Mouse vestibular regeneration and function

小鼠前庭再生和功能

基本信息

批准号：
10058261
负责人：
Alan Gi-Lun Cheng
金额：
$ 59.04万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-12-01 至 2023-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10058261
关键词：
Ablation Acceleration Adult Age Algorithmic Analysis Animals Bioinformatics Candidate Disease Gene Cell Survival Cell physiology Cells Chickens Cochlea Data Data Analyses Dimensions Disease Dizziness Electric Capacitance Electrophysiology (science)Epithelial Equilibrium Evoked Potentials Functional disorder Gene Expression Genes Genetic Goals Hair Cells Histologic Histology Human Impairment In Situ Hybridization Label Legal patent Mammals Maps Measurement Measures Mechanoreceptor Cell Methods Morphology Mus Natural regeneration Organ Physiology Property Recovery Recovery of Function Research Role Sensory Sensory Hair Supporting Cell Synapses Techniques Technology Testing Time Transgenic Mice Transgenic Organisms Utricle structure Vertigo Vestibular Hair Cells Work Zebrafish cell regeneration design effective therapy experimental study gain of function gene discovery gene expression database genetic signature hair cell regeneration improved in vivo insight loss of function mechanotransduction mouse model nerve supply novel overexpression partial recovery prevent ranpirnase regeneration function regeneration model repaired response restoration single-cell RNA sequencing spatiotemporal stem cells transcription factor transcriptome vesicular release

项目摘要

Abstract: Sensory hair cells are required for balance function. Vestibular hair cell degeneration causes balance dysfunction/hypofunction manifested as dizziness and vertigo. While the mammalian cochlea lacks the ability to regenerate lost hair cells, a limited degree of spontaneous regeneration occurs in the utricle, a vestibular organ detecting linear acceleration. Recent studies using fate-mapping techniques have pinpointed supporting cells as precursors of regenerated hair cells. However, it is not clear whether regenerated hair cells are fully functional and if organ function recovers. In preliminary experiments we have characterized hair cell degeneration and regeneration in the mature mouse utricle and also a loss followed by recovery of vestibular evoked potentials (VsEP) in vivo. The first aim of this proposal is to determine if increasing hair cell regeneration improves the recovery of vestibular function. Specifically, regenerated hair cells labeled via fate-mapping are probed via histology and electrophysiology to assess bundle morphology, mechanosensitvity, basolateral currents, and synaptic properties including vesicle release. In parallel, VsEP responses are measured and compared to histologic and electrophysiological measures. Next, by overexpressing Atoh1 via a transgenic approach, we will study the histology and electrophysiology of Atoh1-overexpressing hair cells and also the overall VsEP responses. In the second aim, we will determine if Atoh1 deletion prevents hair cell regeneration and the recovery of VsEP responses. In parallel, fate-mapped, surviving hair cells will be examined for possible repair via histology and electrophysiology. To gain an unbiased insight into the genetic signature of hair cell progenitors and surviving hair cells, the third aim is designed to examine the damaged mature mouse utricle using single cell RNA sequencing technologies. Here the first goal is to discover the genetic landscape of hair cell progenitors and surviving hair cells in the damaged utricle. Secondly, we will examine the gene expression of the undamaged and damaged utricle after Atoh1 overexpression. Lastly, we will use bioinformatic approaches to delineate the trajectory of the spontaneous and Atoh1-enhaced supporting cell-hair cell transition and validate this histologically. In summary, we will apply state-of-the art technologies (vestibular physiology, hair cell physiology, single cell RNA-seq, bioinformatic strategies) to study vestibular hair cell regeneration in transgenic mouse models. We have assembled a team of experts who have worked together to collect promising preliminary data. At the end of this 5-year proposal, we will have 1) determined the relationship between hair cell regeneration and functional recovery and 2) revealed and temporally ordered novel genes during mammalian hair cell regeneration.

抽象的：平衡功能需要感觉毛细胞。前庭毛细胞退化导致平衡功能障碍/机能减退表现为头晕和眩晕。虽然哺乳动物的耳蜗缺乏能力再生丢失的毛细胞，前庭器官椭圆囊发生有限程度的自发再生检测线性加速度。最近使用命运图谱技术的研究已将支持细胞确定为再生毛细胞的前体。然而，尚不清楚再生的毛细胞是否功能齐全以及器官功能是否恢复。在初步实验中，我们已经表征了毛细胞变性和成熟小鼠椭圆囊的再生以及前庭诱发电位恢复后的损失（VsEP）体内。该提案的第一个目的是确定增加毛细胞再生是否可以改善前庭功能恢复。具体来说，通过命运图谱标记的再生毛细胞通过组织学和电生理学来评估束形态、机械敏感性、基底外侧电流和突触特性，包括囊泡释放。同时，测量 VsEP 响应并与组织学和电生理学测量。接下来，通过转基因方法过度表达Atoh1，我们将研究 Atoh1 过表达毛细胞的组织学和电生理学以及整体 VsEP 回应。在第二个目标中，我们将确定 Atoh1 缺失是否会阻止毛细胞再生和恢复 VsEP 反应。与此同时，将通过组织学检查命运图谱上的存活毛细胞是否可能进行修复和电生理学。公正地了解毛细胞祖细胞和存活的遗传特征毛细胞，第三个目标是使用单细胞 RNA 检查受损的成熟小鼠椭圆囊测序技术。这里的第一个目标是发现毛细胞祖细胞的遗传景观和受损的椭圆囊中幸存的毛细胞。其次，我们将检查未受损的基因表达 Atoh1 过度表达后，椭圆囊受损。最后，我们将使用生物信息学方法来描述自发和 Atoh1 增强的支持细胞-毛细胞转变的轨迹并验证这一点组织学上。总之，我们将应用最先进的技术（前庭生理学、毛细胞生理学、单细胞RNA-seq，生物信息学策略）研究转基因小鼠的前庭毛细胞再生模型。我们组建了一个专家团队，他们共同努力收集有希望的初步数据。在这个 5 年提案结束时，我们将 1) 确定毛细胞再生之间的关系和功能恢复以及2）在哺乳动物毛细胞中揭示和按时间顺序排列的新基因再生。