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.gene therapy | adeno-associated virus | mitochondria | Leber hereditary optic neuropathy | blindness

Dehydroepiandrosterone-induced activation of mTORC1 and inhibition of autophagy contribute to skeletal muscle insulin resistance in a mouse model of polycystic ovary syndrome

“…We therefore studied the integrated mitochondrial DNA copy number and the mitochondrial function in the skeletal muscle of the mice. ND4 is a mitochondrial DNA (mtDNA) encoding one subunit of the NADH ubiquinone oxidoreductase which is complex I of the mitochondrial respiratory chain [ 11 ]. The ND4 gene is often deleted.…”

Non‐Viral Gene Delivery via Membrane‐Penetrating, Mannose‐Targeting Supramolecular Self‐Assembled Nanocomplexes

“…[7-15] Antibodies and targeting ligands are introduced to the vectors to allow receptor-mediated uptake by particular target cells. [16, 17] Cell penetrating peptides (CPPs) have been conjugated to or complexed with the nucleic acid cargos to facilitate active cellular internalization and endosomal escape. [18-20] Responsive or degradable polymers have been applied to promote the cytosolic release of the cargo upon internal or external stimuli.…”

Integrative properties of retinal ganglion cell electrical responsiveness depend on neurotrophic support and genotype in the mouse

“…RGC function was assessed by Pattern Electroretinogram (PERG), an electrical signal that specifically depends on the presence of functional RGCs (Chou et al, 2013; Miura et al, 2009; Porciatti, 2015; Porciatti et al, 1996; Xia et al, 2014)and is commonly used in human and experimental models of glaucoma and optic neuropathies (Enriquez-Algeciras et al, 2013; Howell et al, 2007a; Porciatti, 2015; Yu et al, 2015; Yu et al, 2012). Anesthetized mice were gently restrained in a holder allowing unobstructed vision as previously described (Porciatti et al, 2007) and kept at a constant body temperature of 37.0 °C using a rectal probe and fee dback-controlled heating pad (TCAT-2LV; Physitemp Instruments, Inc. Clifton, NJ).…”