Regulation of Inner Ear Stem Cells

描述(申请人提供)：该奖项的主要目标是通过建立一个成功的研究项目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，同时也对同一主题进行基础科学研究，这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儿科耳科，我经常遇到患有听力损失的儿童。作为一名内科医生，我注意到了我们目前的治疗方法中的一个重大缺陷，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，以阐明恢复听力的疗法。为了巩固我以前的研究经验，这个奖项旨在通过实验设计、技术和拨款写作方面的实践培训和密切指导，帮助我发展成为一名独立的调查员。由于Wnt信号在维持其他器官系统中的干细胞种群中起主要作用，我们假设规范的Wnt通路参与了耳蜗祖细胞/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无法在一生中保持再生能力的原因。在这个五年计划的过程中，我们计划进行一系列实验，以检验两个尚未探索的假设：1)WNT反应细胞是内源性耳蜗干/祖细胞，2)WNT激活促进耳蜗支持细胞的增殖。作为通过流式细胞仪分离的单个细胞，不同的耳蜗细胞群，包括Wnt反应细胞和各种耳蜗支承细胞，通过球法评估它们的自我更新能力以及它们在体外产生新的毛细胞和支持细胞的能力。利用器官培养中的增殖和细胞谱系分析标记物，我们将研究Wnt激动剂、有丝分裂原和新霉素诱导的毛细胞丢失对这些细胞的影响。在本奖项的最后，我们将评估和报道规范的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途径是否是听力恢复的有前景的治疗靶点。我们研究的长期目标是维持或激活内源性干细胞的储备，能够通过毛细胞/支持细胞再生来恢复听力和平衡障碍。

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.