Rps14 Regulates The Proliferation And Differentiation Of Inner Ear Stem Cells Into Hair Cells

Sensorineural Hearing Loss(SNHL)is caused by the impairment of the inner ear,cochlear nerve,brainstem auditory pathway or auditory center.According to relevant statistics,the vast majority of patients with hearing impairment in China are suffering from SNHL,which is mostly caused by the pathological changes of the cochlea.And,there is no effective treatment for SNHL in clinic due to nonreproducible hair cells after damage.The application of hearing aids and cochlear implants commonly used in SNHL treatment in clinic are usually of limited by the severity of injury of the auditory pathway,especially the cochlea.The discovery of inner ear stem cells made hair cell regeneration possible.Stem cells have selfrenewal and differentiation potential,including embryonic stem cells and adult stem cells.Studies have shown that Lgr5-positive supporting cells in the inner ear have the properties of inner ear stem cells and can regenerate hair cells after injury.The differentiation of Lgr5-positive supporting cells into hair cells is regulated by various signaling pathways,such as the Wnt and Notch signaling pathways.However,regenerated hair cells still do not have the structural and physiological functions of mature hair cells.Therefore,revealing the process of hair cell development and differentiation and screening new genes that regulate the differentiation of inner ear stem cells into hair cells are very important for functional hair cell regeneration.Earlier,we studied the difference in expression profile between Lgr5-positive and Lgr5-negative supporting cells,and found that Rps14 may have the potential to promote the proliferation and differentiation of inner ear stem cells into hair cells.Therefore,in this study,adeno-associated virus AAV-ie,targeting inner ear stem cells,was used to deliver the exogenous Rps14 into the inner ear stem cells in neonatal mouse cochlea to investigate the proliferation and differentiation of inner ear stem cells.Overexpression of Rps14 promoted the proliferation of stem cells and produced more hair cells in cochlea organoids under two or threedimensional culture conditions.Next,we infected AAV-ie-Rps14 into the mouse cochlea by round window injection,and the overexpression of Rps14 in supporting cells promoted the proliferation ability of the inner ear stem cells,and also the hair cells regeneration.Mechanistically,we used RNAsequence analysis to determine the signaling pathway involved in the proliferation and differentiation of inner ear stem cells regulated by Rps14 overexpression.We found that Rps14 promotes the proliferation of inner ear stem cells by activating the Wnt signaling pathway,regulating the cell cycle factors like Id3,and inducing the regeneration of new hair cells by regulating Notch and TGF-βsignaling pathways.In summary,we found that strength the expression of Rps14 effectively regulated the transformation ability of inner ear stem cells into hair cells.The results of this study will lay a theoretical and clinical foundation for the application of AAV-mediated gene therapy in auditory function restoration.