Genetic Variations Related to Fluorosis Susceptibility

说明(申请人提供)：釉质氟中毒是牙齿形成过程中暴露在过量氟化物中的一种釉质发育缺陷。关于氟化物影响釉质形成的机制，特别是氟化物对成釉细胞功能的影响，仍有许多有待研究。对人类群体和小鼠品系的研究表明，釉质氟中毒个体易感性的一个基因成分。在这些研究中，我们将使用已知对氟化物敏感(A/J)或抗氟剂(129P3/J)的小鼠模型来识别对氟化物的这种可变反应的相关基因。将在这些研究中检验的总体假设是，釉质氟中毒是由于在釉质发育过程中改变釉原蛋白和基质金属蛋白酶-20的相对水平的因素造成的。这一假设将通过以下具体目标进行检验。具体目的1：确定候选基因在A/J和129P3/J小鼠品系中的表达与氟化牙釉质严重程度的相关性。特定目的2：确定氟是否对A/J和129P3/J小鼠的JNK和cJun的磷酸化有不同的影响。具体目的3：比较A/J和129P3/J小鼠品系中釉原蛋白基因的调控区，并确定差异与氟斑牙易感性/抵抗力的关系。我们将使用激光显微解剖显微镜从饮水中给予氟的小鼠的门牙分离出分泌期和成熟期的成釉细胞。实时定量聚合酶链式反应将被用来定量釉原蛋白、基质金属蛋白酶-20和其他候选基因的相对表达，以确定在遗传多样性的小鼠品系中对氟的不同反应。通过测量两个小鼠模型中JNK/c-Jun合成和磷酸化的相对差异，将进一步研究基质金属蛋白酶-20调节因子的遗传差异。将通过比较两种小鼠模型中的启动子序列来研究釉原蛋白基因的多态。我们预计，这些研究将导致识别候选基因，这些基因将形成R01应用程序的基础，以进一步确定人类对氟中毒的遗传易感性。

DESCRIPTION (provided by applicant): Enamel fluorosis is a defect in enamel development seen after exposure to excess fluoride during tooth formation. Much remains to be learned about the mechanisms by which fluoride affects enamel formation, and in particular the effect of fluoride on ameloblast function. Studies of human populations and mouse strains suggest that a genetic component to individual susceptibility to enamel fluorosis. In these studies we will use mouse models known to be either fluoride sensitive (A/J) or fluoride resistant (129P3/J) to identify genes responsible for this variable response to fluoride. The overall hypothesis that will be tested in these studies is that enamel fluorosis results from factors that alter the relative levels of amelogenin and MMP-20 during enamel development. This hypothesis will be tested by the following Specific Aims. Specific Aim 1: To determine the correlation between the expression of the candidate genes and severity of fluorosed enamel in the A/J and 129P3/J mouse strains. Specific Aim 2: To determine whether fluoride differentially effects phosphorylation of JNK and cJun, in A/J and 129P3/J mouse strains. Specific Aim 3: To compare the regulatory regions of the amelogenin gene in both A/J and 129P3/J mouse strains and determine how the differences relate to susceptibility/resistance to Dental fluorosis. We will use laser microdissection microscopy to separate secretory and maturation stage ameloblasts from incisors of mice given fluoride in drinking water. Real-time PCR will be used to quantitate relative expression of amelogenin, MMP-20, and other candidate genes to identify variable response to fluoride in the genetically diverse mouse strains. Genetic differences in regulators of MMP-20 will be further investigated by measuring relative differences in JNK/c-Jun synthesis and phosphorylation in the two mouse models. Polymorphisms in the amelogenin gene will be investigated by comparing promoter sequences in the two mouse models. We anticipate that these studies will result in the identification of candidate genes that will form the basis an R01 application to further determine genetic susceptibility to fluorosis in humans.