Driving the Progeny of Olfactory HBC Stem Cells toward Neuronal Differentiation

项目摘要 成年人嗅上皮更新和再生感觉神经细胞群的能力 神经元依赖于干细胞的持久性和正常功能。老年嗅觉障碍伴随着 由于正常活跃的嗅干的紊乱和最终耗尽而引起的病理变化， 祖细胞，即球状基底细胞（GBC）。在这种情况下，储备干细胞，即 水平基底细胞（HBC）保持休眠，尽管GBC的神经源性衰竭和消失。 相反，如果OE被嗅觉毒素损伤，HBC激活并有助于修复嗅觉神经元。 上皮HBC激活的结果是强烈的上下文依赖性的，并不总是使细胞获得能力。 神经发生的再生。这种失败可能发生在啮齿动物身上， 损伤或老化/神经源性衰竭。大量的间接证据表明，人类也是如此。 OE。一种治疗策略，指导HBCs在神经元再生的背景下沿着神经元再生的途径， 耗尽的OE可能是治疗年龄相关的嗅觉功能障碍的最佳方法。我们有 证明TF p63是调节HBC激活的主开关-p63的急剧下降， 水平是激活的必要条件和充分条件。我们正在测试已知能增强神经元 分化（Aim 1）或抑制非神经元分化（Aim 2）。我们将使用定量蛋白质组学， 定义嗅觉基质细胞的分泌组，这将提名候选机制的复杂 上皮和调节神经发生的固有层细胞之间的相互联系。我们将 使用我们新开发的HBC培养系统，通过定量 在各种因素激活后出现的神经元。然后，我们将通过以下方法验证体外效应： 将活化的/因子处理的HBCs移植到受损的OE中，这提供了一个中性环境， 所有神经元和非神经元的命运都可用于内源性HBCs或已被 有教养我们将在小鼠和人类HBCs上测试疗效，我们将使用一种新的异种移植方法， 测定，以评估人HBCs在体内的后果。完成后，我们将取得很大的成就， 更深入地了解HBCs向神经元分化的过程， 他们可能会恢复神经再生。对小鼠遗传机制的理解， 操纵提供了深刻的分析能力，并在人类将推进我们的努力，旨在确定 用于减轻嗅觉感觉功能障碍，特别是感觉丧失的治疗策略， 伴随着衰老。

PROJECT SUMMARY The capacity of the adult olfactory epithelium (OE) for renewing and regenerating the population of sensory neurons depends on the persistence and proper function of stem cells. Presbyosmia is accompanied by pathological changes due to the disordering and eventual depletion of the normally active olfactory stem and progenitor cells, namely globose basal cells (GBCs). In this setting, the reserve stem cells, namely the horizontal basal cells (HBCs), remain dormant despite the neurogenic exhaustion and disappearance of GBCs. In contrast, if the OE is damaged by an olfactotoxin, the HBCs activate and contribute to the repair of the epithelium. The results of HBC activation are strongly context-dependent and do not always capacitate the rejuvenation of neurogenesis. That failure can occur in rodents as a consequence of either experimental lesion or aging/neurogenic exhaustion. Substantial indirect evidence suggests the same is true of the human OE. A therapeutic strategy that directs HBCs down the pathway toward neuronal regeneration in the setting of an exhausted OE offers possibly the best approach for treating age-related olfactory dysfunction. We have demonstrated that the TF p63 is the master switch that regulates HBC activation – a precipitous decline in p63 levels is necessary and sufficient for activation. We are testing factors that are known to enhance neuronal differentiation (Aim 1) or suppress non-neuronal differentiation (Aim 2). We will use quantitative proteomics to define the secretome of olfactory stromal cells, which will nominate candidate mechanisms for the intricate intercommunication between epithelium and cells of the lamina propria that regulates neurogenesis. We will test their efficacy in vitro using our newly developed HBC culture system by quantifying the proportion of neurons that emerge after activation by the various factors. Then we will validate the in vitro effect by transplanting the activated/factor-treated HBCs into the lesioned OE, which provides a neutral environment in which all neuronal and non-neuronal fates are available to either endogenous HBCs or ones that have been cultured. We will test efficacy on both mouse and human HBCs, and we will use a novel xenotransplantation assay to assess the consequences for human HBCs in vivo. When completed, we will have achieved a much more thorough understanding of the process by which HBCs are directed toward neuronal differentiation so that they might rejuvenate neurogenesis. That understanding of mechanism both in mouse, where genetic manipulations offer profound analytic power, and in humans will advance our efforts aimed at identifying therapeutic strategies for alleviating olfactory sensory dysfunction, particularly the sensory loss which accompanies aging.