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（由申请人提供）：我们已经证明，Notch 信号传导的抑制会导致毛细胞再生，并为因噪声暴露而受损的毛细胞提供部分功能恢复。我们最近发现 Wnt 信号传导提供了推动胚胎祖细胞和成体干细胞走向毛细胞命运所需的刺激。在暴露于 Notch 抑制的细胞中，Wnt 信号传导对于诱导 Atoh1 是必需的，当 Wnt 信号传导被阻断时 Atoh1 上调的缺失就证明了这一点。这表明 Wnt 信号传导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，Wnt控制的基因表达会刺激毛细胞分化，但它似乎也会刺激导致支持细胞更换的基因。这些下游靶标之一 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，以及我们对其操作所需的模型的开发，允许进行一系列关键的实验来测试有关引发毛细胞再生和功能恢复所需的信号通路性质的假设。 尽管我们最近的工作为成人毛细胞替代提供了重要的原理证明，但再生是有限的，并且转分化的支持细胞没有被替代。在这里，我们评估了新生儿和成人耳蜗在毛细胞损伤后对 Notch 抑制的反应。我们假设毛细胞死亡后的 Wnt 信号传导导致新生儿耳蜗部分再生反应，并且通过这些途径的信号传导未达到成人耳蜗启动再生所需的水平。在目标 1 中，我们通过功能获得和丧失研究 Wnt 在响应 Notch 抑制的再生中的作用，检验了关于新生儿耳蜗中 Wnt 信号传导的假设。在目标 2 中，我们测试了成人受损耳蜗中 Notch 抑制或 β-连环蛋白上调（我们可以测量毛细胞再生）是否伴随着再生特征的基因表达变化。在目标 3 中，我们测试了我们的假设，即强制激活 Wnt 与 Notch 抑制相结合将驱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：只有当 Wnt 信号传导活跃时，γ-分泌酶受损和抑制后才会发生再生。