Expression of osteoblastic phenotype in periodontal ligament fibroblasts cultured in three-dimensional collagen gel

Objective : To investigate the influence of a three-dimensional cell culture model on the expression of osteoblastic phenotype in human periodontal ligament fibroblast (hPDLF) cultures. Material and Methods : hPDLF were seeded on bi-dimensional (2D) and three-dimensional (3D) collagen type I (experimental groups) and and on a plastic coverslip (control) for up to 14 days. Cell viability and alkaline phosphatase (ALP) activity were performed. Also, cell morphology and immunolabeling for alkaline phosphatase (ALP) and osteopontin (OPN) were assessed by epifluorescence and confocal microscopy. The expression of osteogenic markers, including alkaline phosphatase, osteopontin, osteocalcin (OC), collagen I (COL I) and runt-related transcription factor 2 (RUNX2), were analyzed using real-time polymerase chain reaction (RT-PCR). Mineralized bone-like nodule formation was visualized by microscopy and calcium content was assessed quantitatively by alizarin red assay. Results : Experimental cultures produced an increase in cell proliferation. Immunolabeling for OPN and ALP in hPDLF were increased and ALP activity was inhibited by three-dimensional conditions. OPN and RUNX2 gene expression was significantly higher on 3D culture when compared with control surface. Moreover, ALP and COL I gene expression were significantly higher in three-dimensional collagen than in 2D cultures at 7 days. However, at 14 days, 3D cultures exhibited ALP and COL I gene expression significantly lower than the control, and the COL I gene expression was also significantly lower in 3D than in 2D cultures. Significant calcium mineralization was detected and quantified by alizarin red assay, and calcified nodule formation was not affected by tridimensionality. Conclusion : This study suggests that the 3D cultures are able to support hPDLF proliferation and favor the differentiation and mineralized matrix formation, which may be a potential periodontal regenerative therapy. .

Enxerto gengival livre e retalho posicionado coronariamente para recobrimento radicular / Free gingival grafts and coronally positioned flap for root coverage

O sucesso do tratamento das retrações gengivais baseia-se no conhecimentode sua etiologia e na avaliação da previsibilidade do recobrimentoradicular. Diversas técnicas têm sido propostas para o tratamento das retraçõesgengivais, a fim de promover possíveis melhorias na estética dosorriso. Este artigo relata um caso clínico de recobrimento radicular deretrações Classe I de Miller por meio do reposicionamento coronal do retalhoapós a colocação do enxerto gengival livre, tendo como resultado osucesso clínico do procedimento, tanto do ponto de vista do recobrimentoradicular como da satisfação da paciente.

Matriz dérmica acelular e matriz derivada do esmalte para recobrimento radicular em fumantes / Acellular dermal matrix grafts and enamel matrix protein for root coverage in smokers

O fumo apresenta uma influência negativa na previsibilidade dos procedimentos de cirurgias plásticas periodontais, principalmente, por meio da deficiência na função de fibroblastos periodontais. A literatura relata bons resultados clínicos de recobrimento radicular com o uso da matriz dérmica celular (MDA) e que a proteína derivada da matriz do esmalte (PDME) contribui para a melhora na atividade fibroblástica. O objetivo desse estudo foi avaliar a MDA associada ou não à PDME para recobrimento radicular em fumantes. Foram selecionados 20 pacientes com retrações gengivais bilaterais Classe I ou II de Miller, > 3 mm. Um grupo foi tratado com MDA e o outro com MDA associada à PDME. Os parâmetros clínicos - profundidade de bolsa a sondagem (PBS), nível clínico de inserção relativo (NCIR), altura da retração gengival (ARG), largura da retração gengival (LRC), quantidade de gengiva queratinizada (QCQ) e espessura da gengiva queratinizada (EGQ) - foram avaliados duas semanas após a terapia períodontal básica (exame inicial) e três meses após os procedimentos cirúrgicos. Par análise estatística, foram usados os testes de Wilcoxon, com nível de significância de 5%. Os dois procedimentos apresentaram resultados favoráveis, com diferenças estatisticamente significantes em todos os parâmetros clínicos após três meses. Não houve diferenças estatisticamente significantes entre os grupos. Concluiu-se que a MDA associada ou não à PMDE pode ser uma alternativa para o recobrimento radicular de retrações gengivais Classe I e II de Miller. Contudo, em pacientes fumantes, a associação da PMDE à MDA parece não mostrar benefícios clínicos adicionais no recobrimento radicular três meses após a cirurgia.

Bone marrow mesenchymal cell adhesion to polished and nitrided titanium surfaces

Aim: To evaluate the adhesion of mouse bone marrow mesenchymal cells (MBMMC) on differenttitanium surfaces. Methods: Grade II titanium discs (ASTM F86) received two different surfacetreatments: polished (S1) and cathodic cage plasma nitriding (S2). MBMMC were cultured ontitanium discs in 24-well cell culture plates, at a density of 1 x 104 cells per well. After 24 h, theadhesion was evaluated using a hemocytometer. Results: The mean adhesion was greater onS2, though without statistically significant difference from S2 (p>0.05). Conclusions: It wasdemonstrated that titanium surface treatment with ionic nitriding in a cathodic cage is biocompatiblesince it preserved the integrity of the cultivated MBMMC for a period of 24 h, allowing theiradhesion.

Adesão de células do ligamento periodontal de ratos a diferentes superfícies de titânio: estudo comparativo in vitro / Adhesion of ligament periodontal cells to different titanium surfaces: an in vitro comparative study

Objetivo: o presente estudo teve como objetivo avaliar, através de experimentos in vitro utilizando cultura celular, a capacidade de adesão das células do ligamento periodontal de ratos sobre as superfícies de titânio polidas e tratadas por nitretação iônica (plasma). Método: foram utilizados discos de titânio grau II ASTM F86 nas dimensões de 15mm de diâmetro por 1,5mm de espessura, os quais receberam diferentes tratamentos de superfície em 2 grupos distintos: polido e nitretado a plasma por gaiola catódica. As células foram isoladas do ligamento periodontal de ratos e cultivadas em meio de cultura alfa-MEM contendo antibióticos e suplementado com 10% de FBS, por 72 horas, em atmosfera úmida com 5% de CO2 a 37ºC. No subcultivo as células foram cultivadas sobre os discos de titânio em uma placa de 24 poços, na densidade de 1 x 104 células por poço, incluindo-se controles positivos sem os discos de titânio. Após 24 horas de cultivo, as células foram submetidas à contagem em câmara de Neubauer. Resultados: os resultados mostraram que a média de adesão celular foi maior na superfície controle (0,62±0,22) do que nos grupos polido (0,46±0,14) e nitretado (0,33±0,10). Foi observada diferença estatisticamente significante entre os grupos controle e nitretado (p=0,04), porém não se observou diferença na adesão celular entre os grupos polido e nitretado. Conclusão: a capacidade de adesão das células do ligamento periodontal de rato a superfícies de titânio não sofreu influência do tratamento de superfície dado ao material.

Objective: the present study aims to evaluate, through experiments in vitro, the adhesion capacity of rats periodontal ligament cells on polishing and treated by ionic nitriding (plasma) titanium surfaces. Method: degree II titanium discs (ASTM F86), 15mm of diameter for 1,5mm of thickness, which had received different treatments from surface in 2 distinct groups (polished and cathodic cage plasma nitriding) were used. The cells were isolated from periodontal ligament of rats and cultivated in alpha-MEM contend antibiotic and supplemented with 10% of FBS, for 72 hours, in humid atmosphere with 5% of CO2 at 37ºC. In the subculture the cells were cultivated on titanium discs in 24-well cell culture plates, with a density of 1 x 104 cells per well, including wells with no discs (control). After 24 hours of cultivation, the cells were counted in a Neubauer chamber. Results: the results had shown that the mean adhesion was greater on the control surface (0.62±0.22) than on polished (0.46±0.14) and nitriding (0.33±0.10) surfaces. Statistical significant difference was observed between the groups control and cathodic cage plasma nitriding (p=0.04), nevertheless no diference was found between polished and nitriding groups. Conclusion: the adhesion capacity of rat periodontal ligament cells on the titanium surfaces was not influenced by the different surface treatments given to the material, since none of these contributed positively in the process of cellular adhesion.

Identificação de células-tronco mesenquimais no ligamento periodontal e perspectivas na regeneração periodontal: revisão de literatura / Identification of mesenchymal stem cells in the periodontal ligament and perspectives for periodontal regeration: literature review

A busca por novas terapias com o intuito de restaurar a antigridade estrutural do periodonto, destruído pela progressão da doença periodontal, tem ganho novas perspectivas com os avanços das pesquisas com células-tronco e a bioengenharia tecidual. Além do embrião, as células-tronco também são encontradas em vários órgãos e tecidos do indivíduo adulto, onde participam homeostase tecidual, gerando novas células devido à renovação fisiológica e abrindo novas perspectivas para a regeneração e recuperação de tecidos e órgãos. Tais populações celulares ou células-tronco adultas tem sido facilmente identificadas pela sua morfologia e localização em alguns tecidos. A ideia de que estas células estão presentes no ligamento periodontal é baseada na capacidade reparadora destas, uma vez que podem se diferenciar em osteoblastos, cementoblastos, fibroblastos e componentes do tecido conjuntivo. Esta revisão de literatura tem como objetivo elucidar a identificação de células-tronco no ligamento periodontal, tendo em vista o interesse no potencial regenerativo dessas células e na sua aplicabilidade na regeneração periodontal.

The search for new therapies to restore the structural integrity of the periodontium, destroyed by the progression of periodontal disease, has gained new perspectives with the advance in stem cell research and tissue bioengineering. In addition to the embryo, stem cells are also found in various organs and tissues of adult individuals, where they take part in tissue homeostasis, generating new cells owing to physiological renewal and opening new perspectives for regenerating and recoveringtissues and organs. These cell populatithe or adult stem cells have been easily identified by their morphology and location in a of tissues. The idea that these cells are present in the periodontal ligament is based on the repairing capacity of these cells, given that they can be differentiated in osteoblasts, cementoblasts, fibroblasts and conjunctive tissue components. This literature review aims to discuss the identifying of stem cells in the periodontal ligament, in light of the interest in the regenerative potential of these cells and in their applicability for periodontal regeneration.

Influence of light-curing units and restorative materials on the micro hardness of resin cements.

Aim: The aim of this study was to evaluate the effect of indirect restorative materials (IRMs) and light-curing units (LCUs) on the micro hardness of dual-cured resin cement. Materials and Methods: A total of 36 cylindrical samples (2 mm thick) were prepared with dual-cured resin cement (Relyx ARC) photo-activated with either a QTH (Optilight Plus) for 40s or a LED (Radii) light-curing unit for 65s. Photo-activation was performed through the 2-mm- thick IRMs and the samples were divided into six groups (n=6) according to the combination of veneering materials (without, ceramic and indirect resin) and LCUs (QTH and LED). In the control group, the samples were light-cured with a QTH unit without the interposition of any restorative material. Vickers micro hardness test was performed on the top and bottom surfaces of each sample (load of 50 g for 15 secs). The data were statistically analyzed using a three-way ANOVA followed by Tukey x s post-hoc test ( P < 0.05). Results: There were no statistically significant differences on the top surface between the light curing-units ( P > 0.05); however, the LED provided greater hardness on the bottom surface when a ceramic material was used ( P < 0.05). The mean hardness in photo-activated samples, in which there was no interposition of indirect materials, was significantly greater ( P < 0.01). Conclusions: It may be concluded that the interposition of the restorative material decreased the micro hardness in the deeper cement layer. Such decrease, however, was lower when the ceramic was interposed and the cement light-cured with LED.