Molecular embryology of the mammalian inner ear

描述（由申请人提供）：我们实验室的长期目标是确定基于基因和细胞的策略，以恢复功能失调的内耳的听力和平衡。短期目标是推进我们对小鼠内耳形态发生、模式形成和细胞命运特化的理解。我们使用小鼠作为模型系统，因为存在一系列不断扩大的天然和诱导基因突变，这些突变可作为人类内耳功能障碍的准确范例。我们的总体技术方法依赖于实验胚胎学，一系列手术，成像，显微注射和分子技术，允许在子宫内进入发育中的小鼠内耳，并对产生听觉和前庭感觉结构的前体细胞进行遗传操作。在这一建议中，我们的目标是：1）在体内绘制小鼠耳泡的命运图2）确定耳前体细胞做出的细胞命运选择的类型和这些选择的时间3）确定神经上皮前体细胞对分化的内耳的克隆贡献。命运图描述了内耳成熟时细胞的不同群体。发育生物学家使用命运图来了解祖基因在发育中的组织或器官中的表达如何驱动成熟并导致形式和功能的获得。不可磨灭的标记前体与已知的基因表达域的交叉可以教会我们识别感觉器官和指定细胞命运所需的分子信号。谱系分析，另一方面，确定分化的细胞个别耳前体，并揭示其在成熟的感觉器官的位置。克隆关系将告诉我们，遗传定义的前体池是否在发育早期被编程，以产生内耳的总体解剖结构和感觉贴片。命运定位和谱系分析的结果是对祖细胞身份和行为的遗传调控的更深入理解。而且这种监管可能比最初设想的要复杂得多。最近的遗传命运作图数据表明，神经管和可能的神经嵴的神经上皮祖细胞贡献的前体细胞的耳上皮细胞产生的感觉和非感觉细胞分化。这些新的数据坚定地挑战了内耳完全从基板外胚层发育的观念，并暗示神经外胚层可能独特地参与模式化和细胞命运规范。对这些遗传上完全可分离的神经上皮前体细胞群的克隆分析将推进我们关于小鼠内耳胚胎起源的核心知识。对内耳祖细胞的命运、谱系和行为的更完整的理解将为旨在恢复听力和平衡的基于基因和细胞的策略的设计提供信息。

DESCRIPTION (provided by applicant): The long term goal of our laboratory is to define gene- and cell-based strategies that restore hearing and balance in the dysfunctional inner ear. The short term goals are to advance our understanding of morphogenesis, pattern formation, and cell fate specification in the mouse inner ear. We use the mouse as a model system because an ever-expanding array of natural and induced genetic mutations exist that serve as accurate paradigms for human inner ear dysfunction. Our overarching technical approach relies on experimental embryology, a palette of surgical, imaging, microinjection and molecular techniques that permit access to the developing mouse inner ear in utero and enable genetic manipulation of precursor cells that give rise to the auditory and vestibular sensory structures. I this proposal, we aim: 1) to fate map the mouse otic vesicle in vivo~ 2) to define the types of cel fate choices otic precursors make and the timing of those choices~ and 3) to define the clonal contributions of neuroepithelial progenitors to the differentiated inner ear. A fate map describes what distinct populations of cells become as the inner ear matures. Developmental biologists use fate maps to understand how progenitor gene expression in a developing tissue or organ drives maturation and results in the acquisition of form and function. Intersection of indelibly labeled precursors with known domains of gene expression can teach us the molecular signals required to pattern sensory organs and specify cell fate. Lineage analysis, on the other hand, identifies the differentiated cells an individual otic precursor makes and reveals their location i the mature sensory organ. Clonal relationships will teach us if genetically defined pools of precursors are programmed early on in development to give rise to gross anatomical structures in the inner ear and to the sensory patches. The output of fate mapping and lineage analysis is a deeper understanding of the genetic regulation of progenitor cell identity and behavior. And that regulation promises to be far more complex that originally envisioned. Recent genetic fate mapping data show that neuroepithelial progenitor cells from the neural tube and likely the neural crest contribute precursor cells to the otic epithelium that generate sensory and nonsensory cells upon differentiation. These novel data firmly challenge the precept that the inner ear develops exclusively from placodal ectoderm and imply that neural ectoderm may uniquely participate in patterning and cell fate specification. Clonal analysis of these geneticall separable neuroepithelial precursor populations will advance our core knowledge about the embryonic origins of the mouse inner ear. A more complete understanding of the fate, lineage, and behavior of inner ear progenitor cells will inform the design of gene- and cell-based strategies aimed at the restoration of hearing and balance.