April 2004, Volume 30, Number 4

Revascularization of Immature Permanent Teeth with Apical Periodontitis: New Treatment Protocol?
Francisco Banchs, DDS, MS, and Martin Trope, DMD

Preliminary Evaluation of Bioactive Glass S53P4 as an Endodontic Medication In Vitro
Matthias Zehnder, DMD, Eva Söderling, PhD, Jukka Salonen, DDS, PhD, and Tuomas Waltimo, DDS, PhD

Acceleration Effect of Human Recombinant Bone Morphogenetic Protein-2 on Differentiation of Human Pulp Cells into Odontoblasts
Takashi Saito, DDS, PhD, Masafumi Ogawa, DDS, Yoshiaki Hata, DDS, PhD, and Kazuhisa Bessho, DDS, PhD

Increased Tgf-β1 Production by Rat Osteoblasts in the Presence of PepGen P-15 in Vitro
Candice Trasatti, DDS, MS, Robert Spears, PhD, James L. Gutmann, DDS, and Lynne A. Opperman, PhD

pH Required to Kill Enterococcus faecalis in Vitro
Christopher P. McHugh, DMD, Ping Zhang, PhD, DDS, Suzanne Michalek, PhD, and Paul D. Eleazer, DDS, MS

Expression of Calcitonin Gene-Related Peptide (CGRP) in Irreversible Acute Pulpitis
Javier Caviedes-Bucheli, MSc, Carolina Camargo-Beltran, DDS, Ana Milena Gomez-la-Rotta, DDS, Sandra Cristina-Trujillo Moreno, DDS, Gloria Cristina Moreno Abello, MSc and John Mario Gonzalez-Escobar, PhD

In Vitro Evaluation of the Accuracy of Three Electronic Apex Locators
C. Lucena-Martín, PhD, DDS, MD, V. Robles-Gijón, PhD, DDS, C. M. Ferrer-Luque, PhD, DDS, MD, and J. M. Navajas-Rodríguez de Mondelo, PhD, DDS, MD

Adhesion of a New Methacrylate Resin-Based Sealer to Human Dentin
Christos Gogos, DDS, PhD, Nickolaos Economides, DDS, MsC, PhD, Christos Stavrianos, DDS, PhD, Ioannis Kolokouris, DDS, PhD, and Ioannis Kokorikos, DDS

K3 Endo, ProTaper, and ProFile Systems: Breakage and Distortion in Severely Curved Roots of Molars
Matthew T. Ankrum, DDS, Gary R. Hartwell, DDS, MS, and John E. Truitt, BS

Influence of Manual Preflaring and Torque on the Failure Rate of ProTaper Rotary Instruments
Elio Berutti, MD, DDS, Alfonso Roberto Negro, MD, DDS, Mario Lendini, DDS, andDamiano Pasqualini, DDS

Dental Alterations Associated with X-Linked ypophosphatemic Rickets
Claudio Maranhão Pereira, DDS, MsC, Cleverton Roberto de Andrade, DDS, MsC, ablo Agustín Vargas, DDS, PhD, Ricardo Della Coletta, DDS, PhD, Oslei Paes de Almeida, DDS, PhD, and Márcio Ajudarte Lopes, DDS, PhD

Intracanal Placement of Calcium Hydroxide: A Comparison of Techniques, Revisited
Craig P. Torres, DDS, Michael J. Apicella, DDS, Peter P. Yancich, DDS, and M. Harry Parker, DDS, MS

Sonicated Extract of Enterococcus faecalis Induces Irreversible Cell Cycle Arrest in Phytohemagglutinin-Activated Human Lymphocytes
WooCheol Lee, DDS, MS, SungSam Lim, DDS, PhD, Ho-Hyun Son, DDS, PhD, andKwang-Shik Bae, DDS, PhD

Revascularization of Immature Permanent Teeth with Apical Periodontitis: New Treatment Protocol?
Francisco Banchs, DDS, MS, and Martin Trope, DMD

A new technique is presented to revascularize immature permanent teeth with apical periodontitis. The canal is disinfected with copious irrigation and a combination of three antibiotics. After the disinfection protocol is complete, the apex is mechanically irritated to initiate bleeding into the canal to produce a blood clot to the level of the cementoenamel junction. The double seal of the coronal access is then made. In this case, the combination of a disinfected canal, a matrix into which new tissue could grow, and an effective coronal seal appears to have produced the environment necessary for successful revascularization.

Preliminary Evaluation of Bioactive Glass S53P4 as an Endodontic Medication In Vitro
Matthias Zehnder, DMD, Eva Söderling, PhD, Jukka Salonen, DDS, PhD, and Tuomas Waltimo, DDS, PhD

Calcium hydroxide is the “gold standard” endodontic medication, but it may fail to eliminate certain facultative bacteria and yeasts found in root canal systems. In this study, standardized bovine dentin blocks infected with Enterococcus faecalis were treated with an aqueous calcium hydroxide or bioactive glass S53P4 (BAG) powder suspension. While calcium hydroxide was ineffective, the BAG suspension eliminated the infection in the sampled dentin layers after 5 days. In a direct exposure test, preincubation with human dentin boosted the BAG-killing efficacy against E. faecalis ATCC29212, Candida albicans CCUG19915, Pseudomonas aeruginosa ATCC9027, Streptococcus sanguis ATCC10556, and S. mutans ATCC25175. BAG placed in the root canals of extracted teeth did not alter rootdentin pH. Consequently, the conditions under which the microbiota were killed were not pHmediated. Further studies are ongoing to help clarify the antimicrobial BAG effect in the presence of dentin.

Predictable pulp capping procedures remain problematic, possibly because of the lack of appropriate stimulating factors for dentin formation. The present study examines the ability of one such stimulating factor, bone morphogenetic protein-2, to accelerate the differentiation of human dental pulp cells into odontoblasts. The number and morphology of cells between groups treated with 0 and 100 ng/ml of human recombinant bone morphogenetic protein-2 (rhBMP-2) did not significantly differ. However, ALPase activity (a marker for biomineralization) in the group stimulated with rhBMP-2 was more than double that of the control group. We then measured the expression of mRNA encoding dentin sialophosphoprotein (DSPP) as a marker of odontoblasts in rhBMP-2– stimulated human pulp cells using a quantitative polymerase chain reaction. The expression of DSPP mRNA in cells stimulated for 24 h by 1000 ng/ml of rhBMP-2 was approximately 20-fold and 5-fold higher than that by stimulated by 10 and 100 ng/ml, respectively. These findings show that rh-BMP-2 promoted the differentiation of human dental pulp cells into odontoblasts but did not affect cell proliferation, suggesting that rhBMP-2 may have therapeutic utility in vital pulp therapy.

Increased Tgf-β1 Production by Rat Osteoblasts in the Presence of PepGen P-15 in Vitro
Candice Trasatti, DDS, MS, Robert Spears, PhD, James L. Gutmann, DDS, and Lynne A. Opperman, PhD

Bone grafting materials may enhance tissue regeneration after endodontic, periodontal, or implant surgery. The differences in physical and biological properties between products may result in different osteoblastic responses. This study was designed to determine whether interleukin-1β and Tgf-β1 production by primary cultures of rat osteoblasts differed when cells were exposed to three grafting materials: BioOss, OsteoGraf N-300, and PepGen P-15. Cells were exposed to materials for 24, 48, and 72 h and were characterized by mineralized nodule formation. Supernatants were collected for Lowry and enzyme-linked immunosorbent assays to assess cytokine production. All groups produced mineralized nodules after 14 days. Statistical analysis revealed no difference in interleukin-1β production between groups, but a significant increase in Tgf-β1 production was noted in the PepGen P-15 group. These results indicate that PepGen P-15 stimulates osteoblasts to express Tgf-β1, which may accelerate repair of bone defects created during periradicular or dental implant surgeries.

pH Required to Kill Enterococcus faecalis in Vitro
Christopher P. McHugh, DMD, Ping Zhang, PhD, DDS, Suzanne Michalek, PhD, and aul D. Eleazer, DDS, MS

Enterococcus faecalis tolerates highly alkaline environments, yet the exact pH required for killing E. faecalis is not known. This study tests growth at 0.5 increments from pH 9.5 to 12. Twelve culture tubes were used in each group. Positive growth was measured using turbidity, a visual scale, and a spectrophotometer. At 24 h, growth was observed in all tubes at pH 9.5 and 10. At 48 h, all pH 10.5 tubes showed growth. At 72 h, six of the pH 11 tubes showed growth. After 7 days, five of the remaining pH 11 tubes were positive. No growth occurred in any of the pH 11.5 or pH 12 tubes. Apparently, pH 10.5 to 11.0 retards growth of E. faecalis, whereas no tubes showed growth at pH 11.5 or greater.

The main goal of this study was to evaluate tissue levels of calcitonin gene-related peptide (CGRP) in human pulpal samples collected from teeth with a clinical diagnosis of acute irreversible pulpitis, normal pulps, and teeth with induced pulpal inflammation. All the pulp tissue was mechanically separated, collagenase digested to release individual cells, and labeled with FITC detection of an anti-CGRP polyclonal antibody. Detection of CGRP was possible in these cells due to a binding of the antibody to CGRP that was itself bound to its cell surface receptor. Flow cytometry analysis indicated that the labeled pulp cells were located in a region of low size and complexity according to their forward (FSC) and side scatter (SSC) properties. Significant statistical differences were found between the percentages of CGRP expression in healthy pulps and pulps with induced inflammation and between healthy pulps and pulps with acute irreversible pulpitis. No significant statistical differences were found between pulps with induced inflammation and pulps with acute irreversible pulpitis. These findings support the hypothesis that the CGRP system is active in human pulpal inflammation and may modulate the inflammatory response.

The accuracy of three electronic apex locators (EALs) (Justy II, Root ZX, and Neosono Ultima EZ) is evaluated, together with the concordance of the measurements obtained by two different operators. Twenty single-root human teeth were used, sectioning the crown to gain access to the root canal. A first operator (A) determined the reference (or control) length (corresponding to the actual length) for each tooth, after which all teeth were measured individually and independently by the other two operators (B and C). The results obtained with each EAL and by each operator were in turn compared with the corresponding control length. The statistical analysis of the results showed EAL reliability in detecting the apex to vary from 80% to 85% and 85% to 90% (depending on the operator) for the Justy II and Neosono systems, respectively, whereas reliability was found to be 85% for the Root ZX device. These results, combined with a high interobserver concordance, suggest electronic root canal measurement to be an objective and acceptably reproducible technique.

The purpose of this study was to compare the shear bond strength of four root canal sealers: Fibrefill (a methacrylate resin sealer), Endion (a glass ionomer sealer), Topseal (an epoxy resin sealer), and CRCS (a calcium hydroxide sealer) to human root canal dentin. The dentin specimens were divided into four groups of 12 specimens each and etched with 3 ml of EDTA 17% before rinsing with NaOCl 2.5% and distilled water. Bond strength was tested using a test machine by subjecting samples to a shear load at a crosshead speed of 0.5 mm/min. The best results were observed in the Fibrefill group, in which the mean shear bond strength was 7.993 ± 2.365 MPa. The values in the Endion (1.816 ± 0.614 MPa) and CRCS (2.224 ± 0.509 MPa) groups were significantly lower than in the Fibrefill and Topseal (6.235 ± 1.428 MPa) groups. Inspection of the fractured surfaces revealed the bond failure to be mainly adhesive to dentin for Fibrefill and cohesive for Endion sealer. CRCS and Topseal sealers showed both adhesive and cohesive failures.

K3 Endo, ProTaper, and ProFile Systems: Breakage and Distortion in Severely Curved Roots of Molars
Matthew T. Ankrum, DDS, Gary R. Hartwell, DDS, MS, and John E. Truitt, BS

It was the aim of this study to investigate the incidence of file breakage and distortion when the ProTaper, K3 Endo, and ProFile systems were used to instrument canals in the severely curved roots of extracted molars. Forty-five roots of extracted mandibular and maxillary molars with curvatures between 40 and 75 degrees were chosen for this study. The canals in group 1 were instrumented with the ProFile system. Roots in group 2 were instrumented with the ProTaper system, and those in group 3 were instrumented with the K3 Endo system. The three systems were used according to the manufacturers’ instructions. The proportion of files distorted was 15.3% for the ProFile group, 2.4% for the ProTaper group, and 8.3% for the K3 Endo group. There was a statistically significant difference between the Pro-File and ProTaper groups (p = 0.0079). The percentage of broken files was 1.7% for the ProFile group, 6.0% for the ProTaper group, and 2.1% for the K3 Endo group. No statistically significant differences were found between these three groups (p = 0.4243). The results of this study showed that these three rotary tapered systems were not significantly different with regard to breakage. There were significantly more distorted files in the ProFile group when compared with the ProTaper group. With regard to distortion, there was no significant difference between the ProTaper and K3 Endo and the ProFile and K3 Endo groups.

We evaluated the influence of manual preflaring and torque on the failure rate of rotary nickel-titanium ProTaper instruments Shaping 1 (S1), Shaping 2 (S2), Finishing 1 (F1), and Finishing 2 (F2). These factors were evaluated using an in vitro method by calculating the mean number of Endo-Training-Blocks shaped before file breakage under different conditions. Group A (S1 on simulators with no preflaring) shaped 10 blocks before failure, group B (S1 on manually preflared simulators) shaped 59 blocks (p < 0.01 versus group A), group C (S2 with low torque) shaped 28 blocks, group D (S2 with high torque) shaped 48 blocks (p < 0.01 versus group C), group E (F1 with low torque) shaped eight blocks, group F (F1 with high torque) shaped 23 blocks (p < 0.01 versus group E), group G (F2 with low torque) shaped four blocks, and group H (F2 with high torque) shaped 11 blocks (p < 0.01 versus group G). Manual preflaring creates a glide path for the instrument tip and is a major determinant in reducing the failure rate of these rotary nickel-titanium files. All instruments worked better at high torque.

The X-linked hypophosphatemic rickets is a rare metabolic disorder characterized by low serum phosphate levels caused by a decreased renal tubular reabsorption of inorganic phosphates. The initial complaints are a delay in the development of walking caused by deformity of the legs. Oral findings include poorly mineralized dentin, enlarged pulp chambers and root canals, and periradicular abscesses in caries-free teeth. We present three patients from the same family with X-linked hypophosphatemic rickets showing bone and dental alterations.

Intracanal Placement of Calcium Hydroxide: A Comparison of Techniques, Revisited
Craig P. Torres, DDS, Michael J. Apicella, DDS, Peter P. Yancich, DDS, and M. Harry Parker, DDS, MS

The goal of this in vitro study was to compare the effectiveness of three different placement techniques of calcium hydroxide paste in a curved canal. An acrylic resin block with a 44-degree curvature was instrumented to a #40 Flex-R file. The canal was filled with radiopaque calcium hydroxide paste using the following techniques: group 1, the paste was injected with an Ultradent tip; group 2, the paste was injected as in group 1 and followed by a size 35 Lentulo to working length; and group 3, the paste was placed with a size 35 Lentulo only, to working length. Each technique was performed 25 times. The radiodensity of the simulated canal was measured at 1, 3, 5, and 7 mm from the canal terminus. At 1 mm, the Lentulo was significantly better than the other two techniques. At 3 mm, the Lentulo technique and the combined technique were better than the Ultradent tip alone.

Sonicated Extract of Enterococcus faecalis Induces Irreversible Cell Cycle Arrest in Phytohemagglutinin-Activated Human Lymphocytes
WooCheol Lee, DDS, MS, SungSam Lim, DDS, PhD, Ho-Hyun Son, DDS, PhD, andKwang-Shik Bae, DDS, PhD

The purpose of this study was to examine whether the sonicated extract of Enterococcus faecalis (SEF) alters the cell cycle transition of lymphocytes and thus regulates the fate of the arrested cells. Human lymphocytes were activated by phytohemagglutinin in the presence or absence of SEF, and cell cycle was assessed by flow cytometry. Seventy-two hours after activation with phytohemagglutinin, cells were activated from G0/G1 to S 6.1%) and G2/M (3.8%) phases of the cell cycle. In contrast, pretreatment with SEF resulted in 90.5% of cells remaining in G0/G1, and cell cycle progression to the S and G2/M phases was consequently inhibited. Caspase assay demonstrated that SEF-treated cells exhibited significantly increased apoptosis (56.7%) compared with phytohemagglutinin alone (28.1%). We propose that if this irreversible cell cycle arrest induced by E. faecalis occurs in vivo, it may result in local immunosuppression and contribute to the pathogenesis of endodontic failure. Our findings that E. faecalis can inhibit lymphocyte responses may be of particular relevance to the pathogenesis of endodontic failure. Although the immunologic mechanism involved in the pathogenesis of persistent periapical lesion is not clearly defined, it is reasonable to predict that the altered immune reaction may be linked to the immunosuppressive potential of E. faecalis or other oral bacteria.

AAE The Root Canal Specialists

April 2004, Volume 30, Number 4