Wnt activation followed by Notch inhibition promotes mitotic hair cell regeneration in the postnatal mouse cochlea

// Wenli Ni 1, 2, * , Shan Zeng 1, 2, * , Wenyan Li 1, 2 , Yan Chen 1, 3, 4 , Shasha Zhang 5, 6 , Mingliang Tang 5, 6 , Shan Sun 1, 3, 4 , Renjie Chai 5, 6 , Huawei Li 1, 2, 3 1 Otorhinolaryngology Department of The Affiliated Eye and ENT Hospital, State Key Laboratory of Medical Neurobiology, Fudan University, Shanghai, PR China 2 Institutes of Biomedical Sciences, Fudan University, Shanghai, PR China 3 Central Laboratory, Affiliated Eye and ENT Hospital of Fudan University, Shanghai, PR China 4 Key Laboratory of Hearing Medicine of The National Health and Family Planning Commission, Shanghai, PR China 5 Key Laboratory for Developmental Genes and Human Disease, Ministry of Education, Institute of Life Sciences, Southeast University, Nanjing, PR China 6 Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, PR China * These authors contributed equally to this work Correspondence to: Huawei Li, email: hwli@shmu.edu.cn Renjie Chai, email: renjiec@seu.edu.cn Keywords: Wnt, Notch, proliferation, regeneration, lineage tracing Received: January 25, 2016      Accepted: June 29, 2016      Published: August 22, 2016 ABSTRACT Hair cell (HC) loss is the main cause of permanent hearing loss in mammals. Previous studies have reported that in neonatal mice cochleae, Wnt activation promotes supporting cell (SC) proliferation and Notch inhibition promotes the trans-differentiation of SCs into HCs. However, Wnt activation alone fails to regenerate significant amounts of new HCs, Notch inhibition alone regenerates the HCs at the cost of exhausting the SC population, which leads to the death of the newly regenerated HCs. Mitotic HC regeneration might preserve the SC number while regenerating the HCs, which could be a better approach for long-term HC regeneration. We present a two-step gene manipulation, Wnt activation followed by Notch inhibition, to accomplish mitotic regeneration of HCs while partially preserving the SC number. We show that Wnt activation followed by Notch inhibition strongly promotes the mitotic regeneration of new HCs in both normal and neomycin-damaged cochleae while partially preserving the SC number. Lineage tracing shows that the majority of the mitotically regenerated HCs are derived specifically from the Lgr5 + progenitors with or without HC damage. Our findings suggest that the co-regulation of Wnt and Notch signaling might provide a better approach to mitotically regenerate HCs from Lgr5 + progenitor cells.