(804) AAVie Tropism is Preserved in Post-Deafened Adult Mouse

Introduction: Damage to spiral ganglion neurons (SGNs) results in auditory neuropathy and neural deafness, with hearing aids and cochlear implants exhibiting limited efficacy. These constraints underscore the necessity for regenerative approaches, including direct neural reprogramming gene therapy. Cochlear glial cells are an appealing target for reprogramming due to their proximity to auditory neurons and ability to respond to injury; however, vectors targeting cochlear glial cells post-injury have not been investigated. We previously identified the adeno-associated virus (AAV) serotype K558Rie variant (AAVie) as a potential vector for targeting cochlear glial cells. Here, we investigated AAVie-mediated cochlear transduction in acutely and chronically deafened mice using a temporally controlled SGN ablation strategy.

Methods: Sox10-Cre mice crossed with Ai reporter mice on a C57 background were used. SGN ablation was conducted by injecting AAVretro-hSyn-DTR in neonatal (P2) mice. SGN ablation was induced by diphtheria toxin (DT) 20 ng/g administration at P30. ABRs were obtained before DT administration. Cochlear tissues were analyzed 1 and 4 weeks after DT to evaluate acute and chronic glia responses. Subsequently, AAVie-GFAP-mScarlet was delivered via the posterior semicircular canal at these time points, and tissues were harvested 2 weeks later. Immunohistochemistry was performed using TUJ1, Sox2, and Sox10.

Results: DT administration resulted in elevated ABR thresholds and approximately a 77% reduction of TUJ1-positive SGN density at 2 weeks. Following acute SGN loss, Sox2-positive glial cells increased in number, indicating activation of a glial response. In both acute and chronic conditions, AAVie-GFAP-mScarlet expression was observed in Sox2- and Sox10-positive glial cells.

Conclusions: Our results demonstrate the proof-of-principle of selectively transducing glial cells following the acute and chronic phases of gliosis with the AAVie vector. These findings provide a foundation for future cochlear glia-based regenerative strategies.