Wnt Signaling in Hair Cell Generation from Supporting Cells

支持细胞生成毛细胞中的 Wnt 信号转导

• 批准号: 9056678
• 负责人: Albert Edge
• 金额: $ 35.49万
• 依托单位: MASSACHUSETTS EYE AND EAR INFIRMARY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-12-01 至 2020-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9056678
• 关键词: Accounting; Adult; Aging; Breeding; Bromodeoxyuridine; Cell Count; Cell Death; Cell Differentiation process; Cell Proliferation; Cell division; Cells; Cessation of life; Characteristics; Cochlea; Detection; Ear; Embryo; Exposure to; Gene Expression; Generations; Genes; Hair Cells; Health; Hearing problem; Immunohistochemistry; In Vitro; Knowledge; Laboratories; Labyrinth; Lead; Measures; Motion; Mus; Natural regeneration; Nature; Necrosis; Neomycin; Neonatal; Newborn Infant; Noise; Pathway interactions; Proteinase 3; Recovery of Function; Regenerative response; Reverse Transcriptase Polymerase Chain Reaction; Role; Series; Signal Pathway; Signal Transduction; Stem cells; Stimulus; Supporting Cell; Tamoxifen; Testing; Toxin; Up-Regulation; Work; adult stem cell; beta catenin; cell injury; equilibration disorder; gamma secretase; hair cell regeneration; immunohistochemical markers; in vivo; inhibitor/antagonist; loss of function; model development; notch protein; overexpression; postnatal; progenitor; ranpirnase; receptor; research study; response; sound; transdifferentiation

 DESCRIPTION (provided by applicant): We have shown that inhibition of Notch signaling leads to regeneration of hair cells and provide partial functional recovery in the adult ear with hair cells damaged by noise-exposure. We have recently discovered that Wnt signaling provides a required stimulus for pushing both embryonic progenitor cells and adult stem cells to a hair cell fate. Wnt signaling is necessary for induction of Atoh1 in cells exposed to Notch inhibition, as demonstrated by an absence of Atoh1 upregulation when Wnt signaling is blocked. This suggests that Wnt signaling is required for hair cell differentiation in the cochlea in response to Notch inhibition. Similar to Notch inhibition, gene expression controlled by Wnt stimulates hair cell dfiferentiation, but it also appears to stimulate genes that lead to the replacement of supporting cells. One of these downstream targets, Lgr5, appears to mark cochlear progenitors capable of postnatal transdifferentiation into new hair cells. Our current knowledge of signaling pathways, and our development of the models needed for their manipulation allow a crucial series of experiments to test hypotheses on the nature of the signaling pathways required to elicit hair cell regeneration and recovery of function. Although our recent work has provided an important proof-of-principle for hair cell replacement in the adult, regeneration was limited and the transdifferentiated supporting cells were not replaced. Here, we assess the response of both the newborn and adult cochlea to inhibition of Notch after hair cell damage. We hypothesize that Wnt signaling after hair cell death leads to a partial regenerative response in the newborn cochlea and that signaling through these pathways does not reach levels necessary to initiate regeneration in the adult cochlea. In Aim 1, we test our hypothesis about Wnt signaling in the newborn cochlea by both gain and loss of function studies for a role of Wnt in regeneration in response to Notch inhibition. In Aim 2 we test whether Notch inhibition or β-catenin upregulation in the damaged cochlea in adults, where we can measure hair cell regeneration, is accompanied by gene expression changes characteristic of regeneration. In Aim 3, we test our hypothesis that forced activation of Wnt in combination with Notch inhibition will drive both supporting cell proliferation and transdifferentiation of new hair cells. We further test the idea that the regeneration we see after damage and inhibition of γ-secretase can only occur if Wnt signaling is active.

 描述(申请人提供)：我们已经证明，抑制Notch信号导致毛细胞再生，并在成年耳朵中提供部分功能恢复，毛细胞因噪音暴露而受损。我们最近发现，Wnt信号提供了将胚胎祖细胞和成年干细胞推向毛细胞命运所需的刺激。在暴露于Notch抑制的细胞中，WNT信号是诱导Atoh1所必需的，当Wnt信号被阻断时，Atoh1没有上调就证明了这一点。这表明Wnt信号是耳蜗毛细胞分化所必需的，以响应 缺口抑制。与Notch抑制类似，由Wnt控制的基因表达可以刺激毛细胞分化，但它似乎也刺激了导致支持细胞替换的基因。这些下游靶点之一，Lgr5，似乎标记了能够在出生后转分化为新毛细胞的耳蜗祖细胞。我们目前对信号通路的了解，以及我们对操纵信号通路所需模型的开发，允许我们进行一系列关键的实验，以检验关于诱导毛细胞再生和功能恢复所需的信号通路的性质的假设。虽然我们最近的工作为成人的毛细胞替代提供了重要的原则证明，但再生有限，转分化的支持细胞没有被替代。在这里，我们评估了毛细胞损伤后抑制Notch的反应，包括新生儿和成人耳蜗组织。我们假设，毛细胞死亡后的Wnt信号会导致新生耳蜗的部分再生反应，而通过这些途径的信号并没有达到启动成年耳蜗组织再生所需的水平。在目标1中，我们通过对Wnt在Notch抑制反应中再生的作用的获得和功能丧失的研究来检验我们关于Wnt信号在新生耳蜗中的假说。在目标2中，我们测试了Notch抑制或β-catenin上调是否伴随着再生特征的基因表达变化。我们可以在成年耳蜗处测量毛细胞再生。在目标3中，我们验证了我们的假设，即强制激活Wnt结合Notch抑制将推动支持细胞增殖和新毛细胞的转分化。我们进一步测试了这样的想法，即我们看到的γ分泌酶损伤和抑制后的再生只有在Wnt信号活跃的情况下才能发生。