Enamel Fluorosis: Mechanisms of Action

描述（由申请人提供）：牙釉质氟中毒是在接触过量氟化物后发生的牙釉质发育缺陷。氟牙釉质比普通牙釉质更多孔，含有更多的蛋白质。氟化物改变牙釉质形成的机制尚不完全清楚。在这一竞争性更新中，我们建议以我们在以前的拨款周期中完成的工作为基础，测试一种新的范式，即将氟中毒定义为氟化物被纳入形成珐琅质矿物时发生的基质相关变化。具体来说，我们认为氟化物加入到发育中的牙釉质晶体中会改变基质/蛋白质/蛋白酶的相互作用，也会降低基质pH值，从而更快地形成矿物质。这些ph相关的影响改变了成釉细胞的功能和调节，以及釉质的成熟。此外，我们提出，当基质蛋白或矿化磷灰石无法结合氟离子（即分泌前阶段）时，氟化物可以直接影响基因表达。深入了解氟化物改变牙釉质形成的机制将使我们能够制定策略，以尽量减少氟化物的负面影响，同时加强氟化物在预防龋齿方面的使用。我们将通过以下三个具体目标来研究氟斑牙的潜在机制。具体目标1：确定氟化物掺入生长的珐琅质矿物对磷灰石/蛋白质相互作用和淀粉原蛋白水解的作用。质谱法将用于确定含氟磷灰石如何在体外和体内改变淀粉原蛋白水解。具体目标2：确定氟相关的牙釉质基质pH值变化如何影响成釉细胞调节和矿物沉积。缓冲牙釉质基质相关pH变化能力降低的小鼠模型将用于将基质pH变化与成釉细胞调节和牙釉质矿化联系起来。特异性目的3：研究氟对牙釉质基质蛋白沉积及pH调节相关基因表达的影响。激光捕获显微解剖将分别收集有氟和没有氟暴露的小鼠分泌前、分泌期和成熟阶段的成釉细胞，用于全转录比较或qPCR分析发育中的牙釉质基质中调节pH值的基因。这些研究将由一个杰出的国际调查小组进行，我们预计在这个资助周期结束时，我们将彻底了解牙釉质发育过程中氟暴露如何导致氟中毒。

DESCRIPTION (provided by applicant): Enamel fluorosis is a defect in enamel development that occurs after exposure to excess fluoride. Fluorotic enamel is more porous, and contains more proteins than sound enamel. The mechanisms by which fluoride alters enamel formation remain incompletely understood. In this competing renewal, we propose to build on work that we have completed in previous grant cycles, to test a new paradigm that defines fluorosis as resulting from matrix related changes that occur as fluoride is incorporated into the forming enamel mineral. Specifically, we propose that incorporation of fluoride into the developing enamel crystals alters matrix/protein/proteinase interactions, and also can reduce matrix pH secondary to more rapid mineral formation. These pH-related effects alter ameloblast function and modulation, and enamel maturation. Furthermore, we propose that when matrix proteins or mineralizing apatite are not available to bind fluoride ions (ie presecretory stage), fluoride can have a direct effect on gene expression. A thorough understanding of the mechanisms by which fluoride can alter enamel formation will allow us to develop strategies to minimize the negative effects of fluoride while enhancing the use of fluoride for caries prevention We will investigate these potential mechanisms of dental fluorosis through the following three specific aims. Specific Aim 1: To determine the role of fluoride incorporation into the growing enamel mineral on apatite/ protein interactions and amelogenin hydrolysis. Mass spectrometry will be used to determine how fluoride-containing apatites alter amelogenin hydrolysis both in vitro and in vivo. Specific Aim 2: To determine how fluoride-related changes in enamel matrix pH affects ameloblast modulation and mineral deposition. Mouse models with a reduced capacity to buffer enamel matrix related pH changes will be used to correlate changes in matrix pH to ameloblast modulation and enamel mineralization. Specific Aim 3: To determine the effects of fluoride on the expression of genes related to enamel matrix protein deposition and pH regulation. Laser capture micro-dissection will be used to separately collect presecretory, secretory and maturation stage ameloblasts from mice with and without fluoride exposure for either whole transcript comparisons, or qPCR analysis of genes that modulate pH in the developing enamel matrix. These studies will be done with an exceptional team of international investigators, and we anticipate that at the completion of this grant cycle, that we will have a thorough understanding of how fluoride exposure during enamel development results in fluorosis. PUBLIC HEALTH RELEVANCE: Enamel fluorosis is a defect in enamel development seen after exposure to excess of fluoride in early childhood. We have shown that the mechanisms responsible for enamel fluorosis include both fluoride related effects in the developing enamel matrix, and altered gene expression. In this proposal we further explore the mechanisms of fluorosis, by analyzing changes in matrix protein hydrolysis, ameloblast modulation and stage specific gene expression in rodent incisor models for enamel fluorosis.