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The Impact of Fluoride on the Rho Signaling Pathway and the Actin Cytoskeleton

氟化物对 Rho 信号通路和肌动蛋白细胞骨架的影响

基本信息

批准号：
7471498
负责人：
Carolyn Gibson
金额：
$ 19.47万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-08-01 至 2010-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7471498
关键词：
Actins Adopted Ameloblasts Animal Model Biochemical Biological Assay Biological Models Cells Cellular Morphology Communities Confocal Microscopy Cytoskeleton Data Defect Dental Dental Enamel Dental Esthetics Dental caries Development Dominant-Negative Mutation Dose Electrons Elevation Environmental Impact Epithelial Cells Exposure to F-Actin Fibroblasts Fluorides G-Protein Signaling Pathway Generations Geographic Locations Goals Histologic Image Incisor Knowledge Lead Link Microscopic Monitor Monomeric GTP-Binding Proteins Morphology Mus Organ Organ Culture Techniques Pathway interactions Play Policies Positioning Attribute Rattus Role Scanning Signal Pathway Sodium Fluoride Staging Testing Thinking Tissues Tooth structure Transgenic Mice Transgenic Organisms United States Water Supply amelogenin dietary excess drinking water environmental agent fluorosis in vivo inhibitor/antagonist mouse model novel public drinking research study response rho transgene expression

项目摘要

DESCRIPTION (provided by applicant): Fluoride is added to drinking water supplied to two-thirds of the communities in the U.S. because of its proven benefit for reduction of dental caries. However, exposure to elevated dietary fluoride during tooth development leads to the enamel structural defect called enamel fluorosis, with dental aesthetic concerns in greater than 12% of residents of some fluoridated communities. Elevated fluoride in drinking water provided to rats led to alterations in the normal ameloblast morphology, and effects on these fluoride-sensitive, enamel-secreting cells are thought to contribute to the enamel structural defects. Using tooth organ culture and specific inhibitors, we have shown that the RhoA pathway is responsible for one of the responses of ameloblasts to sodium fluoride, that of elevation of filamentous actin (F-actin). F-actin localization in ameloblasts normally changes during developmental stages as ameloblast function changes, and we propose that RhoA is central to fluoride-induced cytoskeletal deregulation. We propose to test the central position of the RhoA pathway in the fluoride response with 2 specific aims. In Aim 1, a novel transgenic mouse model will be generated that expresses a dominant-negative RhoA in ameloblasts, the epithelial cell layer that secretes enamel. Histological, confocal and scanning electron microscopic images of wild-type and transgenic mice will be evaluated, and the cellular response to fluoride in drinking water will be analyzed in vivo. In Aim 2, tooth organs from wild-type and RhoADN transgenic mice will be cultured with Rho pathway inhibitors or an activator and analyzed by confocal microscopy and biochemical approaches to monitor the response of RhoA and downstream targets in the pathway to fluoride. These experiments will provide a new animal model for discovery of the mechanism of action for this environmental agent on a small G protein signaling pathway. Fluoride is commonly added to public drinking water, but there are geographic locations where the maximum recommended dose is naturally exceeded. Although elevated fluoride levels lead to an unsightly dental defect known as fluorosis, slightly lower levels are highly beneficial in reducing dental caries. A better understanding of the health-related benefits and problems associated with environmental fluoride will lead to more informed policy decisions and fundamental knowledge about the impact of this environmental agent.

说明(申请人提供)：美国三分之二的社区的饮用水中添加了氟化物，因为氟化物被证明对减少龋齿有好处。然而，在牙齿发育过程中，暴露在高氟饮食中会导致釉质结构缺陷，称为釉质氟中毒，在一些含氟社区，超过12%的居民存在牙齿美容问题。给大鼠提供的饮用水中氟含量升高会导致正常成釉细胞形态的改变，而对这些对氟敏感的、分泌釉质的细胞的影响被认为是导致釉质结构缺陷的原因。利用牙齿器官培养和特异性抑制剂，我们已经证明RhoA通路是成釉细胞对氟化钠的反应之一，即丝状肌动蛋白(F-actin)的升高。随着成釉细胞功能的改变，F-肌动蛋白在成釉细胞中的定位通常会发生变化，我们认为RhoA是氟诱导的细胞骨架去调节的中心。我们建议用两个特定的目标来测试RhoA通路在氟化物反应中的中心位置。在目标1中，将产生一种新的转基因小鼠模型，该模型在成釉细胞中表达显性负RhoA，成釉细胞是分泌釉质的上皮细胞层。将评估野生型和转基因小鼠的组织学、共聚焦和扫描电子显微镜图像，并将在体内分析细胞对饮用水中氟的反应。在目标2中，野生型和RhoADN转基因小鼠的牙齿器官将在Rho途径抑制剂或激活剂中培养，并通过共聚焦显微镜和生化方法进行分析，以监测RhoA及其下游靶标对氟的反应。这些实验将为发现该环境因子在小G蛋白信号通路上的作用机制提供新的动物模型。氟化物通常被添加到公共饮用水中，但在地理位置上，自然会超过最大推荐剂量。尽管氟化物水平升高会导致一种被称为氟斑牙的难看的牙齿缺陷，但略低的氟化物水平对减少龋齿非常有益。更好地了解与健康相关的好处和与环境氟化物相关的问题将导致更明智的政策决定和关于这种环境制剂影响的基本知识。