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Mechanisms regulating afferent innervation in the dental pulp

调节牙髓传入神经支配的机制

基本信息

批准号：
10453569
负责人：
Sarah Peters
金额：
$ 24.36万
依托单位：
OHIO STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10453569
关键词：
Adult Automobile Driving Axon Biological Assay Cells Cementoclast Chronic Disease Coculture Techniques Complex Data Defect Dental Dental Pulp Dental caries Dental crowns Dentin Deposition Development Developmental Gene Embryo Endodontics Enterobacteria phage P1 Cre recombinase Eph Family Receptors Epithelial Exposure to Extracellular Matrix Proteins Future Ganglia Generations Genetic Transcription Goals Image In Vitro Injury Investigation Knockout Mice Laboratories Life Maintenance Mediating Mesenchymal Mesenchyme Modeling Mus Mutant Strains Mice NTN1 gene Natural regeneration Nerve Nerve Fibers Nervous system structure Neurites Neurons Odontoblasts Organ Organism Pain Pain management Paracrine Communication Pathway interactions Pattern Penetration Perinatal mortality demographics Phosphoproteins Plant Roots RNA Reader Recombinants Regulation Reporting Research Role Sensory Signal Transduction Stimulus Structure Structure of trigeminal ganglion Temperature Testing Therapeutic Tissues Tooth Germ Tooth structure Trigeminal nerve structure Viral Visualization afferent nerve axon guidance axonal sprouting base bone experimental study ganglion cell improved in vivo knock-down mRNA sequencing migration mouse model mutant nerve stem cell nerve supply neuroblast neurogenesis neuron development neuron regeneration neurotransmission neurotrophic factor new therapeutic target osteopontin overexpression postnatal postnatal development preservation pressure promoter regenerative approach response skeletal

项目摘要

Project Summary/Abstract Since teeth are exposed to environmental stimuli, tooth innervation is crucial to their protection and usage throughout the life of an organism. The tooth is primarily innervated with sensory nerve fibers from the trigeminal ganglion (TG) that protect the tooth organ by relaying noxious stimuli. The dental pulp (DP) secretes neurotrophic factors to guide axonal penetration and sprouting within the tooth during postnatal development in a highly regulated manner. Research has shown that secreted phosphoprotein, osteopontin (OPN), promotes neuronal migration, proliferation, and survival [2–6]. The long-term goal of this project is to understand the mesenchymal-neuronal signals that promote and maintain sensory innervation of the teeth. The overall objective is to determine the role of DP in regulating tooth innervation during development and regeneration. 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 with significant defects in bones and teeth [7,8]. We performed an mRNA- Seq analysis using control and mutant postnatal day 7 DP and found that neuronal maintenance and developmental genes were most highly regulated, including OPN. Immunofluorescent images indicated reduced innervation throughout the DP in Tgfbr2cko mice. Preliminary experiments with DP and primary TG nerves demonstrated increased axonal sprouting when TG cells were cultured with DP. Guided by these data, we will test our hypothesis with the following two specific aims: 1) test the hypothesis that Tgfbr2 is necessary to promote sensory innervation; 2) test the hypothesis that OPN signaling from the DP guides sensory innervation. In both aims, we propose to cross Tg(Thy1-YFP)16Jrs mice, which express a high level of YFP throughout the nervous system [9] to optimize visualization of the neurons. Under the first aim, a well- characterized neurite outgrowth assay will first be used to co-culture TG neurons with DP where Tgf signals are manipulated in the DP. In the second part of the first aim, we will use an in vivo dental injury model and investigate neuronal regeneration in Tgfbr2cko and WT mice. Under the second aim, we will similarly co-culture TG neurons with OPN-deleted DP cells +/- TGF1 and Tgfbr2-deleted cells supplemented with recombinant OPN to investigate developmental neurogenesis in the DP. We will perform the dental injury assay on OPN-/- mice to examine the mechanisms driving neuronal regeneration. The proposed research is significant because it is expected to advance and expand understanding of how DP cells protect the tooth organ via axonal guidance mechanisms. Such information will enhance our understanding of the complex interplay of mesenchymal-neuronal interactions in the tooth that could serve as a basis for future preventative, therapeutic, and regenerative strategies in endodontics and improve the preservation of teeth.

项目总结/摘要由于牙齿暴露于环境刺激，牙齿神经支配对于牙齿的保护和使用至关重要在一个有机体的一生中。牙齿主要由来自牙齿的感觉神经纤维支配。三叉神经节（TG）通过传递伤害性刺激来保护牙齿器官。牙髓（DP）分泌神经营养因子在出生后发育过程中引导轴突穿透和在牙齿内发芽高度规范的方式。研究表明，分泌的磷蛋白，骨桥蛋白（OPN），促进神经元迁移、增殖和存活[2-6]。本项目的长期目标是了解间充质神经元信号，促进和维持牙齿的感觉神经支配。整体目的探讨DP在牙齿发育和再生过程中对神经支配的调节作用。我们的中心假设是牙齿间充质中的Tgfbr 2通过OPN控制旁分泌信号，牙齿感觉神经支配我们的实验室已经建立了一个小鼠模型，其中Tgfbr 2是条件性的。使用osterix启动子驱动的Cre重组酶（Tgfbr 2cko）在产生成牙本质细胞的间充质中缺失。这些小鼠出生后存活，但骨骼和牙齿有明显缺陷[7，8]。我们做了一个mRNA- 使用对照和突变体出生后第7天DP进行Seq分析，发现神经元维持和发育基因受到最高度的调控，包括OPN。免疫荧光图像显示在Tgfbr 2cko小鼠中整个DP的神经支配减少。DP和初级TG的初步实验当TG细胞与DP一起培养时，神经表现出增加的轴突发芽。在这些数据的指导下，我们将通过以下两个具体目标来检验我们的假设：1）检验Tgfbr 2是必要的假设促进感觉神经支配; 2）测试来自DP的OPN信号传导引导感觉神经支配的假设。神经支配在这两个目标中，我们建议交叉Tg（Thy 1-YFP）16 Jrs小鼠，其表达高水平的YFP [9]以优化神经元的可视化。在第一个目标下，一个很好的- 将首先使用表征的神经突生长测定来共培养TG神经元与DP，其中Tgf β信号传导在DP中被操纵。在第一个目标的第二部分中，我们将使用体内牙损伤模型，研究Tgfbr 2cko和WT小鼠中神经元再生。在第二个目标下，我们将同样共同培养 TG神经元与补充有重组OPN缺失的DP细胞+/- TGF β 1和TGF β 2缺失的细胞 OPN研究DP中的发育神经发生。我们将对OPN-/-进行牙齿损伤测定。小鼠来研究驱动神经元再生的机制。这项研究意义重大，因为它有望推进和扩大DP细胞如何通过轴突保护牙齿器官的理解，指导机制。这些信息将加强我们对以下因素之间复杂相互作用的理解：牙齿中的间充质-神经元相互作用可以作为未来预防，治疗，和再生策略，并改善牙齿的保存。