MOLECULAR REGULATION OF PERIODONTIUM FORMATION

描述（由申请人提供）：了解牙根发育和牙周组织形成的遗传控制是本研究的长期目标。分子和发育生物学的进步已经将许多基因的功能与器官发育的特定阶段联系起来。就牙齿发育而言，据报道有超过300个基因在这一过程中起着一定的作用。正如预期的那样，这些基因的突变会导致牙齿缺陷，其严重程度取决于突变如何影响蛋白质的功能或特定基因调节的过程。与基因功能有关的许多知识来自转基因动物的使用，这使得可以创建遗传疾病的动物模型。在牙齿形成的情况下，许多基因参与早期阶段的牙齿发育和冠的形成已经建立。几年前，我们报道了转录因子核因子I-C （NFI-C）在小鼠体内功能的丧失导致了一种独特的表型，其特征是臼齿非常短或根本没有根。未发现其他表型，冠内未发现缺陷，表明NFI-C基因产物对根形成过程至关重要。NFI- C是第一个也是迄今为止唯一发现的调节牙根发育和牙周组织形成的基因。根的形成与冠的形成一样，依赖于上皮-间质相互作用，导致相应组织的分化和矿化细胞外基质的产生。由于冠和根的主要区别在于其上皮成分，在冠中，上皮成分由星状网和中间层隔开的外层和内层牙釉质上皮组成。根只包含内部和外部的牙釉质上皮，形成被称为Hertwig上皮根鞘（HERS）的结构。基于这一差异和初步数据部分的数据，我们提出验证Nfi-c通过调节HERS的功能来控制牙周组织的发育，进而控制牙根形态发生、成牙细胞分化、牙本质和牙骨质的产生以及牙周韧带的形成的中心假设。我们提出通过实现以下具体目标来验证这一假设：确定NFI-C在HERS分化和成熟中的作用；确定与HERS分化和成熟相关的NFI-C下游基因；确定与根发育相关的细胞中NFI-C的相互作用；确定HERS控制成牙细胞分化的机制，确定HERS控制牙周韧带和牙骨质形成的机制。这些研究将使用NFI-C敲除小鼠以及我们建立的与根形成相关的组织、shRNA和微阵列技术的不朽细胞系。本研究所产生的数据将为正常和异常的牙根和牙周组织形成过程以及参与该过程的调节因子提供急需的信息。这些数据对于了解导致牙根缺损、牙根发育不全和/或牙齿过早脱落的基因突变具有重要的健康意义，这对于建立未来牙周组织再生的基因和组织工程疗法至关重要。

DESCRIPTION (provided by applicant): It is the long-term objective of this study to understand the genetic control of root development and periodontium formation. Advances in molecular and developmental biology have linked the function of numerous genes to specific stages of organ development. In the case of tooth development, more than 300 hundred genes have been reported as having some role in this process. As expected, mutations in these genes can result in dental defects which vary in severity depending on how the mutation affects the function of that protein or what process that particular gene regulates. Much of the knowledge relating a gene to its function is derived from the use of transgenic animals which has allowed the creation of animal models for genetic diseases. In the case of tooth formation, many of the genes involved in the early stages of tooth development and crown formation have been established. A few years ago we reported that the loss of function of the transcription factor Nuclear Factor I-C (NFI-C) in mice results in a unique phenotype characterized by molars with very short or no roots at all. No other phenotype was found and no defects in the crowns were detected indicating that the NFI-C gene product is essential for the process of root formation. NFI- C is the first and up to date only gene that has been found to regulate root development and therefore periodontium formation. Root formation, like crown formation, is dependent on epithelial-mesenchymal interactions resulting in differentiation of the corresponding tissues and production of the mineralized extracellular matrices. Since the main difference between the crown and root is its epithelial component which in the crown is composed by the outer and inner enamel epithelia separated by the stellate reticulum and the stratum intermedium. The root contains only inner and outer enamel epithelia forming the structure known as Hertwig's Epithelial Root Sheath (HERS). Based on this difference and data presented in the preliminary data section, we propose to test the central hypothesis that Nfi-c controls periodontium development by regulating the function of HERS which in turn control root morphogenesis, odontoblast differentiation, dentin and cementum production and periodontal ligament formation. We propose to test this hypothesis by realizing the following Specific Aims: establish the role of NFI-C on HERS differentiation and maturation; determine NFI-C down-stream genes associated with HERS differentiation and maturation; determine NFI-C interactions in cells associated with root development; determine the mechanism by which HERS control odontoblast differentiation and determine the mechanism by which HERS control periodontal ligament and cementum formation. These studies will use the NFI-C knockout mice as well as our established immortal cell lines for the tissues associated with root formation, shRNA and microarray technology. The data generated in this study will provide much needed information related to the process of normal and abnormal root and periodontium formation and the regulatory factors involved in this process. This data is of great health relevance to understand genetic mutations resulting in root defects, root agenesis and/or premature tooth loss which is critical to establish future genetical and tissue engineering therapies for periodontium regeneration. PUBLIC HEALTH RELEVANCE: It is the long-term objective of this study to understand the process of root development and periodontium formation at the morphological, cellular and molecular levels. This includes the mechanisms underlying root dentin, cementum and periodontal ligament formation which lead to root attachment in the healthy periodontium. This application will focus on the role of the transcription factor NFI-C which appears to be essential for periodontium formation. The data generated in this study is of great health relevance to understand genetic defects resulting in root agenesis and periodontium defects which result in premature tooth loss. Additionally, understanding root and periodontium development is critical to establish genetical and tissue engineering therapies for periodontium regeneration.