Investigating the roles of Hh signaling and Gli1+ cells in primary and reparative dentinogenesis in mouse molars

项目概要/摘要 鉴定能够产生分泌成牙本质细胞的间充质干细胞（MSC）群体 牙本质在生长（初级牙本质发生）和修复（修复性牙本质发生）过程中对于牙本质至关重要 改进牙髓治疗、牙本质再生以及最终功能性生物工程的发展 牙齿。尽管在体内鉴定小鼠门牙内 MSC 方面取得了重大进展，但我们知道 更不用说小鼠磨牙中的间充质干细胞了，与门牙不同，它们不会持续生长，而且更容易生长。 与人类的牙列相似。本研究的目的是探讨当前有争议的角色 Hedgehog (Hh) 信号通路和 Gli1（一种转录因子，是激活的下游效应子） 小鼠磨牙初级和修复性牙本质发生中的经典 Hh 信号通路。两个具体目标 已被提议。目标 1 中的实验旨在检验以下假设：在生长的小鼠磨牙中， 牙齿间充质中的 Hh 信号传导和 Gli1 表达细胞产生初级成牙本质细胞。使用血统 追踪和 Gli1-CreERT2； Ai9 报告小鼠我们将检查 Gli1-Cre-tdTomato 表达的贡献 细胞对原代成牙本质细胞的影响以及 HhAntag（一种有效的小分子）抑制 Hh 信号传导的作用 Shh 通路抑制剂对 Gli1-Cre-tdTomato 细胞的激活及其对原代细胞的贡献 成牙本质细胞。目标 2 中的实验旨在检验以下假设：在成年小鼠的磨牙中，Hh 信号传导 牙髓中表达 Gli1 的细胞参与第二代成牙本质细胞的形成 和修复性牙本质发生。我们将对成人磨牙进行活体牙髓暴露实验 检查牙髓暴露后 Shh 配体和 Gli1 的表达。 Gli1-Cre-tdTomato的贡献 在修复性牙本质发生过程中表达细胞至第二代成牙本质细胞及其影响 Hh 信号传导对修复性牙本质发生的抑制以及 Gli1-Cre-tdTomato 表达的贡献 细胞到第二代成牙本质细胞将通过 GLi1- 中的细胞谱系追踪实验进行检查 CreERT2； Ai9记者鼠。拟议的针对小鼠臼齿的研究，这些臼齿不是持续生长的，并且 与人类牙列更加相似将使我们能够更好地理解 Hh 信号和 Gli1 的作用 在原发性和修复性牙本质发生中表达细胞，这是一个以前从未研究过的领域。

PROJECT SUMMARY / ABSTRACT Identification of the mesenchymal stem cell (MSC) population capable of giving rise to odontoblasts secreting dentin during growth (primary dentinogenesis) and repair (reparative dentinogenesis) is critical for the development of improved pulp therapy, regeneration of dentin and ultimately the bioengineering of functional teeth. Despite significant progress in the identification of MSCs within the mouse incisors in vivo, we know much less about MSCs in mouse molars that, unlike incisors, are not continuously growing and are more similar to human dentition. The goal of the present study is to examine the currently controversial roles of the Hedgehog (Hh) signaling pathway and Gli1, a transcription factor that is a downstream effector of the activated canonical Hh signaling pathway in primary and reparative dentinogenesis in mouse molars. Two specific aims have been proposed. Experiments in Aim 1 are designed to test the hypothesis that in growing mice molars, Hh signaling and Gli1 expressing cells in dental mesenchyme give rise to primary odontoblasts. Using lineage tracing and Gli1-CreERT2; Ai9 reporter mice we will examine the contribution of Gli1-Cre-tdTomato expressing cells to primary odontoblasts and the effects of inhibition of Hh signaling by HhAntag, a potent small molecule inhibitor of the Shh pathway, on the activation of Gli1-Cre-tdTomato+ cells and their contribution to primary odontoblasts. Experiments in Aim 2 are designed to test the hypothesis that in adult mice molars, Hh signaling and Gli1 expressing cells in the dental pulp are involved in the formation of second-generation odontoblasts and reparative dentinogenesis. We will perform experimental pulp exposure in adult molars in vivo and examine the expression of Shh ligand and Gli1 after pulp exposure. The contribution of Gli1-Cre-tdTomato expressing cells to second generation of odontoblasts during reparative dentinogenesis and the effects of inhibition of Hh signaling on reparative dentinogenesis and the contribution of Gli1-Cre-tdTomato expressing cells to second generation of odontoblasts will be examined by cell lineage-tracing experiments in GLi1- CreERT2; Ai9 reporter mice. The proposed studies in mouse molars that are not continuously growing and more similar to human dentition will enable us to gain a better understanding of roles of Hh signaling and Gli1 expressing cells in primary and reparative dentinogenesis, an area that has not been studies before.