牙龈卟啉单胞菌菌毛fimA基因型在牙周牙髓联合病变中的分布

目的 分析牙龈卟啉单胞菌菌毛不同fimA基因型在牙周源性牙周牙髓联合病变患牙根管内的分布状况.方法 收集因牙周损害导致的牙周牙髓联合病变患牙43例,开髓揭髓顶后用纸尖采集根管内组织液,提取DNA.采用16SrRNA PCR检测牙龈卟啉单胞菌,并根据各fimA基因型的特异性引物,用聚合酶链反应检测不同fimA基因型菌株的分布.结果 16S rRNA PCR检测牙龈卟啉单胞菌阳性率76.7％.牙龈卟啉单胞菌fimA各基因型在牙龈卟啉单胞菌携带者的检出率分别为:Type Ⅰ fimA为24.2％;Type Ⅰ b fimA为27.2％;Type ⅡfimA为75.8％;Type ⅢfimA为9.1％;TypeⅣfimA为48.5％;Type ⅤfimA未检出;同时检测出Type Ⅰ和Ⅰb fimA为3.0％;Type Ⅱ和ⅣfimA为39.4％;Type Ⅰ、Ⅰb和ⅡfimA为21.2％.Type Ⅱ和Ⅳ fimA基因型的检出率显著高于其他基因型(P＜0.05).结论 牙周源性牙周牙髓联合病变根管内牙龈卟啉单胞菌存在fimA基因多态性.TypeⅡ和ⅣfimA基因型牙龈卟啉单胞菌菌株是牙周牙髓联合病变根管内的主要定植菌,可能与牙周牙髓联合病变的发生、发展相关密切.     

E‐selectin expression induced by Porphyromonas gingivalis in human endothelial cells via nucleotide‐binding oligomerization domain‐like receptors and Toll‐like receptors

Porphyromonas gingivalis, an important periodontal pathogen, has been proved to actively invade cells, induce endothelial cell activation, and promote development of atherosclerosis. Innate immune surveillance, which includes the activity of nucleotide‐binding oligomerization domain (NOD)‐like receptors (NLRs) and Toll‐like receptors (TLRs), are essential for the control of microbial infections; however, the roles of receptor families in P. gingivalis infections remain unclear. Here, we examined the roles of NLRs and TLRs in endothelial cell activation caused by P. gingivalis. Live P. gingivalis and whole cell sonicates were used to stimulate endothelial cells, and both showed upregulation of E‐selectin as well as NOD1, NOD2, and TLR2. In addition, silencing of these genes in endothelial cells infected with P. gingivalis led to a reduction in E‐selectin expression. Porphyromonas gingivalis also induced nuclear factor‐κB (NF‐κB) and P38 mitogen‐activated protein kinase (MAPK) activity in endothelial cells, whereas small interfering RNA targeting NOD1 significantly reduced these signals. Moreover, inhibition of either NOD2 or TLR2 inhibited NF‐κB significantly, but had only a weak inhibitory effect on P38 MAPK signaling. Direct inhibition of NF‐κB and P38 MAPK significantly attenuated E‐selectin expression induced by P. gingivalis in endothelial cells. Taken together, these findings suggest that NOD1, NOD2, and TLR2 play important, non‐redundant roles in endothelial cell activation following P. gingivalis infection.     

The GroEL protein of Porphyromonas gingivalis accelerates tumor growth by enhancing endothelial progenitor cell function and neovascularization

Porphyromonas gingivalis is a bacterial species that causes destruction of periodontal tissues. Additionally, previous evidence indicates that GroEL from P. gingivalis may possess biological activities involved in systemic inflammation, especially inflammation involved in the progression of periodontal diseases. The literature has established a relationship between periodontal disease and cancer. However, it is unclear whether P. gingivalis GroEL enhances tumor growth. Here, we investigated the effects of P. gingivalis GroEL on neovasculogenesis in C26 carcinoma cell‐carrying BALB/c mice and chick eggs in vivo as well as its effect on human endothelial progenitor cells (EPC) in vitro. We found that GroEL treatment accelerated tumor growth (tumor volume and weight) and increased the mortality rate in C26 cell‐carrying BALB/c mice. GroEL promoted neovasculogenesis in chicken embryonic allantois and increased the circulating EPC level in BALB/c mice. Furthermore, GroEL effectively stimulated EPC migration and tube formation and increased E‐selectin expression, which is mediated by eNOS production and p38 mitogen‐activated protein kinase activation. Additionally, GroEL may enhance resistance against paclitaxel‐induced cell cytotoxicity and senescence in EPC. In conclusion, P. gingivalis GroEL may act as a potent virulence factor, contributing to the neovasculogenesis of tumor cells and resulting in accelerated tumor growth.     

三种抛光套装对氧化锆全瓷冠抛光效果比较研究

目的　比较临床常用的3种抛光套装对氧化锆全瓷冠的抛光效果。方法　使用绿色砂石打磨大小相同的圆柱状氧化锆试件45块,随机均分为5组,分别命名如下。SHOFU组：采用Procelain Adjustment Kit PN0301抛光试件;EVE组：采用Rotary Grinding and Polishing Instruments HP321抛光试件;Kerr组：采用Occlubrush抛光刷抛光试件;上釉组：试件表面进行上釉处理;对照组：试件不做处理。测量各组试件表面粗糙度Ra值,在扫描电子显微镜（scanning electron microscope,SEM）下观察试件表面形貌。将牙龈卟啉单胞菌与上述试件混合培养,测定其表面细菌黏附量,并在SEM下观察细菌的黏附情况。结果　各组试件表面粗糙度Ra值及细菌黏附量总的比较,差异具有统计学意义（均P < 0.05）。SHOFU组、EVE组、Kerr组试件表面粗糙度Ra值和细菌黏附量均依次增大,且组间比较差异均有统计学意义（均P < 0.05）。SEM观察显示,SHOFU组、EVE组试件表面见浅细划痕;Kerr组、对照组试件表面见深划痕和密集的凹坑缺陷;上釉组试件表面光滑。细菌在SHOFU组、EVE组、Kerr组、对照组试件表面主要分布在划痕及缺陷周围,其中SHOFU组和EVE组细菌数量较少,Kerr组和对照组数量最多;细菌在上釉组试件表面散在分布,数量最少。结论　3种抛光套装中,Procelain Adjustment Kit PN0301的抛光效果最好。     

大黄素-8-O-β-D-吡喃葡萄糖苷对牙龈卟啉单胞菌唾液酸酶活性及其毒力基因表达影响研究

目的    探讨大黄素-8-O-β-D-吡喃葡萄糖苷对牙龈卟啉单胞菌（P. gingivalis）唾液酸酶活性及其毒力基因表达的影响。方法    使用不同质量浓度的大黄素-8-O-β-D-吡喃葡萄糖苷（0.2、0.5、2、5、10 mg/mL）处理P. gingivalis W83（实验组）,用未加药物的P. gingivalis W83作对照（对照组）,采用荧光法检测大黄素-8-O-β-D-吡喃葡萄糖苷对P. gingivalis唾液酸酶活性的作用。5 mg/mL大黄素-8-O-β-D-吡喃葡萄糖苷作用于P. gingivalis W83,Real-time PCR法检测毒力基因fimA、fimR、fimS、kgp、rgpA和rgpB的表达情况。结果    大黄素-8-O-β-D-吡喃葡萄糖苷对P. gingivalis唾液酸酶活性产生了抑制作用,当其质量浓度为0.2、0.5、2、5、10 mg/mL时,对唾液酸酶活性的抑制率分别为11.4%、32.23%、40.21%、73.54%、84.31%。与对照组比较,实验组（5 mg/mL大黄素-8-O-β-D-吡喃葡萄糖苷处理）的fimA、fimR、fimS、kgp、rgpA和rgpB基因表达均下降,差异均有统计学意义（均P < 0.05）。结论    大黄素-8-O-β-D-吡喃葡萄糖苷可有效抑制P. gingivalis唾液酸酶活性,其抑制作用会降低细菌毒力基因表达,有望成为预防及治疗牙周炎的新型药物。     

Association of Periodontal Diseases and Liver Fibrosis in Patients With HCV and/or HBV infection

Background:

Periodontal disease and systemic health are closely associated. However, there is no data supporting the association between periodontal disease and patients with liver diseases associated with hepatitis C virus (HCV) and/or hepatitis B virus (HBV) infection.

Objectives:

The aim of this study was to evaluate the association between periodontitis and progression of liver diseases in patients with HCV and/or HBV infection.

Patients and Methods:

In this retrospective study, 351 patients with HCV- and/or HBV-related liver diseases underwent screening for periodontal disease using the Salivaster® salivary occult blood test from February 2010 to June 2014. Furthermore, we examined the prevalence of fimbrillin (fimA) genotype of Porphyromonas gingivalis (P. gingivalis) in 28 HCV-infected patients visited at our hospital between January 2013 and June 2014. P. gingivalis with fimA genotype with types I to V was further detected using a PCR method.

Results:

Of 351 patients, 76 patients (group 1) had a strong positive result for salivary occult blood test and 275 patients (group 2) had weak positive or negative test results. Significant factors between the groups were obesity, level of AST, ALT, LDH, ALP, Alb, D.Bil, T.cho, AFP, platelets (Plt), IRI, HOMA-IR, current interferon (IFN) treatment and the daily frequency of tooth brushing. Between-groups analysis indicated that total protein (T.pro) level and liver fibrosis were significant factors. According to multivariate analysis, five factors were associated with periodontal disease as Plt count below 80000, brushing teeth only once a day, current IFN treatment, aged 65 years or older and obesity. The adjusted odds ratios for these five factors were 5.80, 3.46, 2.87, 2.50 and 2.33, respectively, and each was statistically significant. Twenty-eight saliva specimens had positive results for P. gingivalis with fimA genotype types I to V. The prevalence of fimA genotype II was higher in 14 patients with liver cirrhosis or a history of hepatocellular carcinoma treatment (group B, 50.00%) than 14 patients with only hepatitis C (group A, 21.43%).

Conclusions:

Periodontitis might be associated with progression of viral liver disease; hence, controlling oral disease is essential for the prevention and management of liver fibrosis.     

牙龈卟啉单胞菌PG0839基因突变株的构建

目的构建牙龈卟啉单胞菌毒力岛基因PG0839突变菌株,为研究PG0839基因功能提供实验基础。方法扩增1 584 bp PG0839基因片段,对聚合酶链反应（PCR）产物和pUC19载体进行BamH Ⅰ和EcoRⅠ双酶切,连接酶切产物得到质粒pPG0839-1。将2 101 bp erm基因产物插入到pPG0839-1中PG0839基因的EcoRⅤ位点,构建质粒pPG0839-2,作为电穿孔的供体质粒。电穿孔转化于受体菌牙龈卟啉单胞菌W83菌株,红霉素抗性培养基筛选阳性克隆,命名为PG0839基因突变菌株。结果运用插入失活方法构建PG0839基因突变菌株,进而通过酶切、测序、PR和反转录PCR对PG0839基因突变菌株进行验证,证实PG0839基因突变菌株构建成功。结论本实验成功构建PG0839基因突变菌株。     

牙龈卟啉单胞菌对兔血管平滑肌细胞分泌IL-1β、IL-6的影响

目的:观察牙龈卟啉单胞菌(Porphyromonas gingivalis,Pg)上清液对体外培养的兔血管平滑肌细胞分泌IL-1β、IL-6的影响。方法:组织块法培养兔腹主动脉血管平滑肌细胞,并对其细胞来源进行鉴定。用4.3×106 CFU/mL的Pg上清液刺激细胞12、24、48 h后,通过ELISA检测IL-1β、IL-6的水平;同时用RT-PCR检测其mRNA表达的情况。结果:Pg上清液刺激24 h和48 h时能促进血管平滑肌细胞分泌IL-6,分别与同一时间点的对照组相比差异均有统计学意义(P<0.05),而且24 h时,IL-6表达最强,而IL-1β的表达最低,明显低于相同时间的对照组和其他时间点的实验组(P<0.05)。RT-PCR检测显示,4.3×106 CFU/mL的Pg上清刺激血管平滑细胞12、24、48 h后细胞内均有IL-1β、IL-6的基因表达,在24 h时,血管平滑肌细胞的IL-1β基因表达减少,而IL-6基因表达增加。结论:Pg上清液可促进细胞合成和分泌IL-6,在动脉粥样硬化的发生发展过程中可能发挥一定作用。     

牙龈卟啉单胞菌临床菌株的毒力分析

目的:利用啮齿动物模型,比较分析中国人群口腔中牙龈卟啉单胞菌(Porphyromonas gingivalis,P.gingivalis)临床菌株的毒力。方法:采用小鼠脓肿形成实验,将6株从牙周炎患者牙周袋中分离获得的P.gingivalis临床菌株L2、L3、L4、L5、L11和L12分别注射至小鼠后背中央,观察其临床症状,并与国际标准菌株ATCC33277和W83进行比较,分析临床菌株的毒力特性。采用SPSS11.5软件包对数据进行统计学分析。结果:L3、ATCC33277和W83诱发小鼠产生原发性脓肿和轻微的全身反应,L2、L5、L11、L12则诱发小鼠产生远处转移性病变和严重的全身反应。结论:中国牙周炎患者口腔中的P.gingivalis临床菌株与国际标准菌株相比,在诱发实验鼠发生局部脓肿方面,具有一定程度的相似性,但临床菌株间的毒力特征具有较大差异。     

Effect of Streptococcus sanguinis/Porphyromonas gingivalis single and combined biofilms upon platelet aggregation

Oral Diseases (2012) 18, 586–594 Objective: To assess the effect of two oral bacteria Streptococcus sanguinis and Porphyromonas gingivalis upon platelet aggregation. Materials and Methods: Streptococcus sanguinis, P. gingivalis, S. sanguniis + P. gingivalis were added to platelet‐rich plasma and platelet aggregation measured using a platelet aggregometer. Platelets were passed through a flow chamber with S. sanguinis, P. gingivalis or a biofilm of S. sanguinis and P. gingivalis coated with saliva. Platelet adhesion to the chamber was observed under a fluorescence microscope for 15 min. The positive control was platelets treated with adrenaline; the negative control was platelets treated with phosphate‐buffered saline. Results: The mean (± s.e.) aggregation magnitude of S. sanguinis and P. gingivalis was 77.7 ± 7.4% and 79.3 ± 9.9%, respectively. The aggregation magnitude of S. sanguinis + P. gingivalis was 51.3 ± 12.9%, which was significantly lower than that for S. sanguinis/P. gingivalis (P < 0.05). In the flow chamber system, platelets adhered to S. sanguinis/P.gingivalis respectively within 3 min, and reached a plateau at 5–15 min. Under the condition of the S. sanguinis‐ and P. gingivalis‐saliva biofilm, platelet adhesion to the biofilm was significantly reduced at 5–15 min (P < 0.05). Conclusions: In the static or dynamic flow system, platelets adhered to S. sanguinis or P. gingivalis. However, if S. sanguinis was mixed with P. gingivalis, the aggregation magnitude (%) was significantly reduced.