AAV介导Wnt，Atoh1，Gfi1和Pou4f3协同调控内耳干细胞促进功能性毛细胞再生和前庭功能恢复的研究

毛细胞损失是造成前庭功能障碍的主要原因。前庭椭圆囊毛细胞在损伤后仅具有有限的自发性再生能力，且不能再生成熟毛细胞。申请人前期研究表明，在椭圆囊毛细胞损伤后，上调Wnt信号通路可以促进内耳支持细胞增殖，过表达Atoh1可促进支持细胞分化新生毛细胞。但单独调控Wnt或Atoh1却不能促进功能性毛细胞再生和前庭功能修复。本项目在前期研究基础上，拟通过AAV介导Wnt，Atoh1，Gfi1和Pou4f3协同调控内耳干细胞促进功能性毛细胞再生及前庭功能修复。拟实现目标：1.使用Pou4f3-DTR转基因小鼠建立小鼠毛细胞和前庭功能损伤模型。2.建立AAV介导的前庭椭圆囊基因治疗系统。3.以AAV为载体，协同调控Wnt，Atoh1，Gfi1和Pou4f3这4个信号通路。4.研究功能性毛细胞再生和前庭功能修复。本项目将为临床上以AAV为载体，通过基因治疗恢复前庭功能奠定理论和临床应用基础。

Dysfunction or loss of hair cells is the primary causes of balance dysfunction. Limited hair cell regeneration takes place in the mammalian utricles, while few acquires mature hair cell features. Preliminary studies have been showed that upregulation of Wnt signal pathway can induce supporting cell proliferation and overexpression of Atoh1 can promote supporting cells transdifferentiate to hair cells after hair cell damage in mouse utricle. However, no functional hair cell regeneration and vestibular function recovery have been observed by activating Wnt or Atoh1 alone. Based on our previous findings, this proposal will investigate functional hair cell regeneration and vestibular function recovery after AAV-mediated co-regulation of Wnt，Atoh1，Gfi1 and Pou4f3 signal pathways in damaged neonatal and adult mouse vestibular system. First, transgenic mice will be used to establish neonatal and adult mice utricle hair cell and vestibular function damage model. Then, different subtypes of AAV will be screened to build an AAV-mediated high transduction gene delivery system in mouse vestibular system. Next, AAV-β-catenin-GFP and AAV-Atoh1-Gfi1-Pou4f3 will be delivered to inner ear vestibular system. Lastly, functional hair cell regeneration and vestibular function recovery will be examined. This proposal will establish an AAV-mediated gene therapy system to induce hair cell regeneration and function recovery, focusing on translational research from basic science to clinical application with the goal of curing balance disorders resulting from inner ear vestibular hair cell loss.