Developing novel stem cell-based epigenetic approaches to treat hearing loss

项目摘要 听觉毛细胞是一种机械传导受体，通过螺旋状结构将声音信号传递到脑干 神经节神经元（SGN）。然而，毛细胞容易受到一些伤害，包括噪音，爆炸， 衰老和脑外伤成年哺乳动物的毛细胞损失通常是不可逆的， 听力损失和其他内耳疾病。目前，还没有生物学方法来补充人类 毛细胞该项目的长期目标是使用基于干细胞的方法来再生受损的 听觉系统治疗听力损失和其他内耳疾病。在这个方案中，我们将产生人类 感觉毛细胞，并使用基于干细胞的表观遗传方法重建毛细胞-SGN连接。 我们已经鉴定了人椭圆囊上皮来源的前列腺样细胞（HUCs），其表现出以下特征： 前感觉细胞我们在一项初步研究中发现HUCs表现出基因组DNA甲基化。我们应用了 DNA甲基转移酶（DNMT）抑制剂5-氮杂胞苷（5-aza）对HUCs的作用，发现经处理的HUCs 表达毛细胞基因和蛋白质。因此，在本提案中，我们假设DNMT抑制剂5- 氮诱导HUCs分化成新的功能性毛细胞，可以取代受损的毛细胞， 与听觉神经元的突触。为了检验这一假设，提出了三个具体目标： 具体目标1：确定5-aza诱导HUCs进入感觉毛细胞。 具体目标2：研究5-氮杂处理的HUCs在体外的整合和突触形成。 具体目标3：确定5-氮杂处理的HUCs的存活、分化、整合和功能。 在这个提议中，我们将产生人类感觉毛细胞，并使用 基于干细胞表观遗传学方法。表观遗传方法的优点是表达和 基因的表型是遗传改变的，而它们的DNA序列保持不变。长期目标 这项工作的目的是（a）发现和探索创新的再生方法，以改善健康和 福利的军事人员和退伍军人与听力障碍，和（B）加快过渡， 干细胞技术成为治疗听力损失和其他内耳疾病的新标准。成功 这项工作的开展将极大地推动基于干细胞的听力再生研究， 潜在地影响医学科学在今天的战场上的功能，健康和整体经验的状态， 退伍军人及其照顾者和家庭的生活质量。

Project Summary Auditory hair cells are mechanotransduction receptors that convey sound signals to the brainstem via spiral ganglion neurons (SGNs). However, hair cells are vulnerable to a number of insults, including noise, blast, aging, and traumatic brain injury. Adult mammalian hair cell loss is usually irreversible, causing permanent hearing loss and other inner ear disorders. Currently, there are no biological approaches to replenish human hair cells. The long-term goal of this project is to use stem cell-based approaches to regenerate the damaged auditory system to treat hearing loss and other inner ear disorders. In this proposal, we will generate human sensory hair cells and rebuild hair cell-SGN connections using a stem cell-based epigenetic approach. We have identified human utricle epithelia-derived prosensory-like cells (HUCs) that exhibit features of prosensory cells. We found that HUCs demonstrated genomic DNA methylation in a pilot study. We applied the DNA methyltransferase (DNMT) inhibitor 5-azacytidine (5-aza) to HUCs and found that treated HUCs expressed hair cell genes and proteins. Therefore, in this proposal we hypothesize that the DNMT inhibitor 5- aza induces HUCs to differentiate into new functional hair cells that can replace damaged hair cells and form synapses with auditory neurons. To test this hypothesis, three specific aims are proposed: Specific Aim 1: Define the induction of HUCs into sensory hair cells by 5-aza. Specific Aim 2: Investigate the integration and synapse formation of 5-aza-treated HUCs in vitro. Specific Aim 3: Determine the survival, differentiation, integration, and function of 5-aza-treated HUCs in vivo. In this proposal, we will generate human sensory hair cells and rebuild hair cell-neuron connections using a stem cell-based epigenetic approach. The advantage of an epigenetic approach is that expression and phenotype of gene are heritably changed, whereas their DNA sequences remain constant. The long-term aims of this work are (a) to discover and explore innovative regeneration approaches to improve the health and welfare of military personnel and veterans with hearing disturbances, and (b) to accelerate the transition of stem cell technologies into new standards of care to treat hearing loss and other inner ear disorders. Success of this work will significantly promote the research of stem cell-based hearing regeneration, which will potentially affect the state of medical science in today’s battlefield experience on function, wellness, and overall quality of life for veterans as well as their caregivers and families.