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

• 批准号: 8842497
• 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
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8842497
• 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; Metabolism; 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; Relative (related person); Role; Site; Sulfhydryl Compounds; Testing; Time; Tissues; Tooth Exfoliation; Treponema denticola; alveolar bone; 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

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 ?ggt 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（由申请人提供）：在患病的牙周口袋中，谷胱甘肽水平低于健康地点，H2S的量较高。已经假定但尚未证明，这些硫醇化合物的浓度的变化对于牙周炎中看到的组织病理很重要。尽管谷胱甘肽和H2在患病的口袋中扰动的机制尚不清楚，但共识是细菌起着关键作用。因此，我们一直在研究牙周病原体从谷胱甘肽产生H2s的能力，因为这种代谢途径可以改变牙龈缝隙中这两个分子的水平。我们专注于主要的牙周病原体treponema牙本质，并以前表明它可以通过三个步骤酶促途径（GTSP）分解谷胱甘肽与H2S。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在牙周病理学中使用和产生的代谢产物的作用。为此，我们进行了几项初步实验，并具有以下相关结果：（1）牙齿齿状牙齿加谷胱甘肽可以在人类牙龈成纤维细胞和牙周韧带细胞中诱导凋亡。 （2）在体外，牙霉和谷胱甘肽增加了促炎细胞因子促炎细胞因子的牙周韧带合成。 （3）谷胱甘肽加剧了小鼠脓肿模型中牙霉菌引起的病变大小。 （4）牙齿牙杆菌可增强大鼠肺泡骨吸收，并降低牙龈裂纹液的谷胱甘肽水平。 （5）最重要的是，我们最近在GTSP的第一个基因（GGT）中构建了牙霉菌缺失突变体。该突变体不能将谷胱甘肽转化为H2S。因此，我们是独特的定位，以检验以下假设：细菌代谢，特别是牙霉菌GTSP的谷胱甘肽水平降低，而H2S产生的随之增加将在牙周炎中看到的宿主组织损害中起关键作用。在AIM 1中，我们将使用野生型T. denticola和我们的GGT突变体来证明GTSP，尤其是其H2S的产生，可增强牙周细胞中的凋亡，并调节体外宿主细胞产生的细胞因子的水平和模式。最重要的是，将牙霉菌突变体用于两个动物模型（AIM 2），以证明细菌降低谷胱甘肽和增加H2S的能力对于在体内引起组织/骨骼病理学至关重要。肺泡骨损失的大鼠模型也将用于测试我们已经在体外表征的三种GTSP抑制剂的能力，以限制牙霉菌发病机理。拟议的研究的结果将首次证明细菌在尺寸缝隙中影响谷胱甘肽和H2S水平的能力 是牙周组织损伤的重要贡献者。尽管仅使用牙霉菌进行实验，但结果将建立一个新的范式，从而单独或协同的牙周细菌的生理学可以扰动硫醇分子稳态会引起疾病病理学。通过抑制细菌分解代谢途径，在药理学上重新建立正常的牙周袋代谢物水平可能是减少牙周炎宿主损害的创新策略。