Gene Transfer Tools for Delivery and Investigation of Inner Ear MicroRNAs

听力损失会给患者带来巨大的挫败感。在某些情况下，这些人会感到孤独和沮丧，因为他们发现很难与周围的人沟通。目前，治疗听力损失的能力严重依赖于助听器的使用；然而，这些设备并不能将听力恢复到标准水平。这项提案的重点是创造和研究潜在的治疗工具，这些工具可以恢复因毛细胞缺失或丧失而导致听力损失的人的敏锐度和声音检测能力。内耳感觉器官毛细胞下方的支持细胞为毛细胞再生提供了良好的细胞来源。毛细胞和支持细胞在发育过程中由相同的祖细胞池产生，在鸟类中，毛细胞的损失刺激支持细胞分裂并再生更多的毛细胞。支持细胞与毛细胞命运选择的潜在因素可能为治疗耳聋和平衡障碍提供潜在的试剂。先前的研究主要集中在为成年支持细胞提供促毛细胞转录因子Atoh1，以迫使这些细胞转变为毛细胞的命运，但另一种方法是识别并提供反支持细胞因子，以帮助将支持细胞转变为新的命运。目前，miRNA-183家族的成员可能是作为反支持细胞信号的候选者。另一种miRNA miR-9也可能通过降低转录因子HES-1的水平来抑制支持细胞的命运，而转录因子HES-1通常会负向调节促毛基因Atoh1。

Hearing loss can cause great frustration amongst its sufferers. In some cases, these individuals contend with feelings of loneliness and depression because they find it difficult to communicate with the people around them. Currently, the ability to treat hearing loss relies heavily on the use of hearing aids; however, these devices do not restore hearing acuity to standard levels. This proposal focuses on creating and investigating potential therapeutic tools that could restore acuity and detection of sound to people suffering from hearing loss caused by a lack or loss of hair cells. The supporting cells located beneath the hair cells in sensory organs of the inner ear offer an excellent cellular source to target for hair cell regeneration. Hair cells and supporting cells are created from the same progenitor pool during development and in birds, hair cell loss stimulates supporting cells to divide and regenerate additional hair cells. The factors underlying the fate choice of supporting cell vs. hair cell may provide potential reagents for treating deafness and balance disorders. Previous research has focused on supplying adult supporting cells with the pro-hair cell transcription factor, Atoh1, to force those cells to switch to a hair cell fate, but an alternative approach is to identify and supply anti-supporting cell factors to assist in converting supporting cells to a new fate. Presently, members of the miRNA-183 family are likely candidates to serve as anti-supporting cell signals. Another miRNA, miR-9, may also prove to be influential in discouraging a supporting cell fate by decreasing the levels of a transcription factor, HES-1, that normally negatively regulates the pro-hair gene, Atoh1. The goals of this project are to investigate the role and mechanism by which miR-9 and the miR-183 family work during ear development as well as to construct new gene therapy tools encoding both the pro-hair cell factor (Atoh1) and presumed anti-supporting cell factors (miRNAs). Aim 1 will focus on the construction of a dual expression RCAS vector for Atoh1 and the miR-183 family or miR-9, along with an alternate version that encodes only the miRNAs. Aim 2 will involve delivery of these vectors into a chicken otocyst to observe their effects on hair cell development. With this proposal, we hope to elucidate the role of the 183 family of miRNAs and miR-9 in hair cell development along with one of its possible targets, HES-1, and create tools to generate new hair cells that may be used to treat cases of human deafness in the future.