AAE The Root Canal Specialists

AAE Home > JOE > Abstracts > August 2006, Volume 32, Number 8

Detection and Quantitation of E. faecalis by Real-time PCR (qPCR), Reverse Transcription-PCR (RT-PCR), and Cultivation During Endodontic Treatment
John M. Williams, DDS, MS, Martin Trope, DMD, Daniel J. Caplan, DDS, PhD, Diane C. Shugars, DDS, MPH, PhD

Enterococcus faecalis is frequently recovered from refractory endodontic infections and has been implicated in endodontic treatment failures. This study compared real-time quantitative PCR (qPCR) assay to cultivation for E. faecalis detection and quantitation during endodontic treatment. A reverse-transcription PCR (RT-PCR) assay was also developed to detect the bacterium clinically in the viable but nonculturable (VBNC) state. Intra-canal samples (n = 87) were collected upon access, post-instrumentation/irrigation and postcalcium hydroxide treatment from 15 primary and 14 refractory infections involving 29 single-rooted teeth, and analyzed by the three methods. The bacterium was up to three times more prevalent in refractory than primary infections at each collection step. Overall, qPCR detected significantly more E. faecalis-positive teeth and samples than cultivation (p < 0.001). VBNC E. faecalis was detected by RT-PCR in four samples that were negative by cultivation. These findings support qPCR and RT-PCR as more sensitive methods than cultivation for detecting E. faecalis in endodontic infections.

Identification of Enterococcus faecalis in Root-filled Teeth With or Without Periradicular Lesions by Culture-dependent and—Independent Approaches
G.O. Zoletti, DDS, J.F. Siqueira Jr, DDS MSc, PhD, K.R.N. Santos, BS, MSc, PhD

Enterococcus faecalis is the most commonly found species in root-filled teeth evincing recalcitrant periradicular lesions and as a consequence, a role in causation of endodontic treatment failure has been suggested. The purpose of this study was to evaluate the prevalence of this bacterial species in root-filled teeth with or without periradicular lesions. Identification of E. faecalis was carried out by polymerase chain reaction (PCR) or conventional culture procedures. Overall, E. faecalis was detected by species-specific 16S rRNA gene-based PCR in 40/50 teeth (80%), while culture revealed the occurrence of this species in 8/50 teeth (16%). PCR was significantly more effective than culture in detecting this bacterial species (p < 0.001). Of 27 root-filled teeth with no periradicular lesions, E. faecalis was found in 22 cases (81.5%) by PCR and in five cases (18.5%) by culture. Of 23 root-filled teeth with periradicular lesions, E. faecalis was identified in 18 cases (78%) by PCR and in three cases (13%) by culture. Regardless of the identification technique used, no significant difference was observed when comparing the occurrence of E. faecalis in root-filled teeth with and without periradicular lesions (p > 0.05). Although these findings apparently put into question the status of E. faecalis as the main species causing endodontic treatment failure, other related factors still need to be clarified before this assumption turns into certainty.

Predictors of Root Canal Treatment in Teeth With Full Coverage Restorations
Anna Kirakozova, DDS, Daniel J. Caplan, DDS, PhD

This case-control study sought to identify variables predictive of subsequent root canal therapy (RCT) in teeth receiving full coverage restorations. The University of North Carolina School of Dentistry’s computerized treatment database was used to identify all patients receiving a single-unit crown on a nonendodontically treated permanent tooth from January 1, 1998 through December 31, 2002. These patients then were classified either as cases (those whose crowned teeth received RCT before July 1, 2004) or controls (those whose crowned teeth did not). Computerized data, chart entries, and dental radiographs were examined to determine pre-, intra-, and postoperative factors of interest. In the final multivariable logistic regression model that included 66 cases and 71 controls, younger age (p = 0.005) and greater extent of coronal and root destruction (p = 0.020) remained statistically significant predictors of case status, though many potentially important predictors were unavailable through chart review. Patients could benefit if prospective studies were conducted to identify factors predictive of subsequent endodontic involvement in newly crowned teeth.

A Prospective Clinical Study of Mineral Trioxide Aggregate for Partial Pulpotomy in Cariously Exposed Permanent Teeth
Kefah Mahmood Barrieshi-Nusair, BDS, MS, Muawia Abdulla Qudeimat, BDS, MDent, Sci, FRCD(C)

The aim of this study was to evaluate the success of using gray mineral trioxide aggregate (MTA) for partial pulpotomy in cariously exposed young permanent first molars. Thirty-one first permanent molars of 23 patients with a carious exposure were treated using a partial pulpotomy technique. The age of the patients ranged from 7.2 to 13.1 yr with an average of 10 yr. Clinical and radiographic examination revealed a pulpal response within normal limits and normal appearance of the periradicular area respectively. A diagnosis of reversible pulpitis and normal periapex was established. After isolation, caries removal and carious exposure, the exposed pulp tissue was removed with a diamond bur to a depth of 2 to 4 mm. After hemostasis, 2 to 4 mm of gray MTA paste was placed against the fresh wound. The floor of the cavity was covered with a base of glass ionomer. The teeth were restored with amalgam or stainless steel crowns. Teeth were reviewed radiographically and clinically at 3, 6, 12, and 24 month intervals. Twenty-two of the treated teeth did not show any clinical or radiographic signs of failure during the follow-up evaluation period. Six teeth did not respond to vitality testing at the final follow-up period; however, no radiographic signs of failure or clinical symptoms were detected. Gray MTA was a suitable dressing agent for parital pulpotomy in cariously exposed young permanent first molars.

HGF Enhanced Proliferation and Differentiation of Dental Pulp Cells
Ling Ye, PhD, DDS, Li Peng, MS, DDS, Hong Tan, DDS, Xuedong Zhou, PhD, DDS 

Hepatocyte growth factor (HGF) is mesenchymal-derived growth factor acting through a transmembrane tyrosine kinase receptor, c-met. HGF has multiple effects on different cells. However, its function in dentinogenesis remains unclear. In this study, the expression of HGF in human dental pulp cells (DPCs) in vitro was studied by immunostaining and RT-PCR. The effect of HGF on DPCs proliferation was determined by MTT, while its effect on cell differentiation was analyzed using ALPase activity, and further confirmed with ALP and DSPP mRNA and protein expression. Immunostaining revealed that HGF was found mainly in the cytoplasm of DPCs. RT-PCR analysis showed that both HGF and c-met were expressed from the DPCs. Exogenous addition of HGF enhanced proliferation and differentiation of DPCs by up-regulating CREB, ELK-1, and PPAR-γ. U0126, an ERK/MAPK inhibitor, inhibited the effects of HGF on DPCs. It was concluded that HGF stimulated both proliferation and differentiation of DPCs, at least partially through the ERK/MAPK pathway.

Quantitative Analysis of Osteoclast-Specific Gene Markers Stimulated by Lipopolysaccharide
Jin Jiang, DDS, PhD, Haitao Li, DDS, Farshid S. Fahid, DDS, Elizabeth Filbert, DMD, Kamran E. Safavi, DMD, MEd, Larz S. Spangberg, DDS, PhD, Qiang Zhu, DDS, PhD

Lipopolysaccharide (LPS) in the outer layers of Gram-negative bacteria plays an important role in initiating and sustaining periapical lesions. To understand the mechanisms of osteoclastic bone resorption in periapical lesions induced by LPS, we stimulated osteoclast precursors, RAW 264.7 cells with LPS. LPS stimulated osteoclastogenesis when osteoclast precursors were primed with activator for NF-κB ligand (RANKL) as little as 24 h. By employing real-time PCR analysis, we have confirmed that osteoclast-like cells stimulated by LPS express high level of osteoclast-specific gene markers such as TRAP, cathepsin K, and calcitonin receptor. These results suggest that bone-resportive action by LPS is partially independent of RANKL.

Expression of Toll Like Receptor 4 in Normal Human Odontoblasts and Dental Pulp Tissue
Hong-Wei Jiang, MD, Wei Zhang, PhD, Bang-Peng Ren, MS, Jin-Feng Zeng, MD, Jun-Qi Ling, MD 

The aim of the study was to determine the expression of TLR4 in odontoblasts and the dental pulp. Odontoblasts and pulp tissues were collected from freshly extracted human wisdom teeth. Reverse transcription-polymerase chain reaction and Western blotting were performed to detect TLR4 mRNA and protein expression, respectively. Immunohistochemical staining was used to determine the distribution of TLR4 in odontoblasts and the pulp. Scanning electron microscopy (SEM) was applied to observe the morphology of odontoblasts. It was demonstrated that TLR4 mRNA and protein expressions were both present in cells of odontoblast layer and pulp tissues and that TLR4 expression was distributed in odontoblasts and some pulpal vascular endothelial cells. SEM revealed the integrity of the odontoblast cell-layer and the well-preserved morphology of individual odontoblast cells. These findings suggest that TLR4 expressed in odontoblasts may play an important role in the dental immune defense.

Fracture Resistance and Failure Patterns of Endodontically Treated Mandibular Molars Restored Using Resin Composite With or Without Translucent Glass Fiber Posts
Ziad Salameh, Roberto Sorrentino, Federica Papacchini, Hani F. Ounsi, Esam Tashkand, Cecilia Goracci, Marco Ferrari 

The elastic modulus of the restorative material is important in restoring endodontically treated teeth. This study aimed to compare the fracture resistance and failure patterns of 90 mandibular molars restored using resin composites with or without fiber posts, with respect to the number of residual cavity walls. Five restoration types were performed corresponding to different wall defects (groups 1-5). Groups were divided in two subgroups corresponding to the use or absence of fiber posts. Teeth were loaded and resistance of specimens was measured as the axial compressive load to cause fracture and macroscopic fracture patterns were observed. One way ANOVA revealed a significant difference in fracture resistance (p < 0.001). Tukey post hoc test also revealed significant differences between groups as samples restored with fiber posts exhibited mostly restorable fractures. It was concluded that the resistance of endodontically treated mandibular molars restored with composite resins is mainly affected by the number of residual walls. Using fiber-reinforced posts optimized fracture patterns.

Cytotoxicity of a New Root Canal Filling Material on Human Gingival Fibroblasts
Jennifer E. Key, DMD, Firoz G. Rahemtulla, PhD, Paul D. Eleazer, DDS 

This study evaluated the cytotoxicity of the root canal sealing materials Resilon and Epiphany versus gutta-percha, Grossman’s sealer, Thermaseal, and Sealapex. Using human gingival fibroblasts the fibroblasts cultures were incubated for either 1 or 24 h to test the cytotoxicity after freshly mixing or after 24 h of setting. Fibroblasts were then stained with trypan blue, to determine number of dead cells. Data were analyzed using ANOVA and t tests. Resilon was similar to gutta-percha and the control. Epiphany was less cytotoxic than Grossman’s sealer at both the 1 and 24 h time periods. Epiphany was more cytotoxic than Sealapex at the 1-h time period but less cytotoxic at the 24 h time period. These results indicated that Resilon had a lower cytotoxicity and that Epiphany was more cytotoxic than conventional materials.

An In Vitro Assessment of the Sealing Ability of Resilon/Epiphany Using Fluid Filtration
Stephen G. Biggs, DDS, Kenneth I. Knowles, DDS, MS, Jose L. Ibarrola, DDS, MS, David H. Pashley, DMD, PhD

The aim of this in vitro study was to compare the sealing ability of Resilon/Epiphany to gutta-percha and Roth or AH Plus sealers. Leakage of the obturated roots was measured using the fluid filtration technique. There were eight groups of 12 teeth each. Group 1: obturated with Resilon/Epiphany; group 2: obturated with gutta-percha/Roth, allowed to set for 3 wk; group 3: obturated with gutta-percha/Roth; group 4: obturated with Resilon/Epiphany using a single cone; group 5: obturated with Resilon cones without using primer/sealer (positive control); group 6: obturated with gutta-percha/AH Plus; group 7: same as group 6, but allowed to set for 8 h; and group 8: sealed on the outside with three layers of nail varnish (negative control). The results showed that only the positive control (group 5) leaked significantly more (p < 0.05) than the other groups. There was no effect of time on leakage. Resilon/Epiphany was no better than gutta-percha/Roth or gutta-percha/AH Plus at sealing root canals.

A Comparison of Three Gutta-Percha Obturation Techniques to Replicate Canal Irregularities
Jake Collins, BS, MS, Mary P. Walker, DDS, PhD, James Kulild, DDS, MS, Charles Lee, DDS

A split-tooth model with artificially created intracanal wall defects was used to compare three gutta-percha (GP) obturation techniques, cold lateral, warm lateral, and warm vertical. The techniques were evaluated and compared based on defect replication quality as a function of defect location and size. The obturations were evaluated on an ordinal scale, 0 to 4, based on how much each defect was replicated. There was a statistically significantly better result with both warm techniques compared to cold lateral obturation, while there was no significant difference between the warm obturation techniques.

Improving the Performance of EndoRez Root Canal Sealer With a Dual-cured Two-Step Self-etch Adhesive. I. Adhesive Strength to Dentin
Michael D. Doyle, Robert J. Loushine, Kelli A. Agee, William T. Gillespie, R. Norman Weller, David H. Pashley, Franklin R. Tay 

This study examined the hypotheses that the use of a self-etching-adhesive improves the adhesion of EndoRez to dentin, and that both techniques are unaffected by the prolonged use of NaOCl when EDTA is used as the final rinse. A tensile bond testing design was employed with composite cylinders bonded to dentin using: (a) 17% EDTA-2 min/EndoRez; (b) 6.15% NaOCl-1 h/EDTA-2min/EndoRez; (c) EDTA-2 min/Clearfil Liner Bond 2V/EndoRez; and (d) NaOCl-1 h/EDTA-2min/Clearfil Liner Bond 2V/EndoRez. Fractographic analyses of debonded specimens were performed using SEM. Two-way ANOVA and Tukey test revealed higher tensile bond strengths with the self-etching-adhesive-modified EndoRez technique (p < 0.001). Sealer tags pulled away from dentinal tubules when EndoRez was used alone, whereas mixed failures occurred within sealer tags, hybrid layers, adhesive, and sealer in the self-etching-adhesive-modified EndoRez technique. Both techniques were unaffected by the prolonged use of NaOCl (p > 0.05) when EDTA was employed as the final rinse.

Improving the Performance of EndoREZ Root Canal Sealer With a Dual-cured Two-Step Self-etch Adhesive. II. Apical and Coronal Seal
William T. Gillespie, Robert J. Loushine, R. Norman Weller, Annalisa Mazzoni, Michael D. Doyle, Jennifer L. Waller, David H. Pashley, Franklin R. Tay 

This study tested the hypothesis that the seal of EndoREZ may be improved with an adhesive-modified technique. Instrumented single-rooted teeth were filled with: (a) Warm vertical compaction with AH Plus (control); (b). EndoREZ with master cones and passive application of accessory cones; (c) Application of Clearfil Liner Bond 2V before EndoREZ and gutta-percha condensation. Leakage was assessed by fluid filtration at 10 psi before root resection and after 3 to 12 mm apical resections. Statistical analyses revealed significant differences in filling technique and resection lengths. EndoREZ exhibited significantly higher overall leakage, while no difference was found between AH Plus and the adhesive-modified EndoRez technique. Apical resection of EndoREZ to 12 mm exhibited more leakage than all other interactions, but was not significantly different from the same material resected to 9 mm. Although EndoREZ exhibited an acceptable apical seal, its coronal seal may be improved with the use of a dual-cured self-etch adhesive.

A Comparative Study of the Biocompatibility of Three Root-end Filling Materials in Rat Connective Tissue
Shahriar Shahi, DDS, MSc, Saeed Rahimi, DDS, MSc, Mehrdad Lotfi, DDS, MSc, HamidReza Yavari, DDS, MSc, AliReza Gaderian, DDS, MSc

The purpose of this study was to compare the biocompatibility of amalgam, gray MTA and white MTA in the connective tissue of rats. We used 45 Sprague-Dawley rats in this study. The rats were divided into three groups. Root end filling materials were placed in polyethylene tubes and inserted into the rats’ connective tissue through incisions. The rats were sacrificed after 3 days, 1 wk, and 3 wk, respectively. Histologic samples were sectioned in 5-μm thicknesses and stained with hematoxylin and eosin. Kruskal-Wallis test was used for statistical analysis. The results showed that after 3 days, white MTA was more biocompatible than gray MTA and amalgam. After 1 week, gray MTA was more biocompatible than white MTA and Amalgam. After 3 wk, there were no significant differences between experimental groups and the control group.

Effect of CHX on the Decalcifying Effect of 10% Citric Acid, 20% Citric Acid, or 17% EDTA
S. González-López, D. Camejo-Aguilar, P. Sanchez-Sanchez, V. Bolaños-Carmona

The objective of this study was to measure the demineralization capacity of 10% and 20% citric acid and 17% EDTA after three time periods and to determine whether it was modified by the addition of 1% of commercial chlorhexidine (CHX).

Three slices of 2-mm thickness were cut from the cervical third of the root of ten bovine incisors and sectioned into two equal parts, obtaining six specimens per tooth. Specimens were assigned to one of six groups (n = 10) for immersion in 25 ml of the above mentioned solutions or 25 ml of these same solutions plus 1% CHX (Hibimax). At 3, 10, and 15 min of immersion, the concentration of Ca2+ was measured by atomic absorption spectrophometry.

The demineralization effect of all solutions was time-dependent (F = 158,448; p < 0.001), without significant differences among them (p = 0.783). Addition of 1% CHX did not modify the demineralizing capacity of these solutions. In the first three minutes, significantly more [Ca2+] was obtained when 17% EDTA was used in comparison with the other solutions.

Leakage and Sealer Penetration in Smear-free Dentin After a Final Rinse With 95% Ethanol
Richard W. Stevens, DDS, MS, James M. Strother, DDS, MS, Scott B. McClanahan, DDS, MS

Forty extracted maxillary incisors were decoronated, prepared in a crown-down fashion and randomly divided into two groups of 16 roots each. Remaining roots served as controls. Smear layer was removed with 17% EDTA followed by 5.25% NaOCl, the canals in group N were again rinsed with NaOCl before obturation with laterally compacted gutta-percha and Roth’s 801 sealer. The roots in group E were rinsed with 95% ethyl alcohol instead of NaOCl for the final rinse. Leakage was determined using a fluid-flow model. Roots were cleared, split, and sealer penetration into the dentinal tubules was measured under light-microscopy. Group E demonstrated significantly greater sealer penetration (p = 0.002) and significantly less leakage (p = 0.040), than group N. Leakage could not be significantly correlated with sealer penetration (p = 0.725). Under the conditions of this study, we found that a final rinse with 95% ethyl alcohol increased sealer penetration and decreased leakage.

Removal of Separated Files from Root Canals With a New File-removal System: Case Reports
Yoshitsugu Terauchi, DDS, Le O’Leary, DDS, Hideaki Suda, DDS, PhD

There have been many different devices and techniques developed to retrieve instruments fractured during endodontic procedures, but none of them can consistently remove separated instruments from root canals. Iatrogenic accidents such as perforation and canal destruction have been reported during the removal of separated instruments. The file removal process becomes even more difficult when breakage occurs in a curved canal or in the apical third of the canal. Four cases requiring removal of separated files from the apical third of curved canals are presented. All were successfully treated using a newly designed system and technique.

Surgical Root Restoration After External Inflammatory Root Resorption: A Case Report
Geert M.G. Hommez, DDS, PhD, Hilde A.A. Browaeys, MD, Roeland J.G. De Moor, DDS PhD, MSc

External inflammatory root resorption after a jaw fracture is rare. This report describes a case of extensive external root resorption in the middle third of the root of a mandibular right canine after a mandibular fracture involving the tooth socket. Because of delayed treatment and damage to the root caused by a surgical screw, root canal treatment was performed followed by surgical intervention. The resorptive defect was debrided and part of the root was rebuilt with conventionally setting restorative glass ionomer cement. Postoperative follow-up revealed complete healing.