Impact of Oral bacterial interactions on periodontitis

口腔细菌相互作用对牙周炎的影响

• 批准号: 6747953
• 负责人: ERIC C REYNOLDS
• 金额: $ 16.2万
• 依托单位: UNIVERSITY OF MELBOURNE
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-01 至 2005-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6747953
• 关键词: Bacteroides gingivalis; Fusobacterium nucleatum; Treponema; bacteria infection mechanism; bacterial antigens; bacterial proteins; bactericidal immunity; bone disorder; disease /disorder model; host organism interaction; infection; laboratory mouse; mandible /maxilla; mass spectrometry; membrane proteins; microorganism culture; oral bacteria; periodontitis; two dimensional gel electrophoresis; western blottings

DESCRIPTION (provided by applicant): Periodontal diseases are bacterial-associated inflammatory diseases of the supporting tissues of the teeth, and one of the more aggressive forms of chronic periodontitis is characterized by the destruction of the tooth's supporting structures. Chronic periodontitis has a large economic burden and is a major public health problem. Three bacterial species (Porphyromonas gingivalis, Treponema denticola, Bacteroides forsythus) have been shown to be closely associated with chronic periodontitis in humans, and a commensal oral bacterium Fusobacterium nucleatum has been implicated in aiding the colonization of the periodontopathic bacteria. P. gingivalis, T. denticola, B. forsythus and F. nucleatum exist as part of a high density mixed species (polymicrobial) biofilm in the gingival crevice of humans. Recent evidence using rodent abdominal lesion models has indicated that co-infection of B. forsythus or T. denticola with P. gingivalis enhances lesion development. This synergistic interaction that enhances lesion severity, and potentially virulence of the bacteria, has not been studied in an animal periodontitis model. Further, outer membrane proteins that are altered during this interaction also have not been studied. The broad objective of this application is to determine the impact of interactions between P. gingivalis, T. denticola, B. forsythus and F. nucleatum on the level of bone loss and host antibody response in the murine periodontitis model and to characterize the major antigenic outer membrane proteins of these bacteria using 2D PAGE and mass spectrometric methods. Using these techniques, we have already identified over thirty P. gingivalis outer membrane proteins. We have also established a murine periodontitis model based on oral infection of P. gingivalis. These methodologies will be applied to determine the impact of bacterial interactions on the expression of major antigenic outer membrane proteins and on colonization, bone loss and host immune response. The significance of this work is that it will increase our knowledge of the role of bacterial interaction on polymicrobial virulence in the murine periodontitis model and may have a broader significance in the study of human mucosal pathogens. The specific aims of the application are: 1) To determine the extent of alveolar bone loss and level of bacterial colonization in the murine periodontitis model when co-inocula of P. gingivalis, T. denticola, B. forsythus and/or F. nucleatum grown as monocultures are used to orally infect mice compared with mono-inoculation of each species. 2) To determine the extent of alveolar bone loss and bacterial colonization in the murine periodontitis model when polymicrobial cultures of P. gingivalis, T. denticola, B. forsythus and/or F. nucleatum are used to orally infect mice compared with mono- and co-inocula of the bacteria grown as monocultures as in (1) above. 3) To determine the host antibody isotype response to each of the bacteria in the murine periodontitis model when co-inocula or polymicrobial cultures of P. gingivalis, T. denticola, B. forsythus and/or F. nucleatum are used to orally infect mice. 4) To identify the major outer membrane and antigenic proteins of P. gingivalis, T. denticola, B. forsythus and F. nucleatum grown in monoculture and as polymicrobial cultures using 2D-gel electrophoresis, mass spectrometry and Western blotting.

描述（由申请人提供）：牙周病是牙齿支持组织的细菌相关炎症性疾病，慢性牙周炎的更具侵袭性形式之一的特征是牙齿支持结构的破坏。 慢性牙周炎造成巨大的经济负担，是重大的公共卫生问题。 三种细菌（牙龈卟啉单胞菌、齿垢密螺旋体、福赛斯拟杆菌）已被证明与人类慢性牙周炎密切相关，而口腔共生细菌具核梭杆菌则有助于牙周病细菌的定植。 P. gingivalis、T. denticola、B. forsythus 和 F. nucleatum 作为人类牙龈缝隙中高密度混合物种（多种微生物）生物膜的一部分而存在。 最近使用啮齿动物腹部病变模型的证据表明，福赛斯短杆菌或牙菌与牙龈卟啉单胞菌的共同感染会促进病变的发展。 这种增强病变严重程度和细菌潜在毒力的协同相互作用尚未在动物牙周炎模型中进行过研究。 此外，在这种相互作用过程中发生改变的外膜蛋白也尚未被研究。 本申请的主要目标是确定牙龈卟啉单胞菌、齿垢毛菌、连翘短杆菌和具核梭菌之间的相互作用对小鼠牙周炎模型中骨质流失水平和宿主抗体反应的影响，并表征主要抗原使用 2D PAGE 和质谱方法分析这些细菌的外膜蛋白。 利用这些技术，我们已经鉴定了三十多种牙龈卟啉单胞菌外膜蛋白。 我们还建立了基于牙龈卟啉单胞菌口腔感染的小鼠牙周炎模型。 这些方法将用于确定细菌相互作用对主要抗原外膜蛋白表达以及定植、骨质流失和宿主免疫反应的影响。 这项工作的意义在于，它将增加我们对小鼠牙周炎模型中细菌相互作用对多种微生物毒力作用的认识，并可能在人类粘膜病原体的研究中具有更广泛的意义。 本申请的具体目的是： 1) 确定小鼠牙周炎模型中同时接种 P. gingivalis、T. denticola、B. forsythus 和/或 F. nucleatum 时的牙槽骨丢失程度和细菌定植水平与每个物种的单一接种相比，单一培养物用于口服感染小鼠。 2) 与单菌和单菌相比，确定使用 P. gingivalis、T. denticola、B. forsythus 和/或 F. nucleatum 的多种微生物培养物口服感染小鼠时，小鼠牙周炎模型中牙槽骨丢失和细菌定植的程度。如上述（1）中那样作为单一培养物生长的细菌的共接种物。 3) 当使用牙龈卟啉单胞菌、牙菌、福赛斯芽孢杆菌和/或具核梭菌的共接种或多微生物培养物经口感染小鼠时，确定宿主抗体同种型对鼠牙周炎模型中每种细菌的反应。 4) 使用 2D 凝胶电泳、质谱和蛋白质印迹法鉴定单一培养物和多种微生物培养物中生长的 P. gingivalis、T. denticola、B. forsythus 和 F. nucleatum 的主要外膜和抗原蛋白。