p16INK4a in Cochlear Hair Cell Regeneration.

描述（由申请人提供）：听力损失主要由内耳耳蜗感觉毛细胞（HC）受损引起。人类和其他哺乳动物不能替代受损的hc；然而，鸡、鱼和两栖动物可以通过增殖和转分化邻近的支持细胞（SC） （Conwin和Oberholtzer， 1997）。有趣的是，肿瘤抑制基因p16INK4a在具有再生hc能力的非哺乳动物脊椎动物中不存在（Kim et al., 2003; Gil and Peters, 2006）。在哺乳动物中，p16INK4a作为周期蛋白依赖性激酶抑制剂，使细胞处于静止状态。它的表达受年龄和有丝分裂信号的诱导（Zindy et al., 1997）。p16INK4a也被认为在包括神经元在内的成年细胞的体内再生能力中发挥关键作用（Janzen et al., 2006; Molofsky et al., 2006）。通常认为受损的hcc释放信号导致SCs重新进入细胞周期。在非哺乳脊椎动物中，这导致HC再生；然而，在哺乳动物中，我们预测这种有丝分裂活性诱导p16INK4a的表达，p16INK4a使SCs处于静止状态，从而阻止HC再生。是否有可能通过使p16INK4a失活，使老鼠和鸡具有再生hc的能力？为了实现这一目标，我们提出以下中心假设：哺乳动物中p16INK4a的失活将使SCs:(1)对抗生素或基因消融损伤的hcc释放的信号作出反应，(2)重新进入细胞周期，(3)再生hcc。为了验证这一假设，我们提出以下具体目的：具体目的1：测定氨基糖苷类抗生素庆大霉素引起HC损伤后p161nk4a缺失小鼠中SCs的再生能力。特异性目的2：测定p16ink4a缺失小鼠因视网膜母细胞瘤蛋白急性失活导致HC损伤后sc的再生能力。尽管哺乳动物不能自发再生HC，但耳蜗sc的遗传和/或治疗操作可以诱导类似于非哺乳动物脊椎动物的HC再生过程。这项提议是实现最终目标的第一步，即恢复那些暴露于耳毒性药物、大声噪音或其他环境因素的人的听力。此外，出生后HC中视网膜母细胞瘤蛋白急性失活引起的HC损伤是该领域的技术突破。

DESCRIPTION (provided by applicant): Hearing loss is primarily caused by damage to sensory hair cells (HC) in the cochlea of the inner ear. Humans and other mammals cannot replace damaged HCs; however, chicken, fish and amphibians can, by proliferation and transdifferentiation of neighboring supporting cells (SC) (Conwin and Oberholtzer, 1997). Interestingly, the tumor suppressor gene, p16INK4a, is absent in non-mammalian vertebrates that have the capacity to regenerate HCs (Kim et al., 2003; Gil and Peters, 2006). In mammals, p16INK4a acts as a cyclin-dependent kinase inhibitor that keeps cells in a quiescent state. Its expression is induced by age as well as mitogenic signaling (Zindy et al., 1997). p16INK4a is also known to play a critical role in the in vivo regenerative ability of adult cells, including neurons (Janzen et al., 2006; Molofsky et al., 2006). It is commonly thought that damaged HCs release signals that cause SCs to reenter the cell cycle. In non-mammalian vertebrates this results in HC regeneration; however, in mammals, we predict that this mitotic activity induces the expression of p16INK4a which keeps SCs in a quiescent state and thus prevents HC regeneration. Is it possible to give a mouse, the chicken's ability to regenerate HCs by inactivating p16INK4a? To achieve this goal, we propose the following central hypothesis: Inactivation of p16INK4a in mammals will allow SCs: (1) to respond to signals released from HCs damaged by antibiotics or genetic ablation, (2) to reenter the cell cycle and (3) to regenerate HCs. To test this hypothesis, we propose the following specific aims: Specific Aim 1: To determine the regenerative capacity of SCs in p161NK4a-null mice after HC damage caused by the aminoglycoside antibiotic, gentamicin. Specific Aim 2: To determine the regenerative capacity of SCs in p16INK4a-null mice after HC damage caused by acute inactivation of the retinoblastoma protein in postnatal HCs. Although mammals cannot spontaneously regenerate HCs, a process of HC regeneration similar to what occurs in non-mammalian vertebrates could be induced by genetic and/or therapeutic manipulations of cochlear SCs. This proposal is the first step toward the final goal of restoring hearing in those exposed to ototoxic drugs, loud noise or other environmental agents. In addition, HC damage induced by acute inactivation of the retinoblastoma protein in postnatal HCs is a technological breakthrough for the field.