Pou3f4-expressing Otic Mesenchyme Cells: A Novel Influence for Spiral Ganglion Neuron Survival

项目摘要 听觉功能取决于大脑中特定细胞的精确连接模式和正常功能。 耳蜗包括螺旋神经节神经元（SGN）和它们的突触前伙伴毛细胞。赤字 这些细胞的发育、存活或功能是听力障碍和耳蜗植入术疗效的基础。 植入物. SGN特别形成外周和中枢听觉系统之间的传入连接。 SGN对正常听力至关重要，特别容易受到损害。我们最近发现， Pou3f4是一种由耳间充质细胞表达的转录因子，通常是SGN存活所必需的 在出生后的发育过程中。Pou3f4突变导致人类听力损失和小鼠Pou3f4的丧失 模型导致的形态缺陷，在耳间充质和听力障碍，除了减少 神经元存活尽管如此，我们对Pou3f4如何调节这些过程的了解有限，因为 Pou3f4在耳蜗中的转录靶点还不清楚。 如本提案所述，我将首先通过以下方法确定Pou3f4在SGN存活中的功能： 定义了耳间充质细胞和SGN之间的作用机制。第二，我会调查 利用基因治疗，耳间充质细胞促进SGN存活的潜力。关于受指导者 研究培训方面的这一奖项，我将接受额外的培训，在病毒介导的基因治疗， 听觉脑干反应技术，通过共同导师博士罗纳Hertzano。我们预测 Pou3f4的下游靶标包括影响SGN存活的营养或其他环境线索。我会 采用直接的方法来确定这些机制，使用分子技术和肝脏的组合， 成像实验与此同时，我们的合作者将采取一种无偏见的、基于“组学”的方法 识别Pou3f4目标此外，我将在体外和体内重新引入Pou3f4及其靶点， 确定间充质细胞促进SGN存活的程度。我将是第一个定义 耳间充质细胞如何与SGN相互作用以影响正常发育的耳蜗中的神经元存活， 同时也是最早确定耳间充质细胞靶向程度的研究之一 在治疗上有助于促进成人SGN的存活。因此，这项工作探讨了一个新的问题，将 补充其他人正在进行的神经营养素，基因治疗，细胞替代策略和听觉 修复术.本计划的研究、指导和职业发展方面将有助于准备 成功的独立研究生涯。

PROJECT SUMMARY Hearing function depends on the precise connectivity patterns and proper function of specialized cells in the cochlea including spiral ganglion neurons (SGNs) and their presynaptic partners, hair cells. Deficits in the development, survival or function of these cells underlie hearing impairment and the efficacy of cochlear implants. SGNs in particular form the afferent connection between the peripheral and central auditory systems. SGNs are crucial for proper hearing and are particularly vulnerable to damage. We showed recently that Pou3f4, a transcription factor expressed by otic mesenchyme cells, is normally required for SGN survival during postnatal development. Mutations in Pou3f4 cause human hearing loss, and loss of Pou3f4 in mouse models leads to morphological defects in otic mesenchyme and hearing impairment in addition to decreased neuronal survival. Still, we have a limited understanding of how Pou3f4 regulates these processes because the transcriptional targets of Pou3f4 in the cochlea are not well understood. As described in this proposal, I will first seek to determine the function of Pou3f4 in SGN survival by defining the mechanism of action between otic mesenchyme cells and SGNs. Second, I will investigate the potential for otic mesenchyme cells to promote SGN survival using gene therapy. With respect to the mentored research training aspects of this award, I will receive additional training in virus-mediated gene therapy and auditory brainstem response techniques through co-mentorship by Dr. Ronna Hertzano. We predict that downstream targets of Pou3f4 include trophic or other environmental cues that influence SGN survival. I will take a direct approach to determining these mechanisms using a combination of molecular techniques and live imaging experiments. At the same time, our collaborators will be taking an unbiased, “omics”-based approach to identifying Pou3f4 targets. Additionally, I will reintroduce Pou3f4 and its targets both in vitro and in vivo to determine the extent to which mesenchyme cells promote SGN survival. I will be the one of the first to define how otic mesenchyme cells interact with SGNs to impact neuronal survival in the normally developing cochlea, as well as one of the first to determine the extent to which otic mesenchyme cells could be targeted therapeutically to help promote SGN survival in adults. Thus, this work explores a novel question that will complement ongoing work by others on neurotrophins, gene therapy, cell replacement strategies, and auditory prosthetics. The proposed research, mentoring, and career development aspects of this plan will help prepare me for a successful independent research career.