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

项目总结/摘要 牙根是牙齿的关键组成部分，通过牙槽骨锚定到周围的牙槽骨。 牙周膜（PDL）。根及其周围结构的适当形成不仅是至关重要的， 对于牙齿的基本功能，在咀嚼营养摄入，但也为生长和发育， 下面的脸。龋病、牙周病等口腔常见疾病是导致牙齿松动的主要原因 损失，只能通过缺乏功能结构的假体治疗，导致长期受损 预后需要一种有效的方法来再生功能性牙周附着装置， 在牙齿再生治疗方面取得突破。在牙根形成过程中，牙囊（DF） 为成牙骨质细胞、牙周膜（PDL）细胞和牙槽骨隐窝骨提供前体细胞 成骨细胞，以建立功能性牙周附着装置，牙周组织。目前，如何 不同的牙根间充质祖细胞群体一起工作， 牙根仍然未知。在本提案中，我们将定义不同类别的企业如何采取协调一致的行动， 牙根间充质祖细胞对于牙根的正常形成和再生是必需的 并伴有功能性牙周附着装置。在目标1中，我们将定义刺猬狐狸 信号通路调节DF间充质祖细胞。我们假设叉头（福克斯）转录 因子在调节Hedgehog反应性DF间充质细胞的生理功能中起关键作用 祖细胞我们将首先使用单细胞RNA-聚合酶链反应技术揭示Hedgehog反应性Gli 1 + 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体的关系。 第二，我们将使用免疫组织化学方法确定Gli 1+、PTHrP+和Runx 2+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Foxf 1和Foxf 2在牙周组织形成中的作用 使用他们的floxed等位基因。在目标2中，我们将确定Wnt介导的顶根间充质祖细胞的作用 牙根形成的细胞。我们假设趋化因子（C-X-C基序）配体12（CXCL 12）+间充质细胞 根尖区的祖细胞协调牙根和根尖牙周组织的形成， 典型的Wnt信号介导的方式。我们将确定CXCL 12+细胞与 使用离体培养系统，研究了根尖乳头干细胞（SCAP），并确定了其分子机制 a通过比较RNA-seq分析的间充质祖细胞的Wnt介导的细胞命运选择， 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胚中的作用。 牙周再生我们假设不同种类的牙根间充质祖细胞 以协调一致的方式促进再生。我们将定义Gli 1+、PTHrP+、Runx 2+的后代如何 和CXCL 12+祖细胞有助于骨破坏后的牙周再生，通过利用 模拟牙周病的结扎诱导的牙周炎模型和外科牙周缺损模型。

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.