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Multiple Functions of Glutathione Catabolism and H2S in T. denticola Pathogenesis

谷胱甘肽分解代谢和 H2S 在 T. denticola 发病机制中的多种功能

基本信息

批准号：
9043000
负责人：
LIANRUI CHU
金额：
$ 37.38万
依托单位：
UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-05-01 至 2019-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9043000
关键词：
Abscess Affect Alveolar Bone Loss Animal Model Apoptosis Back Bacteria Bacterial Infections Biological Assay Bone Resorption Bone Tissue Catabolism Cells Characteristics Consensus Development Disease Enzymes Fibroblasts Gamma-glutamyl transferase Generations Genes Genetic Gingiva Gingival Crevicular Fluid Glutathione Goals Health Homeostasis Human IL8 gene In Vitro Infection Inflammation Interleukin-6 Investigation Lead Lesion Measures Metabolic Pathway Modeling Mus Outcome Outcome Study Pain Pathogenesis Pathology Pathway interactions Pattern Periodontal Diseases Periodontal Ligament Periodontal Pocket Periodontitis Physiological Physiology Play Positioning Attribute Prevention strategy Production Proteins Rattus Reduced Glutathione Role Site Sulfhydryl Compounds Testing Time Tissues Tooth Exfoliation Treponema denticola alveolar bone bacterial metabolism base bone loss cell injury cell type cystalysin cytokine enzyme activity in vivo inhibitor/antagonist innovation insight mutant novel pathogen research study small molecule soft tissue trait treatment strategy

项目摘要

DESCRIPTION (provided by applicant): In diseased periodontal pockets, glutathione levels are lower than in healthy sites and the amount of H2S is higher. It has been assumed, but not proven, that changes in the concentrations of these thiol-compounds are important for the tissue pathology seen in periodontitis. Although the mechanism by which the levels of glutathione and H2S are perturbed in diseased pockets is unknown, the consensus is that bacteria play a key role. Thus, we have been studying the ability of periodontal pathogens to produce H2S from glutathione since such a metabolic pathway could alter the levels of these two molecules in the gingival crevice. We have focused on the major periodontal pathogen Treponema denticola and have shown previously that it can catabolize glutathione to H2S via a three step enzymatic pathway (GTSP). Our goal is to elucidate the role of such thiol catabolic pathways, and the metabolites they use and produce, in periodontal pathology. To this end, we have conducted several preliminary experiments with the following relevant results: (1) T. denticola plus glutathione can induce apoptosis in human gingival fibroblasts and periodontal ligament cells in vitro. (2) Periodontal ligament cell synthesis of proinflammatory cytokines is increased by T. denticola plus glutathione in vitro. (3) Glutathione exacerbates the lesion size caused by T. denticola in a mouse abscess model. (4) T. denticola enhances alveolar bone resorption in rats and decreases gingival crevicular fluid glutathione levels. (5) Most significantly, we recently constructed a T. denticola deletion mutant in the first gene (ggt) of the GTSP. This mutant cannot convert glutathione into H2S. Thus, we are uniquely positioned to test the hypothesis that the decrease in glutathione levels by bacterial metabolism, specifically the T. denticola GTSP, and the accompanying increase in H2S production will play key roles in the host tissue damage seen in periodontitis. In Aim 1, we will use wild type T. denticola and our Δggt mutant to demonstrate that the GTSP, particularly its generation of H2S, enhances apoptosis in periodontal cells and modulates the levels and pattern of cytokines produced by host cells in vitro. Most importantly, the T. denticola mutant will be used in two animal models (Aim 2) to prove that a bacterium's ability to lower glutathione and increase H2S is critical for causing tissue/bone pathology in vivo. The rat model of alveolar bone loss will also be used to test the ability of three GTSP inhibitors, which we have already characterized in vitro, to limit T. denticola pathogenesis in vivo. The outcomes of the proposed studies will prove, for the first time, that a bacterium's ability to influence glutathione and H2S levels in the subgingival crevice is a significant contributor to periodontal tissue damage. Although the experiments are only being done with T. denticola, the results would establish a new paradigm whereby the physiology of periodontal bacteria, singly or synergistically, could perturb thiol-molecule homeostasis to cause disease pathology. Re-establishing normal periodontal pocket metabolite levels pharmacologically by inhibiting a bacterial catabolic pathway could be an innovative strategy to reduce host damage in periodontitis.

描述(申请人提供)：在患病的牙周袋中，谷胱甘肽水平低于健康部位，而硫化氢的含量较高。人们一直认为，这些硫醇化合物浓度的变化对牙周炎的组织病理学很重要，但还没有得到证实。尽管疾病眼袋中谷胱甘肽和硫化氢水平受到干扰的机制尚不清楚，但人们的共识是细菌发挥了关键作用。因此，我们一直在研究牙周病原体从谷胱甘肽产生硫化氢的能力，因为这样的代谢途径可以改变牙周缝隙中这两个分子的水平。我们重点研究了牙周的主要病原体齿密螺旋体，并表明它可以通过三步酶途径(GTSP)将谷胱甘肽分解为H_2S。我们的目标是阐明这种硫醇分解代谢途径的作用，以及它们使用和产生的代谢物在牙周病理中的作用。为此，我们进行了几个初步的实验，取得了以下相关结果：(1)齿轮虫联合谷胱甘肽在体外可以诱导人牙龈成纤维细胞和牙周膜细胞的凋亡。(2)牙周膜细胞在体外与谷胱甘肽联合作用后，其促炎症细胞因子的合成增加。(3)在小鼠脓肿模型中，谷胱甘肽可加重齿状毛滴虫引起的病变面积。(4)牙周炎可促进大鼠牙槽骨吸收，降低龈沟液中谷胱甘肽水平。(5)最重要的是，我们最近在GTSP的第一个基因(GGT)上构建了一个齿纹夜蛾缺失突变体。该突变体不能将谷胱甘肽转化为硫化氢。因此，我们处于独特的地位来检验这一假说，即细菌代谢，特别是齿状毛滴虫GTSP导致的谷胱甘肽水平的降低，以及伴随而来的硫化氢产量的增加，将在牙周炎的宿主组织损伤中发挥关键作用。在目标1中，我们将使用野生型齿状毛滴虫和我们的ΔGGT突变体来证明GTSP，特别是它产生的H_2S，在体外促进牙周细胞的凋亡，并调节宿主细胞产生的细胞因子的水平和模式。最重要的是，齿状毛滴虫突变体将用于两个动物模型(目标2)，以证明细菌降低谷胱甘肽和增加硫化氢的能力是导致体内组织/骨骼病理的关键。牙槽骨丢失的大鼠模型也将被用来测试我们已经在体外确定的三种GTSP抑制剂在体内限制齿状毛滴虫致病的能力。拟议中的研究结果将首次证明，细菌影响龈下缝隙中谷胱甘肽和硫化氢水平的能力是牙周组织损伤的重要因素。虽然这些实验只在齿轮虫身上进行，但结果将建立一种新的范式，即牙周细菌的生理学，无论是单独的还是协同的，都可以扰乱硫醇分子的动态平衡，从而导致疾病病理。通过抑制细菌分解代谢途径从药理上重建正常的牙周袋代谢物水平可能是减少牙周炎宿主损害的一种创新策略。