Fusobacterium nucleatum mediates endothelial damage and increased permeability following single species and polymicrobial infection.

BACKGROUND: Numerous lines of evidence link periodontal pathobionts and their virulence factors with endothelial damage. Most research has been conducted using single species infections at the exclusion of other periodontal microorganisms that have been identified in vascular tissue. Here, we assessed endothelial infection with either single or mixed periodontal species infection and examined their effect on endothelial damage and permeability. METHODS: Cell surface abundance of platelet endothelial cell adhesion molecule-1 (PECAM-1) or endothelial permeability following infection with Porphyromonas gingivalis, Fusobacterium nucleatum subspecies (ssp) nucleatum, ssp polymorphum or Tannerella forsythia as single or mixed species infection was determined by flow cytometry and a fluorescent dextran permeability assay. Zebrafish embryos were infected systemically with either single or mixed species with mortality and disease measured over time. RESULTS: F. nucleatum ssp nucleatum, ssp polymorphum and P. gingivalis significantly reduced PECAM-1 abundance in single species infection, whereas T. forsythia had no effect. F. nucleatum ssp polymorphum caused considerable mortality and morbidity in a zebrafish systemic infection model. Polymicrobial infection underscored the virulence of F. nucleatum ssp polymorphum in particular with increased endothelial cell death and reduced PECAM-1 abundance in co-infection studies with this organism. When injected systemically into zebrafish in polymicrobial infection, fluorescently labeled bacteria were distributed throughout the vasculature and cardiac region where, in some instances, they co-localized with each other. CONCLUSIONS: These data provide further evidence on the effects of F. nucleatum on endothelium adhesion molecule abundance and permeability while also highlighting the importance of performing polymicrobial infection to study the molecular mechanisms associated with periodontal pathogen-induced vascular damage.

Mechanisms of vascular damage by systemic dissemination of the oral pathogen Porphyromonas gingivalis.

Several studies have shown a clear association between periodontal disease and increased risk of cardiovascular disease. Porphyromonas gingivalis (Pg), a key oral pathogen, and its cell surface-expressed gingipains, induce oedema in a zebrafish larvae infection model although the mechanism of these vascular effects is unknown. Here, we aimed to determine whether Pg-induced vascular damage is mediated by gingipains. In vitro, human endothelial cells from different vascular beds were invaded by wild-type (W83) but not gingipain-deficient (ΔK/R-ab) Pg. W83 infection resulted in increased endothelial permeability as well as decreased cell surface abundance of endothelial adhesion molecules PECAM-1 and VE-cadherin compared to infection with ΔK/R-ab. In agreement, when transgenic zebrafish larvae expressing fluorescently labelled PECAM-1 or VE-cadherin were systemically infected with W83 or ΔK/R-ab, a significant reduction in adhesion molecule fluorescence was observed specifically in endothelium proximal to W83 bacteria through a gingipain-dependent mechanism. Furthermore, this was associated with increased vascular permeability in vivo when assessed by dextran leakage microangiography. These data are the first to show that Pg directly mediates vascular damage in vivo by degrading PECAM-1 and VE-cadherin. Our data provide a molecular mechanism by which Pg might contribute to cardiovascular disease.

The Relationship of Surface Characteristics and Antimicrobial Performance of Pulp Capping Materials.

INTRODUCTION: Pulp capping materials need to be able to protect the pulp but also bond to the overlying restorative materials. Light-curable pulp capping materials bond better to restorative materials and are easier to place than most water-based cements. The aim of this study was to characterize new light-curable tricalcium silicate-based pulp capping materials and compare their surface and antimicrobial properties with clinically available Theracal (Bisco, Schaumburg, IL) and Biodentine (Septodont, Saint-Maur-des-Fossés, France). METHODS: The surface characteristics of 3 light-curable pulp capping materials based on a resin and filled with tricalcium silicate and tantalum oxide radiopacifier and Theracal and Biodentine were assessed by scanning electron microscopy, X-ray diffraction, and contact angle measurement. The radiopacity was measured following ISO 6876 standards. The antimicrobial activity was determined by the direct contact test and the antibiofilm activity by the adenosine triphosphate assay and the confocal laser scanning Live/Dead assay (Invitrogen, Eugene, OR) using a polymicrobial culture. RESULTS: The surface characteristics of the materials varied with the unfilled resin and Biodentine exhibiting a hydrophobic surface. Biodentine showed significantly higher antimicrobial properties in the direct contact test, but this property was absent in the antibiofilm activity tests. The resins filled with tricalcium silicate and Theracal showed higher antimicrobial activity than Biodentine in the adenosine triphosphate and live/dead assays. CONCLUSIONS: The surface characteristics of a material affect its antimicrobial properties. The experimental resin-modified materials exhibited comparable antimicrobial properties with other light-curable pulp capping agents. Further long-term studies on the materials' antimicrobial activity are required to assess whether they can result in better clinical outcomes.

Investigation of Acrylic Resin Disinfection Using Chemicals and Ultrasound.

PURPOSE: Dental prosthetic and orthodontic appliances are transported from the clinic to the laboratory for additions and repairs. These appliances, containing microbes from the oral flora, are a high risk for cross-contamination. The aim of this study was to evaluate the effect of chemical and ultrasound disinfection against two in vitro biofilms and an in vivo formed biofilm grown on unprepared and polished polymethyl methacrylate (PMMA) surfaces. MATERIALS AND METHODS: Rough and polished self-curing PMMA surfaces were infected with strains of both Candida albicans and Streptococcus oralis. After incubation, the samples were treated with different disinfection methods, including ultrasound treatment for both 15 and 30 seconds, and immersion in glutaraldehyde and alcohol-based chemical disinfectants (MD520 and Minuten, respectively). The disinfecting efficacy was assessed by colony forming units (CFU) analysis and by scanning electron microscopy (SEM). Furthermore the adequacy of bacterial elimination of application of 30-second ultrasound and MD520 was assessed on PMMA retrieved from ten volunteers by CFU analyses. ANOVA with p = 0.05 followed by the Tukey post hoc test and the Student t-test was used to analyze the data. RESULTS: The ultrasound treatment for 30 seconds, MD520, and Minuten were the most effective disinfectant methods as they reduced the microbial counts compared to the control (p < 0.05) as shown in the in vitro analyses. S. oralis adhered more to rough acrylic resin surfaces (p < 0.05). Ultrasound treatment was the most effective way to reduce microbial counts on PMMA exposed to oral flora (p = 0.043). CONCLUSION: Ultrasound treatment for 30 seconds was effective against C. albicans, S. oralis, and the oral flora as shown by testing microbial growth on agar plates and SEM.

Antimicrobial and biological activity of leachate from light curable pulp capping materials.

OBJECTIVES: Characterization of a number of pulp capping materials and assessment of the leachate for elemental composition, antimicrobial activity and cell proliferation and expression. METHODOLOGY: Three experimental light curable pulp-capping materials, Theracal and Biodentine were characterized by scanning electron microscopy, energy dispersive spectroscopy and X-ray diffraction. The elemental composition of the leachate formed after 24h was assessed by inductively coupled plasma (ICP). The antimicrobial activity of the leachate was determined by the minimum inhibitory concentration (MIC) against multispecies suspensions of Streptococcus mutans ATCC 25175, Streptococcus gordonii ATCC 33478 and Streptococcus sobrinus ATCC 33399. Cell proliferation and cell metabolic function over the material leachate was assessed by an indirect contact test using 3-(4,5 dimethylthiazolyl-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: The hydration behavior of the test materials varied with Biodentine being the most reactive and releasing the highest amount of calcium ions in solution. All materials tested except the unfilled resin exhibited depletion of phosphate ions from the solution indicating interaction of the materials with the media. Regardless the different material characteristics, there was a similar antimicrobial activity and cellular activity. All the materials exhibited no antimicrobial activity and were initially cytotoxic with cell metabolic function improving after 3days. CONCLUSIONS: The development of light curable tricalcium silicate-based pulp capping materials is important to improve the bonding to the final resin restoration. Testing of both antimicrobial activity and biological behavior is critical for material development. The experimental light curable materials exhibited promising biological properties but require further development to enhance the antimicrobial characteristics.

Clinical relevance of antimicrobial testing results for dental restorative materials.

BACKGROUND: The antimicrobial activity of restorative materials is clinically relevant because all dental materials are subject to an environment containing bacteria. This study aimed to investigate the use of 2 methodologies referred to in the literature to assess antimicrobial properties of restorative materials and investigate whether material properties alter results of these traditional methodologies. METHODS: A number of dental restorative materials - namely, Chemfil Superior®, Spectrum®, Heliobond®, Ionoseal®, Dyract Extra®, Smart Dentin Replacement (SDR®) and Biodentine® - were characterized by scanning electron microscopy (SEM), energy-dispersive spectroscopy (EDS), X-ray diffraction (XRD) analysis, Fourier transform infrared (FT-IR) spectroscopy and pH analysis. Antimicrobial activity was assessed using agar diffusion and biofilm accumulation tests. Key factors affecting results were assessed using analysis of covariance. RESULTS: Biodentine after immediate mixing and Ionoseal aged for 6 weeks resulted in an inhibition zone, while significantly higher McFarland readings were observed in the presence of barium when using materials Ionoseal, Dyract and SDR at 24-hour aging. Through analysis of covariance it was shown that material properties affected methodology results. CONCLUSIONS: Properties of materials affect results of antimicrobial testing, but this may not directly reflect the antimicrobial potential of the material in question. Careful choice of methodology and interpretation of results is important.

Effectiveness of Disinfectants on Antimicrobial and Physical Properties of Dental Impression Materials.

PURPOSE: The aim of this study was to assess the antimicrobial activity of chemical disinfectants on alginate and silicone impression materials. The effect of chemical disinfectants on the dimensional stability of the impression materials was also assessed. MATERIALS AND METHODS: For the microbiologic assessment, impressions of the maxillary arch were taken from 14 participants, 7 using alginate and 7 using an addition silicone. The impressions were divided into three sections. Each section was subjected to spraying with MD 520 or Minuten or no disinfection (control), respectively. Antimicrobial action of the chemical disinfectants was assessed by measuring microbial counts in trypticase soy agar (TSA) media and expressing the results in colony-forming units/cm2. The surface area of the dental impressions was calculated by scanning a stone cast using computer-aided design/computer-assisted manufacture and analyzing the data using a custom computer program. The dimensional stability of the impression materials after immersion in disinfectants was assessed by measuring the linear displacement of horizontally restrained materials using a traveling microscope. The percent change in length over 3 hours was thus determined. RESULTS: Alginate exhibited a higher microbial count than silicone. MD 520 eliminated all microbes as opposed to Minuten. The bacterial growth after Minuten disinfection was almost twice as much for alginate than for addition silicone impressions. The chemical disinfectants affected the alginate dimensional stability. Minuten reduced the shrinkage sustained by alginate during the first hour of storage. CONCLUSIONS: Alginate harbors three times more microorganisms than silicone impression material. Chemical disinfection by glutaraldehyde-based disinfectant was effective in eliminating all microbial forms for both alginate and silicone without modifying the dimensional stability. Alcohol-based disinfectants, however, reduced the alginate shrinkage during the first 90 minutes of setting. The current studies also propose another method to report the surface area based on accurate estimation by 3D image analysis.

Effect of sterilization techniques prior to antimicrobial testing on physical properties of dental restorative materials.

OBJECTIVES: The aim of this study was to investigate any changes to the microstructure and surface properties of selected dental materials after sterilization carried out prior to subjecting them to antimicrobial testing. Initial microbial contamination on the material, as well as other possible sources of contamination were also assessed. METHODS: The materials investigated included dentine replacement materials Chemfil Superior(®), Ionoseal(®), Dyract Extra(®) and SDR(®). The materials were characterized by scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS). The test materials were sterilized using alcohol, steam, ultraviolet light (UV) and ethylene oxide and any changes to these materials were then assessed by SEM, microhardness testing and Fourier transform infrared (FT-IR) spectroscopy. Material microbial levels before treatments were assessed by plate counting technique and turbidity tests. Possible contamination through dispensers was assessed by analysing the CFU/sample. RESULTS: Ethylene oxide affected the microstructure of the Chemfil, Ionoseal and Dyract, resulting in flattening of the SiO stretching vibrations and deposition of chlorine and calcium respectively in Chemfil and Dyract. Varied contamination was demonstrated on all materials when incubated in anaerobic conditions. CONCLUSIONS: The different sterilization techniques affected the microstructure of the materials under investigation. Samples of materials produced in sterile conditions could also be contaminated with bacteria, either from the material itself or through the dispensing apparatus. CLINICAL SIGNIFICANCE: Results of antimicrobial studies cannot be extrapolated clinically as the material sterilization treatment results in changes to material chemistry and microstructure, which could in turn affect the materials' antimicrobial activity.

Antimicrobial properties of conventional restorative filling materials and advances in antimicrobial properties of composite resins and glass ionomer cements-A literature review.

OBJECTIVES: It has been reported that complete caries removal from cavities during restoration of teeth is difficult. Furthermore with the tissue saving approach it is expected that more of the saved affected tissue will possibly harbor more residual bacteria. Antimicrobial restorative filling materials would be ideal to prevent the spread of caries after completion of tooth restoration, thus preventing recurrent decay and eventually restoration failure. This paper reviews the literature on the antimicrobial properties of dental restorative filling materials. METHODS: Pubmed searches on the antibacterial properties of restorative materials were carried out. Keywords were chosen to assess antibacterial properties of conventional filling materials. Methods of introducing antimicrobial agents in restorative materials were also reviewed together with the methodology used to assess antimicrobial activity. RESULTS: 174 articles from 1983 till 2014 were included. SIGNIFICANCE: Adhesive materials have decreased antimicrobial activity when compared to amalgams and zinc oxides. Several techniques have been employed in order to increase the antimicrobial activity of restorative materials. Although antimicrobial activity of restorative materials is important, the introduction of antimicrobial agents/techniques should not be at the expense of other material properties. Environmental changes within a material may affect the bacterial response to the antimicrobial. Bacterial adhesion to the restorative materials should be assessed. Long term assessment of antimicrobial activity is important and is clinically relevant. The use of antimicrobial dental materials is important unless such characteristics are gained to the detriment of other material properties.