Mechanisms regulating afferent innervation in the dental pulp

项目总结/摘要 由于牙齿暴露于环境刺激，牙齿神经支配对其终身保护至关重要， 年龄牙齿主要由三叉神经节（TG）的感觉神经纤维支配，这些感觉神经纤维有助于促进牙齿的生长。 通过传递有害刺激来保护牙齿器官。在出生后发育期间，牙髓（DP）分泌 神经营养因子以高度调节的方式引导轴突穿透和在牙齿内发芽。 转化生长因子β（transforming growth factor β，TGF β）超家族调节牙齿的许多发育过程， 包括矿化和神经支配。一种TGF β信号下游的分泌型磷蛋白，骨桥蛋白 (OPN)是一种主要的细胞外蛋白，调节牙齿矿化，也促进神经突生长。 牙齿损伤后，DP细胞释放神经营养因子，调节神经突生长和牙本质修复。 有趣的是，OPN-/-小鼠在牙髓愈合过程中不形成修复性牙本质，这表明OPN在牙髓愈合过程中起关键作用。 在这个过程中的社会角色。该项目的长期目标是了解保护细胞的细胞机制。 牙齿通过感觉神经支配。总体目标是确定来自牙中膜的信号的作用， 调节牙齿神经分布食糜。我们的中心假设是牙齿间充质中的Tgfbr 2控制着 旁分泌信号通过OPN引导牙齿感觉神经支配。我们实验室建立了一个小鼠模型 其中使用Osterix启动子在产生成牙本质细胞的间充质中有条件地缺失Tgfbr 2- 驱动的Cre重组酶（Tgfbr2cko）。这些小鼠出生后存活，但骨骼有明显缺陷 还有牙齿我们进行了一系列的实验，以确定Tgfbr 2信号是否在DP介导下， 食糜调节它们的分化和在牙齿发育期间引导神经突生长的能力。我们发现 OPN-/-小鼠在P7没有表现出最初假设的神经支配减少，但确实表现出神经支配减少。 轴突定位中断神经突生长在反应性和修复性牙本质中的作用 DP电池的矿化能力是否受损仍然是未知的。在这些数据的指导下， 我们的假设有以下两个具体的目标：1）分析OPN控制的信号级联， 牙间充质在发育过程中调节牙齿感觉神经支配; 2）建立牙齿感觉神经支配的程度， 其中与牙本质修复相关的神经突生长受到Tgfbr 2和/或OPN信号的调节， DP间充质。在第一个目标下，我们将进行对照和OPN-/-DP的mRNA序列分析。 以确定哪些信号可能会破坏传入组织。在第二个目标中，我们将使用 体内牙损伤模型，并研究Tgfbr2cko、OPN-/-和WT小鼠中的神经元再生。的 拟议的研究是重要的，因为它预计将推进和扩大了解如何DP 细胞通过涉及轴突引导的机制保护牙齿器官。这些信息将加强我们的联合国- 了解牙齿中间充质-神经元相互作用的复杂相互作用， 为将来的预防、治疗和再生策略提供基础，以改善牙髓的保存 的牙齿。

Project Summary/Abstract Since teeth are exposed to environmental stimuli, tooth innervation is crucial to their lifelong protection and us- age. The tooth is primarily innervated with sensory nerve fibers from the trigeminal ganglion (TG) that help pro- tect the tooth organ by relaying noxious stimuli. During postnatal development, the dental pulp (DP) secretes neurotrophic factors to guide axonal penetration and sprouting within the tooth in a highly regulated manner. The transforming growth factor β (TGFβ) superfamily regulates many developmental processes in teeth, in- cluding mineralization and innervation. A secreted phosphoprotein downstream of TGFβ signals, osteopontin (OPN), is a major extracellular protein that regulates tooth mineralization and also promotes neurite outgrowth. Following tooth injury, DP cells release neurotrophic factors that modulate neurite growth and dentin repair. Interestingly, OPN-/- mice do not form reparative dentin during pulpal healing, suggesting that OPN plays a cru- cial role in this process. The long-term goal of this project is to understand the cellular mechanisms that protect teeth via sensory innervation. The overall objective is to determine the role of signals from the dental mesen- chyme that regulate tooth innervation. Our central hypothesis is that Tgfbr2 in the dental mesenchyme governs paracrine signaling via OPN to guide tooth sensory innervation. Our laboratory has established a mouse model in which Tgfbr2 is conditionally deleted in odontoblast-producing mesenchyme using an Osterix promoter- driven Cre recombinase (Tgfbr2cko). These mice survive postnatally, but have significant defects in their bones and teeth. We performed a series of experiments to determine whether Tgfbr2 signaling in the DP mesen- chyme regulates their differentiation and ability to guide neurite outgrowth during tooth development. We found that OPN-/- mice do not demonstrate reduced innervation at P7 as originally hypothesized, but do exhibit dis- rupted axonal targeting. The role of neurite outgrowth in regulating reactionary and reparative dentine when the mineralization capacity of the DP cells is compromised remains unknown. Guided by these data, we will test our hypothesis with the following two specific aims: 1) To analyze the OPN-governed signaling cascades in the dental mesenchyme regulating tooth sensory innervation during development; 2) To establish the degree to which the neurite outgrowth associated with dentin repair is regulated by Tgfbr2 and/or OPN signaling in the DP mesenchyme. Under the first aim, we will perform an mRNA Sequence Analysis of control and OPN-/- DP tissue to determine what signals could be disrupting the afferent organization. In the second aim, we will use an in vivo dental injury model and investigate neuronal regeneration in Tgfbr2cko, OPN-/-, and WT mice. The proposed research is significant because it is expected to advance and expand the understanding of how DP cells protect the tooth organ via mechanisms involving axonal guidance. Such information will enhance our un- derstanding of the complex interplay of mesenchymal-neuronal interactions in the tooth that could serve as a basis for future preventive, therapeutic, and regenerative strategies in endodontics to improve the preservation of teeth.