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折感。在某些情况下，这些人与孤独和抑郁的感觉作斗争，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在很大程度上依赖于助听器的使用;然而，这些设备不能将听力恢复到标准水平。该提案的重点是创建和研究潜在的治疗工具，可以恢复因毛细胞缺乏或丧失而导致听力损失的人的敏锐度和声音检测。位于内耳感觉器官中毛细胞下方的支持细胞为毛细胞再生提供了极好的细胞来源。毛细胞和支持细胞在发育过程中由同一祖细胞库产生，在鸟类中，毛细胞损失刺激支持细胞分裂并再生额外的毛细胞。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。以前的研究集中在为成人支持细胞提供前毛细胞转录因子Atoh 1，以迫使这些细胞转换为毛细胞命运，但另一种方法是识别和提供反支持细胞因子，以帮助将支持细胞转换为新的命运。目前，miRNA-183家族的成员可能是作为抗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的水平而被证明在阻止支持细胞的命运方面有影响力，HES-1通常负调节前毛发基因Atoh 1。 该项目的目标是研究miR-9和miR-183家族在耳发育过程中的作用和机制，以及构建编码前毛细胞因子（Atoh 1）和推定的抗支持细胞因子（miRNAs）的新基因治疗工具。目标1将集中于Atoh 1和miR-183家族或miR-9的双表达RCAS载体的构建，沿着仅编码miRNA的替代版本。目的2将涉及这些载体进入鸡耳囊，以观察其对毛细胞发育的影响。有了这个提议，我们希望阐明183家族的miRNAs和miR-9在毛细胞发育中的作用，沿着其可能的靶点之一，HES-1，并创造工具来产生新的毛细胞，这些毛细胞可能在未来用于治疗人类耳聋病例。

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.