Dan-Krister Rechenberg

Potential systematic error in laboratory experiments on microbial leakage through filled root canals: review of published articles

AIM To systematically evaluate whether published studies on microbial leakage through filled root canals in human teeth embedded in a two-chamber system were properly controlled. Specifically, the control for the assumption that leakage should occur through the root canal rather than other routes was investigated. METHODOLOGY A systematic search was conducted using Medline, Biosis, Cochrane, Embase, and Web of Science databases. In addition, the reference lists of review articles pertaining to the topic were searched. No language restriction was applied. Two independent reviewers screened titles and abstracts. All articles deemed appropriate by either reviewer were included in the full-text evaluation. In case of disagreement, a referee arbitrated between the reviewers. RESULTS With 93.8% agreement prior to discussion and arbitration, 67 articles were included. On average, the size of the negative control group was 30% (mean) of the n in the experimental groups (minimum=0.0%, maximum=100%, SD=27%). The majority of studies (57 of 67) used inadequate negative controls. The whole root was covered with the sealing material in these specimens, whilst the root tip was left uncovered in the experimental groups. Consequently, leakage between outer root surface and sealing material was not controlled for. The authors of the remaining 10 communications did not state clearly how negative control assessments were performed. CONCLUSIONS Experimental investigations should be performed to assess the routes of microbial leakage in two-chamber models.

Reduction of Hard-tissue Debris Accumulation during Rotary Root Canal Instrumentation by Etidronic Acid in a Sodium Hypochlorite Irrigant

INTRODUCTION Hard-tissue debris is accumulated during rotary instrumentation. This study investigated to what extent a calcium-complexing agent that has good short-term compatibility with sodium hypochlorite (NaOCl) could reduce debris accumulation when applied in an all-in-one irrigant during root canal instrumentation. METHODS Sixty extracted mandibular molars with isthmuses in the mesial root canal system were selected based on prescans using a micro-computed tomography system. Thirty teeth each were randomly assigned to be instrumented with a rotary system and irrigated with either 2.5% NaOCl or 2.5% NaOCl containing 9% (wt/vol) etidronic acid (HEBP). Using a side-vented irrigating tip, 2 mL of irrigant was applied by 1 blinded investigator to the mesial canals after each instrument. Five milliliters of irrigant was applied per canal as the final rinse. Mesial root canal systems were scanned at high resolution before and after treatment, and accumulated hard-tissue debris was calculated as vol% of the original canal anatomy. Values between groups were compared using the Students t test (α < .05). RESULTS Irrigation with 2.5% NaOCl resulted in 5.5 ± 3.6 vol% accumulated hard-tissue debris compared with 3.8 ± 1.8 vol% when HEBP was contained in the irrigant (P < .05). CONCLUSIONS A hypochlorite-compatible chelator can reduce but not completely prevent hard-tissue debris accumulation during rotary root canal instrumentation.

The receptor activator of NF-κB ligand-osteoprotegerin system in pulpal and periapical disease

AIM To summarize the collective in vitro, in vivo and clinical evidence of the involvement of the receptor activator of NF-κB ligand (RANKL)-osteoprotegerin (OPG) system, a system of two molecules controlling osteoclast differentiation and hard-tissue resorption, in pulpal and periapical pathophysiology. METHODOLOGY A systematic search related to RANKL and/or OPG and pulp or periapical disease was conducted on Medline, Biosis, Cochrane, Embase and Web of Science databases using keywords and controlled vocabulary. No language restriction was applied. Two independent reviewers first screened titles and abstracts and then the full texts that were initially included. The reference lists of the identified publications were examined for additional titles. RESULTS A total of 33 papers were identified. In vitro studies (N = 11) revealed that pulpal cells can be stimulated by various inflammatory agents to produce RANKL, whilst many studies did not consider the RANKL/OPG ratio. Animal studies (N = 9) mostly focused on the time course and development of periapical lesions in relation to the RANKL-OPG system. Levels of RANKL and OPG in the necrotizing pulp were not investigated. Human studies (N = 13) showed a steady-state expression of OPG in the odontoblast layer. Conflicting results have been reported regarding the role of RANKL in active apical periodontitis, again because the correlation of this molecule with its inhibitor (OPG) was often disregarded. CONCLUSIONS There is relatively little information currently available that would highlight the specific role of RANKL and OPG in pulpal and periapical disease. OPG may play a protective role against internal resorption, whilst an increased periapical RANKL/OPG ratio might indicate bone resorption.

Identification of Synergistetes in endodontic infections.

The bacterial phylum Synergistetes consists of Gram-negative anaerobes. Oral Synergistetes are divided in two main clusters, namely A and B. Increasing evidence demonstrates their involvement in etiology of oral infections, including apical periodontitis. This condition causes bone loss around the apex of the tooth, subsequent to pulp inflammation (pulpitis). Although the presence of Synergistetes has been confirmed in endodontic infections by molecular methods, these have not been morphologically identified in the affected apical region, and their prevalence among different endodontic infections has not been determined. Therefore, the aim of this study was to evaluate the prevalence, levels and morphology of oral Synergistetes clusters A and B, in apical root canal samples obtained of teeth with irreversible pulpitis, pulp necrosis and apical periodontitis, or previously root-filled teeth with apical periodontitis. For their detection, fluorescence in situ hybridization and epifluorescence microscopy were used. Synergistetes cluster A was not detected in pulpitis, but was found in both apical periodontitis groups, more frequently and at higher ranges in teeth which were previously root-filled. Microscopically, they appeared as straight or slightly curved long rods. Synergistetes cluster B was not detected in any of the cases. Fusobacteria and Actinomyces, which are well-established taxa in endodontic infections, were detected more frequently and at higher ranges than Synergistetes. In conclusion, Synergistetes cluster A constitutes part of the mixed apical microbiota in apical periodontitis, and may be involved in its pathogenesis.

Influence of different curing approaches on marginal adaptation of ceramic inlays.

PURPOSE To evaluate the effect of different curing protocols on marginal adaptation of ceramic inlays after thermomechanical loading (TML). MATERIALS AND METHODS Forty-eight human molars were randomly divided into 6 groups (n = 8). After Class II cavity preparation (mod), ceramic inlays (Cerec) were fabricated. In groups I to IV, the cavities were conditioned with XP Bond mixed with Self Cure Activator (SCA) and the inlays were placed with the luting composite (LC) Calibra Mix (dual curing). The teeth in groups V and VI were conditioned with XP Bond without SCA and the inlays were placed with Calibra base (only light curing). In groups III, IV and V the adhesive was separately light cured prior to, and in groups II, IV, V and VI, after the inlay insertion. Before and after TML, marginal adaptation was measured using scanning electron microscopy (200X). Continuous margins (% of the total) were compared between groups using analysis of variance (ANOVA). A Bonferroni correction was applied to correct for multiple testing (alpha < 0.005). RESULTS Light curing after inlay insertion improved marginal adaptation on the occlusal interface between LC and enamel significantly, regardless of the LCs curing mode. Separate light polymerization of XP Bond did not result in superior marginal quality. Investigation of the interface between LC and proximal dentin margins showed improved adaptation by dual curing the adhesive and LC, irrespective of light application. CONCLUSION Light curing after inlay insertion showed improved marginal adaptation. Using dual-curing adhesive and LC, advantages in marginal adaptation between LC and dentin were observed.

Biological Markers for Pulpal Inflammation: A Systematic Review.

Background and Objective Pulpitis is mainly caused by an opportunistic infection of the pulp space with commensal oral microorganisms. Depending on the state of inflammation, different treatment regimes are currently advocated. Predictable vital pulp therapy depends on accurate determination of the pulpal status that will allow repair to occur. The role of several players of the host response in pulpitis is well documented: cytokines, proteases, inflammatory mediators, growth factors, antimicrobial peptides and others contribute to pulpal defense mechanisms; these factors may serve as biomarkers that indicate the status of the pulp. Therefore, the aim of this systematic review was to evaluate the presence of biomarkers in pulpitis. Methods The electronic databases of MEDLINE, EMBASE, Scopus and other sources were searched for English and non-English articles published through February 2015. Two independent reviewers extracted information regarding study design, tissue or analyte used, outcome measures, results and conclusions for each article. The quality of the included studies was assessed using a modification of the Newcastle-Ottawa-Scale. Results and Conclusions From the initial 847 publications evaluated, a total of 57 articles were included in this review. In general, irreversible pulpitis was associated with different expression of various biomarkers compared to normal controls. These biomarkers were significantly expressed not only in pulp tissue, but also in gingival crevicular fluid that can be collected non-invasively, and in dentin fluid that can be analyzed without extirpating the entire pulpal tissue. Such data may then be used to accurately differentiate diseased from healthy pulp tissue. The interplay of pulpal biomarkers and their potential use for a more accurate and biologically based diagnostic tool in endodontics is envisaged.

Available chlorine consumption from NaOCl solutions passively placed in instrumented human root canals

AIM To monitor chlorine consumption from nonagitated aqueous sodium hypochlorite (NaOCl) solutions in human root canals using a recently developed assay, which can determine the order of magnitude of available chlorine in small volumes of liquid. METHODOLOGY The root canals of 80 extracted single-rooted human teeth were instrumented to ProTaper Universal F4 and irrigated using 1% NaOCl. Subsequently, canals were irrigated with copious amounts of deionized water to rinse out the residual chlorine. Subsequently, the teeth were sealed externally and placed in a water bath of 37 °C. Root canals were filled with NaOCl of 1%, 2.75%, 5.5%, or distilled water for 1, 10, 100 or 1000 min (n = 5 teeth per solution and time). Consumption of chlorine was measured using paper points pre-impregnated with 15% potassium iodide. Colour change of the paper points was determined photo-electronically, assessing their red value after absorbing solutions from root canals. Measurements were compared to a standard series of NaOCl down to 0.001% (n = 5 paper points per concentration). RESULTS Red values of the paper points inserted into the root canal were affected by initial NaOCl concentration and time (two-way anova, P < 0.05). If NaOCl concentrations above 0.1% are considered to be clinically relevant, then 5.5% NaOCl retained its activity in the root canal for more than 100 min, whereas 1% NaOCl lost its activity between 10 and 100 min. CONCLUSIONS Nonagitated NaOCl solutions can remain biologically active in human root canals for extended time periods.

Comparison of vehicles to collect dentinal fluid for molecular analysis

OBJECTIVES To test the hypothesis that a material with higher water absorption than polyvinylidene fluoride (PVDF) could increase the yield of target molecules from exposed dentine. METHODS In a series of standard tests, different cellulose membranes were compared to a PVDF counterpart for their ability to absorb water and release protein. In a subsequent randomized clinical trial, the cellulose material with the most favourable values was compared to PVDF regarding the levels of MMP-2 that could be collected from exposed dentine of healthy human teeth during filling replacement. MMP-2 levels were determined by enzyme-linked immunosorbent assay (ELISA). Data from the laboratory experiments were compared between materials using the appropriate parametric tests. The frequency of cases yielding quantifiable levels of MMP-2 was compared between materials by Fishers exact test. The level of significance was set at 5%. RESULTS The cellulose membrane with the largest pore size (12-15μm) absorbed significantly (P<0.05) more water than PVDF. It showed a protein release that was similar to that of PVDF, while the cellulose membranes with smaller pore size retained significantly more protein (P<0.05). Using the large-pore cellulose membrane, MMP-2 could be collected at a quantifiable level from the dentine of healthy teeth in 9 of 13 cases, compared to 1 of 13 with the PVDF membrane (P<0.05). CONCLUSIONS Under the current conditions, a large-pore cellulose membrane yielded more of a molecule of diagnostic value compared to a standard PVDF membrane. CLINICAL SIGNIFICANCE Molecular diagnostics of dentinal fluid are hampered by low yields. In the current study, it was shown that cellulose membranes are more useful to collect MMP-2 from dentinal fluid than PVDF membranes.

Bacterial leakage through temporary fillings in core buildup composite material - an in vitro study.

PURPOSE To evaluate the ability of the provisional filling material Cavit-W alone or in combination with different restorative materials to prevent bacterial leakage through simulated access cavities in a resin buildup material. MATERIALS AND METHODS LuxaCore resin cylinders were subdivided into 4 experimental groups (n = 30), plus a positive (n = 5) and a negative (n = 30) control group. One bore hole was drilled through each cylinder, except those in the negative control group (G1). The holes were filled with Cavit-W (G2), Cavit-W and Ketac-Molar (glassionomer cement, G3), Cavit-W and LuxaCore bonded with LuxaBond (G4), Cavit-W and LuxaCore (G5), or left empty (G6). Specimens were mounted in a two-chamber leakage setup. The upper chamber was inoculated with E. faecalis. An enterococci-selective broth was used in the lower chamber. Leakage was assessed for 60 days and compared using Fishers exact test (α < 0.05) corrected for multiple testing. RESULTS Bacteria penetrated specimens in the positive control group within 24 h. All specimens in the negative control group resisted bacterial leakage for 60 days. Twenty-seven specimens in G2, 26 in G3, and 16 specimens in G5 showed bacterial leakage by the end of the experiment. G4 prevented bacterial penetration completely. The statistical comparison revealed significant differences between G4 and all other experimental groups. CONCLUSION Under the current conditions, Cavit-W alone or combined with a glass-ionomer cement did not prevent bacterial leakage through a resin buildup material for two months. In contrast, covering Cavit-W with a bonded resin material resulted in a bacteria-tight seal for two months.

Intracanal Antibiotic Medication for Sustained Root Surface Disinfection–A Laboratory Evaluation

Purpose: To measure the release of an antibiotic mixture of ciprofloxacin, cerfuroxim and metronidazole (TreVitaMix, TVM) through human dentine and to assess the growth inhibition of Fusobacterium nucleatum. Material and Methods: Twenty-four extracted human incisors were scaled and endodontically treated. Root canals were either filled with antibiotic tri-mixture (TVM) or with the carrier material alone (propylene glycol, PG) and were coronally and apically sealed with a flowable composite. Transradicular medicament release was spectrophotometrically measured at 277 nm in simulated body fluid for up to 21 days. In a second part, an agar diffusion assay (F. nucleatum) with representative TVM concentrations as determined in the first part was performed to study the growth inhibition. Samples were anaerobical incubated for 48 h and inhibition zones were measured. Results: TVM was spectrophotometrically detectable in the immersion solution and released in decreasing concentrations up to 21 days (222.5 ± 65.2 mg/ml at day 1 and 35.1 ± 15.6 mg/ml at day 21). In addition, inhibition zones were shown in the agar diffusion assay at representative TVM concentrations. The carrier material showed no antibacterial effect. Conlusion: TVM showed the potential to penetrate through dentine and to inhibit bacterial growth. Therefore, it might have the potential to disinfect the outer root surface in perio-endo lesions, but further research is needed to confirm these observations.