title: A Novel Method Combining Vitreous Aspiration and Intravitreal AAV2/8 Injection Results in Retina-Wide Transduction in Adult Mice.
abstract: OBJECTIVE
  Gene therapies to treat eye disorders have been extensively studied in the past 20 years. Frequently, adeno-associated viruses were applied to the subretinal or intravitreal space of the eye to transduce retinal cells with nucleotide sequences of therapeutic potential. In this study we describe a novel intravitreal injection procedure that leads to a reproducible adeno-associated virus (AAV)2/8-mediated transduction of more than 70% of the retina.
  
  
  METHODS
  Prior to a single intravitreal injection of a enhanced green fluorescent protien (GFP)-expressing viral suspension, we performed an aspiration of vitreous tissue from wild-type C57Bl/6J mice. One and one-half microliters of AAV2/8 suspension was injected. Funduscopy, optical coherence tomography (OCT), laser scanning microscopy of retinal flat mounts, cryosections of eye cups, and ERG recordings verified the efficacy and safety of the method.
  
  
  RESULTS
  The combination of vitreous aspiration and intravitreal injection resulted in an almost complete transduction of the retina in approximately 60% of the eyes and showed transduced cells in all retinal layers. Photoreceptors and RPE cells were predominantly transduced. Eyes presented with well-preserved retinal morphology. Electroretinographic recordings suggested that the new combination of techniques did not cause significant alterations of the retinal physiology.
  
  
  CONCLUSIONS
  We show a novel application technique of AAV2/8 to the vitreous of mice that leads to widespread transduction of the retina. The results of this study have implications for virus-based gene therapies and basic science; for example, they might provide an approach to apply gene replacement strategies or clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in vivo. It may further help to develop similar techniques for larger animal models or humans.
authors: Institute of Human Genetics, Faculty of Medicine and Health Sciences, University of Oldenburg, Oldenburg, Germany.
authors: Romain
authors: R
authors: Da Costa
authors: Institute of Human Genetics, Faculty of Medicine and Health Sciences, University of Oldenburg, Oldenburg, Germany.
authors: Carsten
authors: C
authors: Röger
authors: Visual Neuroscience, Faculty of Medicine and Health Sciences, University of Oldenburg, Oldenburg, Germany.
authors: Jasmin
authors: J
authors: Segelken
authors: Laboratory for Retinal Cell Biology, Department of Ophthalmology, University Hospital Zurich, Zurich, Switzerland 4Neuroscience Center Zurich (ZNZ), University of Zurich, Zurich, Switzerland.
authors: Maya
authors: M
authors: Barben
authors: Laboratory for Retinal Cell Biology, Department of Ophthalmology, University Hospital Zurich, Zurich, Switzerland 4Neuroscience Center Zurich (ZNZ), University of Zurich, Zurich, Switzerland 5Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Zurich, Switzerland.
authors: Christian
authors: C
authors: Grimm
authors: Institute of Human Genetics, Faculty of Medicine and Health Sciences, University of Oldenburg, Oldenburg, Germany 6Research Center Neurosensory Science, University Oldenburg, Oldenburg, Germany.
authors: John
authors: J
authors: Neidhardt
mesh_terms: D000818:Animals; D000229:Dependovirus / Q000235:genetics* / Q000472:pathogenicity; D004195:Disease Models, Animal; D004596:Electroretinography; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000008:administration & dosage*; D058449:Intravitreal Injections; D051379:Mice; D008810:Mice, Inbred C57BL; D018613:Microscopy, Confocal; D010857:Pigment Epithelium of Eye; D012160:Retina / Q000473:pathology / Q000503:physiopathology; D012164:Retinal Diseases / Q000175:diagnosis / Q000628:therapy*; D013396:Suction / Q000379:methods*; D041623:Tomography, Optical Coherence; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes; D014822:Vitreous Body