Lineage reprogramming for hearing loss: development of drug screening and gene therapy approaches

项目摘要/摘要： 由于内侧感觉毛细胞丧失而导致的耳聋和平衡障碍 在美国，耳朵是致残和发病的主要原因。在哺乳动物中，细胞的 内耳的各种感觉上皮细胞以胚胎的形式出现，随后并不是 如果损坏，可再生(Rubel等人，2013年)。毛发死亡引起的听力损失 因此，科尔蒂器官中的细胞是永久性的，旨在逆转听力的治疗 通过刺激毛细胞再生而失去毛细胞是非常必要的。然而， 由于内耳细胞数量较少，且细胞数量较少，因此在内耳细胞上进行实验比较困难。 成年人极难接近。因此，现代细胞和分子技术 分析和药物发现一直很难应用，除了假肢， 感觉神经性耳聋的治疗选择仍然很少。为了克服这些问题， 在寻求治疗毛细胞缺失的新方法时，我们使用了强大的新“直接” “谱系重新编程”技术，最初是为神经元特异性开发的 重新编程(Son等人，2011；Vierbuhen等人，2010)，以产生毛细胞样细胞 在体外，直接来自小鼠和人类的体细胞(成纤维细胞和内耳 支持细胞)。这一进步允许一系列新的实验进入 毛细胞分化的机制、临床前遗传学模型的发展 听力损失(疾病建模)、与以下相关的药物发现高通量筛查 再生和耳毒性，以及基因治疗方法的应用 毛细胞再生的问题。该提案的目的包括：1)发展 改进了重新编程策略，以诱导小鼠和人类毛细胞样细胞。2) 耳毒性药物筛选和遗传性疾病体外模型的建立 听力损失。3)毛细胞临床前模型中的重编程测试 长期耳聋小鼠的再生/替代。

Project Summary/Abstract: Deafness and balance disorders resulting from the loss of sensory hair cells of the inner ear are a major cause of disability and morbidity in the US. In mammals, the cells of the various sensory epithelia of the inner ear arise embryonically and subsequently do not regenerate if damaged (Rubel et al., 2013). Hearing loss resulting from the death of hair cells in the organ of Corti is thus permanent, and treatments aimed at reversing hearing loss through stimulated regeneration of hair cells are badly needed. However, experimentation on the cells of the inner ear is difficult due to their small number and extreme inaccessibility in the adult. As a result, modern techniques of cell and molecular analysis and drug discovery have been difficult to apply, and aside from prosthetics, treatment options for sensorineural deafness remain few. To overcome these problems, and in pursuit of new treatments for hair cell loss, we have used powerful new “direct lineage reprogramming” technologies, originally developed for neuron-specific reprogramming (Son et al., 2011; Vierbuchen et al., 2010), to generate hair cell-like cells in vitro, directly from mouse and human somatic cells (fibroblasts and inner ear supporting cells). This advance allows a new range of experimentation into the mechanisms of hair cell differentiation, the development of preclinical models of genetic hearing loss (disease modeling), high-throughput screening for drug discovery related to regeneration and ototoxicity, and the application of gene therapy approaches to the problem of hair cell regeneration. The Aims of the proposal include: 1) Development of improved reprogramming strategies to induce mouse and human hair cell-like cells. 2) Development of a drug screen for ototoxicity, and an in vitro disease model of genetic hearing loss. 3) A test of reprogramming in a preclinical model of hair cell regeneration/replacement in long-deafened mice.