Effect of Dentin Conditioning with EDTA and Diode Lasers on Expression of Odontoblast-like Cell Markers of Dental Pulp Stem Cells.

Regenerative endodontic procedures rely on the delivery of mesenchymal stem cells into the root canal and on the effect of local growth factors from the dentin and blood clot. The aim of this study was to assess the effect of dentin conditioning with ethylenediamine tetraacetic acid (EDTA) and diode lasers with different wavelengths (808 nm and 980 nm) on the expression of odontoblast-like cell markers. Forty dentin cylinders were divided into four groups according to the irrigation protocol: EDTA, EDTA + 808 nm diode laser, EDTA + 980 nm diode laser, and phosphate-buffered saline as the control group. Dental pulp stem cells were seeded into the previously conditioned cylinders and incubated for 14 days. The quantitative real-time polymerase chain reaction was used to evaluate the expression of dentin sialophosphoprotein (DSPP), dentin morphoprotein-1 (DMP-1), and transforming growth factor-beta 1 (TGF-ß1). Data analysis was performed using the Kruskal-Wallis test. The activation of EDTA with 980 nm and 808 nm diode lasers resulted in lower DSPP and DMP-1 expression than that for EDTA alone (p < 0.05 and p < 0.01, respectively). The expression of TGF was similar among all groups. The highest level of expression of odontoblast-like differentiation markers was observed with EDTA alone. However, the use of an 808 nm diode laser during EDTA irrigation reduced the expression of odontoblastic differentiation markers.

Endocannabinoids Modulate Production of Osteoclastogenic Factors by Stem Cells of the Apical Papilla In Vitro.

INTRODUCTION: Many mediators are produced during pulp inflammation and necrosis, including endocannabinoids (ECbs), which might affect the function of stem cells of the apical papilla (SCAP), cells of paramount importance for root formation, and regenerative endodontic treatment. The aim of this study was to evaluate the production of osteoclastogenesis-related mediators by SCAP modulated by ECbs and lipopolysaccharide (LPS) in vitro. METHODS: SCAP were cultured and treated with ECb anandamide (AEA), 2-arachidonoylglycerol, or N-arachidonoylaminophenol. All groups were incubated in the presence of a vehicle or LPS and the antagonist of transient receptor potential cation channel subfamily V member 1, capsazepine. After 24 hours, the culture medium supernatants were collected for further quantification of tumor necrosis factor alpha, CCL2, macrophage colony-stimulating factor, osteoprotegerin, and receptor activator of nuclear factor kappa B ligand. RESULTS: Small amounts of tumor necrosis factor alpha and receptor activator of nuclear factor kappa B ligand were detected in SCAP supernatants, and none of the experimental conditions altered their production. A down-regulation in constitutive CCL2 production was observed in the AEA group compared with that in the LPS group. The production of macrophage colony-stimulating factor was significantly increased in all groups treated with AEA compared with the control and LPS-treated groups. Osteoprotegerin was significantly increased by AEA alone and by 2-arachidonoylglycerol and N-arachidonoylaminophenol in the presence of LPS and capsazepine. CONCLUSIONS: AEA modulates some of the osteoclastogenic factors produced by SCAP in a bone resorption protective fashion.

Endocannabinoids Regulate Stem Cells of the Apical Papilla via a Cannabinoid Receptor and TRPV1-Independent Mechanism.

INTRODUCTION: Endogenous cannabinoids (endocannabinoids [eCBs]) have been shown to have a multitude of functions including neurotransmission and immune modulatory effects. This study aimed to evaluate if stem cells of the apical papilla (SCAP) express the receptors and enzymes of the endocannabinoid system (ECS) and whether eCBs regulate their proliferation and mineralization potential. METHODS: Gene expression of the main components of the ECS and transient receptor potential vanilloid 1 (TRPV1) was evaluated in SCAP cultures. SCAP were treated with 2 concentrations of eCBs and/or capsazepine, a TRPV1 antagonist. SCAP viability was evaluated after 1, 4, and 7 days. Osteogenic differentiation was assessed after 14 days, and the gene expression of mineralization markers was assessed after 7 days. RESULTS: The enzymes of ECS and TRPV1 but not the cannabinoid receptors (cannabinoid receptors 1 and 2) were expressed in SCAP. Anandamide, 2-arachidonoylglycerol, and N-arachidonoylphenolamine (AM-404) reduced SCAP viability in all experimental periods at the highest concentration compared with the group with no treatment. Anandamide and AM-404 did not inhibit SCAP differentiation potential, but 2-arachidonoylglycerol at the highest concentration did. SCAP treated with AM-404 presented a down-regulation in gene expression of alkaline phosphatase (ALP), dentin matrix protein 1 (DMP-1), and dentin sialophosphoprotein (DSPP) compared with the proliferation medium group but not with control group. CONCLUSIONS: SCAP expressed the genes of the main components of ECS and TRPV1, and eCBs can affect SCAP viability, mineralization, and gene expression.

Cone-beam computed tomography cinematic rendering: clinical, teaching and research applications.

Cone-beam computed tomography (CBCT) is an essential imaging method that increases the accuracy of diagnoses, planning and follow-up of endodontic complex cases. Image postprocessing and subsequent visualization relies on software for three-dimensional navigation, and application of indexation tools to provide clinically useful information according to a set of volumetric data. Image postprocessing has a crucial impact on diagnostic quality and various techniques have been employed on computed tomography (CT) and magnetic resonance imaging (MRI) data sets. These include: multiplanar reformations (MPR), maximum intensity projection (MIP) and volume rendering (VR). A recent advance in 3D data visualization is the new cinematic rendering reconstruction method, a technique that generates photorealistic 3D images from conventional CT and MRI data. This review discusses the importance of CBCT cinematic rendering for clinical decision-making, teaching, and research in Endodontics, and a presents series of cases that illustrate the diagnostic value of 3D cinematic rendering in clinical care.

Cone-beam computed tomography cinematic rendering: clinical, teaching and research applications

Abstract Cone-beam computed tomography (CBCT) is an essential imaging method that increases the accuracy of diagnoses, planning and follow-up of endodontic complex cases. Image postprocessing and subsequent visualization relies on software for three-dimensional navigation, and application of indexation tools to provide clinically useful information according to a set of volumetric data. Image postprocessing has a crucial impact on diagnostic quality and various techniques have been employed on computed tomography (CT) and magnetic resonance imaging (MRI) data sets. These include: multiplanar reformations (MPR), maximum intensity projection (MIP) and volume rendering (VR). A recent advance in 3D data visualization is the new cinematic rendering reconstruction method, a technique that generates photorealistic 3D images from conventional CT and MRI data. This review discusses the importance of CBCT cinematic rendering for clinical decision-making, teaching, and research in Endodontics, and a presents series of cases that illustrate the diagnostic value of 3D cinematic rendering in clinical care.

Aplicação de um novo software de TCFC com filtros específicos para redução de artefatos: série de casos / Application of a new CBCT software with specific filters for blooming artifact reduction: case series

Introdução: As obturações dos canais radiculares e os pinos intrarradiculares geralmente produzem artefatos de contraste branco e possíveis alterações volumétricas nas imagens de tomografia computadorizada de feixe cônico (TCFC). Esses artefatos, induzidos pela maior densidade dos materiais obturadores de canais radiculares, cimentos para colocação de coroa e pinos intracanais, podem potencialmente levar a interpretações incorretas, interferindo no diagnóstico de um volume adquirido de TCFC, principalmente na avaliação de potenciais fraturas e perfurações radiculares. Objetivos: Avaliar em seis casos clínicos os efeitos positivos e negativos dos artefatos de contraste branco no diagnóstico de fraturas radiculares, perfurações radiculares e cimentação de restaurações de porcelana. Métodos: O software e-Vol DX, que possui filtros específicos para redução de artefato de contraste (BAR, Blooming Artifact Reduction), foi usado para melhorar o valor diagnóstico dos volumes adquiridos de TCFC. Conclusões: O em- prego do software e-Vol DX para a reconstrução de imagens de TCFC melhorou a visualização das estruturas anatômicas e reduziu os artefatos. A visualização aprimorada de da- dos nas imagens pode auxiliar na identificação de detalhes essenciais que, em conjunto com os achados clínicos, são úteis para uma correta estruturação do diagnóstico (AU).

Introduction: Root canal fillings and intraradicular posts often create white contrast artifacts and possible volumetric changes on CBCT scans known as blooming artifacts. These alterations could lead to the incorrect interpretations reducing the diagnostic of an acquired CBCT volume, particularly when evaluating potential root fractures and root perforations. Methods and Results: In six clinical cases, the positive and negative effects of beam hardening artifacts on the diagnosis of root fractures, root perforations and porcelain restoration cementation were evaluated. These artifacts, induced by the higher density of root canal filling materials, cements for crown placement and intracanal posts, may potentially lead to inaccurate or false interpretations. A novel software, e-Vol DX, which has specific filters for blooming artifact reduction (BAR), was used to improve the diagnostic value of acquired CBCT volumes. Conclusions: The use of the e-Vol DX software package for the reconstruction of CBCT scans improved visualization of anatomical structures and reduced blooming artifacts. Improved data visualization may help reveal essential details that, in conjunction with clinical findings, are useful to achieve a correct diagnosis (AU).

Development of a New Cone-Beam Computed Tomography Software for Endodontic Diagnosis.

Cone-beam computed tomography (CBCT) has promoted changes in approaches in Endodontics, and enhanced decision-making in complex clinical cases. Despite the technological advancements in CBCT hardware, the interpretation of the acquired images is still compromised by viewing software packages that often have limited navigational tools and lack adequate filters to overcome some challenges of the CBCT technology such as artefacts. This study reviews the current limitations of CBCT and the potential of a new CBCT software package (e-Vol DX, CDT- Brazil) to overcome these aspects and support diagnosing, planning and managing of endodontic cases. This imaging method provide high resolution images due to submillimeter voxel sizes, dynamic multi-plane imaging navigation and ability to change the volume parameters such as slice thickness and slice intervals and data correction applying imaging filters and manipulating brightness and contrast. The main differences between e-Vol DX and other software packages are: compatibility with all current CBCT scanners with the capacity to export DICOM Data, a more comprehensive brightness and contrast library, as other applications, in which adjustments are limited, do not usually support all the DICOM dynamic range features; Custom slice thickness adjustment, often limited and pre-defined in other applications; Custom Sharpening adjustment, often limited in other applications; advanced noise reduction algorithm that enhances image quality; preset imaging filters, dedicated endodontic volume rendering filters with the ability to zoom the image over 1000x (3D reconstructions) without loss of resolution and automatic imaging parameters customization for better standardization and opportunities for research; capture screen resolution of 192 dpi, with a 384 dpi option, in contrast to the 96 dpi of most similar applications. This new CBCT software package may support decision-making for the treatment of complex endodontic cases and improve diagnosis and treatment results. Effective improvement of image quality favors the rational prescription and interpretation of CBCT scans.

Development of a new cone-beam computed tomography software for endodontic diagnosis

Abstract Cone-beam computed tomography (CBCT) has promoted changes in approaches in Endodontics, and enhanced decision-making in complex clinical cases. Despite the technological advancements in CBCT hardware, the interpretation of the acquired images is still compromised by viewing software packages that often have limited navigational tools and lack adequate filters to overcome some challenges of the CBCT technology such as artefacts. This study reviews the current limitations of CBCT and the potential of a new CBCT software package (e-Vol DX, CDT- Brazil) to overcome these aspects and support diagnosing, planning and managing of endodontic cases. This imaging method provide high resolution images due to submillimeter voxel sizes, dynamic multi-plane imaging navigation and ability to change the volume parameters such as slice thickness and slice intervals and data correction applying imaging filters and manipulating brightness and contrast. The main differences between e-Vol DX and other software packages are: compatibility with all current CBCT scanners with the capacity to export DICOM Data, a more comprehensive brightness and contrast library, as other applications, in which adjustments are limited, do not usually support all the DICOM dynamic range features; Custom slice thickness adjustment, often limited and pre-defined in other applications; Custom Sharpening adjustment, often limited in other applications; advanced noise reduction algorithm that enhances image quality; preset imaging filters, dedicated endodontic volume rendering filters with the ability to zoom the image over 1000x (3D reconstructions) without loss of resolution and automatic imaging parameters customization for better standardization and opportunities for research; capture screen resolution of 192 dpi, with a 384 dpi option, in contrast to the 96 dpi of most similar applications. This new CBCT software package may support decision-making for the treatment of complex endodontic cases and improve diagnosis and treatment results. Effective improvement of image quality favors the rational prescription and interpretation of CBCT scans.

Resumo A tomografia computadorizada de feixe cônico (TCFC) promoveu mudanças nas abordagens de Endodontia e melhorou a tomada de decisões em casos clínicos complexos. Apesar dos avanços tecnológicos no hardware da TCFC, a interpretação da imagem adquirida ainda é comprometida pela visualização dos softwares, que muitas vezes têm ferramentas de navegação limitadas e falta de filtros adequados para superar estes desafios, como artefatos. Este estudo analisa as limitações atuais da TCFC e o potencial de um novo software (e-Vol DX, CDT-Brasil) para superar estes aspectos e apoiar o diagnóstico, planejamento e monitoramento de casos endodônticos. Este método de imagem fornece imagens em alta resolução devido a tamanhos submilimétricos de voxel, navegação dinâmica de imagens em vários planos e capacidade de alterar os parâmetros de volume como espessura de corte, intervalos de corte, correção de dados por meio de filtros de imagem, e manipulação do brilho e do contraste. As principais diferenças entre o e-Vol DX e outros software são: compatibilidade com todos os scanners de TCFC atuais com capacidade de exportar dados DICOM, com ajuste de brilho e contraste mais abrangente comparado a outros aplicativos, em que os ajustes são limitados, e geralmente não suportam todos os recursos da faixa dinâmica DICOM; ajuste de espessura de corte personalizado, muitas vezes limitado e pré-definido em outras aplicações; ajuste de nitidez personalizado, muitas vezes limitado em outras aplicações; algoritmo avançado de redução de ruído que melhora a qualidade da imagem; filtros de imagem predefinidos, filtros de para análise de volume do canal radicular com a capacidade de ampliar a imagem em mais de 1000x (reconstruções em 3D) sem perda de resolução, e personalização de parâmetros de imagem automática para melhor padronização e oportunidades de pesquisa; captura com resolução da tela de 192 dpi, com uma opção de 384 dpi, em contraste com os 96 dpi das aplicações similares. Este novo software de TCFC pode apoiar as tomadas de decisões para o tratamento de casos endodônticos complexos e melhorar os resultados do diagnóstico e do tratamento. A melhoria efetiva da qualidade da imagem favorece a prescrição e a interpretação racional das imagens de TCFC.

Infection Control in Teeth with Apical Periodontitis Using a Triple Antibiotic Solution or Calcium Hydroxide with Chlorhexidine: A Randomized Clinical Trial.

INTRODUCTION: This randomized clinical study compared the antibacterial effectiveness of treatment protocols using either a triple antibiotic solution (1 mg/mL) or calcium hydroxide/chlorhexidine paste as interappointment medication in infected canals of teeth with primary apical periodontitis. METHODS: The root canals of single-rooted teeth with apical periodontitis were prepared by using a reciprocating single-instrument technique with 2.5% sodium hypochlorite irrigation and then medicated for 1 week with either a triple antibiotic solution (minocycline, metronidazole, and ciprofloxacin) at 1 mg/mL (n = 24) or a calcium hydroxide paste in 2% chlorhexidine gluconate (n = 23). Samples were taken from the canal at the baseline (S1), after chemomechanical preparation (S2), and after intracanal medication (S3). DNA extracts from clinical samples were evaluated for total bacterial reduction using a 16S ribosomal RNA gene-based quantitative polymerase chain reaction assay. RESULTS: All S1 samples were positive for the presence of bacteria, and counts were substantially reduced after treatment procedures (P < .01). Bacterial levels in S2 and S3 samples did not significantly differ between groups (P > .05). S2 to S3 reduction was 97% in the antibiotic group and 39% in the calcium hydroxide/chlorhexidine group; only the former reached statistical significance (P < .01). There were significantly more quantitative polymerase chain reaction-negative S3 samples in the antibiotic group than in the calcium hydroxide group (P < .05). CONCLUSIONS: Interappointment medication with a triple antibiotic solution at the concentration of 1 mg/mL significantly improved root canal disinfection, and its effects were at least comparable with the calcium hydroxide/chlorhexidine paste. Effectiveness and easy delivery of the antibiotic solution make it an appropriate medicament as part of a disinfecting protocol for conventional nonsurgical endodontic treatment and possibly regenerative endodontic procedures.

Stem cells of the apical papilla regulate trigeminal neurite outgrowth and targeting through a BDNF-dependent mechanism.

Regenerative endodontic procedures have become a valuable alternative for the treatment of immature teeth with pulp necrosis. In addition to resolution of periradicular pathosis and promotion of continued root development, positive vitality testing has been observed in some regenerative clinical cases. Importantly, the positive response to electric stimulation of the regenerated tissue requires targeting of periradicular axons into the previously empty root canal space. However, the mechanism by which this process occurs is largely unknown. Since stem cells of the apical papilla (SCAP) have been proposed to populate the root canal following regenerative endodontic procedures, we hypothesized that SCAP regulate neurite outgrowth and axonal targeting. To test this hypothesis, we established primary co-cultures of human SCAP and rat trigeminal neurons, and performed neurite outgrowth assays using ELISA and confocal microscopy to determine the effect of increasing concentration of SCAP on the total neurite outgrowth and axonal targeting. In addition, we evaluated whether SCAP evoked axonal targeting in vivo using a matrigel subcutaneous implant assay. Data were analyzed by ANOVA with Bonferroni's post hoc test, and significance was set at p<0.05. The results demonstrated that SCAP release a soluble factor that regulates neurite outgrowth from cultured trigeminal neurons. Next, we demonstrated that this effect is completely abolished by pretreatment with a neutralizing antibody to brain-derived neurotrophic factor (BDNF), but not by antibodies to other neurotrophins. Further, SCAP release BDNF in a concentration-dependent manner as detected by ELISA, and trigger directed axonal targeting both in vitro and in vivo as demonstrated by microfluidic and matrigel implant experiments, respectively. Collectively, these results suggest that SCAP may be responsible for the chemical signal driving axons to target regenerated tissue via a BDNF-dependent mechanism.

Lipopolysaccharide from Porphyromonas gingivalis sensitizes capsaicin-sensitive nociceptors.

Although odontogenic infections are often accompanied by pain, little is known about the potential mechanisms mediating this effect. In this study we tested the hypothesis that trigeminal nociceptive neurons are directly sensitized by lipopolysaccharide (LPS) isolated from an endodontic pathogen, Porphyromonas gingivalis. In vitro studies conducted with cultures of rat trigeminal neurons demonstrated that pretreatment with LPS produced a significant increase in the capsaicin-evoked release of calcitonin gene-related peptide (CGRP) when compared with vehicle pretreatment, thus showing sensitization of the capsaicin receptor, TRPV1, by LPS. Furthermore, confocal microscopic examination of human tooth pulp samples showed the colocalization of the LPS receptor (toll-like receptor 4, TLR4) with CGRP-containing nerve fibers. Collectively, these results suggest the direct sensitization of nociceptors by LPS at concentrations found in infected canal systems as one mechanism responsible for the pain associated with bacterial infections.