RESUMO
INTRODUCTION: Periodontal diseases are among the most common diseases of the oral cavity in the worldwide population. The prevention of gingivitis and periodontitis is based on the removal of bacterial plaque from the teeth with use of toothpaste containing active substances. Noteworthy is the ethanolic extract of Brazilian green propolis (EEP-B), which, due to the high content of artepillin C, has anti-inflammatory, antibacterial, or immunostimulatory effects. Little is known about interactions between EEP-B and gingival fibroblasts within the oral cavity. The purpose of the article is to determine the role of acidic fibroblast growth factor (aFGF-1), E-selectin, and ligand of CD40 (CD40L) secreted by human gingival fibroblasts (HGF-1) in the gingiva. MATERIAL AND METHODS: We performed our experiments on gingival fibroblasts (HGF-1), which are an ideal in vitro model for studying the processes taking place within the gingiva. We incubated cells with EEP-B or artepillin C at the concentrations of 1 µg/ml and 10 µg/ml. The aFGF-1, E-selectin, and CD40L were detected using the Bio-Plex Magnetic Luminex Assay and the Bio-Plex 200 System. RESULTS: Ethanolic extract of Brazilian green propolis and artepillin C increased the levels of aFGF-1 secreted by HGF-1. Moreover, EEP-B decreased the levels of E-selectin in both tested concentrations, which was not proved for artepillin C. No changes in the concentration of CD40L released by HGF-1 were observed. CONCLUSIONS: The obtained results may suggest that EEP-B, due to the mixture of various compounds including flavonoids, accelerates the wound healing effects and has anti-inflammatory activity.
RESUMO
Heparin-based silver nanoparticles (AgHep-NPs) and gold nanoparticles (AuHep-NPs) were produced by a photochemical method using silver nitrate and chloroauric acid as metal precursors and UV light at 254 nm. UV-Vis spectroscopy graphs showed absorption for AgHep-NPs and AuHep-NPs at 420 nm and 530 nm, respectively. TEM revealed a pseudospherical morphology and a small size, corresponding to 10-25 nm for AgHep-NPs and 1.5-7.5 nm for AuHep-NPs. Their antifungal activity against Candida albicans, Issatchenkia orientalis (Candida krusei), and Candida parapsilosis was assessed by the microdilution method. We show that AgHep-NPs were effective in decreasing fungus density, whereas AuHep-NPs were not. Additionally, the viability of human gingival fibroblasts was preserved by both nanoparticle types at a level above 80%, indicating a slight cytotoxicity. These results are potentially useful for applications of the described NPs mainly in dentistry and, to a lesser extent, in other biomedical areas.
Assuntos
Antifúngicos , Candida/crescimento & desenvolvimento , Citotoxinas , Fibroblastos/metabolismo , Gengiva/metabolismo , Ouro , Nanopartículas Metálicas/química , Processos Fotoquímicos , Prata/química , Antifúngicos/síntese química , Antifúngicos/química , Antifúngicos/farmacologia , Linhagem Celular , Citotoxinas/síntese química , Citotoxinas/química , Citotoxinas/farmacologia , Ouro/química , Ouro/farmacologia , HumanosRESUMO
Background: The aim of this study was to evaluate the cytotoxicity of cyanoacrylate adhesives in an indirect contact assay in human gingival fibroblast (FGH) and oral osteoblasts (GO) lineages. Methods: Cover glasses were glued with adhesives following the ISO 10993-2012 protocol. The groups were: C (control with cells and regular Dulbecco Modified Eagle Medium; LC (liquid ethyl-cyanoacrylate); GC (ethyl-cyanoacrylate gel); EGC (easy gel [ethyl-cyanoacrylate]); and D (Dermabond [octyl-cyanoacrylate]). Each cell linage was plated in the sixth passage using 104 cells. Cell viability was measured by the MTT test at 24, 48, 72, and 96 hours. Data were analyzed by two-way analysis of variance complemented by the Tukey test, with p < 0.05 being significant. Results: Dermabond stimulated osteoblast viability at 72 h (p < 0.05). All other groups were similar to the control cells (p > 0.05). For the fibroblasts, there was no difference in the groups, including the control except that EGC was cytotoxic for these cells (p < 0.05). Conclusions: Ethyl-cyanoacrylate gel and liquid forms available on the general chemical market were not cytotoxic for oral osteoblasts and fibroblasts in most cases. However, the easy gel form was cytotoxic for fibroblasts.
Assuntos
Acetatos/toxicidade , Fibroblastos/efeitos dos fármacos , Gengiva/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Adesivos Teciduais/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Gengiva/citologia , HumanosRESUMO
BACKGROUND AND OBJECTIVE: Transfection of cementum protein 1 (CEMP1) into human gingival fibroblasts (HGFs) notably increases cell metabolism and results in overexpression of molecules related to biomineralization at transcriptional and protein levels. Therefore, HGF-CEMP1 cells are considered as putative cementoblasts. This represents a significant advance in periodontal research because cementum neoformation is a key event in periodontal regeneration. In addition, it is well known that important changes in cell metabolism and protein expression are related to nucleolar structure and the function of this organelle, which is implicated in ribosome biogenesis. The aim of this study was to determine the effect of transfecting CEMP1 gene in human HGF on the ultrastructure of the nucleolus. MATERIAL AND METHODS: Cells were processed using the conventional technique for transmission electron microscopy, fixed with glutaraldehyde, postfixed with osmium tetraoxide, and embedded in epoxy resin. Semi-thin sections were stained with Toluidine blue and observed by light microscopy. Thin sections were stained with uranyl acetate and lead citrate. For ribonucleoprotein detection, the staining method based on the regressive effect of EDTA was used. In addition, the osmium ammine technique was used for specific staining of DNA. RESULTS: The results obtained in this study suggest that transfection of CEMP1 into HGFs does not produce changes in the general nucleolar ultrastructure because the different components of the organelle are present as fibrillary centers, and dense fibrillar and granular components compared with the control. CONCLUSION: The transfection of CEMP1 into HGFs allows these cells to perform cementoblast-like functions without alteration of the ultrastructure of the nucleolus, evaluated by the presence of the different compartments of this organelle involved in ribosomal biogenesis.
Assuntos
Fibroblastos/efeitos dos fármacos , Fibroblastos/ultraestrutura , Gengiva/citologia , Proteínas/farmacologia , Transfecção , Humanos , Microscopia Eletrônica de Transmissão , Coloração e RotulagemRESUMO
Introduction: In recent decades, dental implants have become one of the best options for comprehensive dental restoration; their placement is a multidisciplinary task that requires a solid understanding of biological, periodontal, surgical and prosthetic principles. Objective: The aim of this study was to quantify in vitro the adhesion and proliferation of human gingival fibroblasts (HGF) response on titanium (Ti) and zirconia (Zr) surfaces. Methodology: Samples of Ti and Zr were observed under atomic force microscopy (AFM). HGFs were inoculated in each sample to determine adhesion and cell proliferation. The reagent MTT was mixed with medium DMEM and inoculated in each plate; formazan was dissolved with dimethyl sulfoxide and analyzed at 540nm in a microplate spectrophotometer. The test was performed with three independent experiments. Data were analyzed with Kolmogorov-Smirnov tests (Lilliefors), Kruskal-Wallis tests and Mann-Whitney test comparisons. Results: Topography of the Zr plates showed greater roughness (Ra= 0.39 microns) than Ti (Ra= 0.049 microns). Quantification of HGF adhesion was significantly higher (p<0.05) in Ti, while proliferation showed no statistically significant differences between the groups. Conclusion: It is noteworthy that, even though Ti initially showed increased cell adhesion on the surface, after 24h Zr samples showed similar proliferation; this demonstrates that both surfaces have a comparable biological response.
Introducción: En las últimas décadas, los implantes dentales se han posicionado como una de las mejores opciones de restauración dental integral; su colocación es una tarea multidisciplinaria que requiere una sólida comprensión de los principios biológicos, periodontales, quirúrgicos y protésicos. Objetivo: El objetivo de este estudio fue cuantificar in vitro la adhesión y proliferación de la respuesta de fibroblastos gingivales humanos (HGF) en superficies de titanio (Ti) en contraste con superficies de zirconia (Zr). Metodología: Se observaron las muestras de Ti y Zr bajo microscopía de fuerza atómica (AFM). Los HGF fueron inoculados en cada muestra para determinar la adhesión y proliferación celular. El reactivo MTT se mezcló con medio DMEM y se inoculó en cada placa, el formazán se disolvió con dimetilsulfóxido y se analizó a 540nm en un espectrofotómetro de microplaca. El ensayo se realizó con tres experimentos independientes. Los datos fueron analizados con pruebas de Kolmogorov-Smirnov (Lilliefors), pruebas de Kruskal-Wallis y comparaciones de Mann-Withney. Resultados: La topografía de las placas de Zr mostraron una mayor rugosidad (Ra=0.39 micrones) en comparación con las de Ti (Ra=0.049 micrones). La cuantificación de la adhesión de HGF fue significativamente mayor (p<0.05) en el Ti, mientras que la proliferación no mostró diferencias estadísticamente significativas entre los grupos. Conclusión: Es importante mencionar que, a pesar de que el Ti mostró inicialmente una mayor adhesión celular sobre la superficie, después de 24 hrs las muestras de Zr mostraron una proliferación similar; lo que demuestra que ambas superficies poseen una respuesta biológica comparable.
Assuntos
Materiais Biocompatíveis , Adesão Celular , Proliferação de Células , Implantes Dentários , Teste de Materiais , Propriedades de Superfície , Titânio/química , Zircônio/químicaRESUMO
The purpose of this study is to compare the cytotoxic effect of threematerials, which have been used for treating dental hypersensitivity. Materialand method: In vitro study. Clinpro (3M Co, St. Paul, MN. USA), Seal & Protect(Dentsply, DeTrey GmbH. Germany) and UltraEZ (Ultradent Products,Inc., S. South Jordan UT. USA) were used at concentrations of 0.1, 0.05, 0.01 and 0.001g/ml on human gingival fibroblasts. Furthermore, Clinpro and Seal & Protect were applied to this cell culture as polymerized disks. Toxicity was assessed at 24 and 48 hours by the use of the cell viability assay (MTT). Statistical analysis for cell viability was performed using two-way ANOVA and Tukeys post hoc test. Statistical significance was set at 5 percent. Results: Seal & Protect and Clinpro were found to be highly toxic at 24 and 48 hours, reaching 70 percent toxicity at concentrations over 0.01g/ml. Seal & Protect and Clinpro polymerized disks were toxic at 24 and 48 hours. UltraEZ showed an increased between 46 percent and 67 percent in cell viability at 24 hours and between 8 percent and 45 percent at 48 hours. Statistical analysis showed differences between these three desensitizers when comparing concentration and control group (p<0.05). Discussion: UltraEZ did not have a cytotoxic effect and may be considered a compatible and safe material, whereas polymerized and non-polymerized Clinpro and Seal & Protect should be used with caution...
Introduccion: El proposito de este estudio es comparar el efecto citotoxico de tres materiales que se han utilizado para el tratamiento de la hipersensibilidad dental. Material y metodo: Estudio in-vitro. Los desensibilizantes dentinarios Clinpro (3M ESPE), Seal&Protect (Dentsply) yUltraEZ (Ultradent) fueron utilizados a concentraciones de 0,1; 0,05; 0,01 y 0,001 g/ml sobre cultivos celulares de fibroblastos gingivales humanos. Ademas, Clinpro y Seal&Protect se aplicaron a este cultivo celular como discos polimerizados. La toxicidad se evaluo a 24 y 48 horas mediante ensayo de viabilidad (MTT). El analisis estadistico para la viabilidad celular se realizo mediante ANOVA de dos vias seguido de analisis Tukey. La significancia estadistica se fijo al 5 por ciento. Resultados: Clinpro y Seal&Protect resultaron ser altamente toxicos a las 24 y 48 horas, alcanzando un 70 por ciento de toxicidad aconcentraciones superiores de 0,01 g/ml. Los discos polimerizadosde Clinpro y Seal&Protect fueron toxicos a 24 y 48 horas. UltraEZ produjo un aumento de la viabilidad celular entre un 46 por ciento y 67 por ciento a las 24 horas y entre un 8 por ciento y 45 por ciento a las 48 horas. El analisis estadistico mostro diferencias entreestos tres desensibilizantes al comparar la concentracion y su grupo control (p<0,05). Discusion: UltraEZ no tuvo efecto citotoxico y puede ser considerado como un material compatible y seguro para ser utilizado, mientras que Clinpro y Seal&Protect en su estado polimerizado y no polimerizado deberian ser utilizados con precaucion...
Assuntos
Humanos , Dessensibilizantes Dentinários/toxicidade , Gengiva , Fibroblastos , Sensibilidade da Dentina/terapia , Análise de Variância , Sobrevivência Celular , Fatores de TempoRESUMO
Fisetin is an anti-inflammatory flavonoid; however, its anti-inflammatory mechanism is not yet understood. In this study, we evaluated the anti-inflammatory effect of fisetin and its association with mitogen-activated protein kinase (MAPK) and nuclear factor kappa-beta pathways in human gingival fibroblasts (HGFs) treated with lipopolysaccharide (LPS) obtained from Porphyromonas gingivalis. The cell signaling, cell viability, and cyclooxygenase-2 (COX-2) expression of HGFs treated with various concentrations (0, 1, 5, 10, and 15 µM) of fisetin were measured by cell viability assay (MTT), Western blotting, and reverse transcriptase polymerase chain reaction analysis on COX-2. We found that fisetin significantly reduced the synthesis and expression of prostaglandin E2 in HGFs treated with LPS. Activation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 MAPK was suppressed consistently by fisetin in HGFs treated with LPS. The data indicate that fisetin inhibits MAPK activation and COX-2 expression without affecting cell viability. These findings may be valuable for understanding the mechanism of the effect of fisetin on periodontal disease.
Assuntos
Anti-Inflamatórios/farmacologia , Flavonoides/farmacologia , Lipopolissacarídeos/farmacologia , Anti-Inflamatórios/química , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/efeitos dos fármacos , Dinoprostona/metabolismo , Fibroblastos/efeitos dos fármacos , Flavonoides/química , Flavonóis , Gengiva/citologia , Gengiva/efeitos dos fármacos , Humanos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Estrutura Molecular , NF-kappa B/metabolismo , Porphyromonas gingivalis/citologia , Transdução de Sinais/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
The aim of this study was to evaluate the viability and biomechanical properties of artificial human oral mucosa stroma (HOMS) subjected to cryopreservation with different cryoprotectant solutions. Artificial HOMS based on a fibrin-agarose matrix with human gingival fibroblasts cultured 7 days in vitro were cryopreserved with three cryoprotectant solutions: (A) TC-199 Medium, DMSO 15%, albumin; (B) DMEM, FCS, DMSO 10%; (C) QC Medium, glycerol. As controls, artificial HOMS not subjected to cryopreservation (CF) and HOMS cryopreserved without cryoprotectant solution (CS) were used. Histological analysis by light microscopy showed that solutions A and B preserved a pattern of porosity similar to values in CF. Based on the number of intact cells in the fibrin-agarose matrix, substitutes preserved with solution B showed the best results. Cell proliferation detected with PCNA immunochemical methods showed that the cell proliferation index was highest in substitutes cryopreserved with solution B. The reculture method and cell viability analyses with Live & Dead(®) revealed increased number of viable in cells preserved with solution B. Artificial stroma substitutes in CS control samples showed the greatest alterations in microstructure and cell proliferation. Analysis of the biomechanical properties showed that substitutes cryopreserved with different solutions had adequate rheological parameters (yield stress, elastic modulus and viscous modulus) and were therefore suitable for use in regenerative medicine. These results establish effective methods of cryopreservation for all experimental situations and suggest that solution B (DMEM, FCS, DMSO 10%) was the best cryoprotectant for the cryopreservation of an artificial oral human mucosa substitute based on a fibrin-agarose matrix.
Assuntos
Materiais Biocompatíveis/química , Criopreservação/métodos , Fibroblastos/citologia , Gengiva/citologia , Mucosa Bucal/química , Sobrevivência Celular , Células Cultivadas , Fibrina/química , Humanos , Reologia , Sefarose/química , Alicerces Teciduais/químicaRESUMO
The acellular dermal matrix (ADM) was introduced in periodontology as a substitute for the autogenous grafts, which became restricted because of the limited source of donor's tissue. The aim of this study was to investigate, in vitro, the distribution, proliferation and viability of human gingival fibroblasts seeded onto ADM. ADM was seeded with human gingival fibroblasts for up to 21 days. The following parameters were evaluated: cell distribution, proliferation and viability. Results revealed that, at day 7, fibroblasts were adherent and spread on ADM surface, and were unevenly distributed, forming a discontinuous single cell layer; at day 14, a confluent fibroblastic monolayer lining ADM surface was noticed. At day 21, the cell monolayer exhibited a reduction in cell density. At 7 days, about to 90 percent of adherent cells on ADM surface were cycling while at 14 and 21 days this proportion was significantly reduced. A high proportion of viable cell was detected on AMD surface both on 14 and 21 days. The results suggest that fibroblast seeding onto ADM for 14 days can allow good conditions for cell adhesion and spreading on the matrix; however, migration inside the matrix was limited.
A matriz dérmica acelular (MDA) foi introduzida na Periodontia como um substituto para enxertos autógenos, os quais se tornaram restritos devido à quantidade limitada de tecido doador. O objetivo deste estudo foi verificar, in vitro, a distribuição, proliferação e viabilidade de fibroblastos gengivais humanos cultivados em MDA. Fibroblastos gengivais foram cultivados sobre MDA por até 21 dias. Os seguintes parâmetros foram avaliados: distribuição, proliferação e viabilidade celular. Os resultados revelaram que, aos 7 dias, os fibroblastos estavam aderidos e espraiados na superfície da MDA, e estavam distribuídos de forma desigual, formando uma camada celular descontínua; aos 14 dias, uma monocamada confluente de fibroblastos revestindo a superfície da MDA foi observada. Aos 21 dias, a monocamada celular exibiu uma redução na densidade celular. Aos 7 dias, cerca de 90 por cento das células aderidas na superfície da MDA estavam no ciclo celular, enquanto que aos 14 e 21 dias esse número reduziu significativamente. Uma maior proporção de células viáveis foi detectada na superfície da MDA tanto aos 14 quanto aos 21 dias. Os resultados sugerem que fibroblastos cultivados sobre a MDA por 14 dias permitem boas condições de adesão e espraiamento das células sobre a matriz, porém, a migração de células para o interior da matriz foi limitada.