Regulation of Inner Ear Stem Cells

描述（由申请人提供）：这个奖项的主要目标是通过建立一个成功的研究计划来促进我作为外科医生科学家的职业生涯。我长期以来对治疗听力障碍的兴趣，以及同时对同一主题进行基础科学研究，促使我决定成为一名儿科耳鼻喉科科学家。由于我的临床实践主要集中在小儿耳科，我经常遇到患有听力损失的儿童。作为一名医生，我注意到我们目前治疗方法的一个主要缺陷，即我们无法逆转感音神经性听力损失。作为一名科学家，我希望建立一个研究项目，为恢复听力的疗法提供帮助。在我以前的研究经验的基础上，这个奖项旨在帮助我通过实践培训和对实验设计，技术和拨款写作的密切指导，发展成为一名独立的研究人员。由于Wnt信号在维持其他器官系统中的干细胞群体中起着重要作用，我们假设经典Wnt通路参与耳蜗祖细胞/干细胞的维持。同样，我们假设Wnt信号或反应性的丧失是哺乳动物耳蜗在一生中无法维持再生能力的原因。在五年计划的过程中，我们建议进行一系列实验，旨在测试两个未探索的假设：1）Wnt反应细胞是内源性耳蜗干/祖细胞，和2）Wnt激活促进耳蜗支持细胞的增殖。作为通过流式细胞术分离的单细胞，通过球体测定评估不同耳蜗细胞群（包括Wnt应答细胞和各种耳蜗支持细胞）的自我更新能力以及它们在体外产生新毛细胞和支持细胞的能力。在器官型培养物中使用增殖和细胞谱系分析的标记物，我们将研究Wnt激动剂、有丝分裂原和新霉素诱导的毛细胞损失对这些细胞的影响。在本奖项结束时，我们将评估和报告经典Wnt信号在调节内源性Wnt反应细胞和耳蜗支持细胞中的作用，以及Wnt通路是否是听力恢复的有希望的治疗靶点。我们研究的长期目标是维持或激活能够通过毛细胞/支持细胞再生恢复听力和平衡障碍的内源性干细胞储备。

DESCRIPTION (provided by applicant): The main goal of this award is to further my career as a surgeon-scientist by establishing a successful research program. My longstanding interest in treating hearing disorders and, in parallel, performing basic science research on the same topic has fueled my decision to become a pediatric otolaryngologist-scientist. Since my clinical practice focuses on pediatric otology, I am frequently confronted with children suffering from hearing loss. As a physician, I have noticed a major deficiency in our current treatment approach where we cannot reverse sensorineural hearing loss. As a scientist, I hope to establish a research program that will shed light on therapies that restore hearing. To build on my previous research experience, this award is designed to help me develop into an independent investigator through hands-on training and close guidance on experimental designs, techniques, and grant writing. Because Wnt signaling plays major roles in maintaining stem cell populations in other organ systems, we hypothesize that canonical Wnt pathway is involved in the maintenance of cochlear progenitor/stem cells. Likewise, we hypothesize that loss of Wnt signaling or responsiveness is responsible for the inability of the mammalian cochlea to maintain regenerative capacity throughout life. Over the course of a five-year plan, we propose to conduct a series of experiments designed to test two unexplored hypotheses: 1) Wnt responsive cells are endogenous cochlear stem/progenitor cells, and 2) Wnt activation promotes proliferation in cochlear supporting cells. As single cells isolated via flow cytometry, different cochlear cell populations, including Wnt responsive cells and various cochlear supporting cells, are assessed for their ability to self-renew by sphere assays and their ability to generate new hair cells and supporting cells in vitro. Using markers for proliferation and cell lineage analysis in organotypic cultures, we will investigate the effects of Wnt agonists, mitogens, and neomycin-induced hair cell loss on these cells. At the end of this award, we will have evaluated and reported the roles of canonical Wnt signaling in regulating the endogenous Wnt responsive cells and cochlear supporting cells, and whether the Wnt pathway is a promising therapeutic target for hearing restoration. The long-term goal of our research is to maintain or activate a reserve of endogenous stem cells capable of restoring hearing and balance disorders through hair cell/supporting cell regeneration.