miR-204靶向 TMPRSS3调控耳蜗螺旋神经节神经元存活和功能的实验研究

Sensorineural hearing loss (SNHL) is the most common cause of hearing impairment. One of the essential steps to prevent progressive hearing loss is to protect spiral ganglion neurons (SGNs) from ongoing degeneration. However, there is no effective treatment to prevent the progressive degeneration or loss of spiral ganglion neurons. MicroRNAs and TMPRSS3 (transmembrane protease, serine 3) have been reported to be involved in development of SGNs and genesis of SNHL. By using bioinformatic algorithms, we identified a putative miR-204 binding site within 3′-UTR of TMPRSS3 gene, which was confirmed by our preliminary experiment. To study whether miR-204 regulates SGNs survival by targeting TMPRSS3, We will employ in situ hybridization, TUNEL assay, LIVE/DEAD assay, siRNA etc techniques to investigate the role of miR-204 and TMPRSS3 in the survival of SGNs. By finishing this project, we will show whether miR-204 plays a role in SNHL by control the expression of TMPRSS3. Our results will provide new potential therapeutic target in prevention and treatment of SNHL.

感音神经性耳聋(SNHL)是最常见的听力受损原因。已知耳蜗螺旋神经节神经元(SGNs)的进行性丢失是SNHL听力进一步恶化的关键环节，然而对如何改善SGNs的进行性丢失却仍无有效方法。有资料表明microRNAs 和跨膜蛋白酶TMPRSS3与SGNs的发育和SNHL的发生密切相关。我们通过生物信息学软件和前期实验发现TMPRSS3的3'-UTR序列具有miR-204结合位点，但miR-204是否能够通过TMPRSS3调控SGNs尚不清楚。本项目拟采用原位杂交、TUNEL染色、LIVE/DEAD染色、siRNA、病毒载体、荧光素酶报告系统分析等实验技术来探索miR-204和TMPRSS3对SGNs神经元存活的影响以及进一步阐明两者之间的靶向关系和可能机制。本研究将进一步揭示miR-204和TMPRSS3与SNHL的相关性，为有效防治SNHL提供新思路和和药物作用的新靶点。

感音神经性耳聋的典型特征是螺旋神经节神经元（SGNs）的变性，TMPRSS3与SGNs的发育和SNHL的发生有紧密联系，而miR-204-5p可通过抑制TMPRSS3(transmembrane protease, serine 3)的表达来降低SGNs的细胞活性。申请人发现了一个miR-204-5p的重要上游调控因子lncRNA-EBLN3P，可通过竞争性结合miR-204-5p以及调节TMPRSS3的表达来修复受损螺旋神经节的功能，能够成为感音神经性耳聋潜在的治疗靶点。此外，申请人通过RNA二代测序的方法挖掘TMPRSS3的潜在靶基因调节群，重点考察了TMPRSS3下游circRNA表达谱的改变，根据circRNA测序的结果，TMPRSS3的过表达诱导了更多的circRNA表达上调，而circ-Slc41a2（chr10：82744115 | 82767120）就是表达上调最显著的circRNA之一。该结果提示，circ-Slc41a2是TMPRSS3的下游靶基因。本项目按计划开展了系统深入的研究，取得了预期的成果。在本项目的支持下，已发表SCI论文2篇。