Introduction: Macrophages are highly activated by endodontic contents. This study investigated th... more Introduction: Macrophages are highly activated by endodontic contents. This study investigated the correlation between different clinical signs/symptoms and radiographic features according to the levels of interleukin (IL)-1b, tumor necrosis factor a (TNF-a), IL-6, IL-10, prostaglandin E 2 (PGE 2 ), and their networks produced by endodontic content-stimulated macrophages collected from primary endodontic infection with apical periodontitis (PEIAP). Methods: Samples were taken from 21 root canals with PEIAP by using paper points. The presence of exudate (EX), pain on palpation (POP), tenderness to percussion (TTP), and the size of the radiographic lesion (SRL) were recorded. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate (LAL) assay for endotoxin measurement. Raw 264.7 macrophages were stimulated with bacterial contents during 24 hs. The amounts of IL-1b, TNF-a, IL-6, IL-10 and PGE 2 were measured by enzyme-linked immunosorbent assay. Log-based data were correlated by multiple logistic regression (P < .05). Results: Bacteria and endotoxin were detected in 100% of the samples. IL-6 and TNF-a were positively correlated with SRL and EX, respectively (P < .05). Clinical signs/symptoms and radiographic findings were set as dependent variables for EX-positive correlations between PGE 2 , IL-1b, and TNF-a (P < .05), whereas IL-6 and PGE 2 were positively correlated to each other in POP but negatively correlated in SRL (P < .05). When POP and TTP-POP were set as dependent variables, different cytokine networks were found. Conclusions: Our findings suggest different roles for each cytokine in the development of apical periodontitis, whose effects of overlapping networks depend on the signs/symptoms and radiographic features found in endodontic infection. (J Endod 2012;38:740-745)
Introduction: Periradicular tissue chronic stimulation by endotoxin may cause apical periodontiti... more Introduction: Periradicular tissue chronic stimulation by endotoxin may cause apical periodontitis. This study investigated the microbial profile and the levels of endotoxin found in primary root canal infection with apical periodontitis, determined their antigenicity against macrophages through the levels of interleukin (IL)-1b and tumor necrosis factor a (TNF-a), and evaluated their relationship with clinical and radiographic findings. Methods: Samples were taken from 21 root canals with primary endodontic infection and apical periodontitis with paper points. PCR technique (16S rDNA) was used for the detection of the target bacteria. Limulus Amebocyte Lysate (LAL) was used to measure endotoxin. The amounts of IL-1ß/ TNF-alpha in macrophages supernatants were measured by enzyme-linked immunosorbent assay -Duoset-kit (ELISA). Results: Prevotella nigrescens (13/21), Porphyromonas endodontalis (6/21), and Treponema socranskii (6/21) were the most frequently detected gram-negative bacterial species. The presence of the sinus tract (2/21) was related to the detection of Filifactor alocis (p < 0.05), whereas a tooth with a radiolucent area $2 mm was related to the detection of Treponema denticola. A correlation was found between the number of gram-negative bacteria and the levels of IL-1b/TNF-a (p < 0.05). Increased levels of endotoxin were followed by TNF-a release (p < 0.05). Higher levels of IL-1b (p < 0.05) and endotoxin contents were related to the larger size of the radiolucent area. Conclusion: The antigenicity of the endodontic contents is not only related to the amount of endotoxin found in the root canal but also to the number of different species of gram-negative bacteria involved in the infection. Moreover, a larger size ($2 mm) of the radiolucent area was related
Introduction: Root canal contents are potent stimuli for proinflammatory cytokines involved in ap... more Introduction: Root canal contents are potent stimuli for proinflammatory cytokines involved in apical periodontitis. This study investigated target gram-negative bacterial species and endotoxins in primary endodontic infection with apical periodontitis, determined their antigenicity against macrophages through the levels of PGE 2 , and evaluated their relationship with clinical findings. Methods: Samples were taken from 21 root canals with primary infection and apical periodontitis by using paper points. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate assay for endotoxin measurement. Levels of prostaglandin E2 (PGE 2 ) were measured by enzyme-linked immunosorbent assay (Duoset Kit; R&D, Minneapolis, MN). Results: Prevotella nigrescens (13/21), Fusobacterium nucleatum (6/21), and Porphyromonas endodontalis (6/21) were the most frequently observed species. A positive association was found between F. nucleatum and P. endodontalis (P < .05). A correlation was found between the number of gram-negative bacterial species and the levels of endotoxins, as well as PGE 2 (P < .05). Higher levels of endotoxin were detected in teeth with exudation, whereas elevated levels of PGE 2 were found in teeth with tenderness to percussion and pain on palpation. Conclusions: Our findings imply an additive effect between the number of gram-negative bacterial species involved in endodontic infection regarding the induction of proinflammatory cytokine by macrophage cells. Moreover, teeth with clinical symptomatology were related to higher levels of endotoxins and PGE 2 secretion. (J Endod 2011;37:602-607)
Introduction: Although Enterococcus faecalis is a member of the normal microbiota, it is also a m... more Introduction: Although Enterococcus faecalis is a member of the normal microbiota, it is also a major cause of nosocomial infections. Some strains of E. faecalis produce capsule, which contributes to pathogenesis through evasion of host defenses, and its production is dependent on the capsule (cps) operon polymorphism. This study investigated cps locus polymorphism in distinct lineages of E. faecalis isolated from canals of root-filled teeth with periapical lesions. Methods: Twenty-two E. faecalis isolates were evaluated regarding the cps operon polymorphism and genetic diversity. The 3 known CPS types were determined by polymerase chain reaction. This information was correlated with multilocus sequence typing data, which were used to define genetic lineages. Results: cpsA and cpsB were the only detected genes within the cps operon in 62.5% of E. faecalis strains (14/22), indicative of genotype CPS 1, which lacks capsule expression. The essential genes in the cps operon for capsule production were detected in the remaining strains, whereas 3 belonged to genotype CPS 5 and 5 strains to genotype CPS 2. A total of 14 sequence types (STs) were resolved in 22 E. faecalis isolates. Comparison with the E. faecalis international multilocus sequence typing database revealed that 9 STs were previously found, and that the 5 STs were novel. Conclusions: Certain E. faecalis genotypes from canals of root-filled teeth with periapical lesions belong to lineages associated with capsule expression and production of multiple virulence factors, which might account for their increased pathogenic potential. (J Endod 2012;38:58-61)
Introduction: This clinical study was conducted to measure the endotoxin levels in infected root ... more Introduction: This clinical study was conducted to measure the endotoxin levels in infected root canals (RCs) and exudates related to acute apical abscesses (AAAs). In addition, the effectiveness of RC procedures in reducing the endotoxin levels in RCs was monitored. Methods: Paired samples of infected RCs and exudates from AAAs were collected from 10 subjects by using paper points. RCs samples were collected before (RCS1) and after chemomechanical preparation (CMP) (RCS2), after 17% EDTA (RCS3), and after 30 days of intracanal medication (Ca[OH] 2 + chlorhexidine) (RCS4). A turbidimetric kinetic limulus amebocyte lysate assay was used for the measurement of endotoxins. Results: Endotoxins were detected in 100% of the baseline samples of AAAs and RCs (RCS1) with median values of 175 EU/mL and 41.5 EU/mL, respectively (P < .05). After CMP (RCS2), endotoxins were reduced to a median value of 0.54 EU/mL (P < .05). Subsequent irrigation with EDTA (RCS3) failed to present a significant effectiveness in reducing the endotoxin levels (median= 0.37 EU/mL) (P = .07). However, intracanal medication for 30 days (RCS4) reduced endotoxins to median values of 0.03 EU/mL (P < .01). Conclusions: The present study revealed a strong association between the high levels of endotoxins found in AAAs and RCs collected from the same tooth. Moreover, the effectiveness of CMP in reducing the endotoxin levels from RCs in acute endodontic infection was improved by the use of RC medication. (J Endod 2014;40:177-181) A pical periodontitis is the result of an inflammatory response to bacteria and their
This study assessed the prevalence and microbial interactions of Fusobacterium nucleatum and Fuso... more This study assessed the prevalence and microbial interactions of Fusobacterium nucleatum and Fusobacterium necrophorum in primary endodontic infections from a Brazilian population and their antimicrobial susceptibility to some antibiotics by the E-test. One hundred ten samples from infected teeth with periapical pathologies were analyzed by culture methods. Five hundred eighty individual strains were isolated; 81.4% were strict anaerobes. F. nucleatum was found in 38 root canals and was associated with Porphyromonas gingivalis, Prevotella spp., and Eubacterium spp. F. necrophorum was found in 20 root canals and was associated with Peptostreptococcus prevotii. The simultaneous presence of F. nucleatum and F. necrophorum was not related to endodontic symptoms (p Ͼ 0.05). They were 100% susceptible to amoxicillin, amoxicillin/ clavulanate, and cephaclor. Fusobacterium spp. is frequently isolated from primary-infected root canals of teeth with periapical pathologies. Amoxicillin is a useful antibiotic against F. nucleatum and F. necrophorum in endodontic infections and has been prescribed as the first choice in Brazil. (J Endod 2008;34:1451-1456
Introduction: Procedural accidents have a negative effect on healing and might contribute to the ... more Introduction: Procedural accidents have a negative effect on healing and might contribute to the persistence of infections in inaccessible apical areas, requiring surgical intervention. This report describes a case of persistent apical periodontitis of a lower left first molar associated with the sinus tract and a periapical lesion that required nonsurgical endodontic retreatment and apical surgery for resolution. Methods: The tooth had received endodontic treatment 3 years ago and had to be retreated using the crown-down technique with chemical auxiliary substance (2% chlorhexidine gel), foramen patency, and enlargement and was filled in a single appointment. The occlusal access cavity was immediately restored with composite resin. After 1 month, it could be observed that the sinus tract persisted and, radiographically, the lesion remained unaltered. Therefore, endodontic microsurgery was indicated. Apical microsurgery was performed under magnification with the use of a dental operating microscope including apicectomy, root end with ultrasound, and sealing with mineral trioxide aggregate. A microbiological sample was collected from the apical lesion. The resected distal root apex was observed by scanning electron microscopy. Results: The following species were detected: Actinomyces naeslundii and Actinomyces meyeri, Propionibacterium propionicum, Clostridium botullinum, Parvimonas micra, and Bacteroides ureolyticus; scanning electron microscopic analysis revealed bacterial biofilm surrounding the apical foramen and external radicular surface. Gutta-percha overfilling at the apex because of a zip caused during initial endodontic treatment could be observed. A 6-month follow-up showed apparent radiographic periapical healing, which progressed after 24 months. Conclusion: Gram-positive anaerobic bacteria and extraradicular biofilm seem to participate in the maintenance of persistent periapical pathology, and endodontic retreatment followed by periapical microsurgery proved to be a successful alternative in the resolution of persistent extraradicular infections. (J Endod 2011;37:1696-1700
This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pu... more This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pulp necrosis and apical periodontitis before and after chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and to investigate the possible correlation of endotoxin and cultivable bacteria with the presence of clinical symptomatology. Twenty-four root canals were selected. Samples were collected before (s1) and after chemomechanical preparation (s2). Culture techniques were used to determine the colony-forming unit. A limulus amebocyte lysate (LAL) assay was used to quantify endotoxins (lipopolysaccharide, LPS). LPS and bacteria were detected in 100% of the initial samples (s1), with a median concentration of 139 endotoxin units/mL and 2.64 ϫ 10 5 colony-forming units/mL, respectively. Higher levels of LPS were found in teeth with clinical symptomatology (p Ͻ .05). At s2, mean endotoxin reduction of 59.99% and mean bacterial load reduction of 99.78% were found. Our findings indicated that chemomechanical preparation with 2.5% NaOCl was moderately effective against bacteria but less effective against endotoxins in root canal infection. Furthermore, a statistically significant association was found between higher levels and clinical symptomatology. (J Endod 2008;34:268 -272)
Introduction: This clinical study was conducted to compare the levels of endotoxins (lipopolysacc... more Introduction: This clinical study was conducted to compare the levels of endotoxins (lipopolysaccharides [LPSs]) found in primary and secondary endodontic infections with apical periodontitis by correlating LPS contents with clinical/radiographic findings. In addition, the presence of target gram-negative anaerobic bacteria was also investigated. Methods: Samples were taken from 15 root canals with primary infections and 15 with secondary infections by using paper points. The limulus amebocyte lysate assay was used to quantify endotoxins, and the polymerase chain reaction technique (16S rDNA) was used for bacterial investigation. Results: Endotoxins were detected in 100% of the root canal samples collected from primary (15/15) and secondary (15/15) infections with median values of 7.49 EU/mL and 3.96 EU/mL, respectively (P < .05). The median value of endotoxins found in the presence of clinical symptoms was significantly higher than in asymptomatic teeth with primary infections (P < .05). A positive correlation was found between endotoxin contents and a larger size of the radiolucent area (>3 mm) (P < .05). Prevotella nigrescens (10/15, 4/15), Fusobacterium nucleatum (5/15, 1/15), Treponema denticola (3/15, 1/15), and Treponema socranskii (5/15, 1/15) were detected in teeth with primary and secondary infections, respectively. P. endodontalis was present only in teeth with primary infections (5/15). Conclusions: Teeth with primary endodontic infections had higher contents of endotoxins and a more complex gram-negative bacterial community than teeth with secondary infections. Moreover, the levels of endotoxins were related to the severity of bone destruction in periapical tissues as well as the development of clinical features in teeth with primary infections. (J Endod 2012;38:1082-1086
Introduction: The aim of this study was to evaluate the influence of different irrigation protoco... more Introduction: The aim of this study was to evaluate the influence of different irrigation protocols on resin-based sealer bond strength to dentin and the associated failure pattern. Methods: One hundred eighty single-rooted premolars were used. Roots were divided into 18 groups according to the irrigation protocol used and instrumented using 5.25% NaOCl or 2% chlorhexidine (CHX) gel as chemical auxiliary substances and 17% EDTA or 37% phosphoric acid for smear layer removal. Finally, CHX solution was used as the final irrigant in some groups. Root canals were filled with either gutta-percha and AH Plus (Dentsply, Petropolis, RJ, Brazil) (GP/AH) or Resilon/Real Seal SE (SybronEndo, Orange, CA). Bond strength was measured by the push-out test. Additionally, failure patterns were classified as adhesive, cohesive, and mixed. Data were statistically analyzed by Kruskal-Wallis and Mann-Whitney U tests. Results: In the GP/AH groups, high bond strength values were obtained when NaOCl was associated with phosphoric acid with (6.9 AE 3.9) and without (7.4 AE 3.8) a final flush with CHX. However, when CHX gel was used, the association with EDTA with (7.5 AE 3.2) and without (6.0 AE 2.3) a final flush with CHX solution showed better results. The GP/AH groups mainly showed a cohesive failure pattern. Evaluation of Resilon/Real Seal SE showed that higher bond strength values were reached when CHX was associated with phosphoric acid with (5.2 AE 3.0) and without (4.2 AE 2.0) a final flush with CHX solution. The failure patterns were mainly adhesive. Conclusions: Irrigation protocols influenced the bond strength of resin sealers to dentin. (J Endod 2013;39:689-692)
Introduction: This clinical study was conducted to compare the efficacy of chemomechanical prepar... more Introduction: This clinical study was conducted to compare the efficacy of chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and 2% chlorhexidine (CHX) gel on eliminating oral bacterial lipopolysaccharide (LPS) in teeth with pulp necrosis and apical periodontitis. Methods: Fifty-four root canals were selected. Samples were collect before (s1) and after chemomechanical preparation (s2). Teeth were randomly divided into groups: GI, 2.5% NaOCl (n = 27) and GII, CHX gel (n = 27). Limulus amebocyte lysate assay was used to quantify endotoxins. Results: Endotoxin was present in 100% of the samples investigated, with a median value of 272 endotoxin units (EU)/mL (GI) and of 152.46 EU/mL (GII). As a result of chemomechanical preparation, LPS content was reduced to a median value of 86 EU/mL (GI) and 85 EU/mL (GII). Higher percentage value of endotoxin reduction was found in GI (P < .05). Conclusions: The 2.5% NaOCl and 2% CHX gel were not effective in eliminating endotoxin from the primarily infected root canals. (J Endod 2009;35:1350-1353
Introduction: Acute primary endodontic infections are polymicrobial infections that affect both t... more Introduction: Acute primary endodontic infections are polymicrobial infections that affect both the root canal (RC) system and apical tissues. It is known that these communities cannot be detected by conventional culturing methods. The aim of this study was to examine the profile of microbial communities in necrotic RCs and acute apical abscesses (AAAs) using an open-ended molecular approach to compare the diversity and composition of the microbiota of these two communities. Methods: Paired samples of RC and PA exudates were collected from 20 subjects and analyzed by terminal restriction fragment length polymorphism (t-RFLP). The number of peaks, the peak areas, and the community diversity were compared between RCs and PAs. The similarity of the microbial profile of each pair of RCs and PAs was assessed by computing the number of shared peaks and the Bray-Curtis Similarity Index. Results: A total of 103 and 75.5 unique fragments (t-RFs) were detected in RC and PA samples, respectively. RCs and PAs were not different in the number of species or in the community diversity; however, very few species were shared between RC and PA samples. No single t-RF fragment was detected in all samples, and the majority was detected in only one sample. Low diversity of species was observed in the RCs of smokers. Subjects with previous pain showed fewer species and greater community diversity. Conclusion: The microbial profiles of the RC and PA communities are distinct and diverged between all subjects, suggesting that acute endodontic infections are microbiologically heterogeneous. (J Endod
This study examined similarities in the microbiology of initial, and subsequent, infections in di... more This study examined similarities in the microbiology of initial, and subsequent, infections in different dental root canals in the same patient. In the initial infections, 61.5% of root canals had anaerobes present compared with 54% of root canals treated subsequently. Commercial identification kits were employed in speciation of isolates. Both sets of dental root canals had in common Peptostreptococcus micros, Eubacterium lentum and Veillonella parvula as species very commonly isolated. In addition Prevotella spp. tended to be present in both initial and subsequently infected teeth in the same patient (P=0.01); this was true of Gram-negative bacteria in general (P=0.03). A significant association was observed between the type of tooth infected at an initial and subsequent infection in the same mouth (P=0.01). In conclusion, there is an association between bacteria isolated from initial endodontically infected teeth and subsequent infected root canals in a patient.
This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pu... more This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pulp necrosis and apical periodontitis before and after chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and to investigate the possible correlation of endotoxin and cultivable bacteria with the presence of clinical symptomatology. Twenty-four root canals were selected. Samples were collected before (s1) and after chemomechanical preparation (s2). Culture techniques were used to determine the colony-forming unit. A limulus amebocyte lysate (LAL) assay was used to quantify endotoxins (lipopolysaccharide, LPS). LPS and bacteria were detected in 100% of the initial samples (s1), with a median concentration of 139 endotoxin units/mL and 2.64 ϫ 10 5 colony-forming units/mL, respectively. Higher levels of LPS were found in teeth with clinical symptomatology (p Ͻ .05). At s2, mean endotoxin reduction of 59.99% and mean bacterial load reduction of 99.78% were found. Our findings indicated that chemomechanical preparation with 2.5% NaOCl was moderately effective against bacteria but less effective against endotoxins in root canal infection. Furthermore, a statistically significant association was found between higher levels and clinical symptomatology. (J Endod 2008;34:268 -272)
Procedural accidents have a negative effect on healing and might contribute to the persistence of... more Procedural accidents have a negative effect on healing and might contribute to the persistence of infections in inaccessible apical areas, requiring surgical intervention. This report describes a case of persistent apical periodontitis of a lower left first molar associated with the sinus tract and a periapical lesion that required nonsurgical endodontic retreatment and apical surgery for resolution.The tooth had received endodontic treatment 3 years ago and had to be retreated using the crown-down technique with chemical auxiliary substance (2% chlorhexidine gel), foramen patency, and enlargement and was filled in a single appointment. The occlusal access cavity was immediately restored with composite resin. After 1 month, it could be observed that the sinus tract persisted and, radiographically, the lesion remained unaltered. Therefore, endodontic microsurgery was indicated. Apical microsurgery was performed under magnification with the use of a dental operating microscope including apicectomy, root end with ultrasound, and sealing with mineral trioxide aggregate. A microbiological sample was collected from the apical lesion. The resected distal root apex was observed by scanning electron microscopy.The following species were detected: Actinomyces naeslundii and Actinomyces meyeri, Propionibacterium propionicum, Clostridium botullinum, Parvimonas micra, and Bacteroides ureolyticus; scanning electron microscopic analysis revealed bacterial biofilm surrounding the apical foramen and external radicular surface. Gutta-percha overfilling at the apex because of a zip caused during initial endodontic treatment could be observed. A 6-month follow-up showed apparent radiographic periapical healing, which progressed after 24 months.Gram-positive anaerobic bacteria and extraradicular biofilm seem to participate in the maintenance of persistent periapical pathology, and endodontic retreatment followed by periapical microsurgery proved to be a successful alternative in the resolution of persistent extraradicular infections.
The aim of this work was to characterize the by-products formed in the associations between the m... more The aim of this work was to characterize the by-products formed in the associations between the most commonly used irrigants in endodontic practice through electrospray ionization quadrupole time-of-flight mass spectrometry analyses. Sodium hypochlorite (NaOCl) (0.16%, 1%, 2.5%, and 5.25%) was associated with 2% chlorhexidine (CHX) solution and gel, 17% EDTA, 10% citric acid, 37% phosphoric acid, saline solution, ethanol, and distilled water. CHX solution and gel were also associated with all above mentioned irrigants. The solutions were mixed in a 1:1 ratio, and electrospray ionization quadrupole time-of-flight mass spectrometry was used to characterize the precipitates when formed. CHX produced an orange-brown precipitate when associated with NaOCl from 1%-5.25% and an orange-white precipitate when associated with 0.16% NaOCl. When associated with EDTA, CHX produced a white milky precipitate, and when associated with saline solution and ethanol, a salt precipitation was produced. No precipitation was observed when CHX was associated with citric acid, phosphoric acid, or distilled water. In the NaOCl associations, precipitation occurred only when CHX was present. The orange-brown precipitate observed in the association between CHX and NaOCl occurs because of the presence of NaOCl, an oxidizing agent causing chlorination of the guanidino nitrogens of the CHX. The precipitates formed in the reaction of CHX with EDTA, saline solution, and ethanol were associated with acid-base reactions, salting-out process, and lower solubility, respectively. NaOCl associated with EDTA, citric acid, and phosphoric acid leads mainly to chlorine gas formation. Intermediate flushes with distilled water seem to be appropriate to prevent or at least reduce formation of by-products.
Three solutions of EDTA-a 15% concentration of the alkaline salt, a 15% concentration of the acid... more Three solutions of EDTA-a 15% concentration of the alkaline salt, a 15% concentration of the acid salt, and a 25% concentration of the alkaline saltwere evaluated for smear layer removal in root canal systems. All solutions were adjusted to pH 7.1 using either NaOH or HCl. When the EDTA solutions were alternately used for root canal irrigation with 5.25% NaOCl, they completely removed the smear layer in the middle and coronal thirds of canal preparations, but were less effective in the apical third. None of the EDTA solutions by themselves were effective at completely removing the smear layer at any level. The alkaline tetrasodium salt, pH adjusted with HCl, is more cost effective and performed equally as well as the more commonly used disodium salt.
Endodontic therapy is indicated for cases of traumatic tooth dislocation associated with pulp nec... more Endodontic therapy is indicated for cases of traumatic tooth dislocation associated with pulp necrosis and/or inflammatory resorption. Here we describe the management of a maxillary left lateral permanent incisor that suffered lateral luxation, leading to pulp necrosis and root resorption, in a 13-year-old boy. The traumatized tooth was treated successfully by intracanal medication with calcium hydroxide, 2% chlorhexidine gel and zinc oxide for 12 months without any need to change the dressing, followed by conventional root canal filling. The postoperative course was uneventful and a stable clinical outcome was obtained with evidence of periapical lesion repair and stabilization of the resorption process.
Introduction: Macrophages are highly activated by endodontic contents. This study investigated th... more Introduction: Macrophages are highly activated by endodontic contents. This study investigated the correlation between different clinical signs/symptoms and radiographic features according to the levels of interleukin (IL)-1b, tumor necrosis factor a (TNF-a), IL-6, IL-10, prostaglandin E 2 (PGE 2 ), and their networks produced by endodontic content-stimulated macrophages collected from primary endodontic infection with apical periodontitis (PEIAP). Methods: Samples were taken from 21 root canals with PEIAP by using paper points. The presence of exudate (EX), pain on palpation (POP), tenderness to percussion (TTP), and the size of the radiographic lesion (SRL) were recorded. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate (LAL) assay for endotoxin measurement. Raw 264.7 macrophages were stimulated with bacterial contents during 24 hs. The amounts of IL-1b, TNF-a, IL-6, IL-10 and PGE 2 were measured by enzyme-linked immunosorbent assay. Log-based data were correlated by multiple logistic regression (P < .05). Results: Bacteria and endotoxin were detected in 100% of the samples. IL-6 and TNF-a were positively correlated with SRL and EX, respectively (P < .05). Clinical signs/symptoms and radiographic findings were set as dependent variables for EX-positive correlations between PGE 2 , IL-1b, and TNF-a (P < .05), whereas IL-6 and PGE 2 were positively correlated to each other in POP but negatively correlated in SRL (P < .05). When POP and TTP-POP were set as dependent variables, different cytokine networks were found. Conclusions: Our findings suggest different roles for each cytokine in the development of apical periodontitis, whose effects of overlapping networks depend on the signs/symptoms and radiographic features found in endodontic infection. (J Endod 2012;38:740-745)
Introduction: Periradicular tissue chronic stimulation by endotoxin may cause apical periodontiti... more Introduction: Periradicular tissue chronic stimulation by endotoxin may cause apical periodontitis. This study investigated the microbial profile and the levels of endotoxin found in primary root canal infection with apical periodontitis, determined their antigenicity against macrophages through the levels of interleukin (IL)-1b and tumor necrosis factor a (TNF-a), and evaluated their relationship with clinical and radiographic findings. Methods: Samples were taken from 21 root canals with primary endodontic infection and apical periodontitis with paper points. PCR technique (16S rDNA) was used for the detection of the target bacteria. Limulus Amebocyte Lysate (LAL) was used to measure endotoxin. The amounts of IL-1ß/ TNF-alpha in macrophages supernatants were measured by enzyme-linked immunosorbent assay -Duoset-kit (ELISA). Results: Prevotella nigrescens (13/21), Porphyromonas endodontalis (6/21), and Treponema socranskii (6/21) were the most frequently detected gram-negative bacterial species. The presence of the sinus tract (2/21) was related to the detection of Filifactor alocis (p < 0.05), whereas a tooth with a radiolucent area $2 mm was related to the detection of Treponema denticola. A correlation was found between the number of gram-negative bacteria and the levels of IL-1b/TNF-a (p < 0.05). Increased levels of endotoxin were followed by TNF-a release (p < 0.05). Higher levels of IL-1b (p < 0.05) and endotoxin contents were related to the larger size of the radiolucent area. Conclusion: The antigenicity of the endodontic contents is not only related to the amount of endotoxin found in the root canal but also to the number of different species of gram-negative bacteria involved in the infection. Moreover, a larger size ($2 mm) of the radiolucent area was related
Introduction: Root canal contents are potent stimuli for proinflammatory cytokines involved in ap... more Introduction: Root canal contents are potent stimuli for proinflammatory cytokines involved in apical periodontitis. This study investigated target gram-negative bacterial species and endotoxins in primary endodontic infection with apical periodontitis, determined their antigenicity against macrophages through the levels of PGE 2 , and evaluated their relationship with clinical findings. Methods: Samples were taken from 21 root canals with primary infection and apical periodontitis by using paper points. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate assay for endotoxin measurement. Levels of prostaglandin E2 (PGE 2 ) were measured by enzyme-linked immunosorbent assay (Duoset Kit; R&D, Minneapolis, MN). Results: Prevotella nigrescens (13/21), Fusobacterium nucleatum (6/21), and Porphyromonas endodontalis (6/21) were the most frequently observed species. A positive association was found between F. nucleatum and P. endodontalis (P < .05). A correlation was found between the number of gram-negative bacterial species and the levels of endotoxins, as well as PGE 2 (P < .05). Higher levels of endotoxin were detected in teeth with exudation, whereas elevated levels of PGE 2 were found in teeth with tenderness to percussion and pain on palpation. Conclusions: Our findings imply an additive effect between the number of gram-negative bacterial species involved in endodontic infection regarding the induction of proinflammatory cytokine by macrophage cells. Moreover, teeth with clinical symptomatology were related to higher levels of endotoxins and PGE 2 secretion. (J Endod 2011;37:602-607)
Introduction: Although Enterococcus faecalis is a member of the normal microbiota, it is also a m... more Introduction: Although Enterococcus faecalis is a member of the normal microbiota, it is also a major cause of nosocomial infections. Some strains of E. faecalis produce capsule, which contributes to pathogenesis through evasion of host defenses, and its production is dependent on the capsule (cps) operon polymorphism. This study investigated cps locus polymorphism in distinct lineages of E. faecalis isolated from canals of root-filled teeth with periapical lesions. Methods: Twenty-two E. faecalis isolates were evaluated regarding the cps operon polymorphism and genetic diversity. The 3 known CPS types were determined by polymerase chain reaction. This information was correlated with multilocus sequence typing data, which were used to define genetic lineages. Results: cpsA and cpsB were the only detected genes within the cps operon in 62.5% of E. faecalis strains (14/22), indicative of genotype CPS 1, which lacks capsule expression. The essential genes in the cps operon for capsule production were detected in the remaining strains, whereas 3 belonged to genotype CPS 5 and 5 strains to genotype CPS 2. A total of 14 sequence types (STs) were resolved in 22 E. faecalis isolates. Comparison with the E. faecalis international multilocus sequence typing database revealed that 9 STs were previously found, and that the 5 STs were novel. Conclusions: Certain E. faecalis genotypes from canals of root-filled teeth with periapical lesions belong to lineages associated with capsule expression and production of multiple virulence factors, which might account for their increased pathogenic potential. (J Endod 2012;38:58-61)
Introduction: This clinical study was conducted to measure the endotoxin levels in infected root ... more Introduction: This clinical study was conducted to measure the endotoxin levels in infected root canals (RCs) and exudates related to acute apical abscesses (AAAs). In addition, the effectiveness of RC procedures in reducing the endotoxin levels in RCs was monitored. Methods: Paired samples of infected RCs and exudates from AAAs were collected from 10 subjects by using paper points. RCs samples were collected before (RCS1) and after chemomechanical preparation (CMP) (RCS2), after 17% EDTA (RCS3), and after 30 days of intracanal medication (Ca[OH] 2 + chlorhexidine) (RCS4). A turbidimetric kinetic limulus amebocyte lysate assay was used for the measurement of endotoxins. Results: Endotoxins were detected in 100% of the baseline samples of AAAs and RCs (RCS1) with median values of 175 EU/mL and 41.5 EU/mL, respectively (P < .05). After CMP (RCS2), endotoxins were reduced to a median value of 0.54 EU/mL (P < .05). Subsequent irrigation with EDTA (RCS3) failed to present a significant effectiveness in reducing the endotoxin levels (median= 0.37 EU/mL) (P = .07). However, intracanal medication for 30 days (RCS4) reduced endotoxins to median values of 0.03 EU/mL (P < .01). Conclusions: The present study revealed a strong association between the high levels of endotoxins found in AAAs and RCs collected from the same tooth. Moreover, the effectiveness of CMP in reducing the endotoxin levels from RCs in acute endodontic infection was improved by the use of RC medication. (J Endod 2014;40:177-181) A pical periodontitis is the result of an inflammatory response to bacteria and their
This study assessed the prevalence and microbial interactions of Fusobacterium nucleatum and Fuso... more This study assessed the prevalence and microbial interactions of Fusobacterium nucleatum and Fusobacterium necrophorum in primary endodontic infections from a Brazilian population and their antimicrobial susceptibility to some antibiotics by the E-test. One hundred ten samples from infected teeth with periapical pathologies were analyzed by culture methods. Five hundred eighty individual strains were isolated; 81.4% were strict anaerobes. F. nucleatum was found in 38 root canals and was associated with Porphyromonas gingivalis, Prevotella spp., and Eubacterium spp. F. necrophorum was found in 20 root canals and was associated with Peptostreptococcus prevotii. The simultaneous presence of F. nucleatum and F. necrophorum was not related to endodontic symptoms (p Ͼ 0.05). They were 100% susceptible to amoxicillin, amoxicillin/ clavulanate, and cephaclor. Fusobacterium spp. is frequently isolated from primary-infected root canals of teeth with periapical pathologies. Amoxicillin is a useful antibiotic against F. nucleatum and F. necrophorum in endodontic infections and has been prescribed as the first choice in Brazil. (J Endod 2008;34:1451-1456
Introduction: Procedural accidents have a negative effect on healing and might contribute to the ... more Introduction: Procedural accidents have a negative effect on healing and might contribute to the persistence of infections in inaccessible apical areas, requiring surgical intervention. This report describes a case of persistent apical periodontitis of a lower left first molar associated with the sinus tract and a periapical lesion that required nonsurgical endodontic retreatment and apical surgery for resolution. Methods: The tooth had received endodontic treatment 3 years ago and had to be retreated using the crown-down technique with chemical auxiliary substance (2% chlorhexidine gel), foramen patency, and enlargement and was filled in a single appointment. The occlusal access cavity was immediately restored with composite resin. After 1 month, it could be observed that the sinus tract persisted and, radiographically, the lesion remained unaltered. Therefore, endodontic microsurgery was indicated. Apical microsurgery was performed under magnification with the use of a dental operating microscope including apicectomy, root end with ultrasound, and sealing with mineral trioxide aggregate. A microbiological sample was collected from the apical lesion. The resected distal root apex was observed by scanning electron microscopy. Results: The following species were detected: Actinomyces naeslundii and Actinomyces meyeri, Propionibacterium propionicum, Clostridium botullinum, Parvimonas micra, and Bacteroides ureolyticus; scanning electron microscopic analysis revealed bacterial biofilm surrounding the apical foramen and external radicular surface. Gutta-percha overfilling at the apex because of a zip caused during initial endodontic treatment could be observed. A 6-month follow-up showed apparent radiographic periapical healing, which progressed after 24 months. Conclusion: Gram-positive anaerobic bacteria and extraradicular biofilm seem to participate in the maintenance of persistent periapical pathology, and endodontic retreatment followed by periapical microsurgery proved to be a successful alternative in the resolution of persistent extraradicular infections. (J Endod 2011;37:1696-1700
This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pu... more This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pulp necrosis and apical periodontitis before and after chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and to investigate the possible correlation of endotoxin and cultivable bacteria with the presence of clinical symptomatology. Twenty-four root canals were selected. Samples were collected before (s1) and after chemomechanical preparation (s2). Culture techniques were used to determine the colony-forming unit. A limulus amebocyte lysate (LAL) assay was used to quantify endotoxins (lipopolysaccharide, LPS). LPS and bacteria were detected in 100% of the initial samples (s1), with a median concentration of 139 endotoxin units/mL and 2.64 ϫ 10 5 colony-forming units/mL, respectively. Higher levels of LPS were found in teeth with clinical symptomatology (p Ͻ .05). At s2, mean endotoxin reduction of 59.99% and mean bacterial load reduction of 99.78% were found. Our findings indicated that chemomechanical preparation with 2.5% NaOCl was moderately effective against bacteria but less effective against endotoxins in root canal infection. Furthermore, a statistically significant association was found between higher levels and clinical symptomatology. (J Endod 2008;34:268 -272)
Introduction: This clinical study was conducted to compare the levels of endotoxins (lipopolysacc... more Introduction: This clinical study was conducted to compare the levels of endotoxins (lipopolysaccharides [LPSs]) found in primary and secondary endodontic infections with apical periodontitis by correlating LPS contents with clinical/radiographic findings. In addition, the presence of target gram-negative anaerobic bacteria was also investigated. Methods: Samples were taken from 15 root canals with primary infections and 15 with secondary infections by using paper points. The limulus amebocyte lysate assay was used to quantify endotoxins, and the polymerase chain reaction technique (16S rDNA) was used for bacterial investigation. Results: Endotoxins were detected in 100% of the root canal samples collected from primary (15/15) and secondary (15/15) infections with median values of 7.49 EU/mL and 3.96 EU/mL, respectively (P < .05). The median value of endotoxins found in the presence of clinical symptoms was significantly higher than in asymptomatic teeth with primary infections (P < .05). A positive correlation was found between endotoxin contents and a larger size of the radiolucent area (>3 mm) (P < .05). Prevotella nigrescens (10/15, 4/15), Fusobacterium nucleatum (5/15, 1/15), Treponema denticola (3/15, 1/15), and Treponema socranskii (5/15, 1/15) were detected in teeth with primary and secondary infections, respectively. P. endodontalis was present only in teeth with primary infections (5/15). Conclusions: Teeth with primary endodontic infections had higher contents of endotoxins and a more complex gram-negative bacterial community than teeth with secondary infections. Moreover, the levels of endotoxins were related to the severity of bone destruction in periapical tissues as well as the development of clinical features in teeth with primary infections. (J Endod 2012;38:1082-1086
Introduction: The aim of this study was to evaluate the influence of different irrigation protoco... more Introduction: The aim of this study was to evaluate the influence of different irrigation protocols on resin-based sealer bond strength to dentin and the associated failure pattern. Methods: One hundred eighty single-rooted premolars were used. Roots were divided into 18 groups according to the irrigation protocol used and instrumented using 5.25% NaOCl or 2% chlorhexidine (CHX) gel as chemical auxiliary substances and 17% EDTA or 37% phosphoric acid for smear layer removal. Finally, CHX solution was used as the final irrigant in some groups. Root canals were filled with either gutta-percha and AH Plus (Dentsply, Petropolis, RJ, Brazil) (GP/AH) or Resilon/Real Seal SE (SybronEndo, Orange, CA). Bond strength was measured by the push-out test. Additionally, failure patterns were classified as adhesive, cohesive, and mixed. Data were statistically analyzed by Kruskal-Wallis and Mann-Whitney U tests. Results: In the GP/AH groups, high bond strength values were obtained when NaOCl was associated with phosphoric acid with (6.9 AE 3.9) and without (7.4 AE 3.8) a final flush with CHX. However, when CHX gel was used, the association with EDTA with (7.5 AE 3.2) and without (6.0 AE 2.3) a final flush with CHX solution showed better results. The GP/AH groups mainly showed a cohesive failure pattern. Evaluation of Resilon/Real Seal SE showed that higher bond strength values were reached when CHX was associated with phosphoric acid with (5.2 AE 3.0) and without (4.2 AE 2.0) a final flush with CHX solution. The failure patterns were mainly adhesive. Conclusions: Irrigation protocols influenced the bond strength of resin sealers to dentin. (J Endod 2013;39:689-692)
Introduction: This clinical study was conducted to compare the efficacy of chemomechanical prepar... more Introduction: This clinical study was conducted to compare the efficacy of chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and 2% chlorhexidine (CHX) gel on eliminating oral bacterial lipopolysaccharide (LPS) in teeth with pulp necrosis and apical periodontitis. Methods: Fifty-four root canals were selected. Samples were collect before (s1) and after chemomechanical preparation (s2). Teeth were randomly divided into groups: GI, 2.5% NaOCl (n = 27) and GII, CHX gel (n = 27). Limulus amebocyte lysate assay was used to quantify endotoxins. Results: Endotoxin was present in 100% of the samples investigated, with a median value of 272 endotoxin units (EU)/mL (GI) and of 152.46 EU/mL (GII). As a result of chemomechanical preparation, LPS content was reduced to a median value of 86 EU/mL (GI) and 85 EU/mL (GII). Higher percentage value of endotoxin reduction was found in GI (P < .05). Conclusions: The 2.5% NaOCl and 2% CHX gel were not effective in eliminating endotoxin from the primarily infected root canals. (J Endod 2009;35:1350-1353
Introduction: Acute primary endodontic infections are polymicrobial infections that affect both t... more Introduction: Acute primary endodontic infections are polymicrobial infections that affect both the root canal (RC) system and apical tissues. It is known that these communities cannot be detected by conventional culturing methods. The aim of this study was to examine the profile of microbial communities in necrotic RCs and acute apical abscesses (AAAs) using an open-ended molecular approach to compare the diversity and composition of the microbiota of these two communities. Methods: Paired samples of RC and PA exudates were collected from 20 subjects and analyzed by terminal restriction fragment length polymorphism (t-RFLP). The number of peaks, the peak areas, and the community diversity were compared between RCs and PAs. The similarity of the microbial profile of each pair of RCs and PAs was assessed by computing the number of shared peaks and the Bray-Curtis Similarity Index. Results: A total of 103 and 75.5 unique fragments (t-RFs) were detected in RC and PA samples, respectively. RCs and PAs were not different in the number of species or in the community diversity; however, very few species were shared between RC and PA samples. No single t-RF fragment was detected in all samples, and the majority was detected in only one sample. Low diversity of species was observed in the RCs of smokers. Subjects with previous pain showed fewer species and greater community diversity. Conclusion: The microbial profiles of the RC and PA communities are distinct and diverged between all subjects, suggesting that acute endodontic infections are microbiologically heterogeneous. (J Endod
This study examined similarities in the microbiology of initial, and subsequent, infections in di... more This study examined similarities in the microbiology of initial, and subsequent, infections in different dental root canals in the same patient. In the initial infections, 61.5% of root canals had anaerobes present compared with 54% of root canals treated subsequently. Commercial identification kits were employed in speciation of isolates. Both sets of dental root canals had in common Peptostreptococcus micros, Eubacterium lentum and Veillonella parvula as species very commonly isolated. In addition Prevotella spp. tended to be present in both initial and subsequently infected teeth in the same patient (P=0.01); this was true of Gram-negative bacteria in general (P=0.03). A significant association was observed between the type of tooth infected at an initial and subsequent infection in the same mouth (P=0.01). In conclusion, there is an association between bacteria isolated from initial endodontically infected teeth and subsequent infected root canals in a patient.
This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pu... more This clinical study was conducted to quantify endotoxins and cultivable bacteria in teeth with pulp necrosis and apical periodontitis before and after chemomechanical preparation with 2.5% sodium hypochlorite (NaOCl) and to investigate the possible correlation of endotoxin and cultivable bacteria with the presence of clinical symptomatology. Twenty-four root canals were selected. Samples were collected before (s1) and after chemomechanical preparation (s2). Culture techniques were used to determine the colony-forming unit. A limulus amebocyte lysate (LAL) assay was used to quantify endotoxins (lipopolysaccharide, LPS). LPS and bacteria were detected in 100% of the initial samples (s1), with a median concentration of 139 endotoxin units/mL and 2.64 ϫ 10 5 colony-forming units/mL, respectively. Higher levels of LPS were found in teeth with clinical symptomatology (p Ͻ .05). At s2, mean endotoxin reduction of 59.99% and mean bacterial load reduction of 99.78% were found. Our findings indicated that chemomechanical preparation with 2.5% NaOCl was moderately effective against bacteria but less effective against endotoxins in root canal infection. Furthermore, a statistically significant association was found between higher levels and clinical symptomatology. (J Endod 2008;34:268 -272)
Procedural accidents have a negative effect on healing and might contribute to the persistence of... more Procedural accidents have a negative effect on healing and might contribute to the persistence of infections in inaccessible apical areas, requiring surgical intervention. This report describes a case of persistent apical periodontitis of a lower left first molar associated with the sinus tract and a periapical lesion that required nonsurgical endodontic retreatment and apical surgery for resolution.The tooth had received endodontic treatment 3 years ago and had to be retreated using the crown-down technique with chemical auxiliary substance (2% chlorhexidine gel), foramen patency, and enlargement and was filled in a single appointment. The occlusal access cavity was immediately restored with composite resin. After 1 month, it could be observed that the sinus tract persisted and, radiographically, the lesion remained unaltered. Therefore, endodontic microsurgery was indicated. Apical microsurgery was performed under magnification with the use of a dental operating microscope including apicectomy, root end with ultrasound, and sealing with mineral trioxide aggregate. A microbiological sample was collected from the apical lesion. The resected distal root apex was observed by scanning electron microscopy.The following species were detected: Actinomyces naeslundii and Actinomyces meyeri, Propionibacterium propionicum, Clostridium botullinum, Parvimonas micra, and Bacteroides ureolyticus; scanning electron microscopic analysis revealed bacterial biofilm surrounding the apical foramen and external radicular surface. Gutta-percha overfilling at the apex because of a zip caused during initial endodontic treatment could be observed. A 6-month follow-up showed apparent radiographic periapical healing, which progressed after 24 months.Gram-positive anaerobic bacteria and extraradicular biofilm seem to participate in the maintenance of persistent periapical pathology, and endodontic retreatment followed by periapical microsurgery proved to be a successful alternative in the resolution of persistent extraradicular infections.
The aim of this work was to characterize the by-products formed in the associations between the m... more The aim of this work was to characterize the by-products formed in the associations between the most commonly used irrigants in endodontic practice through electrospray ionization quadrupole time-of-flight mass spectrometry analyses. Sodium hypochlorite (NaOCl) (0.16%, 1%, 2.5%, and 5.25%) was associated with 2% chlorhexidine (CHX) solution and gel, 17% EDTA, 10% citric acid, 37% phosphoric acid, saline solution, ethanol, and distilled water. CHX solution and gel were also associated with all above mentioned irrigants. The solutions were mixed in a 1:1 ratio, and electrospray ionization quadrupole time-of-flight mass spectrometry was used to characterize the precipitates when formed. CHX produced an orange-brown precipitate when associated with NaOCl from 1%-5.25% and an orange-white precipitate when associated with 0.16% NaOCl. When associated with EDTA, CHX produced a white milky precipitate, and when associated with saline solution and ethanol, a salt precipitation was produced. No precipitation was observed when CHX was associated with citric acid, phosphoric acid, or distilled water. In the NaOCl associations, precipitation occurred only when CHX was present. The orange-brown precipitate observed in the association between CHX and NaOCl occurs because of the presence of NaOCl, an oxidizing agent causing chlorination of the guanidino nitrogens of the CHX. The precipitates formed in the reaction of CHX with EDTA, saline solution, and ethanol were associated with acid-base reactions, salting-out process, and lower solubility, respectively. NaOCl associated with EDTA, citric acid, and phosphoric acid leads mainly to chlorine gas formation. Intermediate flushes with distilled water seem to be appropriate to prevent or at least reduce formation of by-products.
Three solutions of EDTA-a 15% concentration of the alkaline salt, a 15% concentration of the acid... more Three solutions of EDTA-a 15% concentration of the alkaline salt, a 15% concentration of the acid salt, and a 25% concentration of the alkaline saltwere evaluated for smear layer removal in root canal systems. All solutions were adjusted to pH 7.1 using either NaOH or HCl. When the EDTA solutions were alternately used for root canal irrigation with 5.25% NaOCl, they completely removed the smear layer in the middle and coronal thirds of canal preparations, but were less effective in the apical third. None of the EDTA solutions by themselves were effective at completely removing the smear layer at any level. The alkaline tetrasodium salt, pH adjusted with HCl, is more cost effective and performed equally as well as the more commonly used disodium salt.
Endodontic therapy is indicated for cases of traumatic tooth dislocation associated with pulp nec... more Endodontic therapy is indicated for cases of traumatic tooth dislocation associated with pulp necrosis and/or inflammatory resorption. Here we describe the management of a maxillary left lateral permanent incisor that suffered lateral luxation, leading to pulp necrosis and root resorption, in a 13-year-old boy. The traumatized tooth was treated successfully by intracanal medication with calcium hydroxide, 2% chlorhexidine gel and zinc oxide for 12 months without any need to change the dressing, followed by conventional root canal filling. The postoperative course was uneventful and a stable clinical outcome was obtained with evidence of periapical lesion repair and stabilization of the resorption process.
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Papers by Brenda Gomes