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

项目摘要 牙周炎被认为具有很强的遗传背景，但我们对牙周炎的决定因素的了解 对这种炎性疾病的易感性仍然是不完全的，并阻碍了有效治疗的发展。 个体化的医学方法来预防和治疗。同样， 控制疾病的进展，从牙周健康状态到牙龈炎再到严重的牙周炎， 仍然没有被充分理解。新出现的证据表明，表观遗传调控在以下方面发挥着重要作用： 具有炎症成分的多种病理状况。本项目将采用 遗传/表观遗传策略研究牙周炎易感性和发病机制。在我们的第一个目标中，我们将 检查早期鉴定的遗传位点是否与牙周炎的程度和严重程度相关， 在华盛顿高地/因伍德社区，某些细菌物种的定殖可以复制 老龄化项目（WHICAP），一个以社区为基础的老年人队列， 通过下一代测序评估的数据和龈下微生物概况。我们将列出 候选基因和简单核苷酸多态性，包括统计学显著性和提示性 GWAS信号，我们将提出交叉验证和精细映射。在我们的第二个目标，我们 假设单倍型依赖等位基因特异性DNA甲基化（hap-ASM）和甲基化 患者外周血和牙龈组织中关键细胞类型的数量性状基因座（mQTL） 将识别牙周炎易感性的真阳性GWAS信号。在一个新的队列中， 牙周炎患者，我们将产生细胞特异性hap-ASM和mQTL的全基因组图谱， 基于微阵列和下一代亚硫酸氢盐测序，并将这些地图与上述和 目标1的亚阈值GWAS峰值。鉴定在两种类型的数据中得分为正的基因座将允许 我们确定真正的调节单倍型和特定的调节序列元件，介导中间- 牙周炎易感性的个体差异。在我们的第三个目标中，我们将测试映射的假设， 来自健康和牙周炎影响部位的牙龈组织细胞中基因特异性DNA甲基化模式 将揭示牙周炎发病相关的表观遗传调控基因和途径。在 与目标2中相同的患者，我们将评估激光照射后CpG甲基化和羟甲基化的变化。 从牙周炎影响的牙龈和健康牙龈中捕获显微解剖的上皮细胞和成纤维细胞， 使用免疫组织化学标记和基因表达分析来验证我们的发现。我们希望我们 一种新的结合遗传学/表观遗传学的方法将提高我们识别牙周炎易感者的能力。 个人，将提高我们对牙周炎发病机制中组织局部反应的理解， 将最终为牙周炎预防和治疗的个性化药物设计提供信息。 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