Optimizing atoh1‐induced vestibular hair cell regeneration

Abstract

Objectives/Hypothesis Determine the optimal design characteristics of an adenoviral vector to deliver atoh1 and induce regeneration of vestibular hair cells. Study Design Evaluation of a mouse model of intra-labyrinthine gene delivery. Tissue culture of mouse and human macular organs. Methods Macular organs from adult C57Bl/6 mice were treated with binding modified and alternate adenovectors expressing green fluorescent protein (gfp) or luciferase (L). Expression of marker genes was determined over time to determine vector transfection efficiency. The inner ear of adult mice was then injected with modified vectors. Expression of gfp and distribution of vector DNA was followed. Hearing and balance function was evaluated in normal animals to ensure safety of the novel vector designs. An optimized vector was identified and tested for its ability to induce hair cell regeneration in a mouse vestibulopathy model. Finally this vector was tested for its ability to induce hair cell regeneration in human tissue. Results Ad5 serotype based vectors were identified as having a variety of different binding capacities for inner ear tissue. This makes it difficult to limit the dose of vector due to entry into non-targeted cells. Screening of rare adenovector serotypes demonstrated that Ad28 based vectors were ideally suited for delivery to supporting cells and therefore useful for hair cell regeneration studies. Utilization of an Ad28 based vector to deliver atoh1 to a mouse model of vestibular loss resulted significant functional recovery of balance. This vector was also capable of transfecting human macular organs and inducing regeneration of human vestibular hair cells in vitro. Conclusions Improvement in vector design can lead to more specific cell based delivery and reduction of non specific delivery of the trans gene leading to the development of optimized molecular therapeutics to induce hair cell regeneration. Level of Evidence N/A Controlled basic science study.

Spatial and Age-Dependent Hair Cell Generation in the Postnatal Mammalian Utricle

“…Previous studies have showed that delivery of Atoh1 into the sensory and nonsensory region of vestibular organs could induce the formation of ectopic hair cells in vitro [30][31][32]. In the current study, by using the triallelic transgenic mice, we tested the effect of Atoh1 induction on hair cell generation in the vestibule in vivo.…”

“…Previously, it is reported that hair cells could be generated in the vestibule at a later stage [9-11, 20-22, 32]. In these studies, hair cell ablation was performed prior to infection of viral vectors containing an Atoh1 expression cassette [20,32]. It is possible that the loss of hair cells eliminates inhibitory factors or activates factors to endow the competency of hair cell generation in the vestibule.…”

Control of hair cell development by molecular pathways involving Atoh1, Hes1 and Hes5

“…Atoh1, a member of the bHLH family of transcription factors and a close homologue of the Drosophila proneural gene atonal, plays a significant role in development of various tissues and organs, such as CNS, intestine, and inner ear (Atkinson et al, 2014;Ben-Arie et al, 1997;Bermingham et al, 1999;Isaka et al, 1999;Ren et al, 2014;Staecker et al, 2014;Yang et al, 2001). During neural development, it involves not only in proneural development but also in neurogenesis as well (Akazawa et al, 1995;Bermingham et al, 1999).…”

Intracochlear Drug Injections through the Round Window Membrane: Measures to Improve Drug Retention

“…With applications by intracochlear injection into the cochlear apex [Salt et al 2006] or by injections through the RW membrane [Salt et al 2007, Hahn et al 2012], concentrations in perilymph were found to be higher, with lower variation and the substances were distributed more uniformly along scala tympani. Effective deliveries of viral vectors have been accomplished with single intracochlear injection procedures [Stover et al 1999; Kawamoto et al, 2001; Staecker et al, 2014]. Nevertheless, previous studies have suggested that the efficiency of delivery with injections though the RW membrane will be influenced by fluid leakage at the injection site [Salt et al, 2007].…”