Osteogenic differentiation and reconstruction of mandible defects using a novel resorbable membrane: An in vitro and in vivo experimental study.

To evaluate the cellular response of both an intact fish skin membrane and a porcine-derived collagen membrane and investigate the bone healing response of these membranes using a translational, preclinical, guided-bone regeneration (GBR) canine model. Two different naturally sourced membranes were evaluated in this study: (i) an intact fish skin membrane (Kerecis Oral®, Kerecis) and (ii) a porcine derived collagen (Mucograft®, Geistlich) membrane, positive control. For the in vitro experiments, human osteoprogenitor (hOP) cells were used to assess the cellular viability and proliferation at 24, 48, 72, and 168 h. ALPL, COL1A1, BMP2, and RUNX2 expression levels were analyzed by real-time PCR at 7 and 14 days. The preclinical component was designed to mimic a GBR model in canines (n = 12). The first step was the extraction of premolars (P1-P4) and the 1st molars bilaterally, thereby creating four three-wall box type defects per mandible (two per side). Each defect site was filled with bone grafting material, which was then covered with one of the two membranes (Kerecis Oral® or Mucograft®). The groups were nested within the mandibles of each subject and membranes randomly allocated among the defects to minimize potential site bias. Samples were harvested at 30-, 60-, and 90-days and subjected to computerized microtomography (µCT) for three-dimensional reconstruction to quantify bone formation and graft degradation, in addition to histological processing to qualitatively analyze bone regeneration. Neither the intact fish skin membrane nor porcine-based collagen membrane presented cytotoxic effects. An increase in cell proliferation rate was observed for both membranes, with the Kerecis Oral® outperforming the Mucograft® at the 48- and 168-hour time points. Kerecis Oral® yielded higher ALPL expression relative to Mucograft® at both 7- and 14-day points. Additionally, higher COL1A1 expression was observed for the Kerecis Oral® membrane after 7 days but no differences were detected at 14 days. The membranes yielded similar BMP2 and RUNX2 expression at 7 and 14 days. Volumetric reconstructions and histologic micrographs indicated gradual bone ingrowth along with the presence of particulate bone grafts bridging the defect walls for both Kerecis Oral® and Mucograft® membranes, which allowed for the reestablishment of the mandible shape after 90 days. New bone formation significantly increased from 30 to 60 days, and from 60 to 90 days in vivo, without significant differences between membranes. The amount of bovine grafting material (%) within the defects significantly decreased from 30 to 90 days. Collagen membranes led to an upregulation of cellular proliferation and adhesion along with increased expression of genes associated with bone healing, particularly the intact fish skin membrane. Despite an increase in the bone formation rate in the defect over time, there was no significant difference between the membranes.

The Collaborative Cross-Mouse Population for Studying Genetic Determinants Underlying Alveolar Bone Loss Due to Polymicrobial Synergy and Dysbiosis.

Dysbiosis of oral microbiota is associated with the initiation and progression of periodontitis. The cause-and-effect relationship between genetics, periodontitis, and oral microbiome dysbiosis is poorly understood. Here, we demonstrate the power of the collaborative cross (CC) mice model to assess the effect of the genetic background on microbiome diversity shifts during periodontal infection and host suitability status. We examined the bacterial composition in plaque samples from seven different CC lines using 16s rRNA sequencing before and during periodontal infection. The susceptibility/resistance of the CC lines to alveolar bone loss was determined using the micro-CT technique. A total of 53 samples (7 lines) were collected before and after oral infection using oral swaps followed by DNA extraction and 16 s rRNA sequencing analysis. CC lines showed a significant variation in response to the co-infection (p < 0.05). Microbiome compositions were significantly different before and after infection and between resistant and susceptible lines to periodontitis (p < 0.05). Gram-positive taxa were significantly higher at the resistant lines compared to susceptible lines (p < 0.05). Gram-positive bacteria were reduced after infection, and gram-negative bacteria, specifically anaerobic groups, increased after infection. Our results demonstrate the utility of the CC mice in exploring the interrelationship between genetic background, microbiome composition, and periodontitis.

Avaliação da perda óssea utilizando diferentes parâmetros na análise microtomográfica: estudo em ratos / Bone loss assessment using different parameters in the microtomographic analysis: study in rats

Introdução: o desafio no uso do Micro-Ct tem sido estabelecer e padronizar padrões adequados para escaneamento e tratamento das imagens, para que se obtenha o máximo desempenho do equipamento, e permitir a comparação dos achados entre diferentes estudos. Objetivo: o presente estudo tem como objetivo comparar a porcentagem de volume ósseo em regiões com perda óssea periodontal utilizando diferentes metodologias para definição da área a ser analisada (ROI). Material e método: dez ratos foram submetidos à indução de doença periodontal, e, após a eutanásia, as mandíbulas foram escaneadas com cortes de 9 µ e 18 µm de espessura, com passo de rotação de 0.3mm. As imagens foram reconstituídas utilizando o software NRecon, e em seguida, utilizando o software CTAnalyser - CTAn, foram definidas as áreas de interesse (ROI) ao redor dos segundos molares. A primeira área de interesse (ROI1) foi definida em um padrão retangular que se restringiu às regiões interproximais e furca, onde a área total de tecido ósseo foi obtida somando os resultados dos 3 ROIs. A segunda área (ROI2) foi definida no sentido corono/apical ao redor do segundo molar até suas limitações proximais com os dentes vizinhos, onde a porcentagem de tecido ósseo pertencente às raízes foram excluídas das avaliações. As análises foram realizadas com cinco diferentes valores de thresholds (130-50, 130-60, 130-70, 130-80, 130-90 e 130-100). Resultado: a análise entre os diferentes ROIs demonstrou que em ambas as análises foi observada a tendência a menor porcentagem de tecido ósseo quanto maior o contraste de tons de cinza utilizado, no entanto, nos resultados obtidos no ROI2 essa diferença não foi estatisticamente significante. Conclusão: com os resultados obtidos pode se concluir que a utilização de diferentes thresholds para quantificação óssea, em áreas onde houve doença periodontal, pode trazer resultados divergentes; a definição da área de interesse interfere com os resultados obtidos e que a obtenção de uma área de interesse com a remoção das raízes mostrou-se menos susceptível à variação dos parâmetros de escaneamento.

Introduction: the challenge in the use of Micro-Ct has been to establish and standardize adequate standards for scanning and image processing to obtain the maximum performance of the equipment and to allow the comparison of findings between different studies. Objective: this study aims to compare the percentage of bone volume in regions with periodontal bone loss using different methodologies to define the area to be analysed (ROI). Material and method: ten rats were submitted to periodontal disease induction, and, after euthanasia, the mandibles were scanned with slices of 9µ and 18 µm thickness, with a rotation step of 0.3mm. The images were reconstructed using the NRecon software, and then, using the CTAnalyser - CTAn software, the areas of interest (ROI) around the second molars were defined. The first area of interest (ROI1) was defined in a rectangular pattern that was restricted to the interproximal and furcation regions, where the total area of bone tissue was obtained by adding the results of the 3 ROIs. The second area (ROI2) was defined in the coronal/apical direction around the second molar to its proximal limitations with the neighboring teeth, in which the percentage of bone tissue belonging to the roots was excluded from the evaluations. Analyses were performed with five different threshold values (130-50, 130-60, 130-70, 130-80, 130-90, and 130-100). Result: the analysis between the different ROIs showed that, in both analyses, there was a tendency towards a lower percentage of bone tissue the greater the grayscale contrast used. However, in the results obtained in ROI2, this difference was not statistically significant. Conclusion: with the results obtained, it can be concluded that the use of different thresholds for bone quantification, in areas where there was a periodontal disease, can bring divergent results; the definition of the area of ​​interest interferes with the results obtained; and that obtaining an area of interest with the removal of the roots, proved to be less susceptible to the variation of the scanning parameters.

Internal Lower Incisor Morphology revealed by Computerized Microtomography.

This study evaluated the internal morphology of lower incisors using computerized microtomography (micro-CT) images. Eighty-nine lower incisors were scanned by micro-CT and reconstructed with NRecon software. 2D parameters (perimeter, root length, circularity and canal diameter) and 3D parameters (volume, surface area and structure model index) were evaluated with CTAn and CTVol software. The results are presented descriptively. It was found that 89.9% of the canals had a single main root canal (type I), followed by type II (6.7%) and III (3.4%), while 5.6% of the specimens presented lateral canals and 1.1% had an apical delta. Mean volume and surface area were 31.80mm³ and 90.58mm², respectively. The most prevalent shape of the root canal at CEJ level was circular (41.6%) and 1mm from the apex, 73% of the samples were classified as oval. Lower incisors with internal anatomical variations may offer a high degree of technical complexity and may result in treatment failure.

Este estudo avaliou a morfologia interna dos dentes incisivos inferiores utilizando imagens de microtomografia computadorizada (micro-TC). 89 incisivos inferiores foram escaneados or micro-CT e reconstruídos com o programa NRecon. Parâmetros 2D - perímetro, comprimento da raiz, circularidade e diâmetro do canal - e parâmetros 3D - volume, área superficial e índice de modelo de estrutura - foram avaliados com os softwares CTAn e CTVol. Os resultados foram apresentados de forma descritiva. 89,9% dos canais apresentaram um único canal radicular principal (tipo I), seguido pelo tipo II (6,7%) e III (3,4%). 5,6% dos espécimes apresentavam canais laterais e 1,1% delta apical. O volume médio e a área de superfície foram 31,80mm³ e 90,58mm², respectivamente. A forma mais prevalente do canal radicular no nível da CEJ foi circular (41,6%) e a 1 mm do ápice, 73% das amostras foram classificadas como ovais. Incisivos inferiores com variações anatômicas internas podem oferecer um alto grau de complexidade técnica e resultar em falha do tratamento.

Internal Lower Incisor Morphology revealed by Computerized Microtomography / Morfologia interna de incisivos inferiores reveladas por microtomografía computadorizada

ABSTRACT This study evaluated the internal morphology of lower incisors using computerized microtomography (microCT) images. Eightynine lower incisors were scanned by microCT and reconstructed with NRecon software. 2D parameters (perimeter, root length, circularity and canal diameter) and 3D parameters (volume, surface area and structure model index) were evaluated with CTAn and CTVol software. The results are presented descriptively. It was found that 89.9% of the canals had a single main root canal (type I), followed by type II (6.7%) and III (3.4%), while 5.6% of the specimens presented lateral canals and 1.1% had an apical delta. Mean volume and surface area were 31.80mm³ and 90.58mm², respectively. The most prevalent shape of the root canal at CEJ level was circular (41.6%) and 1mm from the apex, 73% of the samples were classified as oval. Lower incisors with internal anatomical variations may offer a high degree of technical complexity and may result in treatment failure.

RESUMO Este estudo avaliou a morfologia interna dos dentes incisivos inferiores utilizando imagens de microtomografia computado rizada (microTC). 89 incisivos inferiores foram escaneados por microCT e reconstruídos com o programa NRecon. Parâmetros 2D perímetro, comprimento da raiz, circularidade e diâmetro do canal e parâmetros 3D volume, área superficial e índice de modelo de estrutura foram avaliados com os softwares CTAn e CTVol. Os resultados foram apresentados de forma descritiva. 89,9% dos canais apresentaram um único canal radicular principal (tipo I), seguido pelo tipo II (6,7%) e III (3,4%). 5,6% dos espécimes apresentavam canais laterais e 1,1% delta apical. O volume médio e a área de superfície foram 31,80mm³ e 90,58mm², respectivamente. A forma mais prevalente do canal radicular no nível da CEJ foi circular (41,6%) e a 1 mm do ápice, 73% das amostras foram classificadas como ovais. Incisivos inferiores com variações anatômicas internas podem oferecer um alto grau de complexidade técnica e resultar em falha do tratamento.

Mesiolingual Canal Prevalence in Maxillary First Molars assessed through Different Methods.

AIM: Evaluate the prevalence of mesiolingual canal prevalence orifice in mesiobuccal roots of maxillary first molars using five methods of visualization. MATERIALS AND METHODS: About 73 first permanent maxillary molars were analyzed. Visual clinical analysis of the presence of the fourth canal was performed using a straight end-odontic exploratory probe (EXDG16®) and a K10 manual file (SybronEndo®). Dental elements that were not located on the fourth canal were analyzed with the aid of a magnifying glass (Zeiss®) with a 2.5-fold increase and those teeth in which the fourth canal was not found went through the examination with clinical surgical microscope (OPTO®) with magnification of 20 times with both the explorer and endodontic file. Next, a periapical radiography of the teeth was performed in the teeth in which the mesiolingual canal was not yet found to observe the presence or absence of the fourth canal. Afterward, the teeth in which the canal was not yet located were scanned using the microtomography equipment (SkyScan®), at 100 kV and 100 µA, with an isotropic resolution of 16 µm. RESULTS: The mesiolingual canal was located in 70 teeth (95.8%) and in only 3 teeth it was not identified. CONCLUSION: The visual method in the fourth canal search has limitations, whereas the composite magnifying glass, the clinical surgical microscopy, and the computerized microtomography are efficient methods for locating the fourth canal in the upper first molars. CLINICAL SIGNIFICANCE: The anatomical complexity of the first maxillary molars is one of the factors that leads to high failure rates in the endodontic treatments of this group of teeth. In most clinical situations, the mesiolingual canal goes unnoticed by professionals, since conventional radiographs do not always allow the visualization of all root canals. Determining an effective method for locating the mesiolingual canal is of paramount importance to the success of endodontic treatment.