Dental Follicle: A Central Regulator of Tooth Root Formation and Regeneration

项目摘要/摘要 牙根是牙齿的重要组成部分，通过牙根固定在周围的牙槽骨上 牙周膜(PDL)。根部及其周围结构的适当形成不仅至关重要 对于牙齿在咀嚼中的基本功能，对于营养的摄入，也是对生长发育的重要作用 脸部较低。常见的牙病，如龋齿和牙周病是牙齿的病因。 丢失，只能通过缺乏功能结构的假体来治疗，导致长期受损 预后。需要一种有效的方法来再生具有功能的牙周附着装置 实现牙齿再生治疗的突破。在牙根形成过程中，牙囊(DF) 为成牙骨质细胞、牙周膜细胞和牙槽隐骨提供前体细胞 成骨细胞建立起功能性牙周附着器官--牙周组织。目前，如何 不同群体的牙根间充质祖细胞共同作用并产生高功能 牙根仍不清楚。在这项提案中，我们将定义不同类别的 牙根间充质祖细胞是牙根正常形成和再生所必需的 配有功能齐全的牙周附着器。在目标1中，我们将定义刺猬-狐狸如何 调控DF间充质祖细胞的途径。我们假设叉头(狐狸)转录 因子在调节Hedgehog反应性DF间充质的生理功能中起关键作用 祖细胞。我们将首先利用单细胞RNA揭示刺猬反应Gli1 DF细胞的多样性。 SEQ分析，并确定它们与PTHrP DF细胞和其他前体细胞群的关系。 第二，我们将确定Gli1，PTHrP和Runx2细胞在牙周组织形成中的功能 诱导细胞消融实验，进一步检测Foxf1和Foxf2在牙周组织形成中的作用 利用它们的等位基因。在目标2中，我们将确定Wnt介导的根尖根间充质祖细胞的作用 牙根形成中的细胞。我们假设趋化因子(C-X-C基序)配体12(CXCL12)间充质 根尖区的祖细胞协调牙根和根尖牙周组织的形成 规范的Wnt信号传导方式。我们将确定CXCL12细胞与 尖端乳头干细胞(SCAP)的体外培养系统，并确定其分子机制 通过比较RNA-SEQ分析Wnt介导的间充质祖细胞的细胞命运选择 通过现场验证。在目标3中，我们将确定牙根间充质祖细胞在 牙周再生。我们假设不同类别的牙根间充质祖细胞 以协调一致的方式为再生作出贡献。我们将定义Gli1、PTHrP、Runx2的后代如何 和CXCL12祖细胞有助于骨破坏后的牙周再生，通过利用 模拟牙周病的牙周炎模型和外科牙周缺损症模型。

PROJECT SUMMARY/ABSTRACT The tooth root is a critical component of the tooth, anchored to surrounding alveolar bones through the periodontal ligament (PDL). Appropriate formation of the root and its surrounding structure is essential not only for fundamental functions of the tooth in mastication for nutrition intake, but also for growth and development of the lower face. Prevalent dental diseases such as caries and periodontal diseases are the etiology for tooth loss, which can be treated only by prostheses lacking functional structures, leading to compromised long-term prognosis. An effective approach to regenerate a functional periodontal attachment apparatus is needed to achieve a breakthrough in dental regenerative therapies. During tooth root formation, the dental follicle (DF) provides precursor cells for cementoblasts, periodontal ligament (PDL) cells and alveolar cryptal bone osteoblasts to establish the functional periodontal attachment apparatus, the periodontium. Currently, how diverse populations of dental root mesenchymal progenitor cells work together and generate highly functional tooth roots remains unknown. In this proposal, we will define how concerted actions of distinct classes of dental root mesenchymal progenitor cells are essential for proper formation and regeneration of the tooth root accompanied by a functional periodontal attachment apparatus. In Aim 1, we will define how Hedgehog-Fox pathway regulates DF mesenchymal progenitor cells. We hypothesize that the Forkhead (Fox) transcription factors play a key role in regulating physiological functions of Hedgehog-responsive DF mesenchymal progenitor cells. We will first reveal the diversity of Hedgehog-responsive Gli1+ DF cells using single cell RNA- seq analyses, and determine their relationships with PTHrP+ DF cells and other precursor cell populations. Second, we will determine the function of Gli1+, PTHrP+ and Runx2+ cells in periodontium formation using inducible cell ablation experiments, and further test the function of Foxf1 and Foxf2 in periodontium formation using their floxed alleles. In Aim 2, we will identify Wnt-mediated roles of apical root mesenchymal progenitor cells in tooth root formation. We hypothesize that chemokine (C-X-C motif) ligand 12 (CXCL12)+ mesenchymal progenitor cells in the apical root area orchestrate formation of the tooth root and the apical periodontium in a canonical Wnt signaling-mediated manner. We will determine the relationship between CXCL12+ cells and stem cells for apical papilla (SCAP) using ex vivo culture system, and define molecular mechanisms underlying a Wnt-mediated cell fate choice of mesenchymal progenitor cells by a comparative RNA-seq analysis followed by in situ validation. In Aim 3, we will determine actions of dental root mesenchymal progenitor cells in periodontal regeneration. We hypothesize that distinct classes of dental root mesenchymal progenitor cells contribute to regeneration in a concerted manner. We will define how descendants of Gli1+, PTHrP+, Runx2+ and CXCL12+ progenitor cells contribute to periodontal regeneration after bone destruction, by utilizing a ligature-induced periodontitis model mimicking periodontal diseases, and a surgical periodontal defect model.