牙龈卟啉单胞菌及其牙龈蛋白酶K对青少年牙龈健康的影响

目的：分析牙龈卟啉单胞菌（P. gingivalis）及牙龈蛋白酶K（Kgp）对青少年牙龈健康的影响。方法???收集12~17岁青少年牙龈正常组（50例）、牙龈炎症指数Ⅰ级组（25例）和牙龈炎症指数Ⅱ级组（32例）的3组龈沟液标本,应用16S?rDNA?PCR技术检测各样本中的P. gingivalis及Kgp。3组P. gingivalis和Kgp阳性检出率的比较采用χ2检验;P. gingivalis和Kgp的相对含量与牙龈炎症指数之间的关系采用Spearman相关分析。结果???P. gingivalis在牙龈炎症指数Ⅰ级组的检出率大于牙龈正常组,牙龈炎症指数Ⅱ级组的检出率大于牙龈炎症指数Ⅰ级组和牙龈正常组,牙龈炎症指数Ⅰ级组、Ⅱ级组与牙龈正常组间有明显差异（P<0.01）,牙龈炎症指数Ⅰ级组与Ⅱ级组间差异具有统计学意义（P<0.05）。Kgp在牙龈炎症指数Ⅰ级组的检出率大于牙龈正常组,牙龈炎症指数Ⅱ级组的检出率大于牙龈炎症指数Ⅰ级组和牙龈正常组,牙龈炎症指数Ⅰ级组与牙龈正常组间差异具有统计学意义（P<0.05）,牙龈炎症指数Ⅱ级组与牙龈正常组间有明显差异（P<0.01）。P. gingivalis和Kgp的检出率随着牙龈炎症指数的增加而升高。结论???P. gingivalis和Kgp与青少年牙龈健康密切相关。     

牙周炎症微环境对牙龈上皮屏障影响研究进展

牙周病是一种常见的慢性炎症性疾病，与牙周致病菌、黏膜和免疫宿主细胞之间复杂的相互作用有关。牙龈卟啉单胞菌（Porphyromonas gingivalis，P. gingivalis）在牙周病中被认为是重要的致病菌之一，与黏膜和免疫细胞相互作用，导致炎症细胞因子的产生。牙龈上皮是口腔微生物群的物理和免疫屏障，在牙周组织感染炎症后激活免疫反应中起着至关重要的作用。文章就牙龈上皮细胞在P. gingivalis刺激炎症细胞形成的炎症微环境中的生物学行为研究做一综述。     

牙龈卟啉单胞菌诱导的牙周微环境失调引发牙周炎的作用机制研究进展

牙周炎作为一种由多种微生物诱发的慢性炎症性疾病,其造成的牙周组织损伤常与宿主免疫反应所引发的炎症相关。研究发现,补体系统参与牙周炎的发生与发展,牙龈卟啉单胞菌（P.gingivalis）可以利用补体成分破坏宿主的免疫防御,造成牙周微环境失调,进而诱发牙周炎的发生。因此,探究P.gingivalis利用补体诱发牙周微环境失调的作用机制,可以为今后牙周治疗的防治提供新的途径。     

牙龈卟啉单胞菌感染对人主动脉内皮细胞炎症反应的影响研究

目的　研究牙龈卟啉单胞菌（Porphyromonas gingivalis，P.gingivalis）感染对人主动脉内皮细胞（human aortic endothelial cells，HAECs）表达炎症因子的影响并探讨其可能的分子机制。方法　在体外以感染复数（multiplicity of infection，MOI）分别为0、25∶1、100∶1和200：1的P.gingivalis刺激HAECs 24 h后，分别采用实时荧光定量PCR、酶联免疫吸附试验和免疫印迹试验检测单核细胞趋化因子-1（monocyte chemotactic protein 1，MCP-1）、白细胞介素-1β（interleukin 1β，IL-1β）、Toll样受体（Toll like receptor，TLR）-2和TLR-4的表达水平；在体外以MOI分别为0和200∶1的P.gingivalis刺激HAECs 30 min后，采用凝胶迁移实验检测核因子κB（nuclear factor κB，NF-κB）的活化水平。结果　P.gingivalis刺激组（MOI分别为25∶1、100∶1和200∶1）的HAECs中MCP-1、IL-1β、TLR-2和TLR-4的表达均显著高于未刺激组（MOI为0）（均P < 0.05），且提示这种上调作用呈P.gingivalis刺激剂量依赖性；P.gingivalis刺激组（MOI为200∶1）的HAECs中NF-κB活化水平显著高于未刺激组（MOI为0）（P < 0.05）。结论　P.gingivalis可能通过TLRs-NF-κB信号通路上调HAECs中炎症因子的表达，导致血管内皮功能紊乱，提示P. gingivalis可能促进动脉粥样硬化的发生和发展。     

他米巴罗汀对牙龈卟啉单胞菌抗原刺激RAW264.7细胞的影响

目的 探讨他米巴罗汀（Tamibarotene,Am80）对经牙龈卟啉单胞菌（Porphyromonas gingivalis,P.gingivalis）抗原刺激后的小鼠单核巨噬细胞RAW264.7的影响。方法 采用P.gingivalis抗原刺激RAW264.7细胞,分别用不同浓度的Am80进行干预;相差显微镜下观察各组细胞生长情况;MTS法检测不同浓度Am80对抗原刺激RAW264.7细胞的增殖情况;酶联免疫吸附法（ELISA）检测细胞培养上清中抗炎因子IL-10的含量。结果 10 nmol/L Am80能够显著抑制P.gingivalis抗原刺激RAW264.7细胞的增殖分化（P<0.001）;Am80（10 nmol/L）致抗原刺激的细胞IL-10的分泌量显著增加（P<0.05）;倒置相差显微镜下观察显示Am80抑制抗原刺激细胞的增殖分化。结论 Am80（10 nmol/L）时可显著促进P.gingivalis抗原诱导小鼠巨噬细胞分泌抗炎因子IL-10,并可能由此抑制P.gingivalis抗原诱导的炎症反应。     

新型抗菌物质对牙龈卟啉单胞菌抗菌作用的研究进展

龈卟啉单胞菌（Porphyromonas gingivalis,P. gingivalis）是一种重要的牙周致病菌,研究和开发P. gingivalis的抗菌物质是牙周治疗的重要内容。传统抗生素类药物可诱发细菌耐药现象及菌群失调;化学类抗菌药物常引起着色、味觉改变等副反应。克服传统抗菌物质副反应的新型抗菌物质越来越受到重视。近年来的研究发现植物提取物、脂类物质、人工合成抗菌肽、金属纳米粒子、群体感应抑制剂等新型抗菌物质对P. gingivalis具有一定的抗菌作用。这些新型抗菌物质可通过作用于细菌的细胞膜、生物膜、毒力因子、群体感应系统等发挥其抗菌作用。文章就各种新型抗菌物质对P. gingivalis的抗菌作用及相关机制做一综述。     

牙龈卟啉单胞菌与不良妊娠结局的相关研究进展

龈卟啉单胞菌(Porphyromonas gingivalis, P.gingivalis)是一种革兰阴性厌氧菌,是牙周炎尤其是慢性牙周炎病变区或活动部位最主要的病原菌。研究表明,牙周炎与多种不良妊娠结局(adverse pregnancy outcomes, APO)有关,这一过程涉及多种病理生理机制,而P.gingivalis作为关键牙周致病菌之一在其中扮演重要角色。目前我们对P.gingivalis与APO的关系及其作用机制的认识仍然十分有限。该文拟通过回顾近年来发现的P.gingivalis与APO之间相关的直接或间接的证据,对P.gingivalis与APO的联系和作用机制作一综述。     

两种fimA基因型牙龈卟啉单胞菌刺激下口腔上皮细胞ICAM-1的表达

目的比较2种fimA基因型牙龈卟啉单胞菌（Porphyromonas gingivalis,P．gingivalis）刺激下口腔上皮细胞ICAM-1的表达水平。方法实验组用P．gingivalis ATCC 33277（Ⅰ型菌毛组）,W83（Ⅳ型菌毛组）,47A-1（Ⅳ型菌毛组）分别与KB细胞（ATCC CCL17）共同孵育24h;对照组为未受P.gingivalis刺激的KB细胞。分别在1h、3h、6h和24h收集细胞,运用流式细胞仪检测KB细胞膜上ICAM-1的动态表达。结果P．gingivalis刺激细胞后3h、6h和24h,实验组ICAM-1的表达水平均高于对照组;2种fimA基因型P．gingivalis调节KB细胞表达ICAM-1的方式相似,Ⅳ型菌毛组的调节作用强于Ⅰ型菌毛组。结论P.gingivalis上调口腔上皮细胞表达ICAM-1的水平与其fimA基因型相关,提示P.gingivalis致病性与其fimA基因型相关。     

DHA对人牙龈成纤维细胞生物学活性及炎症因子表达的作用

目的 探索二十二碳六烯酸（docosahexaenoic acid,DHA）对人牙龈成纤维细胞（human gingival fibroblasts,HGFs）生物学活性及炎症因子表达的作用。方法 分别采用活死细胞染色法、荧光染色法、流式细胞术观察DHA对细胞活性、细胞形态、细胞周期的影响;DHA预处理HGFs后,利用牙龈卟啉单胞菌（Porphyromonas gingivalis,P.gingivalis）脂多糖（lipopolysaccharides,LPS）或热灭活P.gingivalis刺激细胞,定量PCR和ELISA观察白介素-6（interleukin-6,IL-6）、IL-8、IL-1β基因和蛋白表达的变化;通过定量PCR和Western blot检测DHA对血红素氧合酶-1（heme oxygenase-1,HO-1）基因和蛋白表达的作用。结果 200 μmol/L DHA可降低HGFs的细胞活性,但100 μmol/L DHA不影响细胞活性及形态,对细胞周期也无明显影响（P＞0.05）;DHA可抑制P.gingivalis LPS或热灭活P.gingivalis诱导HGFs表达的IL-6、IL-8、IL-1β mRNA以及IL-6、IL-8蛋白（P＜0.05）,并可显著上调HGFs表达的HO-1 mRNA及蛋白（P＜0.05）。结论 DHA可显著抑制HGFs的炎症因子表达,并且不影响其生物学活性,其抗炎作用可能与HO-1上调相关。     

Ⅰ、ⅣfimA型牙龈卟啉单胞菌对内皮细胞与平滑肌细胞共培养体系产生内皮素-1及一氧化氮的影响

［摘要］ 目的 研究不同fimA基因型牙龈卟啉单胞菌（Porphyromonas gingivalis,P.gingivalis）刺激人脐静脉血管内皮细胞（human umbilical vein endothelial cells,HUVECs）及人脐动脉血管平滑肌细胞（human umbilical artery smooth muscle cells,HUASMCs）共培养体系产生内皮素-1（endothelin-1,ET-1）和一氧化氮（NO）的水平。方法 用ⅠfimA型P.gingivalis（ATCC 33277）和ⅣfimA型P.gingivalis（W83）分别刺激HUVECs-HUASMCs共培养体系,于2、8、24、48 h时收集细胞培养上清液,酶联免疫反应检测ET-1的含量,硝酸还原酶法测定NO的含量。各组均设阴性对照组（纯培养基）及阳性对照组（1 μg/mL E.coli-LPS）。结果 HUVECs-HUASMCs共培养体系在Ⅰ、ⅣfimA型P.gingivalis刺激作用下产生ET-1和NO的量以及ET-1/NO水平,与阴性及阳性对照组比较存在差异。ⅠfimA型P.gingivalis刺激细胞共培养模型分泌ET-1和NO情况的总趋势与阴性对照组相似、而ⅣfimA型P.gingivalis的刺激作用总趋势则与阳性对照组相似,ⅣfimA型P.gingivalis较ⅠfimA型P.gingivalis刺激共培养细胞可分泌更多的ET-1、而NO量减少,ⅣfimA型P.gingivalis感染48 h的细胞共培养模型表现出明显的ET-1/NO的失衡。结论 不同fimA型P.gingivalis刺激共培养模型后产生ET-1及NO的情况及ET-1/NO的水平有明显差异,可能与其本身毒力相关,ⅣfimA型P.gingivalis比ⅠfimA型P.gingivalis更易引起内皮功能紊乱。     

Deficiency of iNOS contributes to Porphyromonas gingivalis-induced tissue damage

Periodontitis is a chronic inflammatory disease that results in extensive soft and hard tissue destruction of the periodontium. Porphyromonas gingivalis possesses an array of virulence factors and has been shown to induce expression of inducible nitric oxide synthase (iNOS) in inflammatory cells. The aim of this study was to investigate the effect of eliminating iNOS in a murine model of P. gingivalis infection. This was achieved by utilizing a P. gingivalis-induced skin abscess model, and an alveolar bone loss model employing an oral infection of P. gingivalis in iNOS knockout mice. The results indicated that iNOS knockout mice exhibit more extensive soft tissue damage and alveolar bone loss in response to P. gingivalis infection compared to wild-type mice. The local immune response to P. gingivalis in iNOS knockout mice was characterized by increased numbers of polymorphonuclear monocytes, while the systemic immune response was characterized by high levels of interleukin-12. The iNOS is required for an appropriate response to P. gingivalis infection.     

Porphyromonas gingivalis lipopolysaccharide displays functionally diverse interactions with the innate host defense system

Periodontitis is a bacterially induced chronic inflammatory disease and a major cause of tooth loss in the world. The tissue damage and alveolar bone resorption characteristic of the disease are believed to be due to a destructive innate host response to a pathogenic subgingival biofilm. Porphyromonas gingivalis, a Gram-negative bacterium, is a member of this mixed microbial community that has been designated an etiologic agent of periodontitis. The innate host response to lipopolysaccharide (LPS) obtained from P. gingivalis is unusual in that different studies have reported that it can be an agonist for Toll-like receptor (TLR) 2 as well as an antagonist or agonist for TLR4. In addition, human monocytes respond to this LPS by secreting a variety of different inflammatory mediators, while endothelial cells do not. We have examined highly purified preparations of P. gingivalis LPS and found that they activate both TLR2 combined with TLR1 and TLR4 in transiently transfected human embryonic kidney (HEK) 293 cells. We have further demonstrated that highly purified P. gingivalis LPS preparations contain at least 3 major different lipid A species. We speculate that P. gingivalis lipid A structural heterogeneity contributes to the unusual innate host response to this LPS and its ability to interact with different TLR molecules.     

The role of cytokines in a Porphyromonas gingivalis-induced murine abscess model

INTRODUCTION: Porphyromonas gingivalis is an important periodontopathic bacterium that is strongly associated with periodontal disease and is part of human dental plaque. Periodontal disease results from the interaction of the host with bacterial products, and T-cell-derived cytokines remain critical in the immunoregulation of periodontal disease. METHODS: The aim of this study was to examine the role of T helper type 1 [interleukin-12p40 (IL-12p40), interferon-gamma, tumour necrosis factor (TNF)] and type 2 (IL-4, IL-10) cytokines in the immune response to a subcutaneous challenge with P. gingivalis using a well-established murine abscess model, in genetically modified cytokine-specific knockout mice. RESULTS: IL-12p40(-/-) mice exhibited more advanced tissue destruction and a reduced inflammatory cell infiltrate after subcutaneous P. gingivalis challenge. Deficiency of IL-4 or IL-10 did not result in increased susceptibility to P. gingivalis-mediated tissue destruction. Furthermore, TNF deficiency appeared to reduce local tissue destruction. Interestingly, serum-specific antibodies suggested a strong T helper type 2 response. CONCLUSION: The results of our study indicate an important role for IL-12 in a primary P. gingivalis subcutaneous challenge.     

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?? Periodontitis is a polymicrobial community-induced chronic inflammatory disease?? and the destruction of the periodontium is associated with the inflammation of host immune response. Evidences from studies indicate that the complement system involves in the pathogenesis of periodontitis. P.gingivalis can manipulate the complement system to subvert host immunity and remodel a symbiotic microbiota into a dysbiotic state?? resulting in the develoment of periodontitis. As a result?? further understanding of the mechanism of P.gingivalis in manipulating the complement system in the pathogenesis of periodontitis may provide a new approach to the treatment of periodontitis.     

Tumor necrosis factor modulates fibroblast apoptosis, PMN recruitment, and osteoclast formation in response to P. gingivalis infection.

P. gingivalis is an important oral pathogen, which has been closely linked to periodontal disease as well as lesions of endodontic origin. Both infections are associated with a decrease in fibroblast numbers, formation of an inflammatory infiltrate, and bone resorption. The goal of this study was to investigate the role that the host response plays in the capacity of P. gingivalis to stimulate fibroblast apoptosis, PMN recruitment, and osteoclastogenesis. This was accomplished by the use of an in vivo calvarial model in mice with targeted deletion of TNF receptors p55 and p75 and matched wild-type mice. The results indicate that P. gingivalis induces fibroblast apoptosis in vivo and establish for the first time that this involves the stimulation of a host response. Moreover, bacteria-stimulated PMN recruitment and osteoclastogenesis were also dependent upon the host response. The results suggest that much of the damage caused by P. gingivalis infection, including fibroblast apoptosis, at least under some circumstances, results from stimulation of the host response rather than the direct effect of bacterial products. Furthermore, this may represent a more general mechanism by which bacterial challenge induces apoptosis of matrix-producing cells through the induction of TNF.     

P. gingivalis and E. coli lipopolysaccharides exhibit different systemic but similar local induction of inflammatory markers

BACKGROUND: Porphyromonas gingivalis is a Gram-negative bacterium that is an important etiologic agent of human adult periodontitis. The goal of the study was to test the hypothesis that two isoforms of P. gingivalis lipopolysaccharide (PgLPS), PgLPS(1435)(/1449) and PgLPS(1690), exhibit differences in their capacity to stimulate systemic versus local responses compared to Escherichia coli lipopolysaccharide (LPS). METHODS: LPS was inoculated into the scalp of mice, and the response was measured locally at the site of inoculation and systemically in the heart/aorta. Vascular cell adhesion molecule (VCAM)-1 was assessed at the protein level by enzyme-linked immunosorbent assay, and VCAM-1, E-selectin, and intercellular adhesion molecule (ICAM)-1 were assessed at the RNA level of the RNase protection assay. Serum tumor necrosis factor-alpha (TNF-alpha) levels were also measured. RESULTS: E. coli LPS and both isoforms of P. gingivalis LPS were relatively potent in stimulating the expression of inflammatory markers, with E. coli LPS being more potent. In contrast, when the systemic response was measured in the heart/aorta, E. coli LPS, but not P. gingivalis LPS, significantly induced inflammatory markers. At moderate to low doses (1 and 10 microg per injection), serum TNF-alpha levels were minimally induced by P. gingivalis LPS compared to E. coli LPS. CONCLUSIONS: Both forms of P. gingivalis LPS stimulated an inflammatory response when injected into connective tissue but were minimally stimulatory when a systemic response was measured. In contrast, E. coli LPS was a potent stimulus at the systemic and local levels.     

Effects of Porphyromonas gingivalis on the central nervous system: activation of glial cells and exacerbation of experimental autoimmune encephalomyelitis

BACKGROUND: Several studies have suggested that peripheral inflammation may be involved in the etiology of multiple sclerosis (MS), a demyelinating disease of the central nervous system (CNS). T-cells activated in the periphery enter the CNS, leading to demyelination and axonal loss. We hypothesized that peripheral infection by Porphyromonas gingivalis can affect pathological processes in the CNS and aggravate MS. METHODS: Glial cells derived from rat brains were cultured and stimulated with P. gingivalis or P. gingivalis lipopolysaccharide (LPS). Secretion of nitric oxide (NO) and prostaglandin E2 (PGE2) was determined. In addition, we examined the proliferation of lymphocytes harvested from P. gingivalis-immunized mice in response to stimulation by echephalitogenic proteins. The effect of peripheral inflammation induced by P. gingivalis on the clinical course of the disease was tested in experimental autoimmune encephalomyelitis (EAE), a mouse model used for the study of MS. RESULTS: P. gingivalis LPS and heat-killed bacteria induced secretion of the proinflammatory mediators NO and PGE2 by CNS glial cells. Lymphocytes derived from P. gingivalis-immunized mice proliferated in the presence of the echephalitogenic protein myelin basic protein. Injection of P. gingivalis into subcutaneous chambers in mice, followed by EAE induction led to aggravation of the disease. CONCLUSIONS: The present study provides evidence that infection with a periodontal pathogen, such as P. gingivalis, may play a role in the pathogenesis of CNS inflammatory disorders such as MS.