cells, which is associated with reduced amount of COX holoenzyme, and of several COX subunits, including mitochondrially encoded cytochrome c oxidase 1 (MTCO1), MTCO2, COX4, and COX5A. This reduction is not paralleled by consistent downregulation in expression of the corresponding mRNAs. Tissue-specific ablation of Ethe1 causes COX deficiency in targeted organs, suggesting that failure in neutralizing endogenous, tissue-specific production of H(2)S is sufficient to cause the biochemical defect but neither to determine a clinical impact nor to induce the biomarker profile typical of EE. The mechanism by which H(2)S causes COX deficiency consists of rapid heme a inhibition and accelerated long-term degradation of COX subunits. However, the pleiotropic devastating effects of H(2)S accumulation in EE cannot be fully explained by the sole defect of COX in critical tissues, but are likely consequent to several toxic actions on a number of enzymatic activities in different tissues, including endothelial lining of the small vessels, leading to multiorgan failure.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1089/ars.2010.3520DOI Listing
July 2011

Loss of ETHE1, a mitochondrial dioxygenase, causes fatal sulfide toxicity in ethylmalonic encephalopathy.

Nat Med 2009 Feb 11;15(2):200-5. Epub 2009 Jan 11.

Pierfranco and Luisa Mariani Center for Research on Children's Mitochondrial Disorders, Institute of Neurology Carlo Besta-Istituto di Ricovero e Cura a Carattere Scientifico Foundation, Milan, Italy.

Ethylmalonic encephalopathy is an autosomal recessive, invariably fatal disorder characterized by early-onset encephalopathy, microangiopathy, chronic diarrhea, defective cytochrome c oxidase (COX) in muscle and brain, high concentrations of C4 and C5 acylcarnitines in blood and high excretion of ethylmalonic acid in urine. ETHE1, a gene encoding a beta-lactamase-like, iron-coordinating metalloprotein, is mutated in ethylmalonic encephalopathy. In bacteria, ETHE1-like sequences are in the same operon of, or fused with, orthologs of TST, the gene encoding rhodanese, a sulfurtransferase. In eukaryotes, both ETHE1 and rhodanese are located within the mitochondrial matrix. We created a Ethe1(-/-) mouse that showed the cardinal features of ethylmalonic encephalopathy. We found that thiosulfate was excreted in massive amounts in urine of both Ethe1(-/-) mice and humans with ethylmalonic encephalopathy. High thiosulfate and sulfide concentrations were present in Ethe1(-/-) mouse tissues. Sulfide is a powerful inhibitor of COX and short-chain fatty acid oxidation, with vasoactive and vasotoxic effects that explain the microangiopathy in ethylmalonic encephalopathy patients. Sulfide is detoxified by a mitochondrial pathway that includes a sulfur dioxygenase. Sulfur dioxygenase activity was absent in Ethe1(-/-) mice, whereas it was markedly increased by ETHE1 overexpression in HeLa cells and Escherichia coli. Therefore, ETHE1 is a mitochondrial sulfur dioxygenase involved in catabolism of sulfide that accumulates to toxic levels in ethylmalonic encephalopathy.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/nm.1907DOI Listing
February 2009