Hair Cell Development in the Mammalian Cochlea

Corti器官起源于位于内耳内的一群细胞，称为前感觉细胞。 基于现有的数据，前感觉细胞被认为是唯一有能力发展为毛细胞和相关的支持细胞。 然而，指定前感觉域的因素仍然未知。 最近的数据表明，notch信号通路可能在前感觉特化中发挥作用。 notch通路是一种高度保守的信号级联，在多种组织和多种物种中使用，包括苍蝇、蠕虫、小鼠和人类。 哺乳动物的基因组包含四个notch基因，每个都可以部分补偿任何其他notch基因的损失。 因此，为了确定notch信号在前感觉形成中的完整作用，我们使用cre-lox方法来特异性删除内耳中的Rbp-j（所有notch信号的关键组分）。 结果表明，notch信号的缺失对前感觉形成有深远的影响，包括几乎所有内耳感觉细胞的丧失。 然而，对这些小鼠内耳的彻底检查表明，在没有notch的情况下，一些前感觉细胞仍然形成，但这些细胞无法保持前感觉身份，随后失去了作为毛细胞和支持细胞发育的能力。 作为上述研究的一部分，我们检查了正常和Rbp-j缺失内耳中基因表达的总体变化。 在这些情况下下调的基因中有胰岛素样生长因子信号通路的成员。 为了确定这些基因是否发挥作用，在调节的prosensory身份的缺口信号，我们的特点是表达模式的胰岛素样生长因子（Igf 1和Igf 2），胰岛素样生长因子受体（Igfr 1和igfr 2）和胰岛素样生长因子结合蛋白（Igfbp 1 -5）。 这些基因中的每一个都被发现在发育中的内耳的特定区域中表达。 基于表达模式，我们使用RNAi下调内耳外植体培养物中Igfbp 5的表达。 结果表明，特异性激活的notch途径，这表明一个相互之间存在的notch和Ifgbp 5的信号循环。Igfb 5的具体作用正在确定中。 最后，我们实验室以前的工作已经证明，Sox 2，一个参与前感觉特化的转录因子，也有能力诱导耳蜗内的一些非感觉细胞发育为神经元。 这一发现使我们研究是否其他已知在发育中的内耳神经元中表达的转录因子可能具有类似的能力。 特别是，两个转录因子，Neurogenin 1和NeuroD 1，表达在非神经元细胞内的内耳使用基因转移。 这两种因素都能够诱导神经元标记物的表达和神经元表型。 此外，这些细胞的电生理学评估表明，它们表达的离子通道和电特性与神经元表型一致。 这些结果表明，它可能是可能的诱导内耳神经元的形成，利用基因转移。 这种方法可用于开发用于诱导新的内耳神经元的疗法，其中这些细胞由于基因突变或创伤而丢失。

The organ of Corti arises from a population of cells, referred to as prosensory cells, that are located within the inner ear. Based on existing data, prosensory cells are believed to be uniquely competent to develop as both hair cells and associated supporting cells. However, the factors that specify the prosensory domain remain unknown. Recent data has suggested that the notch signaling pathway could play a role in prosensory specification. The notch pathway is a highly conserved signaling cascade that is utilized in multiple tissues and in multiple species including flies, worms, mice and humans. The mammalian genome contains four notch genes, each of which can partially compensate for the loss of any other notch gene. Therefore, to determine the complete role of notch signaling in prosensory formation, we used a cre-lox approach to specifically delete Rbp-j (a key component of all notch signaling) in the inner ear. Results indicated that deletion of notch signaling has a profound effect on prosensory formation that includes the loss of nearly all inner ear sensory cells. However, a thorough examination of the inner ears from these mice indicated that some prosensory cells still form in the absence of notch, but that these cells fail to maintain a prosensory identity and subsequently lose the ability to develop as hair cells and supporting cells. As part of the study described above, we examined overall changes in gene expression in normal and Rbp-j deleted inner ears. Among the genes that were down-regulated under these circumstances were members of the insulin-like growth factor signaling pathway. To determine whether these genes play a role in the regulation of prosensory identity by notch signaling, we characterized the expression patterns for both the insulin-like growth factors (Igf1 and Igf2), the insulin-like growth factor receptors (Igfr1 and igfr2) and the insulin-like growth factor binding proteins (Igfbp1-5). Each of these genes was found to be expressed in a specific region of the developing inner ear. Based on the patterns of expression, we used Rnai to down-regulate Igfbp5 expression in explant cultures of the inner ear. Results indicated specific activation of the notch pathway, suggesting that a reciprocal signaling loop exists between notch and Ifgbp5. The specific effects of Igfb5 are in the process of being determined. Finally, previous work from our laboratory had demonstrated that Sox2, a transcription factor involved in prosensory specification also had the ability to induce some non-sensory cells within the cochlea to develop as neurons. This finding lead us to examine whether other transcription factors that are known to be expressed in developing inner ear neurons might have a similar ability. In particular, two transcription factors, Neurogenin1 and NeuroD1, were expressed in non-neuronal cells within the inner ear using gene transfer. Both of these factors were able to induce the expression of neuronal markers and a neuronal phenotype. Moreover, electrophysiological assessments of these cells indicated that they expressed ion channels and electrical characteristics consistent with neuronal phenotypes. These results suggest that it may be possible to induce the formation of inner ear neurons using gene transfer. Such an approach could be used to develop therapies for the induction of new inner ear neurons in individuals in which these cells have been lost as a result of genetic mutation or trauma.