Resumo O objetivo deste estudo foi avaliar in vitro o efeito do hidróxido de cálcio [Ca(OH)2], clorexidina gel a 2% (CHX) em macrófagos (RAW 264.7) na produção de citocinas pró-inflamatórias e óxido nítrico após pré-tratamento com ácido lipoteicóico (LTA) de Enterococcus faecalis. Quarenta e oito dentes humanos uniradiculares foram instrumentados com R25.08 (RECIPROC) e esterilizados por irradiação gama. LTA foi inoculado no canal radicular de cada espécime por 96 horas. Os espécimes foram instrumentados com 40.06 e 50.05 (RECIPROC) e medicados com: I) solução salina apirogênica (SS); II) CHX 2%; III) Ca(OH)2 + SS; ou IV) Ca(OH)2 + CHX durante 14 dias. Três amostras (S) foram realizadas do canal radicular de cada espécime em: S1) imediatamente após a instrumentação; S2) após ácido etilenodiaminotetracético (EDTA); S3) após a retirada da medicação intracanal. Foi realizada a quantificação subsequente de citocinas (IL-1β, TNF-α, MIP-1α, IP-10, G-CSF e IL-6) por imunoensaio (ELISA) e de óxido nítrico pelo método de Griess. Os dados foram submetidos a um teste de normalidade e então analisados com ANOVA one-way e teste de Tukey com nível de significância de 5% usando GraphPad Prism 6. Ca(OH)2 + SS e Ca(OH)2 + CHX apresentaram níveis mais baixos de TNF -α, TNF-α, IL-6, G-CSF e óxido nítrico. Ca(OH)2 + SS foi o mais eficaz na redução de MIP-1α. CHX foi eficaz na redução de IL-6 e G-CSF. Sendo assim a associação de hidróxido de cálcio e clorexidina é eficaz na redução das citocinas TNF-α, IL-1β, IL-6, G-CSF e óxido nítrico.

Abstract The aim of this study was to evaluate in vitro the effect of calcium hydroxide [Ca(OH)2], 2% chlorhexidine gel (CHX) on macrophages (RAW 264.7) to produce pro-inflammatory cytokines and nitric oxide after pretreatment with lipoteichoic acid (LTA) of Enterococcus faecalis. Forty-eight human single-rooted teeth were instrumented with R25.08 (RECIPROC) and sterilized by gamma irradiation. LTA was inoculated in the root canal of each specimen for 96 hours. Specimens were instrumented with 40.06 and 50.05 (RECIPROC) and medicated with: I) Pyrogen-free saline solution (SS); II) 2% CHX gel; III) Ca(OH)2 + SS; or IV) Ca(OH)2 + CHX for 14 days. Three samples (S) were performed of the root canal of each specimen at: S1) immediately after instrumentation; S2) after Ethylenediaminetetraacetic acid (EDTA); S3) after intracanal medication removal. Subsequent quantification of cytokines (IL-1β, TNF-α, MIP-1α, IP-10, G-CSF and IL-6) by immunosorbent assay (ELISA) and nitric oxide by the Griess method was carried-out. Data were submitted to a normality test and then analyzed with one-way ANOVA and Tukey test with a significance level of 5% using GraphPad Prism 6. Ca(OH)2 + SS and Ca(OH)2 + CHX presented lower levels of TNF-α, TNF-α, IL-6, G-CSF and nitric oxide. Ca(OH)2 + SS was the most effective in reducing MIP-1α. CHX was effective in reducing IL-6 and G-CSF. Therefore, the combined intracanal medication of calcium hydroxide and chlorhexidine is effective in reducing the cytokines TNF-α, IL-1β, IL-6, G-CSF and nitric oxide.

Resumo O objetivo deste estudo foi avaliar o hipoclorito de sódio (NaOCl), água de cal (LW) e polimixina B (PMB) como soluções irrigadoras sobre MMP-3, MMP-8 e MMP-9. Trinta e três pacientes com periodontite apical de dentes unirradiculares foram tratados de acordo com três grupos experimentais (n= 11): grupo 1: 2,5% NaOCl foi usado como solução irrigadora; grupo-2: NaOCl a 2,5% para as duas primeiras limas e LW: [0,14% Ca(OH)2] para as duas últimas limas; group-3: 2.5% NaOCl para as duas primeiras limas e PMB para as duas últimas limas. A associação de Ca(OH)2 e CHX foi utilizada como medicação intracanal em todos os grupos. Quatro amostras de canais radiculares (S) foram coletadas: S1) imediatamente após a cirurgia de acesso; S2) após o preparo biomecânico; S3) após aplicação do EDTA; e S4) após a remoção da medicação intracanal. Após a quantificação das MMP-3, MMP-8 e MMP-9, os dados foram analisados pelos testes de Friedman e Kruskal-Wallis e completados pelo teste de Dunn (5%). Independentemente da solução irrigadora utilizada, não houve diferença na redução de MMP-3 ou MMP-8 (P= 0,5273, P= 0,7048 respectivamente). No entanto, na redução de MMP-9 (P= 0,0246) o grupo NaOCl foi o mais eficaz, seguido pelo grupo NaOCl+LW e grupo NaOCl+PMB, respectivamente. A medicação intracanal [Ca(OH)2 + CHX] com o NaOCl e NaOCl+LW foi eficaz na redução de MMP-8 (P<0,0001, P= 0,0025) e MMP-9 (P= 0,0007, P= 0,0047) respectivamente, mas não para o grupo de NaOCl+PMB que não foi eficaz na redução de MMP-8 ou MMP-9 (P= 0,1718, P= 0,1953), respectivamente. NaOCl e NaOCl+LW foram eficazes na redução dos níveis de MMP-9, e esta efetividade pode ser melhorada pelo uso da medicação intracanal [Ca(OH)2+CHX] na redução dos níveis de MMP-8 e MMP-9.

Abstract This study aimed to evaluate sodium hypochlorite (NaOCl), limewater (LW), and Polymyxin B (PMB) as irrigants over MMP-3, MMP-8 and MMP-9. Thirty-three patients with apical periodontitis of single-rooted teeth were treated according to three-experimental groups (n=11): group-1: 2.5% NaOCl was used as irrigant; group-2: 2.5% NaOCl for the first two files and LW: [0.14% Ca(OH)2] for the last two files; group-3: 2.5% NaOCl for the first two files and PMB for the last two files. The association of Ca(OH)2 and CHX was used as an intracanal medication in all groups. Four root canal samplings (S) were collected: S1) immediately after access cavity; S2) after biomechanical preparation; S3) after EDTA application; and S4) after removal of the intracanal medication. After quantification of MMP-3, MMP-8, and MMP-9, the data were analyzed by Friedman and Kruskal-Wallis tests and completed by Dunn test (5%). Regardless the used irrigant, there was no difference in reducing MMP-3 or MMP-8 (P=0,5273, P=0,7048 respectively). However, in reducing MMP-9 (P=0,0246) the NaOCl group was the most effective followed by NaOCl+LW group and NaOCl+PMB group respectively. The intracanal medication [Ca(OH)2 + CHX] with the NaOCl and NaOCl+LW was effective in reducing MMP-8 (P<0,0001, P=0,0025) and MMP-9 (P=0,0007, P=0,0047) respectively, but not for the group of NaOCl+PMB which was not effective in reducing MMP-8 or MMP-9 (P=0,1718, P=0,1953) respectively. NaOCl and NaOCl+LW were effective in reducing MMP-9 levels, and this effectivity could be improved by the use of the intracanal medication [Ca(OH)2 + CHX] in reducing MMP-8 and MMP-9 levels.

Resumo Este estudo foi realizado para investigar o perfil microbiano e os níveis de endotoxina de lesões endoperiodontais de origem periodontal. Amostras periodontais e endodônticas foram obtidas de bolsas periodontais e canais radiculares necróticos de 10 dentes com lesões endoperiodontais. A investigação de 40 espécies bacterianas diferentes foram determinadas em cada amostra endodôntica e periodontal usando o método de hibridização de DNA-DNA (checkerboard) e o ensaio cinético cromogênico LAL foi usado para quantificação de endotoxinas. O teste exato de Fisher correlacionou as espécies bacterianas com a microbiota endodôntica ou periodontal. Os níveis de endotoxina (EU/mL) encontrados nas amostras do canal radicular e na bolsa periodontal foram comparados pelo teste de Wilcoxon (p<0,05). Bactérias e unidades de LPS foram encontradas em 100% das amostras endodônticas e periodontais. As espécies E. faecium, P. acnes, G. morbillorum, C. sputigena e L. buccalis foram fortemente correlacionadas com a microbiota endodôntica e P. nigrescens com a microbiota periodontal. P. intermedia, P. endodontalis e V. parvula foram mais prevalentes em ambas microbiotas endodôntica e periodontal. Os níveis de endotoxina na bolsa periodontal (89600 EU/mL) foram significativamente maiores do que no canal radicular (2310 EU/mL). Concluiu-se que a microbiota presente nos tecidos periodontal e endodôntico é semelhante, com maior prevalência de espécies do complexo laranja e maior nível de endotoxina nas bolsas periodontais.

Abstract This study was carried out to investigate the microbial profile and endotoxin levels of endodontic-periodontal lesions of periodontal origin. Periodontal and endodontic samples were taken from periodontal pockets and necrotic root canals of 10 teeth with endodontic-periodontal lesions. Evidencing of 40 different bacterial species were determined in each endodontic and periodontal sample using the checkerboard DNA-DNA hybridization method and Kinetic chromogenic LAL assay was used for quantification of endotoxins. Fisher’s exact test correlated the bacterial species with the endodontic or periodontal microbiota. The endotoxin levels (EU/mL) found in samples of the root canal and periodontal pocket were compared by the Wilcoxon test (p<0.05). Bacteria and LPS units were found in 100% of the endodontic and periodontal samples. The species E. faecium, P. acnes, G. morbillorum, C. sputigena and L. buccalis were strongly correlated with the endodontic microbiota and P. nigrescens with the periodontal microbiota. P. intermedia, P. endodontalis and V. parvula were more prevalent in both endodontic and periodontal microbiots. The endotoxin levels in the periodontal pocket (89600 EU/mL) were significantly higher than in the root canal (2310 EU/mL). It was concluded that the microbiota present in the periodontal and endodontic tissues is similar, with a higher prevalence of species of the orange complex and a higher level of endotoxin in the periodontal pockets.

Resumo O presente estudo objetivou comparar a eficácia do QMiX e do EDTA 17% associado à irrigação ultrassônica passiva (PUI) ou agitação manual (MA) na redução de E. faecalis, E. coli e LPS de canais radiculares. Quarenta dentes humanos unirradiculares foram divididos aleatoriamente em quatro grupos (n = 10), de acordo com o protocolo final de irrigação: EDTA+MA, QMiX+MA, EDTA+PUI, QMiX+PUI. Coletas das amostras foram obtidas a partir do conteúdo do canal radicular imediatamente antes do preparo (inicial-S1), após a instrumentação (S2), após o protocolo final de irrigação (S3) e 7 dias após a instrumentação e irrigação final (S4). A eficácia antimicrobiana e o conteúdo de endotoxina foram analisados por procedimento de cultura (UFC/mL) e ensaio LAL (EU/mL), respectivamente. Os resultados foram analisados estatisticamente pelo teste de Kruskal-Wallis e Friedman (α = 5%). QMiX+MA e QMiX+PUI reduziram 100% das bactérias E. coli e E. faecalis e também preveniram a recolonização de E. faecalis em S4. O EDTA reduziu significativamente E. coli, mas não foi eficaz na redução de E. faecalis. Todos os protocolos reduziram EU/mL quando comparados com S1, no entanto, no S4 houve uma redução significativa de EU/mL apenas nos grupos QMiX+MA e QMiX+PUI em relação a S3 e S2, respectivamente. A irrigação final com QMiX associada a MA ou PUI apresentou eficácia antibacteriana superior em relação ao EDTA, eliminando 100% das cepas de E. coli e E. faecalis. Além disso, QMiX+PUI reduziu 97,61% do conteúdo inicial de LPS.

Abstract The present study aimed to compare the effectiveness of QMiX and 17% EDTA associated to passive ultrasonic irrigation (PUI) or manual agitation (MA) on the reduction of E. faecalis, E. coli and LPS from root canals. Forty single rooted human teeth were randomly divided into four groups (n=10), according to the final irrigation protocol: EDTA+MA, QMiX+MA, EDTA+PUI, QMiX+PUI. Sample collections were obtained from the root canal content immediately before preparation (baseline-S1), after instrumentation (S2), after final irrigation protocol (S3) and 7 days after instrumentation and final irrigation (S4). The antimicrobial effectivity and on endotoxin content were analyzed by culture procedure (CFU/mL) and LAL assay (EU/mL), respectively. The results were statistically analyzed by Kruskal-Wallis and Friedman test (α=5%). QMiX+MA and QMiX+PUI reduced 100% of E. coli and E. faecalis bacteria and also prevented E. faecalisregrowth at S4. EDTA significantly reduced E. coli, but it was not effective in reducing E. faecalis. All protocols reduced EU/mL when compared to S1, however at S4 there was a significant reduction of EU/mL only in the QMiX+MA and QMiX+PUI groups in relation to S3 and S2, respectively. Final irrigation with QMiX associated with MA or PUI had superior antibacterial efficacy compared to EDTA, eliminating 100% of E. coli and E. faecalis strains. In addition, QMiX+PUI reduced 97.61% of the initial content of LPS.

Objetivo O presente estudo teve como objetivo avaliar as alterações morfológicas e químicas na dentina geradas por diferentes métodos de esterilização, através de microscopia eletrônica de varredura (MEV) e da análise por espectrometria de energia dispersiva de raios-X. Material e método Seccionaram-se 5 dentes humanos em 4 amostras, as quais foram divididas em 3 espécimes cada, totalizando 60 espécimes. Os espécimes foram separados conforme os grupos de esterilização: calor úmido sob pressão; radiação gama cobalto 60; controle (sem esterilização). Após a esterilização, os 60 espécimes foram analisados por MEV, usando 3 aumentos: 1500X, 5000X, 10000X. As imagens foram analisadas por 3 examinadores previamente calibrados, que distribuíram escores referentes às alterações observadas nos túbulos dentinários: 0 = sem alteração morfológica, 1 = baixa obliteração dos túbulos dentinários, 2 = moderada obliteração, 3 = completa obliteração. A composição química da dentina foi avaliada por meio do processo EDS, com 15 kV de incidência e penetração de 1 μm. Os dados obtidos foram submetidos ao teste estatístico de Kruskall-Wallis com nível de significância de 5%. Resultado De acordo com os resultados obtidos, verificou-se que a esterilização tanto por autoclave quanto por radiação gama cobalto 60 não provocou alterações significativas na morfologia dos túbulos dentinários e na constituição química da dentina. Conclusão Concluiu-se que ambos os métodos podem ser utilizados para a esterilização de dentes em pesquisa in vitro.

Aim The present study evaluated the morphological and chemical changes of dentin produced by different sterilization methods, using scanning electron microscopy (SEM) and energy-dispersive X-ray spectrometry (EDS) analysis. Material and method Five human teeth were sectioned into 4 samples, each divided into 3 specimens. The specimens were separated into sterilization groups, as follows: wet heat under pressure; cobalt 60 gamma radiation; and control (without sterilization). After sterilization, the 60 specimens were analyzed by SEM under 3 magnifications: 1500X, 5000X, and 10000X. The images were analyzed by 3 calibrated examiners, who assigned scores according to the changes observed in the dentinal tubules: 0 = no morphological change; 1, 2 and 3 = slight, medium and complete obliteration of the dentinal tubules. The chemical composition of dentin was assessed by EDS, with 15 kV incidence and 1 μm penetration. Result The data obtained were submitted to the statistical tests of Kruskall-Wallis and ANOVA. It was observed that both sterilization methods – with autoclave and with cobalt 60 gamma radiation – produced no significant changes to the morphology of the dentinal tubules or to the chemical composition of dentin. Conclusion Both methods may thus be used to sterilize teeth for research conducted in vitro.

The purpose of this study was to evaluate the effectiveness of glycolic propolis (PRO) and ginger (GIN) extracts, calcium hydroxide (CH), chlorhexidine (CLX) gel and their combinations as ICMs (ICMs) against Candida albicans, Enterococcus faecalis, Escherichia coli and endotoxins in root canals. Material and Methods After 28 days of contamination with microorganisms, the canals were instrumented and then divided according to the ICM: CH+saline; CLX, CH+CLX, PRO, PRO+CH; GIN; GIN+CH; saline. The antimicrobial activity and quantification of endotoxins by the chromogenic test of Limulus amebocyte lysate were evaluated after contamination and instrumentation at 14 days of ICM application and 7 days after ICM removal. Results and Conclusion After analysis of results and application of the Kruskal-Wallis and Dunn statistical tests at 5% significance level, it was concluded that all ICMs were able to eliminate the microorganisms in the root canals and reduce their amount of endotoxins; however, CH was more effective in neutralizing endotoxins and less effective against C. albicans and E. faecalis, requiring the use of medication combinations to obtain higher success.

OBJECTIVE: The purpose of this study was to evaluate the effcacy of auxiliary chemical substances and intracanal medications on Escherichia coli and its endotoxin in root canals. MATERIAL AND METHODS: Teeth were contaminated with a suspension of E. coli for 14 days and divided into 3 groups according to the auxiliary chemical substance used: G1) 2.5% sodium hypochlorite (NaOCl); G2) 2% chlorhexidine gel (CLX); G3) pyrogenfree solution. After, these groups were subdivided according to the intracanal medication (ICM): A) Calcium hydroxide paste (Calen®), B) polymyxin B, and C) Calcium hydroxide paste+2% CLX gel. For the control group (G4), pyrogen-free saline solution was used without application of intracanal medication. Samples of the root canal content were collected immediately after biomechanical preparation (BMP), at 7 days after BMP, after 14 days of intracanal medication activity, and 7 days after removal of intracanal medication. The following aspects were evaluated for all collections: a) antimicrobial activity; b) quantifcation of endotoxin by the Limulus Amebocyte Lysate test (LAL). Results were analyzed by the Kruskal-Wallis and Dunn's tests at 5% signifcance level. RESULTS: The 2.5% NaOCl and CLX were able to eliminate E. coli from root canal lumen and reduced the amount of endotoxin compared to saline. CONCLUSIONS: It was concluded that 2.5% NaOCl and CLX were effective in eliminating E. coli. Only the studied intracanal medications were to reduce the amount of endotoxin present in the root canals, regardless of the irrigant used.

OBJECTIVE: The purpose of this study was to evaluate the action of sodium hypochlorite (NaOCl) associated with an intracanal medication against Candida albicans and Enterococcus faecalis inoculated in root canals. MATERIAL AND METHODS: Thirty-six human single-rooted teeth with single root canals were used. The canals were contaminated with C. albicans and E. faecalis for 21 days and were then instrumented with 1% NaOCl. The roots were divided into 3 groups (n=12) according to the intracanal medication applied: calcium hydroxide paste, 2% chlorhexidine (CHX) gel, and 2% CHX gel associated with calcium hydroxide. The following collections were made from the root canals: a) initial sample (IS): 21 days after contamination (control), b) S1: after instrumentation, c) S2: 14 days after intracanal medication placement; S3: 7 days after intracanal medication removal. The results were analyzed statistically by the Kruskal-Wallis test at 5% significance level. RESULTS AND CONCLUSIONS: Both 1% NaOCl irrigation and the intracanal medications were effective in eliminating E. faecalis and C. albicans inoculated in root canals.

OBJECTIVE: To evaluate the efficacy of 16% carbamide peroxide gel (CP16%), tetrahydrate sodium perborate (SP) and mixture (CP16% + SP), in walking bleaching of non-vital discolored teeth. MATERIALS AND METHODS: Sixty single-rooted human premolars with intact crowns were used and initial color was assessed using Vita shade guide and standardized photos. The teeth were stained using rabbit fresh blood for 18 days and photos of discolored teeth and color evaluation were performed. The teeth were divided into 4 groups (n= 15), according to bleaching agent used: G1) CP16% gel; G2) CP16% gel + SP; G3) SP + distilled water; G4: control. The bleaching agents were replaced twice at 7-day intervals for 21 days. All teeth were evaluated by two endodontists at days 0, 7, 14 and 21 and the color changes were assessed using Vita shade guide and standardized photos. The results were analyzed by Kruskal-Wallis and Dunn's tests (p=0.05). RESULTS: The experimental groups presented statistically similar bleaching results (p>0.05) at the end of 7, 14 and 21 days. These groups presented significantly higher bleaching efficacy than control group (G4) (p<0.05). The mixture CP16% + SP promoted return of original color in 100% of teeth at the end of 21 days. CONCLUSION: It was concluded that three bleaching agents were effective in bleaching of stained teeth with blood products, especially at the end of 21 days.

O objetivo desta pesquisa foi avaliar in vitro a capacidade e o tempo necessário para a endotoxina se difundir pelos túbulos dentinários em direção ao cemento. Foram utilizados 30 dentes humanos unirradiculados, que tiveram suas coroas e seus ápices seccionados, padronizando-se o tamanho em 15 mm. Os dentes foram instrumentados até a lima K30 e impermeabilizados externamente com adesivo epóxi, deixando-se 10 mm de raiz exposta (terço médio). Os espécimes foram acondicionados em tubos plásticos e submetidos à radiação gama cobalto 60. Após a radiação, foram divididos em 2 grupos (n = 15): G1) foi inoculada uma solução de endotoxina de Escherichia coli no canal radicular dos espécimes e 1 mL de água apirogênica foi colocado no interior dos tubos; G2 (controle): foi inoculada água apirogênica nos canais radiculares e 1 mL de água apirogência foi colocado em cada tubo. Após 30 min, 2 h, 6 h, 12 h, 24 h, 48 h, 72 h e 7 dias, a água do interior dos tubos foi removida e substituída por outra. A alíquota removida foi testada para se detectar presença de endotoxina através da produção de anticorpos (IgM) em cultura de linfócitos B, pois a endotoxina é um ativador policlonal dessas células. Os resultados foram submetidos à análise estatística ANOVA (5%) e teste de Tukey, em que foi verificado que a água removida dos tubos após 24 h, 48 h, 72 h e 7 dias induziu maior produção de anticorpos em relação aos demais grupos, com diferença significante (p < 0,05). Assim, a endotoxina foi capaz de se difundir pelos túbulos dentinários em direção ao cemento, atingindo a região externa da raiz após 24 h.

The aim of this study was to evaluate the ability of endotoxin to diffuse through dentinal tubules towards the cement and to observe the period of time needed for it to reach the external root surface. Thirty single-rooted human teeth had their crowns and apices removed in order to standardize the root length to 15 mm. Teeth were instrumented until #30 K-file and made externally impermeable with epoxy adhesive, leaving 10 mm of the exposed root (middle third). The specimens were placed in plastic vials and irradiated (60Co gamma-rays). Then, they were divided into 2 groups (n = 15): G1) Escherichia coli endotoxin was inoculated into the root canal of the specimens and 1 ml of pyrogen-free water was put in the tubes; G2) (control): pyrogen-free water was inoculated into the root canals and 1 ml of pyrogen-free water was put in each tube. After 30 min, 2 h, 6 h, 12 h, 24 h, 48 h, 72 h and 7 days, the water of the tubes was removed and replaced. The removed aliquot was tested for the presence of endotoxin. Considering that the endotoxin is a B-lymphocyte polyclonal activator, at each experimental period, B-lymphocyte culture was stimulated with a sample of water removed from each tube and antibody (IgM) production was detected by ELISA technique. The results of IgM production were higher in groups of 24 h, 48 h, 72 h and 7 days in relation to the other studied groups, with statistically significant differences (ANOVA and Tukey's test p < 0.05). Endotoxin was able to diffuse through the dentinal tubules towards the cement, reaching the external root surface after the period of 24 h.