title: Consequences of zygote injection and germline transfer of mutant human mitochondrial DNA in mice.
abstract: Considerable evidence supports mutations in mitochondrial genes as the cause of maternally inherited diseases affecting tissues that rely primarily on oxidative energy metabolism, usually the nervous system, the heart, and skeletal muscles. Mitochondrial diseases are diverse, and animal models currently are limited. Here we introduced a mutant human mitochondrial gene responsible for Leber hereditary optic neuropathy (LHON) into the mouse germ line using fluorescence imaging for tissue-specific enrichment in the target retinal ganglion cells. A mitochondria-targeted adeno-associated virus (MTS-AAV) containing the mutant human NADH ubiquinone oxidoreductase subunit 4 (ND4) gene followed by mitochondrial-encoded mCherry was microinjected into zygotes. Female founders with mCherry fluorescence on ophthalmoscopy were backcrossed with normal males for eight generations. Mutant human ND4 DNA was 20% of mouse ND4 and did not integrate into the host genome. Translated human ND4 protein assembled into host respiratory complexes, decreasing respiratory chain function and increasing oxidative stress. Swelling of the optic nerve head was followed by progressive demise of ganglion cells and their axons, the hallmarks of human LHON. Early visual loss that began at 3 mo and progressed to blindness 8 mo after birth was reversed by intraocular injection of MTS-AAV expressing wild-type human ND4. The technology of introducing human mitochondrial genes into the mouse germ line has never been described, to our knowledge, and has implications not only for creating animal models recapitulating the counterpart human disorder but more importantly for reversing the adverse effects of the mutant gene using gene therapy to deliver the wild-type allele.
authors.affiliation: Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami FL 33136;
authors.forename: Hong
authors.initials: H
authors.lastname: Yu
authors.affiliation: Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami FL 33136;
authors.forename: Rajeshwari D
authors.initials: RD
authors.lastname: Koilkonda
authors.affiliation: Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami FL 33136;
authors.forename: Tsung-Han
authors.initials: TH
authors.lastname: Chou
authors.affiliation: Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami FL 33136;
authors.forename: Vittorio
authors.initials: V
authors.lastname: Porciatti
authors.affiliation: Hussman Institute of Human Genomics, University of Miami Miller School of Medicine, Miami, FL 33136;
authors.forename: Arpit
authors.initials: A
authors.lastname: Mehta
authors.affiliation: Miami Project to Cure Paralysis, University of Miami Miller School of Medicine, Miami, FL 33136;
authors.forename: Ian D
authors.initials: ID
authors.lastname: Hentall
authors.affiliation: Department of Ophthalmology, University of Florida College of Medicine, Gainesville, FL 32603;
authors.forename: Vince A
authors.initials: VA
authors.lastname: Chiodo
authors.affiliation: Department of Ophthalmology, University of Florida College of Medicine, Gainesville, FL 32603;
authors.forename: Sanford L
authors.initials: SL
authors.lastname: Boye
authors.affiliation: Department of Ophthalmology, University of Florida College of Medicine, Gainesville, FL 32603;
authors.forename: William W
authors.initials: WW
authors.lastname: Hauswirth
authors.affiliation: Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville, FL 32603.
authors.forename: Alfred S
authors.identifier: http://orcid.org/0000-0002-4192-9727
authors.initials: AS
authors.lastname: Lewin
authors.affiliation: Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami FL 33136; jguy@med.miami.edu.
authors.forename: John
authors.initials: J
authors.lastname: Guy
mesh_terms: D000818:Animals; D001369:Axons; D001921:Brain / Q000473:pathology; D004272:DNA, Mitochondrial / Q000235:genetics*; D004579:Electron Transport; D018014:Gene Transfer Techniques; D005854:Germ Cells; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D009154:Mutation; D009245:NADH Dehydrogenase / Q000235:genetics; D018384:Oxidative Stress; D012162:Retinal Degeneration / Q000235:genetics; D015053:Zygote
chemical_list: D004272:DNA, Mitochondrial; C098483:NADH dehydrogenase subunit 4; D009245:NADH Dehydrogenase
keywords: Leber hereditary optic neuropathy; adeno-associated virus; blindness; gene therapy; mitochondria