A combined genetic/epigenetic approach to study periodontitis susceptibility and pathobiology

项目摘要 牙周炎被认为有很强的遗传背景，但我们对牙周炎决定因素的了解 这种炎症性疾病的易感性仍然不完全，阻碍了有效的发展 个体化的医学方法防治它。同样，分子基础 控制疾病的发展，从牙周健康状态到牙周炎再到严重牙周炎， 仍然没有得到充分的理解。新出现的证据表明，表观遗传调控在 有炎症性成分的各种病理情况。该项目将采用一种联合的 研究牙周炎易感性和发病机制的遗传/表观遗传学策略。在我们的第一个目标中，我们将 检查早期发现的基因座是否与牙周炎的程度和严重程度相关，或者与 某些细菌物种的定居可以在华盛顿高地/因伍德社区复制 老龄化项目(WHICAP)，一个以社区为基础的老年人队列，有可用的临床牙周 通过下一代测序评估的数据和牙龈下微生物图谱。我们将列出一份清单， 候选基因和简单核苷酸多态，包括统计上有意义的和有提示意义的 Gwas信号，我们将提出进行交叉验证和精细绘图。在我们的第二个目标，我们 假设单倍型依赖的等位基因特异性DNA甲基化(HAP-ASM)和甲基化的作图 患者外周血和牙周组织中关键细胞类型的数量性状基因座 对于牙周炎的易感性，与牙周炎的联合检测将识别真正阳性的GWAs信号。在一个新的队列中 牙周炎患者，我们将使用以下工具制作细胞特异性HAP-ASM和mQTL的全基因组图谱 基于微阵列的和新一代亚硫酸氢盐测序，并将与以上和 低于阈值的GWAS峰来自目标1。识别在两种类型的数据中得分为正的基因座将允许 美国将确定真正的调控单倍型和特定的调控序列元件 患牙周炎的个体差异。在我们的第三个目标中，我们将测试映射的假设 牙周健康和牙周炎患者牙周组织细胞中基因特异性DNA甲基化模式的研究 将揭示与牙周炎发病机制相关的表观遗传调控基因和途径。在 与目标2相同的患者，我们将评估激光治疗中CpG甲基化和羟甲基化的变化。 从受牙周炎影响的牙周组织和健康牙周组织中捕获显微解剖的上皮细胞和成纤维细胞，并将 使用免疫组织化学标记物和基因表达分析来验证我们的发现。我们希望我们的 新的遗传/表观遗传学相结合的方法将提高我们识别牙周炎易感性的能力 将加深我们对牙周炎发病机制中组织局部反应的理解，以及 最终将指导设计一种个性化的医学方法来预防和治疗牙周炎。 1

Project Summary Periodontitis is thought to have a strong genetic background, but our knowledge on the determinants of susceptibility to this inflammatory disease is still incomplete and hinders the development of effective personalized medicine approaches to its prevention and treatment. Likewise, the molecular underpinnings governing the progression of the disease, from states of periodontal health to gingivitis to severe periodontitis, are still inadequately understood. Emerging evidence suggests an important role of epigenetic regulation in diverse pathologic conditions that have an inflammatory component. This project will adopt a combined genetic/epigenetic strategy to study periodontitis susceptibility and pathogenesis. In our first aim, we will examine if earlier identified genetic loci associated with the extent and severity of periodontitis or with colonization by certain bacterial species can be replicated in the Washington Heights/Inwood Community Aging Project (WHICAP), a community based cohort of elderly individuals with available clinical periodontal data and subgingival microbial profiles assessed through next generation sequencing. We will produce a list of candidate genes and simple nucleotide polymorphisms, including both statistically significant and suggestive GWAS signals, which we will bring forward for cross-validation and fine mapping. In our second aim, we hypothesize that mapping of haplotype-dependent allele-specific DNA methylation (hap-ASM) and methylation quantitative trait loci (mQTLs) in key cell types in the peripheral blood and in the gingival tissues of patients with periodontitis will identify true-positive GWAS signals for periodontitis susceptibility. In a new cohort of periodontitis patients, we will produce genome-wide maps of cell-specific hap-ASM and mQTLs using microarray-based and next generation bisulfite sequencing, and will overlap these maps with the supra- and sub-threshold GWAS peaks from aim 1. Identification of loci that score positively in both types of data will allow us to pinpoint bona fide regulatory haplotypes and specific regulatory sequence elements that mediate inter- individual differences in susceptibility to periodontitis. In our third aim, we will test the hypothesis that mapping of gene-specific DNA methylation patterns in gingival tissue cells from healthy and periodontitis-affected sites will reveal epigenetically regulated genes and pathways relevant to the pathogenesis of periodontitis. In the same patients as in Aim 2, we will assess changes in CpG methylation and hydroxymethylation in laser- capture micro-dissected epithelial cells and fibroblasts from periodontitis-affected vs. healthy gingiva, and will validate our findings using immunohistochemical markers and gene expression assays. We expect that our novel, combined genetic/epigenetic approach will improve our ability to identify periodontitis-susceptible individuals, will enhance our understanding of the tissue-localized responses in periodontitis pathogenesis, and will ultimately inform the design of a personalized medicine approach to periodontitis prevention and treatment. 1