Molecular Basis For The Morphogenesis Of The Inner Ear

内耳形态发生的分子基础

• 批准号: 7593329
• 负责人: Doris Wu
• 金额: $ 183.45万
• 依托单位: NATIONAL INSTITUTE ON DEAFNESS AND OTHER COMMUNICATION DISORDERS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7593329
• 关键词: Activator Appliances; Alleles; Animal Model; Ankylosis; Area; Auditory; Bone Morphogenetic Proteins; Branchial arch structure; Chickens; Cochlear duct; Complex; Conductive hearing loss; Defect; Development; Disease; Distal; Dorsal; Ear; Ear ossicles; Embryo; Embryonic Development; Equilibrium; Erinaceidae; Exons; Floor; Genes; Genetic; Goals; Human; Knock-out; Knockout Mice; Labyrinth; Manuscripts; Modeling; Molecular; Morphogenesis; Mus; Mutant Strains Mice; Mutation; Organ; Paper; Patients; Phenotype; Positioning Attribute; Proteins; Publishing; Role; Saccule structure; Sensory; Signal Transduction; Skeletal system; Source; Specific qualifier value; Stapes; Stapes Surgery; Structure; Syndrome; Testing; Tympanic cavity; Vestibule; base; bone; deafness; design; hearing impairment; hindbrain; malformation; middle ear; mutant; otoconia; outcome forecast; prospective; sample fixation; sound

Our major accomplishments are in the following areas: 1) Inner ear analyses of Noggin knockout mouse embryos (manuscript published) Mutations of the NOGGIN gene in humans are associated with several autosomal dominant disorders such as symphalangism and multiple synotoses. These syndromes are characterized by skeletal defects and synotoses, which include conductive hearing loss. Noggin encodes a protein that antagonizes activities of Bone morphogenetic proteins (BMP). To understand the role of Noggin in ear development, we analyzed the ear phenotypes of Noggin homozygous mice. We published a paper this year describing the inner ears of Noggin knockout embryos. We show that the inner ear phenotypes observed in Noggin knockout embryos are largely due to a misalignment of the otocysts with the hindbrain during early embryogenesis. We propose that this misalignment disrupted the normal hindbrain signaling that is required for proper inner ear morphogenesis. We tested this hypothesis by generating similar misalignment in chicken embryo surgically as what was observed in Noggin -/- embryos. The resulting inner ear phenotypes in chicken resemble those in Noggin mutants, in particularly the saccule and cochlear duct malformations. These results indicate that the inner ear phenotypes observed in Noggin mutants are largely attributed to improper formation of the body axis. 2) Noggin heterozygous mice an animal model for conductive hearing loss (manuscript submitted) We conducted a study in the Noggin heterozygous mice. We show that the Noggin heterozygous mice suffer from conductive hearing loss, similar to the human patients with NOGGIN mutations. Since Noggin is a single exon gene, our results strongly suggest that the autosomal dominant disorders associated with NOGGIN mutations in humans are due to haploinsufficiency of NOGGIN. We discovered that the heterozygous mice with hearing loss have an ectopic bone fragment connecting one of the middle ear ossicles, the stapes, to the wall of the tympanum. This fixation most likely impedes the mobility of the stapes during sound conduction leading to hearing loss. We concluded from our analyses that the ectopic bone fragment is not due to de novo synthesis. Most likely, the extra bone formed by an incomplete separation of the prospective stapes from its precursor in the second branchial arch due to unopposed BMP activities. Since the long-term prognosis is not ideal for patients with NOGGIN mutations who underwent corrective surgery for stapes ankylosis, these Noggin mutant mice will continue to serve as a good model for exploring better strategies to alleviate hearing loss. 3) Roe of Sonic hedgehog (Shh) and Gli proteins in inner ear development (manuscript published) From the analyses of mouse mutants with various genetic combinations of mutant alleles such as Shh, Gli2, Gli3, and Gli3 delta699, we show that several mechanisms are involved in Shh signal transduction in the inner ear. The formation of the distal cochlear duct, which is positioned closest to the source of Shh in the notchord and floor plate, requires robust Gli2 and Gli3 activator functions. The proximal region of the cochlear duct requires less Shh signaling to antagonize Gli3 repressors. The dorsal vestibule of the inner ear requires little Shh signaling but Gli3 repressor is important for the formation of this region. Thus the graded Shh signaling from the ventral midline specifies various inner ear structures along the dorsal-ventral axis.

我们的主要成就体现在以下几个方面： 1）Noggin敲除小鼠胚胎的内耳分析（手稿已发表） 人类中 NOGGIN 基因的突变与多种常染色体显性遗传疾病有关，例如并指症和多合音。 这些综合征的特点是骨骼缺陷和听力损失，其中包括传导性听力损失。 Noggin 编码一种拮抗骨形态发生蛋白 (BMP) 活性的蛋白质。 为了了解 Noggin 在耳朵发育中的作用，我们分析了 Noggin 纯合小鼠的耳朵表型。 我们今年发表了一篇论文，描述了 Noggin 敲除胚胎的内耳。 我们发现，在 Noggin 敲除胚胎中观察到的内耳表型很大程度上是由于早期胚胎发生过程中耳囊与后脑的错位所致。 我们认为这种错位破坏了正常内耳形态发生所需的正常后脑信号传导。 我们通过手术在鸡胚胎中产生与在 Noggin -/- 胚胎中观察到的类似的错位来测试这一假设。 由此产生的鸡内耳表型与 Noggin 突变体相似，特别是球囊和耳蜗管畸形。 这些结果表明，在Noggin突变体中观察到的内耳表型很大程度上归因于身体轴的不正确形成。 2）Noggin杂合小鼠传导性听力损失动物模型（已提交手稿） 我们对 Noggin 杂合小鼠进行了一项研究。 我们发现 Noggin 杂合小鼠患有传导性听力损失，与携带 NOGGIN 突变的人类患者类似。 由于 Noggin 是单外显子基因，我们的结果强烈表明，与人类 NOGGIN 突变相关的常染色体显性遗传疾病是由于 NOGGIN 单倍体不足所致。 我们发现患有听力损失的杂合子小鼠有一个异位骨碎片将中耳小骨之一（镫骨）连接到鼓室壁。 这种固定很可能会阻碍声音传导过程中镫骨的活动性，从而导致听力损失。 我们从分析中得出结论，异位骨碎片不是由于从头合成造成的。 最有可能的是，由于 BMP 活动不受阻碍，预期镫骨与其前体在第二鳃弓中不完全分离而形成额外的骨。 由于接受镫骨强直矫正手术的NOGGIN突变患者的长期预后并不理想，因此这些Noggin突变小鼠将继续作为探索更好的缓解听力损失策略的良好模型。 3) Sonic Hedgehog (Shh) 的 Roe 和 Gli 蛋白在内耳发育中的作用（手稿已发表） 通过对具有 Shh、Gli2、Gli3 和 Gli3 delta699 等突变等位基因的各种遗传组合的小鼠突变体的分析，我们发现内耳中的 Shh 信号转导涉及多种机制。 远端耳蜗管位于最靠近脊索和底板中的 Shh 源，其形成需要强大的 Gli2 和 Gli3 激活剂功能。 耳蜗管的近端区域需要较少的 Shh 信号来拮抗 Gli3 阻遏物。 内耳背侧前庭几乎不需要 Shh 信号传导，但 Gli3 阻遏物对于该区域的形成很重要。 因此，来自腹侧中线的分级Shh信号指定了沿背腹轴的各种内耳结构。