Tobacco-induced alterations to P. gingivalis-host interactions

描述(申请人提供)：牙周炎是一种破坏性的传染病，与多种致命性和衰弱的慢性系统性疾病有关，在美国每年消耗数十亿美元的医疗费用。吸烟者对牙周炎的易感性更高；更有可能感染牙周关键病原体牙龈卟啉单胞菌；与不吸烟者相比，这种细菌的数量更多。尽管如此，吸烟者的临床炎症反应一直在减少。我们发现，当香烟烟雾提取物(CSE)暴露于纯化的人单核细胞时，Pggivalis的促炎反应明显受到抑制，当细胞传回到没有CSE的新鲜培养液中时，Pggivalis的炎症诱导潜力恢复。然而，烟草对牙龈假单胞菌毒力决定因素编码基因调控的影响基本上是未知的。我们假设CSE代表一种环境压力，牙龈假单胞菌通过改变基因表达来适应这种压力。我们认为，这种适应性反应可能涉及应激相关基因的诱导、毒力决定因素和细菌表面成分的改变。我们还假设，这种香烟烟雾诱导的牙龈假单胞菌的生理变化将包括编码关键病原体识别决定因素的基因的活性改变，这将解释烟雾暴露的牙龈假单胞菌炎症反应减弱的原因。我们的目标是在原代人类单核细胞炎症模型中识别、分离和测试CSE改变的基因产物。通过转录组分析产生的初步数据和几个微生物决定因素的结构特征有力地支持了我们的假设。我们已经确定了几个在CSE暴露中调节失调的基因、基因产物和细菌结构，这些基因、基因产物和细菌结构可能在牙龈假单胞菌参与天然免疫系统的过程中发挥关键作用。这些包括菌毛蛋白、脂多糖、主要外膜抗原和其他已知具有炎症潜在性的牙龈假单胞菌成分，如DNAJ、GRPE和RAGA。这些研究代表了一个新的研究领域，将提供一些第一批信息，说明牙龈假单胞菌如何在分子水平上对香烟烟雾做出反应，并可能为吸烟者的疾病发病机制提供独特的见解。因此，这些研究将开辟新的领域，并为未来的研究提供基础，这些研究将阐明烟草如何影响病原体-宿主反应相互作用的机制。 公共卫生相关性：革兰氏阴性厌氧菌牙龈卟啉单胞菌是慢性牙周炎的病原体，慢性牙周炎是一种由烟草引起和/或加重的破坏性疾病。吸烟者比不吸烟者更容易感染牙龈假单胞菌，并且这些细菌细胞的数量更多。然而，吸烟者的临床炎症也有所减少，与此相一致的是，暴露在烟雾中的牙龈假单胞菌表现出从免疫细胞中非法产生炎症反应的能力降低。这种炎症潜能降低的机制是完全未知的。因此，我们将通过对培养在香烟烟雾提取物(CSE)条件培养液中的牙龈假单胞菌进行微阵列分析，找出受烟草暴露影响的牙龈假单胞菌基因。我们还将研究CSE对已知在激活内毒素和菌毛等先天免疫细胞中起关键作用的细胞结构特征的影响。烟草改变的细菌结构和CSE失调基因产物的炎症潜力将在新鲜分离的人类单核细胞中进行测试。这项研究将提供一些第一批信息，说明牙龈假单胞菌如何在分子水平上对香烟烟雾做出反应，并可能为吸烟者的疾病发病机制提供独特的见解。

DESCRIPTION (provided by applicant): Periodontitis is a destructive, infectious disease that has been associated with multiple fatal and debilitating chronic systemic diseases and consumes billions of dollars in health costs annually in the US. Tobacco smokers exhibit increased susceptibility to periodontitis; are more likely to be infected with the key periodontal pathogen Porphyromonas gingivalis; and to harbor higher numbers of this bacterial species relative to non-smokers. Despite this, smokers consistently exhibit reduced clinical inflammation. We show that the pro-inflammatory response of purified human monocytes is significantly suppressed when P. gingivalis is exposed to cigarette smoke extracts (CSE) and that the inflammation inducing potential of P. gingivalis is restored when cells are sub-cultured back into fresh medium without CSE. However, the influence of tobacco on the regulation of genes encoding P. gingivalis virulence determinants is essentially unknown. We hypothesize that CSE represents an environmental stress and that P. gingivalis responds by altering gene expression to adapt to this stress. We believe that this adaptive response may involve the induction of stress-related genes, virulence determinants and alterations of bacterial surface components. We also hypothesize that such cigarette smoke-induced physiological changes in P. gingivalis will include altered activities of genes encoding key pathogen recognition determinants, which will explain the reduced inflammatory response of smoke-exposed P. gingivalis. It is our aim to identify, isolate and test CSE-altered gene products in a primary human monocyte model of inflammation. Preliminary data generated through transcriptome analyses and the structural characterization of several microbial determinants are strongly supportive of our hypotheses. We have identified several genes, gene products, and bacterial structures that are dysregulated on CSE exposure that may play pivotal roles in the engagement of the innate immune system by P. gingivalis. These include fimbrial proteins, LPS, major outer membrane antigens, and other P. gingivalis components of known inflammatory potential, such as dnaJ, grpE and ragA. These studies represent a novel area of investigation and will provide some of the first information illustrating how P. gingivalis responds at the molecular level to cigarette smoke and may provide unique insights into disease pathogenesis in smokers. Thus, these studies will break new ground and provide the foundation for future studies that will elucidate mechanisms explaining how tobacco influences pathogen-host response interactions. PUBLIC HEALTH RELEVANCE: The Gram negative anaerobic bacterium, Porphyromonas gingivalis, is a causative agent of chronic periodontitis, a destructive tobacco-induced and/or exacerbated disease. Smokers are more likely to be infected by P. gingivalis and to harbor higher numbers of these bacterial cells than non-smokers. However, smokers also exhibit reduced clinical inflammation and, in keeping with this, smoke-exposed P. gingivalis exhibits a reduced capacity to illicit an inflammatory response from immune cells. The mechanisms underlying this reduced inflammatory potential are entirely unknown. Therefore, we will identify P. gingivalis genes that are dysregulated by tobacco exposure by microarray analysis of P. gingivalis cultured in cigarette smoke extract (CSE)-conditioned medium. We will also examine the effects of CSE on the characteristics of cellular structures known to play critical roles in the activation of innate immune cells, such as LPS and fimbriae. The inflammatory potential of tobacco-altered bacterial structures and the products of CSE-dysregulated genes will be tested in freshly isolated human monocytes. This study will provide some of the first information illustrating how P. gingivalis responds at the molecular level to cigarette smoke and may provide unique insights into disease pathogenesis in smokers.