协同调控Sox2基因和Wnt信号通路促进耳蜗毛细胞损伤后再生及存活的机制研究

At present, we have yet found an effective way to biologically achieve cochlear hair cell regeneration and its electrophysiological functional recovery. Previous studies have repeatedly reported that Sox2 and Wnt signaling pathway play crucial roles in hair cell differentiation and cell cycle regulation during cochlear development. However, their functions during cochlear hair cell regeneration after damage remain to be determined. Our preliminary data have shown that through gene editing， stabilization of β-catenin alone enhances the degree of supporting cell proliferation, but fail to increase the number of transitional cell formation. Decreased Sox2 expression level in neonatal cochlea after damage promotes hair cell regeneration through both mitotic proliferation and direct transdifferentiation. Therefore, in the present research proposal, based on the purpose of pharmaceutical exploration, we will seek to answer the following questions: (1) Whether the mitotic proliferation and transitional cell formation in the damaged cochlea can be further promoted by the co-regulation of Sox2 and Wnt in vitro and in vivo with local drug delivery. (2) Dissociate the molecular mechanisms of the interaction between Sox2 and Wnt in directing hair cell regeneration. (3) Whether the co-regulation of Sox2 and Wnt can promote the survival and maturation of the regenerated hair cells. (4) Whether the co-regulation of Sox2 and Wnt can improve the audiologic function after hair cell damage. Our research findings will provide insights guiding future designs of combinatorial approaches to stimulate mammalian cochlear regeneration to reverse hearing loss.

目前尚未找到实现耳蜗毛细胞损伤后再生及听功能修复的有效办法。既往研究报道Sox2基因和Wnt信号通路在耳蜗毛细胞分化和细胞周期调控过程中均发挥了重要作用。然而它们在毛细胞损伤后再生过程中的调控机制尚不清楚。申请人前期研究发现：通过基因编辑技术在耳蜗中过表达Wnt信号通路可以在毛细胞损伤后促进支持细胞增殖，但不改变其转分化水平；下调 Sox2基因表达可以从增殖和转分化两方面显著促进新生小鼠耳蜗毛细胞损伤后再生。本项目将在前期研究发现的基础上，从基因治疗药物研发的角度出发，通过离体培养的新生小鼠耳蜗毛细胞损伤模型和活体小鼠耳蜗圆窗注射局部给药技术来研究：下调Sox2基因协同Wnt信号通路过表达是否可以从增殖和转分化两方面促进耳蜗毛细胞损伤后再生及成熟分化、同时延长再生毛细胞存活时间并部分修复受损的听功能。我们的研究成果将为毛细胞损伤导致感音神经性聋的基因治疗提供新靶点和理论依据。

内耳毛细胞的发育异常或不可逆损失是导致感音神经性聋的主要原因之一。因此，探明耳蜗毛细胞分化命运决定的分子机制并探索如何利用分子调控手段促进毛细胞损伤后再生成为耳聋研究领域的关键问题。在本课题中，我们重点研究并探明了Sox2基因和Wnt信号通路在耳蜗感觉上皮细胞分化命运决定和细胞周期调控过程中的具体功能及相互调控关系。进一步阐明Sox2了基因在耳蜗毛细胞损伤后再生过程中的核心作用并摸索其最适表达水平。初步探索通过协同调控Wnt信号通路、Sox2基因作为新靶点来扩增再生毛细胞前体细胞并提高其转分化水平，进而促进耳蜗毛细胞损伤后再生及存活，最终实现受损听功能修复的新策略。我们的后续研究将更完善的阐明Sox2基因和Notch信号通路在耳蜗发育及毛细胞再生过程中的相互作用机制，并为探索促进毛细胞损伤后再生的新策略及寻找感音神经性聋治疗新靶点提供理论依据。同时，我们还发现毛细胞在不同外界刺激因素下发生损伤和死亡的机制不尽相同，那么保护毛细胞的策略则需要更加精准，我们初步探索和总结了保护毛细胞的策略并探明了其具体机制，研究成果具有较高的临床价值及广阔的应用前景。