The role of intracellular calcium in ototoxin-mediated hair cell death

细胞内钙在耳毒素介导的毛细胞死亡中的作用

• 批准号: 8366054
• 负责人: Robert Esterberg
• 金额: $ 5.57万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8366054
• 关键词: Adverse drug effect; Adverse effects; Affect; Aminoglycoside Antibiotics; Aminoglycosides; Apoptotic; Buffers; Calcium; Cell Death; Cell Death Signaling Process; Cells; Cessation of life; Chelating Agents; Cisplatin; Cues; Cytoplasm; DNA; Data; Dose; Dyes; Effectiveness; Equilibrium; Event; Exhibits; Exposure to; Fishes; Future; Genetic; Hair Cells; Health; Homeostasis; Human; Image; Imagery; Ionophores; Kinetics; Laboratories; Labyrinth; Lead; Life; Location; Mammals; Mediating; Membrane; Methods; Monitor; Movement; Neomycin; Nuclear; Organelles; Oxygen; Pathway interactions; Pharmaceutical Preparations; Physical condensation; Platinum; Population; Predisposition; Production; Relative (related person); Research; Research Personnel; Resistance; Role; Sensory Receptors; Signal Transduction; Source; Surface; Survival Analysis; System; Therapeutic Agents; Time; Transgenic Organisms; Work; Zebrafish; anti-cancer therapeutic; base; blocking factor; cell injury; cost; extracellular; hearing impairment; insight; killings; lateral line; neuromast; ototoxicity; ototoxin; photolysis; prevent; programs; research study; response; sensory system; sound; stem; therapy design; time use; tool

DESCRIPTION (provided by applicant): Hair cells are the primary sensory receptors of the inner ear, and are responsible for detecting sound as well as balance. A number of factors are attributable to hearing loss; this includes drugs that elicit ototoxicity through mechanisms that are largely unclear. Our lab is investigating the causative role of these drugs in hair cell death in hopes of identifying protective strategies. Because it is challenging to study ototoxin-induced hair cell death in mammals, we employ the zebrafish system as a tool to study such events. In addition to hair cells of the inner ear, zebrafish also possess hair cells on their surface in a sensory system termed the lateral line. While their external location enable zebrafish to sense environmental cues, we as researchers can easily observe and image intracellular events with little perturbation. Because lateral line hair cells resemble hair cells of the mammalian inner ear both structurally and in their response to ototoxins, zebrafish are ideal for the study of signaling and cell death pathways that affect hair cells. We have begun to examine such pathways in earnest, and have started to focus on the role of Ca2+ signaling in hair cell death. Despite its well-known role in many intracellular pathways, including pro-apoptotic ones, Ca2+ has not been extensively examined in hair cells following exposure to ototoxic agents. I propose to examine the role of intracellular Ca2+ in hair cell death utilizing the many benefits of the zebrafish lateral line system. I have taken advantage of previous dose-response characterizations performed in our lab to treat zebrafish with doses of ototoxin that kill approximately 50% of lateral line hair cells, and quantify differences in Ca2+ levels between hair cells that die and those that survive. My initial analysis has identified that intracellular Ca2+ levels are significantly higher in dying hair cells exposed to either the aminoglycoside neomycin or the anticancer therapeutic cisplatin. The work proposed here aims to reveal subcellular events that lead to this increase, as well as to better understand why a seemingly uniform population of cells exhibits a differential cell death response. I will utilize genetically encoded Ca2+ indicators targeted to various subcellular compartments within the hair cell to visualize these events in real-time. Much of this work is expected to define additional steps within the pathway of hair cell death. These studies should provide new insight into methods of ototoxin action as well as the potential to block hair cell death during ototoxin exposure. Pharmacological agents exist that are capable of regulating the subcellular flow of Ca2+ within the cell, and understanding how these ototoxins affect intracellular Ca2+ holds promise for future therapies designed to protect human hair cells from death.

描述（由申请人提供）：毛细胞是内耳的主要感觉受体，负责检测声音以及平衡。许多因素可归因于听力损失;这包括通过基本上不清楚的机制引起耳毒性的药物。我们的实验室正在研究这些药物在毛细胞死亡中的致病作用，希望找到保护性策略。由于研究哺乳动物耳毒素诱导的毛细胞死亡具有挑战性，我们采用斑马鱼系统作为研究此类事件的工具。除了内耳的毛细胞外，斑马鱼表面的感觉系统中也有毛细胞，称为侧线。虽然它们的外部位置使斑马鱼能够感知环境线索，但我们作为研究人员可以很容易地观察和成像细胞内事件，而几乎没有干扰。由于侧线毛细胞在结构上和对耳毒素的反应上都类似于哺乳动物内耳的毛细胞，因此斑马鱼是研究影响毛细胞的信号传导和细胞死亡途径的理想动物。我们已经开始认真研究这些途径，并开始关注Ca 2+信号在毛细胞死亡中的作用。尽管其众所周知的作用，在许多细胞内的途径，包括促凋亡的，Ca 2+还没有被广泛研究暴露于耳毒性剂后的毛细胞。我建议研究的作用，利用斑马鱼侧线系统的许多好处，在毛细胞死亡的细胞内Ca 2+。我利用我们实验室以前进行的剂量反应表征，用杀死大约50%侧线毛细胞的耳毒素剂量治疗斑马鱼，并量化死亡和存活的毛细胞之间Ca 2+水平的差异。我的初步分析已经确定，细胞内Ca 2+水平显着较高，在垂死的毛细胞暴露于氨基糖苷类新霉素或抗癌治疗顺铂。本文提出的工作旨在揭示导致这种增加的亚细胞事件，以及更好地理解为什么看似均匀的细胞群体表现出差异性细胞死亡反应。我将利用遗传编码的Ca 2+指标，针对毛细胞内的各种亚细胞区室，实时可视化这些事件。预计这项工作的大部分将确定毛细胞死亡途径中的其他步骤。这些研究将为耳毒素作用的方法以及在耳毒素暴露期间阻断毛细胞死亡的潜力提供新的见解。存在能够调节细胞内Ca 2+的亚细胞流动的药理学试剂，并且了解这些耳毒素如何影响细胞内Ca 2+为未来设计用于保护人类毛细胞免于死亡的疗法带来希望。