Inner ear gene transfection in neonatal mice using adeno-associated viral vector: a comparison of two approaches.
Local gene transfection is a promising technique for the prevention and/or correction of inner ear diseases, particularly those resulting from genetic defects. Adeno-associated virus (AAV) is an ideal viral vector for inner ear gene transfection because of its safety, stability, long-lasting expression, and its high tropism for many different cell types. Recently, a new generation of AAV vectors with a tyrosine mutation (mut-AAV) has demonstrated significant improvement in transfection efficiency. A method for inner ear gene transfection via the intact round window membrane (RWM) has been developed in our laboratory. This method has not been tested in neonatal mice, an important species for the study of inherited hearing loss. Following a preliminary study to optimize the experimental protocol in order to reduce mortality, the present study investigated inner ear gene transfection in mice at postnatal day 7. We compared transfection efficiency, the safety of the scala tympani injection via RWM puncture, and the trans-RWM diffusion following partial digestion with an enzyme technique. The results revealed that approximately 47% of inner hair cells (IHCs) and 17% of outer hair cells (OHCs) were transfected via the trans-RWM approach. Transfection efficiency via RWM puncture (58% and 19% for IHCs and OHCs, respectively) was slightly higher, but the difference was not significant.
Department of Otolaryngology, Affiliated Sixth People's Hospital of Shanghai Jiao Tong University, Otolaryngology Institute of Shanghai Jiao Tong University, Shanghai, China.
Li
L
Xia
Shankai
S
Yin
Jian
J
Wang
D000818:Animals; D000831:Animals, Newborn; D000229:Dependovirus; D005822:Genetic Vectors / Q000008:administration & dosage / Q000235:genetics*; D007150:Immunohistochemistry; D007267:Injections / Q000379:methods; D007759:Labyrinth Diseases / Q000235:genetics / Q000628:therapy*; D051379:Mice; D018613:Microscopy, Confocal; D012405:Round Window, Ear / Q000601:surgery*; D012533:Scala Tympani / Q000601:surgery*; D014162:Transfection / Q000379:methods*