Mechanisms that regulate hair cell survival

 描述(申请人提供)：当听觉毛细胞(HC)丢失时，它们在人类或其他成熟哺乳动物中不会被取代，导致永久性听力损失。相比之下，听觉HC的再生自然发生在非哺乳动物身上，从而允许听力功能的恢复。我们最近观察到新生小鼠耳蜗会在损伤后自发再生，然而，这些再生细胞中的大多数都死亡了。此外，许多研究表明，外耳HCs比内耳HCs更容易受到噪声或耳毒性药物的损害。然而，使HC变得更脆弱的机制尚不清楚。在正常条件下，分化完成后调控HC存活的途径也缺乏了解。拟议的研究将调查HC在出生后成熟和成年期、HC再生期间以及噪声暴露后应激HC中的存活情况。我们将关注一个基因，Pou4f3，一种转录因子，从分化开始在HCS中表达。虽然Pou4f3在维持发育期间的HC存活方面的作用已经被发现，但它在出生后成熟、衰老、再生和应激性HC中的作用还没有被探索过。初步数据显示，Pou4f3在成体毛囊细胞中的缺失会导致细胞死亡，这表明成熟的毛囊细胞仍然需要Pou4f3的表达才能存活。此外，POU4F3免疫组织化学染色结果显示，许多成体OHC的POU4F3表达水平降低，而IHC保持强POU4F3表达。这些数据表明，POU4F3的完全缺失会导致HC死亡，但POU4F3水平的降低足以维持HC的存活。目的1将通过在新生儿、青少年和成人HCS中删除Pou4f3来进一步研究这一点。此外，大多数在新生小鼠耳蜗处自发形成的再生HCs不表达POU4F3，我们假设这会导致细胞死亡。为了支持这一假设，另一项研究在支持细胞中异位表达Atoh1，将它们转化为HCs。这些新形成的HCs存活至少3个月，并且确实表达POU4F3。这些数据暗示Pou4f3在分化过程中对HC存活的调节，Aim 2将通过异位表达Pou4f3来挽救新生小鼠耳蜗中再生的HC。由于已知Pou4f3可调节几个与细胞凋亡有关的基因，成年小鼠毛囊细胞中POU4F3表达水平的降低可能会使这些细胞在应激条件下更加脆弱，并解释了毛细胞损伤易感性增加的原因。我们将在目标3中通过过度表达Pou4f3来测试这一假设，以保护毛细胞免受噪音诱导的损伤。共同提出的研究将探讨Pou4f3‘S在正常、未受损伤的正常耳蜗组、新生小鼠耳蜗组的自发HC再生期间以及噪声暴露后应激组的HC存活调节中所起的作用。这些目标的完成将促进我们对多种条件下调节HC存活的机制的理解，这些机制可以用于开发保护HC的药物。

 DESCRIPTION (provided by applicant): When auditory hair cells (HCs) are lost, they are not replaced in humans or other mature mammals, resulting in permanent hearing loss. In contrast, regeneration of auditory HCs naturally occurs in non-mammals, allowing for recovery of hearing function. We have recently observed that the neonatal mouse cochlea can spontaneously regenerate its HCs after damage; however, the majority of these regenerated cells died. In addition, many studies have shown that outer HCs (OHCs) are more susceptible than inner HCs (IHCs) to damage caused by noise or ototoxic drugs. Yet the mechanism that makes OHCs more vulnerable is not understood. There is also a lack of understanding of the pathways that regulate HC survival under normal conditions after differentiation is complete. Proposed studies will investigate HC survival during postnatal maturation and adulthood, during HC regeneration, and in stressed HCs following noise exposure. We will focus on one gene, Pou4f3, a transcription factor that is expressed in HCs beginning with differentiation. While the role of Pou4f3 in maintaining HC survival during development was discovered previously, its role in postnatal maturation, aging, regeneration, and in stressed HCs has not been explored. Preliminary data show that deletion of Pou4f3 in adult OHCs causes cell death which suggests that mature OHCs still require Pou4f3 expression to survive. In addition, POU4F3 immunostaining results show that many adult OHCs have decreased levels of POU4F3 expression, while IHCs retained strong POU4F3 expression. These data suggest that complete deletion of POU4F3 causes HCs to die, but reduced levels of POU4F3 are enough to maintain HC survival. Aim 1 will investigate this further by deleting Pou4f3 in neonatal, juvenile, and adul HCs. In addition, the majority of regenerated HCs that spontaneously form in the neonatal mouse cochlea do not express POU4F3 and we hypothesize that this causes cell death. In support of this hypothesis, another study ectopically expressed Atoh1 in supporting cells to convert them into HCs. These newly formed HCs survived at least 3 months and did express POU4F3. These data implicate Pou4f3 in the regulation of HC survival during the differentiation process and Aim 2 will rescue regenerated HCs in the neonatal mouse cochlea by ectopic expression of Pou4f3. Since Pou4f3 is known to regulate several genes involved in apoptosis, decreased levels of POU4F3 expression in OHCs of adult mice may make these cells more vulnerable under stressful conditions and account for the increased damage susceptibility of OHCs. We will test this hypothesis in Aim 3 by over-expressing Pou4f3 to protect OHCs from noise-induced damage. Collectively proposed studies will investigate Pou4f3's role in the regulation of HC survival during maturation and adulthood in the normal, undamaged cochlea, during spontaneous HC regeneration in the neonatal mouse cochlea, and in stressed HCs following noise exposure. Completion of these aims will advance our understanding of the mechanisms that regulate HC survival under multiple conditions, which could be used to develop drugs to protect HCs.