Epigenetic regulation in the postnatal mammalian inner ear

描述(由申请人提供)：细胞重新编程为治疗学、疾病研究和发育过程提供了巨大的潜力。然而，通过异位表达已定义的转录因子直接重新编程是一个缓慢而低效的过程，大多数细胞无法重新编程。在听觉领域，异位表达 Atoh1等转录因子已被用于将哺乳动物的支持细胞转化为表达许多内源性毛细胞标记的细胞。然而，将支持细胞转化为毛细胞的重新编程过程可能不仅仅是遗传转化，还包括表观遗传转化。使用诱导多能干细胞(IPSCs)的研究表明，它们保留了其起源的体细胞的表观遗传记忆。IPSCs保留的表观遗传记忆可能会干扰其分化为其他类型细胞的潜力。此外，与来自幼年鼠的IPSCs相比，来自老龄鼠的IPSCs具有更低的再编程潜力。尽管异位表达转录因子(Atoh1)可以将新生儿支持细胞转化为毛细胞样细胞，但出生后后期细胞可塑性的丧失可能会极大地影响这一方法的临床应用。据推测，支持细胞到毛细胞的最佳重新编程需要完全表观遗传重新编程到内源性毛细胞，但目前还没有开发出完全表观遗传重新编程发生的技术。然而，组蛋白脱乙酰酶抑制剂、组蛋白甲基转移酶抑制剂和DNA甲基化抑制剂等表观遗传疗法已经用于IPSCs以提高重编程效率。为了更好地理解哺乳动物毛细胞再生是否需要表观遗传重编程，以及表观遗传疗法是否可能在毛细胞再生中发挥作用，还需要进一步的分析。这项研究的长期目标是更好地了解出生后内耳发育过程中发生的表观遗传修饰，并最终帮助增加成熟哺乳动物内耳的细胞可塑性。因此，我们提出以下中心假设：哺乳动物内耳在新生儿发育过程中的表观遗传修饰会导致细胞可塑性的丧失，表观遗传疗法的使用将增加支持细胞的转分化潜力，使其成为毛细胞。为了验证这一假说，我们提出了以下具体目标：目的1.测试表观遗传因子在新生小鼠内耳发育过程中的差异表达；目的2.测试DNA甲基化在小鼠内耳出生后发育过程中是否发生改变或对异位Atoh1表达的反应；以及目的3.测试表观遗传疗法是否影响小鼠内耳毛细胞的再生

DESCRIPTION (provided by applicant): Cellular reprogramming offers tremendous potential for therapeutics, disease studies, and developmental processes. However, direct reprogramming through ectopic expression of defined transcription factors is a slow and inefficient process with most cells failing to reprogram. In the auditory field, ectopic expression of transcription factors such as Atoh1 has been used to convert mammalian supporting cells into cells that express many endogenous hair cell markers. However, the reprogramming process of transforming supporting cells into hair cells may not be solely about genetic transformation, but also epigenetic transformation. Studies using induced pluripotent stem cells (iPSCs) have shown that they retain the epigenetic memory of their somatic cell of origin. The epigenetic memory retained by iPSCs can interfere with their potential for differentiation into other cell types. Additionally, iPSCs derived from aged mice have a decreased potential for reprograming compared to iPSCs derived from juvenile mice. Although, ectopic expression of transcription factors (Atoh1) can convert neonatal supporting cells into hair cell-like cells, loss of cellular plasticity at later postnatal ages could largely impact clinical application of this method. Presumably, the optimal reprogramming of supporting cells to hair cells would require complete epigenetic reprogramming to that of an endogenous hair cell, but current techniques have not yet been developed for complete epigenetic reprogramming to occur. However, epigenetic therapeutics such as histone deacetylase inhibitors, histone methyltransferase inhibitors, and DNA methylation inhibitors have been used on iPSCs to increase reprogramming efficiency. Further analysis is needed to better understand whether epigenetic reprogramming is required for mammalian hair cell regeneration and whether epigenetic therapeutics may have a role in hair cell regeneration. The long term goal of this research is to better understand the epigenetic modifications that occur during postnatal inner ear development, and ultimately to help increase cellular plasticity in the mature mammalian inner ear. Thus, we propose the following central hypothesis: Epigenetic modifications during neonatal development in the mammalian inner ear cause a loss in cellular plasticity and use of epigenetic therapeutics will increase the trans-differentiation potential of supporting cells to become hair cells. To test this hypothesis, we propose the following specific aims: Aim 1. Test whether epigenetic factors are differentially expressed during neonatal development in the mouse inner ear; Aim 2. Test whether DNA methylation is modified in the mouse inner ear during postnatal development or in response to ectopic Atoh1 expression; and Aim 3. Test whether epigenetic therapeutics affect hair cell regeneration in the mouse inner ear