Fibra de carbono ativada: avaliação das interações biológicas in vitro / Activated carbon fiber: evaluation of biological interactions in vitro

Devido a constante necessidade de desenvolver materiais biocompatíveis com propriedades osteocondutores e osteoindutoras, a presente tese conta com o desenvolvimento de dois estudos in vitro com fibra de carbono obtida a partir de fibra PAN têxtil, incorporada com diferentes íons de metais, na osteogênese com vistas à compreensão das necessidades da engenharia tecidual no desenvolvimento desse biomaterial com adequadas propriedades biológicas. As células foram obtidas dos fêmures de 09 ratos machos adultos (Wistar) pesando 300g, com 90 dias.Estudo 1: A partir da preparação da fibras foram obtidos corpos de prova de 4 mm de diâmetro e 2 mm de altura, dos seguintes grupos: fibra de carbono não ativada (FCNA), fibra carbono ativada (FCA) e fibra carbono ativada com prata (FCAAg). Após plaqueamento (n=5) em meio suplementado (MTS) e meio suplementado osteogênico (MTSO) foram analisados: viabilidade celular, conteúdo de proteína total (PT), atividade de fosfatase alcalina (ALP), interaçãocelular e formação de nódulos de mineralização. Foi avaliada a formação de biofilme nos corpos de prova, utilizando cepas de S. aureus, P. aeruginosa e E. coli. Na viabilidade celular, houve diferença estatística entre grupo controle celular (C) e FCA-MTS, FCAAg-MTS e FCAAg-MTSO. Em PT, não houvediferença, na ALP houve diferença entre C-MTS e as fibras, C-MTSO se mostrou semelhante. Em nódulos, houve diferença entre C-MTS e C-MTSO e as fibras do MTSO. Houve redução de formação de biofilme do S. aureus na FCAAg.Estudo 2: Foram obtidos corpos de prova da mesma dimensão do estudo 1 (n=5) dos seguintes grupos: fibra carbono ativada com prata (FCAAg), fibra carbono ativada com ouro (FCAAu), fibra carbono ativada com cobre (FCACu), fibra carbono ativada com paládio (FCAPd) e fibra carbono ativada com platina (FCAPt). Foram quantificadas a proliferação celular, viabilidade celular, formação de nódulos de mineralização, conteúdo de PT e ALP. Todas as amostras mostraram-se semelhantes quanto a proliferação celular, com exceção do grupo FCAAg comparado ao grupo controle (C). Sobre viabilidade celular, C obteve maior viabilidade que os outros grupos, e FCA obteve maior taxa que os grupos FCAAg, FCACu, FCAPt, sendo semelhante aos grupos FCAAu e FCAPd. Já os grupos FCAAu e FCAPd apresentaram diferença aos grupos FCAAg e FCACu. Na análise de expressão de PT apenas houve diferença entre FCA e FCAAu, sendo FCAAu com menor expressão de produção de PT. Na avaliação da ALP os grupos FCAAg e FACu mostraram diferença estatística e inferior com os grupos C, FCAAu, FCAPd e FCAPt, além disso, o grupo FCA mostrou menor taxa que C.Conclusões: As fibras utilizadas de base para a incorporação dos íons demonstraram grande potencial para uso como scaffold para reparação óssea, isso porque em ambos os estudos, na forma ativada e não ativada, as fibras apresentaram viabilidade celular e quantificação de cálcio satisfatórias. Sendo a versão não ativada mais econômica no que diz respeito ao tempo e custo de preparação. Mais estudos devem ser empregados a fim de assegurar sua segurança clínica em relação à citotoxicidade da incorporação de íons de ouro e paládio.(AU)

Due to the constant need to develop biocompatible materials with osteoconductive and osteoinductive properties, this thesis involves the development of two in vitro studies with carbon fiber obtained from textile PAN fiber, incorporated with different metal ions, in osteogenesis with a view to understanding the needs of tissue engineering in the development of this biomaterial with adequate biological properties. The cells were obtained from the femurs of 9 adult male rats (Wistar) weighing 300g, aged 90 days. Study 1: From the fiber preparation, specimens measuring 4 mm in diameter and 2 mm in height were obtained from the following groups: non-activated carbon fiber (FCNA), activated carbon fiber (FCA) and silver-activated carbon fiber (FCAAg). After plating (n=5) in supplemented medium (MTS) and supplemented osteogenic medium (MTSO), cell viability, total protein content (PT), alkaline phosphatase (ALP) activity, cell interaction and formation of mineralization nodules were analyzed. . Biofilm formation was evaluated in the specimens, using strains of S. aureus, P. aeruginosa and E. coli. In cell viability, there was a statistical difference between the cell control group (C) and FCAMTS, FCAAg-MTS and FCAAg-MTSO. In PT, there was no difference, in ALP there was a difference between C-MTS and fibers, C-MTSO was similar. In nodules, there was a difference between C-MTS and C-MTSO and MTSO fibers. There was a reduction in S. aureus biofilm formation on FCAAg. Study 2: Specimens of the same size as in study 1 (n=5) were obtained from the following groups: carbon fiber activated with silver (FCAAg), carbon fiber activated with gold (FCAAu), carbon fiber activated with copper (FCACu), palladium-activated carbon fiber (FCAPd) and platinum-activated carbon fiber (FCAPt). Cell proliferation, cell viability, formation of mineralization nodules, PT and ALP content were quantified. All samples were similar in terms of cell proliferation, with the exception of the FCAAg group compared to the control group (C). Regarding cell viability, C obtained higher viability than the other groups, and FCA obtained a higher rate than the FCAAg, FCACu, FCAPt groups, being similar to the FCAAu and FCAPd groups. The FCAAu and FCAPd groups showed differences to the FCAAg and FCACu groups. In the analysis of PT expression, there was only a difference between FCA and FCAAu, with FCAAu having lower expression of PT production. In the ALP assessment, the FCAAg and FACu groups showed a lower statistical difference compared to the C, FCAAu, FCAPd and FCAPt groups, in addition, the FCA group showed a lower rate than C. Conclusions: The fibers used as the basis for the incorporation of ions demonstrated great potential for use as a scaffold for bone repair, because in both studies, in activated and non-activated form, the fibers showed satisfactory cell viability and calcium quantification. The non-activated version is moreeconomical in terms of preparation time and cost. More studies must be carried out to ensure its clinical safety in relation to the cytotoxicity of the incorporation of gold and palladium ions. (AU)

Efeito da curcumina e três análogos ­ DMAD, DMAM E RI75 ­ sobre viabilidade, diferenciação e expressão gênica em pré-osteoblastos / Effect of curcumin and three of its analogues - DMAD, DMAM and RI75 - on pre-osteoblast cells' viability, differentiation, and gene expression

A curcumina, encontrada nos rizomas da cúrcuma (Curcuma longa L.), tem sido amplamente estudada devido aos seus potenciais benefícios à saúde que, entre outros, incluem propriedades anti-inflamatórias, antioxidantes e cicatrizantes. No entanto, devido à sua baixa biodisponibilidade e farmacocinética desfavorável, compostos análogos são desenvolvidos e estudados para obter melhores características biofarmacêuticas e aumentar os efeitos biológicos. Neste trabalho, avaliamos a atividade da curcumina e três de seus análogos sintéticos (DMAD, DMAM e RI75) sobre a viabilidade e diferenciação de uma linhagem celular pré-osteoblástica (MC3T3-E1). A expressão dos genes fator de crescimento endotelial vascular (vegf), do ligante 12 da quimiocina de motivo C-X-C (cxcl12) e do fator de transcrição runt- related 2 (runx2) também foi avaliada. As células foram estimuladas com curcumina e os três análogos usando concentrações de 10, 30 ou 50 µM. A curcumina e os análogos testados apresentaram nenhuma, ligeira ou moderada citotoxicidade celular em comparação com o controle negativo, após 24, 48 e 72 horas, nas concentrações testadas. Os resultados de atividade de mineralização para a curcumina e o análogo DMAD não apresentaram diferença estatística com o grupo controle, enquanto os análogos DMAM e RI75 apresentaram menor atividade de mineralização. Nenhuma das substâncias apresentou expressão gênica diferencial de cxcl12, um ativador de células endoteliais e inflamatórias. Por outro lado, em comparação com o controle, o análogo de curcumina RI75 mostrou regulação positiva de runx2 e vegf, ambos relacionados ao reparo tecidual. Os resultados sugerem que os análogos DMAD, DMAM e RI75 apresentam citotoxicidade semelhante ou inferior à curcumina natural e maior atividade biológica, com destaque para o análogo RI75, sendo substâncias com potencial promissor para biomodificações de materiais.

Curcumin, found in the rhizomes of turmeric (Curcuma longa L.), has been widely studied due to its potential health benefits which, among others, include anti inflammatory, antioxidant and healing properties. However, due to their low bioavailability and unfavorable pharmacokinetics, analogous compounds are developed and studied to obtain better biopharmaceutical characteristics and increase biological effects. In this work, we evaluated the activity of curcumin and three of its synthetic analogues (DMAD, DMAM and RI75) on the viability and differentiation of a pre-osteoblastic cell line (MC3T3-E1). The expression of the genes vascular endothelial growth factor (vegf), C-X-C motif chemokine ligand 12 (cxcl12) and runt related transcription factor 2 (runx2) was also evaluated. Cells were stimulated with curcumin and the three analogues using concentrations of 10, 30 or 50 µM. Curcumin and the analogues tested showed no, slight or moderate cellular cytotoxicity compared to the negative control, after 24, 48 and 72 hours, at the concentrations tested. The mineralization activity results for curcumin and the DMAD analogue showed no statistical difference with the control group, while the DMAM and RI75 analogues showed lower mineralization activity. None of the substances showed differential gene expression of cxcl12, an activator of endothelial and inflammatory cells. On the other hand, compared to the control, the curcumin analogue RI75 showed upregulation of runx2 and vegf, both related to tissue repair. The results suggest that the DMAD, DMAM and RI75 analogues present similar or lower cytotoxicity compared to natural curcumin and greater biological activity, especially the RI75 analogue, being, therefore, substances with promising potential for biomodification of materials.

Cytotoxicity Comparison of a Calcium Silicate-Based Resin Cement versus Conventional Self-Adhesive Resin Cement and a Resin-Modified Glass Ionomer: Cell Viability Analysis

Abstract Objective: To compare the cytotoxicity level of a new calcium silicate-based resin cement (TheraCem) with two commonly used cements, including a conventional self-adhesive resin cement (Panavia SA) and a resinmodified glass ionomer cement (FujiCem2), on the human gingival fibroblast cells after 24 and 48 hours. Material and Methods: Twelve discs of each cement type were fabricated. The extract of cement disks was made by incubating them in the cell medium. Human gingival fibroblast cells were cultured and exposed to cement extracts for 24 h and 48 h. MTT assay was performed on extracts and optical density and cell viability rates were calculated by the spectrophotometer device at 570 nm. Data were analyzed using ANOVA and Tukey HSD tests. Results: The cell viability rates after 24 hours and 48 hours were as follows: TheraCem: 89.24% and 85.46%, Panavia SA: 49.51% and 46.57% and FujiCem2: 50.63% and 47.36%. TheraCem represented the highest cell viability rate. However, no significant difference was noted between Panavia SA and FujiCem2. Time had no significant effect on cell viability. Conclusion: TheraCem exhibited the best results among three tested cements and was considered non-toxic. Panavia SA and FujiCem2 were not significantly different regarding the cell viability rate. Time had no significant effect on the cytotoxicity level of cements (AU).

Biological properties of experimental methacrylate-based sealers containing calcium phosphates

Abstract This study aimed to evaluate, in vitro and in vivo, the biocompatibility of experimental methacrylate-based endodontic sealers containing α-tricalcium phosphate (α-TCP) or nanostructured hydroxyapatite (HAp). Experimental methacrylate-based dual-cure sealers with the addition of α-TCP or HAp, at 10%wt were formulated and compared to AH Plus (AHP). Cell viability was assessed by 3-(4,5-dimethyl-thiazoyl)-2,5-diphenyl-tetrazolium bromide (MTT), and sulforhodamine B (SRB). Sealers were implanted in rats' subcutaneous tissue and histologically evaluated. Bioactivity was assessed by alkaline phosphatase enzyme activity (ALP) and Alizarin Red (AR), using apical papillary cells (SCAPs), and by the bone deposition measured in surgical cavities on rats' femur filled with AH Plus or α-TCP. In both viability assays, HAp and AHP sealers were similar, and α-TCP presented lower viability compared to the others at MTT assay (p<0.05). A gradual decrease of the inflammatory response according to the periods was observed and AHP was the only that presented giant cells (7-day period). Collagen fibers condensation increased according to the periods, with no differences among sealers. There was an increase at ALP activity and mineralized nodules deposition according to periods. HAp and α-TCP presented higher values for ALP activity at 5 days and at 5, 10, and 15 days for AR and were different from AHP (p<0.05). α-TCP presented superior values at 10 and 15 days compared to HAp and AHP for AR (p<0.05). At 90 days, α-TCP and control (empty cavity) showed high bone deposition compared to AHP (p<0.05). α-TCP and HAp, in a methacrylate-based sealer, presented biocompatibility and bioactivity, with the potential to be used as endodontic sealers in clinical practice. Further investigations are required to gain information on the physicochemical properties of these sealers formulation before its clinical implementation.

Resumo O objetivo deste estudo foi avaliar a biocompatibilidade de cimentos endodônticos experimentais à base de metacrilato contendo fosfato α-tricálcico ou hidroxiapatita nanoestruturada in vitro e in vivo. Cimentos experimentais de cura dual à base de metacrilato com a adição de fosfato de α-tricálcico (α-TCP) ou hidroxiapatita (HAp), a 10% em peso, foram formulados e comparados com AH Plus (AHP). Viabilidade celular foi avaliada por brometo de 3- (4,5-dimetil-tiazoil) -2,5-difenil-tetrazólio (MTT) e sulforodamina B (SRB). Cimentos foram implantados no tecido subcutâneo dos ratos e avaliados histologicamente. Bioatividade foi avaliada pela atividade da enzima fosfatase alcalina (ALP) e Alizarin Red (AR) utilizando células da papila apical (SCAPs) e pela deposição óssea, medida em cavidades cirúrgicas no fêmur de ratos preenchidos com AH Plus e α-TCP. Nos dois ensaios de viabilidade, HAp e AHP não apresentaram diferenças estatísticas, α-TCP apresentou menores resultados de viabilidade para o ensaio MTT (p <0,05). Resultados histológicos mostraram que houve uma diminuição do conteúdo inflamatório de acordo com os períodos, e o AHP foi o único grupo que apresentou células gigantes (período de 7 dias). A condensação das fibras colágenas aumentou conforme os períodos, sem diferenças entre os grupos. Houve aumento da atividade da ALP e deposição de nódulos mineralizados de acordo com os períodos. HAp e α-TCP apresentaram maiores valores para a atividade de ALP em 5 dias e em 5, 10 e 15 dias para AR, com diferença para o AHP (p <0,05). O α-TCP apresentou valores superiores aos 10 e 15 dias quando comparado ao HAp e AHP para AR (p <0,05). Aos 90 dias, α-TCP e controle (cavidade vazia) apresentaram maior deposição de tecido ósseo quando comparado ao AHP (p <0,05). α-TCP e HAp, presentes nos cimentos à base de metacrilato, apresentaram biocompatibilidade e potencial para serem utilizados como seladores endodônticos na prática clínica. Investigações adicionais são necessárias para obter informações sobre as propriedades físico-químicas dessas formulações de cimentos antes de sua implementação clínica.

Avaliação do efeito sinérgico entre risedronato sistêmico e genisteína local sobre o reparo peri-implantar em ratas com deficiência de estrógeno e síndrome metabólica / Evaluation of the synergistic effect between systemic Risedronate and local Genistein on peri-implant repair in strogen deficient rats and metabolic syndrome

O objetivo deste estudo foi investigar a ação sinérgica do risedronato de sódio sistêmico e da genisteína administrada localmente, através da funcionalização de implantes, de ratas submetidas a ovariectomia e com hábitos de mimetizam a síndrome metabólica. A parte in vitro deste estudo foi executado em 2 etapas. Na primeira etapa, foi realizada a funcionalização da superfície de discos/implantes com genistína na concentração de 100 µM pela técnica layer by layer (lbl). Na segunda etapa foram feitos testes biológicos em culturas de células, para avaliar as propriedades da superfície funcionalizada, quanto às respostas osteogênicas. Para a cultura de células foram utilizadas células mesenquimais diferenciadas em osteoblastos, isoladas de fêmures de ratos. Após a validação pelos testes executados nas superfícies funcionalizadas, foi realizado estudo in vivo (3ª etapa). Para tanto, no dia 0, as ratas Wistar adultas jovens, fêmeas (n=64) foram divididas em 4 grupos: 1- SHAM (n= 16), animais foram submetidos à ovariectomia (OVX) fictícia e dieta balanceada. 2- SHAM Síndrome Metabólica (SM) (n=16), animais foram submetidos à ovariectomia fictícia e dieta de cafeteria. 3- OVX SM (n=16), animais foram submetidos à ovariectomia bilateral e dieta de cafeteria. 4- OVX SM Risedronato (RIS) (n=16), animais foram submetidos à ovariectomia bilateral, dieta de cafeteria e tratadas com risedronato de sódio. Em cada grupo há 2 subgrupos: A- implantes convencionais e B- implantes funcionalizados com genisteína. No dia 30, foi iniciado o tratamento medicamentoso com risedronato de sódio, na concentração de 0,35mg/kg, ou apenas solução salina, via gavagem, 1 vez por semana. Passados 60 dias da medicação (dia 90), todos os animais foram submetidos à cirurgia para exodontia dos 1os molares superiores bilateralmente e, imediatamente, no alvéolo da raiz mesial, foi instalado os implantes com superfície convencional ou funcionalizada. Os animais foram eutanasiados aos 28 dias (dia 118) após a instalação dos implantes para mensuração do torque de falha na interface osso implante em N/cm. Os dados foram submetidos ao teste de homocedasticidade (Shapiro Wilk). Houve a confirmação de distribuição normal dos dados amostrais e na sequência, foi realizado o teste paramétrico ANOVA One Way or Two Way, seguido do pós teste de Tukey, com o nível de significância de 5% (p< 0,05). Concluiu-se que, a concentração de 100 µM da genisteína manteve a viabilidade celular e resultados favoráveis quanto a genotoxicidade. A dieta de cafeteria e a ovariectomia bilateral mimetizam a síndrome metabólica e a predisposição para osteoporose por deficiência de esteroides gonadais. E, a ação sinérgica entre fármaco sistêmico (risedronato de sódio) e genisteína local foi promissora para a melhora no processo de reparo periimplantar, principalmente no grupo SHAM e OVX SM RIS(AU)

The aim of this study was to investigate the synergistic action of systemic risedronate sodium and locally administered genistein, through implant functionalization, of rats submitted to ovariectomy and with habits mimicking the metabolic syndrome. The in vitro part of this study was performed in 2 steps. In the first step, the surface functionalization of discs/implants was performed with genistein at a concentration of 100 µM by the layer by layer (lbl) technique. In the second step biological tests were performed in cell cultures to evaluate the properties of the functionalized surface for osteogenic responses. For the cell culture, mesenchymal cells differentiated into osteoblasts, isolated from rat femurs, were used. After validation by tests performed on the functionalized surfaces, the in vivo study (third test) was performed. For this purpose, on day 0, young adult female Wistar rats (n=64) were divided into 4 groups: 1- SHAM (n=16), animals were submitted to sham ovariectomy (OVX) and balanced diet. 2- SHAM Metabolic Syndrome (MS) (n=16), animals were submitted to sham ovariectomy and cafeteria diet. 3- OVX SM (n=16), animals underwent bilateral ovariectomy and cafeteria diet. 4- OVX SM Risedronate (RIS) (n=16), animals underwent bilateral ovariectomy, cafeteria diet and treated with risedronate sodium. In each group there are 2 subgroups: A- conventional implants and B- implants functionalized with genistein. On day 30, drug treatment was started with risedronate sodium, at a concentration of 0.35 mg/kg, or just saline solution, via gavage, once a week. After 60 days of medication (day 90), all animals underwent surgery to extract the 1st upper molars bilaterally, and implants with conventional or functionalized surfaces were immediately installed in the mesial root alveolus. The animals were euthanized at 28 days (day 118) after implant installation to measure the failure torque at the implant-bone interface in N/cm. The data were submitted to the homoscedasticity test (Shapiro Wilk). The normal distribution of the sample data was confirmed and then the parametric One Way or Two Way ANOVA test was performed, followed by Tukey's post-test, with a significance level of 5% (p< 0.05). It was concluded that, the concentration of 100 µM of genistein maintained cell viability and favorable results regarding genotoxicity. The cafeteria diet and bilateral ovariectomy mimic the metabolic syndrome and predisposition to osteoporosis by gonadal steroid deficiency. And, the synergistic action between systemic drug (risedronate sodium) and local genistein was promising for the improvement in the periimplant repair process, especially in the SHAM and OVX SM RIS groups(AU)

Evaluation of physical and biological properties of conventional glass ionomer cements / Avaliação das propriedades físicas e biológicas de cimentos de ionômero de vidro convencionais

Objetivo: avaliar as propriedades físicas e biológicas dos cimentos de ionômero de vidro convencionais (CIVs). Metodologia: foram avaliados os seguintes CIVs: Fuji IX (GC Europe, Bélgica), Ketac Molar (3M ESPE, Estados Unidos), Maxxion R (FGM, Brasil) e Vitro Molar (Nova DFL, Brasil). O tempo de presa, a alteração dimensional, a radiopacidade, a sorção e a solubilidade em água foram avaliados para todos os materiais. A resistência à compressão foi analisada em intervalos de 1h, 24h, 7 dias e 28 dias; e liberação de íons fluoreto em 3h, 24h e 72h. A viabilidade celular foi avaliada após 24 e 48 horas com células de fibroblastos. A análise estatística foi realizada por meio do software SigmaPlot 12 (Systat Inc, San Jose, CA, EUA), com nível de significância estabelecido em α = 0,05. Resultado: apenas o Fuji IX teve um tempo de presa dentro da faixa recomendada pela Especificação Padrão da ADA 96 (2012), não superior a 6 minutos. Vitro Molar e Maxxion R apresentaram radiopacidade que não estava de acordo com a especificação ADA 96 (2012). Maxxion R e Vitro Molar mostraram uma alteração dimensional estatisticamente semelhante. Quanto às propriedades mecânicas, o Fuji IX foi o único CIV que apresentou aumento da resistência à compressão durante o período de avaliação de 28 dias. O Ketac Molar apresentou a maior viabilidade celular, enquanto o Maxxion R apresentou citotoxicidade severa e o maior valor cumulativo de liberação de flúor. Conclusão: Fuji IX e Ketac Molar apresentaram as propriedades físicas e biológicas mais adequadas entre os CIVs avaliados.(AU)

Objective: to evaluate the physical and biological properties of conventional glass ionomer cements (GICs). Methodology: the following GICs were evaluated: Fuji IX (GC Europe, Belgium), Ketac Molar (3M ESPE, United States), Maxxion R (FGM, Brazil) and Vitro Molar (Nova DFL, Brazil). Setting time, dimensional change, radiopacity, water solubility and water absorption were evaluated for all materials. Compressive strength was analyzed after intervals of 1h, 24h, 7 days and 28 days; and release of fluoride ions at 3 am, 24 am and 72 am. Cell viability was assessed after 24 and 48 hours with fibroblast cells. Statistical analysis was performed using SigmaPlot 12 software (Systat Inc, San Jose, CA, USA), with a significance level set at α = 0.05. Result: only the Fuji IX had an adjustment time within the range recommended by the Standard Specification of ADA 96 (2012) of not exceeding 6 minutes. Vitro Molar and Maxxion R had radiopacity that was not in accordance with the ADA 96 (2012) specification. Maxxion R and Vitro Molar showed a statistically similar dimensional change. As for mechanical properties, Fuji IX was the only GIC that showed an increase in compressive strength during the evaluation period of 28 days. Ketac Molar showed the highest cell viability, while Maxxion R showed severe cytotoxicity and the highest cumulative fluoride release value. Conclusion: Fuji IX and Ketac Molar showed the most appropriate physical and biological properties among the evaluated GICs.(AU)

Effect of a novel bioceramic root canal sealer on the angiogenesis-enhancing potential of assorted human odontogenic stem cells compared with principal tricalcium silicate-based cements

Abstract Objective: This study evaluated the angiogenesis-enhancing potential of a tricalcium silicate-based mineral trioxide aggregate (ProRoot MTA), Biodentine, and a novel bioceramic root canal sealer (Well-Root ST) in human dental pulp stem cells (hDPSCs), human periodontal ligament stem cells (hPLSCs), and human tooth germ stem cells (hTGSCs). Methodology: Dulbecco's modified Eagle's medium was conditioned for 24 h by exposure to ProRoot MTA, Biodentine, or Well-Root ST specimens (prepared according to the manufacturers' instructions). The cells were cultured in these conditioned media and their viability was assessed with 3-(4,5-dimethyl-thiazol-2-yl)-5-(3-carboxy-methoxy-phenyl)-2-(4-sulfo-phenyl)-2H tetrazolium (MTS) on days 1, 3, 7, 10, and 14. Angiogenic growth factors [platelet-derived growth factor (PDGF), basic fibroblast growth factor (FGF-2), and vascular endothelial growth factor (VEGF)] were assayed by sandwich enzyme-linked immunosorbent assay (ELISA) on days 1, 7, and 14. Human umbilical vein endothelial cell (HUVEC) migration assays were used to evaluate the vascular effects of the tested materials at 6-8 h. Statistical analyses included Kruskal-Wallis, Mann-Whitney U, and Friedman and Wilcoxon signed rank tests. Results: None of tricalcium silicate-based materials were cytotoxic and all induced a similar release of angiogenic growth factors (PDGF, FGF-2, and VEGF) (p>0.05). The best cell viability was observed for hDPSCs (p<0.05) with all tricalcium silicate-based materials at day 14. Tube formation by HUVECs showed a significant increase with all tested materials (p<0.05). Conclusion: The tricalcium silicate-based materials showed potential for angiogenic stimulation of all stem cell types and significantly enhanced tube formation by HUVECs.

Cytotoxicity of Calcium Hydroxide on Human Umbilical Cord Mesenchymal Stem Cells

Abstract Objective: To examine the cytotoxicity of calcium hydroxide on human umbilical cord mesenchymal stem cells (HUCMSC) to understand the characteristics for use in regenerative dentistry procedures especially regenerative endodontics. Material and Methods: HUCMSC was isolated, cultured, and confirmed by flow cytometry. The biological characteristics, such as cell morphology, proliferation, and protein expression, were screened. To check the cytotoxicity, HUCMSC was cultured and divided into two groups, the control group (cultured in minimum essential medium (MEM) alpha) and calcium hydroxide group (cultured in MEM alpha and calcium hydroxide). Methyl-thiazole-tetrazolium (MTT) assay was done on different concentrations of calcium hydroxide (0.39 to 25 µg/mL) and the cells were observed and counted. One-way ANOVA test was used with a significance level set at 5%. Results: Flow cytometric analysis confirmed positive of CD73, CD90, CD105, negative of CD45 and CD34. A significant difference was found between the concentration of 6.25 and 3.125 µg/mL (p=0.004). There was no significant difference among 6.25, 12.5 and 25 µg/mL concentrations. There was also no significant difference among 0.39, 0.78, 1.56, and 3.125 µg/mL concentrations. Conclusion: Even though calcium hydroxide is a medicament of choice in clinical endodontics, it decreases the viability of HUCMSC. The lower the concentration of calcium hydroxide, the higher the viability of HUCMSC.

Evaluating of cytotoxic effects of highly esthetic dental composites / Avaliação dos efeitos citotóxicos de resinas compostas altamente estéticas

Objective: Dental composites developed by using nanotechnology in the field of dentistry are widely used in the treatment of anterior and posterior teeth. This study aimed to investigate the cytotoxic effects of dental composites of different particle size on L929 mouse fibroblast cell line by extract test method in vitro. Material and Methods: Composite samples of 8 x 2 mm diameter were prepared by polymerizing with led light device by using glass mod in a sterile cabinet. Composite samples of which surface areas were calculated according to ISO standards (3 cm2 / ml), were incubated for 24 and 72 hours, at 37 o C. cell viability was assessed by 3-[4,5-dimethylthiazole-2- yl]-2,5-diphenyltetrazolium bromide (MTT) assay and cell death was evaluated by the lactate dehydrogenase (LDH) leakage assay. Results: The 1:1 extracts of the composites at the end of 24 hours (except for nanoceramic composite) showed no toxic effect. When the cell viability results of the 1:1 extracts of the composite samples at the end of 72 hours were statistically analyzed, significant differences were found comparing to the control group (p < 0.05). Conclusion: It was observed that the type and size of the filler were effective on the toxicity of the composites, and the composites containing Bis-GMA, TEGDMA, UDMA and Bis EMA monomers in their organic matrix showed acceptable cell viability (70%) as specified by ISO. However, the composites with PEGDMA and BPA monomers in their organic matrix showed poor cell viability, which is below the acceptable level of 70%, and were found to have a toxic effect. (AU)

Objetivo: As resinas compostas desenvolvidas pela nanotecnologia no campo da odontologia são amplamente utilizadas no tratamento de dentes anteriores e posteriores. Este estudo teve como objetivo investigar os efeitos citotóxicos de resinas compostas de diferentes tamanhos de partículas na linha celular de fibroblastos de camundongos L929 pelo método de teste de extrato in vitro. Material e Métodos: Amostras compostas de 8 x 2 mm de diâmetro foram preparadas por polimerização com dispositivo de luz led usando um molde de vidro em um gabinete estéril. Amostras de resinas cujas áreas de superfície foram calculadas de acordo com os padrões ISO (3 cm2 / ml), foram incubadas por 24 e 72 horas, a 37 o C. A viabilidade celular foi avaliada pelo ensaio de brometo de 3- [4,5-dimetiltiazol-2- il] -2,5-difeniltetrazólio (MTT) e a morte celular foi avaliada pelo ensaio de infiltração de lactato desidrogenase (LDH). Resultados: Os extratos 1: 1 dos compósitos ao final de 24 horas (exceto o composto nanocerâmico) não apresentaram efeito tóxico. Quando os resultados de viabilidade celular dos extratos 1: 1 das amostras compostas ao final de 72 horas foram analisados, estatisticamente, foram encontradas diferenças significativas em relação ao grupo controle (p < 0,05). Conclusão: Observou-se que o tipo e tamanho da carga foram eficazes na toxicidade dos compósitos, e os compósitos contendo os monômeros Bis-GMA, TEGDMA, UDMA e Bis EMA em sua matriz orgânica apresentaram viabilidade celular aceitável (70%) como especificado pela ISO. No entanto, os compósitos com monômeros PEGDMA e BPA em sua matriz orgânica apresentaram baixa viabilidade celular, que está abaixo do nível aceitável de 70%, e foram encontrados como tendo um efeito tóxico. (AU)

Avaliação da citotoxicidade e resposta tecidual de soluções irrigadoras experimentais a partir de biovidro e biovidro modificado com cobalto / Evaluation of cytotoxicity and tissue response of experimental irrigating solutions from bioglass and bioglass modified with cobalt

Introdução: Novas formulações de materiais vítreos apresentaram atividade antimicrobiana e osteoindutora. Objetivos: O objetivo deste estudo foi avaliar a citotoxicidade e a biocompatibilidade de soluções produzidas a partir de novas formulações de biovidros, o F18 (biovidro experimental) e o F18 com Cobalto (F18-Co; biovidro experimental dopado com cobalto), comparadas à água de cal, obtida a partir de solução com hidróxido de cálcio (Ca(OH)2). Material e métodos: O F18 foi preparado e moído, e parte deste foi dopada com cobalto. As soluções foram preparadas com cada material (1:10 de pó para água), formando os grupos F18, F18-Co e Ca(OH)2. Células L929 foram cultivadas, e a viabilidade celular avaliada a partir das soluções e de suas diluições (½, », 1/8 e 1/16 diluição) pelo teste MTT, após 24 e 48 horas. Para avaliar a biocompatibilidade, tubos de polietileno foram preenchidos com esponjas de fibrina embebidas nas soluções não diluídas, e tubos embebidos em soro fisiológico serviram de controle. Os tubos foram implantados no dorso de 16 ratos. Após 7 e 30 dias (n = 8), os ratos foram eutanasiados, e os tubos com o tecido circundante foram processados para coloração de hematoxilina-eosina (H.E.) e análise da inflamação através de escores. Os dados paramétricos (citotoxicidade) foram submetidos aos testes de normalidade para definir o teste estatístico a ser empregado, e os dados não-paramétricos (H.E.), foram avaliados pelos testes de Kruskal Wallis e Dunn (p < 0,05). Resultados: As soluções não diluídas dos materiais, e as diluições de ½ e », reduziram a viabilidade celular em 24 h (p < 0,05). As diluições de 1/8 e 1/16 do F18 e F18Co apresentaram viabilidade celular semelhante ao controle (p > 0,05), o que não ocorreu com Ca(OH)2 (p < 0,05), que foi citotóxico. Em 48 h, apenas as soluções não diluídas e diluições de ½ e » do F18 foram similares ao controle (p > 0,05), e as demais foram citotóxicas. Mas as diluições de 1/8 e 1/16 do F18Co teve um aumento na viabilidade celular comparadas às soluções do Ca(OH)2 (p < 0,05), e foram semelhantes ao controle (p > 0,05). Aos 7 dias, controle, F18 e F18-Co apresentaram inflamação moderada, e Ca(OH)2, severa (p > 0,05); a cápsula fibrosa foi espessa. Aos 30 dias, controle e F18-Co apresentaram inflamação leve comparados ao F18, com inflamação moderada (p < 0,05); Ca(OH)2 teve inflamação leve ( p > 0,05); a cápsula fibrosa foi fina na maior parte dos espécimes . Conclusões: Soluções experimentais de F18 e F18 dopado com cobalto são citocompatíveis, diferentemente da solução de Ca(OH)2; todas as soluções apresentaram biocompatibilidade(AU)

Introduction: New formulations of vitreous materials showed antimicrobial and osteoinductive activity. Objectives: The aim of this study was to evaluate the cytotoxicity and biocompatibility of solutions produced from new formulations of bioglass, F18 (experimental bioglass) and F18 with Cobalt (F18-Co; experimental bioglass doped with cobalt), compared to lime water, obtained from a solution with calcium hydroxide (Ca(OH)2). Material and methods: The F18 was prepared and ground, and part of it was doped with cobalt. The solutions were prepared with each material (1:10 powder to water), forming groups F18, F18-Co and Ca(OH)2. L929 cells were cultured, and cell viability assessed from solutions and from its dilutions (½, », 1/8 and 1/16 dilution) by the MTT test, after 24 and 48 hours. For biocompatibility analysis, polyethylene tubes were filled with fibrin sponges embedded in the non-diluted solutions, and tubes embedded in saline solution served as controls. The tubes were implanted on the dorsum of 16 rats. After 7 and 30 days (n = 8), the rats were euthanized, and the tubes with the surrounding tissue were processed for staining of hematoxylin-eosin (H.E.) and analysis of inflammation through scores. The parametric data (cytotoxicity) were subjected to normality tests to define the statistical test to be used, and the nonparametric data (H.E.), were evaluated by the Kruskal Wallis and Dunn tests (p < 0.05). Results: Undiluted solutions of the materials, and dilutions of ½ and », reduced cell viability in 24 h (p < 0.05). The 1/8 and 1/16 dilutions of F18 and F18-Co showed cell viability similar to the control (p > 0.05), which did not occur with Ca(OH)2 (p <0.05), which was cytotoxic. At 48 h, only undiluted solutions and dilutions of ½ and » of F18 were similar to the control (p > 0.05), and the others were cytotoxic. However, the 1/8 and 1/16 dilutions of F18-Co had an increase in cell viability compared to Ca(OH)2 solutions (p <0.05), and were similar to the control (p > 0.05). At 7 days, control, F18 and F18-Co showed moderate inflammation, and Ca(OH)2, severe (p > 0.05); the fibrous capsule was thick. At 30 days, control and F18-Co showed mild inflammation compared to F18, with moderate inflammation (p < 0.05); Ca(OH)2 had mild inflammation (p > 0.05); the fibrous capsule was thin in most specimens. Conclusions: Experimental solutions of F18 and F18 doped with cobalt are cytocompatible, unlike the Ca(OH) 2 Keywords: solution; all solutions showed biocompatibility(AU)

Effect of Ions Released and pH of Two Glass Ionomer Cements in Human Gingival Fibroblasts / Efecto de la liberación de iones y pH de dos cementos de ionómero de vidrio en fibroblastos gingivales humanos

Abstract 20. Conventional glass ionomer cements are used as dental provisional restorative materials, which present several advantages such as adhesion to the tooth mineral phase among others. On the other hand, the knowledge about biological property of glass ionomers shows various approaches and results. In this work, it was studied the in vitro biological response of human gingival fibroblasts in contact with commercial cements of glass ionomer: Mirafil® and Ionglass® and with their extracts, according to ISO 10993. The extracts of the cements, in which the cells were cultured, were adjusted at different concentrations ranging 0.1% to 100%. The cellular metabolic activity of gingival fibroblasts was measured using the Alamar Blue® reagent. The results showed a significant effect on the cellular metabolic activity correlated with the concentration of liberated ions (Al³+ and Ca²+) for both ionomers, as well as the pH variations of the culture media. This could mean that the cellular metabolic activity is substantially influenced by ions and pH of the cell culture.

Resumen 24. Los cementos de ionómero de vidrio convencionales se utilizan como materiales de restauración provisional para uso dental, los cuales presentan varias ventajas como la adhesión a la fase mineral de los dientes. Por otro lado, las propiedades biológicas de los ionómeros de vidrio muestran diversos enfoques y resultados. En éste trabajo se estudió la respuesta biológica in vitro de fibroblastos gingivales humanos en contacto con cementos comerciales de ionómero de vidrio: Mirafil® e Ionglass® y con sus respectivos extractos según la norma ISO 10993. Los extractos de los cementos en los que se cultivaron las células estaban en diferentes concentraciones: de 0.1% a 100%. La actividad metabólica celular se midió usando el reactivo Alamar Blue®. Los resultados mostraron un efecto significativo sobre la actividad metabólica celular correlacionada con la concentración de iones liberados (Al³+ y Ca²+) para ambos ionómeros, así como las variaciones de pH de los medios de cultivo. Ello podria explicar la influencia por los iones y el pH del cultivo celular en la actividad metabólica celular.

Efeito do substituto ósseo particulado associado ou não ao MTA na citotoxicidade, resposta tecidual e reparo ósseo em defeitos críticos em calvária de ratos / Effect of particulate bone substitute associated or not to MTA on cytotoxicity, tissue response and bone repair in critical defects in rats calvaria

A utilização de biomateriais que visam devolver o volume ósseo perdido após a perda dentária tem se expandido. O osso é um tecido conjuntivo altamente especializado, possuindo dinâmica aposicional onde o equilíbrio entre neoformação e reabsorção envolve a interação de fatores endócrinos, parácrinos e autócrinos. Diversas associações de materiais diversos e substitutos ósseos têm sido estudadas, porém, o MTA (Agregado Trióxido Mineral) ainda carece de informações acerca da sua utilização como substituto ósseo ou em associação com demais substitutos. Assim, o objetivo deste estudo foi avaliar a influência da presença do MTA Angelus Branco® nas proprorções de 5%, 10% e 15% em associação com substituto ósseo de Hidroxiapatita e ß- Tricálcio Fosfato na citotoxicidade, resposta tecidual e reparo ósseo, em defeito crítico em calvária de ratos. Para tanto, utilizou-se cultura celular da SAOS-2 para avaliação citológica e ensaio MTT do contato direto e eluentes. Também foram utilizados 112 ratos machos Wistar distribuídos em 7 grupos e avaliados em 2 tempos (7 e 28 dias). Após eutanasiados foram submetidos à microtomografia e por coloração de hematoxilina e eosina para análise histológica e histomorfométrica. A análise da homocedasticidade foi realizada pelo teste Shapiro-Wilk, para distinção dos dados paramétricos e não paramétricos. Para análise dos dados paramétricos, foi realizada análise de variância One-Way (ANOVA One-Way) e realizado o pós-teste de Tukey para os parâmetros microtomográficos e para a citotoxicidade foi aplicado o teste de correção de Bonferroni. Para os dados não paramétricos utilizou-se o Kruskal-Wallis e Mann Whitney. Adotou-se o nível de significância de p <0,05. Os parâmetros volume ósseo (BV), porcentagem de volume ósseo (BV/TV), espessura do trabeculado ósseo (Tb.Th) e número de trabéculas (Tb.N) não apresentaram diferenças estatísticas. Sendo o grupo Osteosynt® +15% MTA o grupo com maiores valores médios para os parâmetros BV, BV/TV, Tb.Th e Tb.N. A análise histomorfométrica para neoformação óssea e reabsorção dos biomateriais não apresentaram diferenças estatísticas. De acordo com os resultados, pode-se concluir que, estatisticamente, a adição de MTA nas 3 proporções testadas não demonstrou efetividade quando comparada ao uso isolado dos substitutos ósseos Bio-Oss® e Osteosynt® e coágulo. No entanto, foi evidenciada tendências à superioridade numérica quanto ao volume ósseo neoformado da associação Osteosynt® + 15% MTA aos 7 dias e Osteosynt® + 5% MTA e Osteosynt® + 10% MTA nas análises aos 28 dias(AU)

The use of biomaterials that aim to return lost bone volume after tooth loss has expanded. Bone is a highly specialized connective tissue with appositional dynamics where the balance between neoformation and resorption involves the interaction of endocrine, paracrine and autocrine factors. Several associations of different materials and bone substitutes have been studied. However, the MTA (Mineral Trioxide Aggregate) still lacks information about its use as bone substitute or in association with other substitutes. Thus, the aim of this study was to evaluate the influence of presence of MTA Angelus Branco® in the 5%, 10% and 15% proportions in association with Hydroxyapatite and ß-Tricalcium Phosphate bone substitute in cytotoxicity, tissue response and bone repair in critical defect in rat calvaria. For this, SAOS-2 cell culture was used for cytological evaluation and MTT assay of direct contact and eluents. We also used 112 male Wistar rats distributed into 7 groups and evaluated at 2 distinct times (7 and 28 days). After euthanasia, the specimens were submitted to microtomography and hematoxylin and eosin staining for histological and histomorphometric analysis. The analysis of homoscedasticity was performed by the Shapiro-Wilk test for distinction of parametric and non-parametric data. For analysis of parametric data, one-way analysis of variance (One-Way ANOVA) was performed and Tukey's post-test was performed for the microtomographic parameters and for the cytotoxicity the Bonferroni correction test was applied. For non-parametric data we used the Kruskal-Wallis and Mann Whitney. The significance level of p <0.05 was adopted. The parameters bone volume (BV), bone volume percentage (BV / TV), bone trabecular thickness (Tb.Th) and number of trabeculae (Tb.N) showed no statistical differences. The Osteosynt® + 15% MTA group was the group with the highest mean values for the parameters BV, BV / TV, Tb.Th and Tb.N. Histomorphometric analysis for bone neoformation and biomaterial resorption showed no statistical differences. According to the results, it can be concluded that, statistically, the addition of MTA in the 3 tested proportions did not show effectiveness when compared to the isolated use of the Bio-Oss® and Osteosynt® bone substitutes and clot. However, trends in numerical superiority regarding the newly formed bone volume of the association Osteosynt® + 15% MTA at 7 days and Osteosynt® + 5% MTA and Osteosynt® + 10% MTA in the analyzes at 28 days were evidenced(AU)

Composite-derived monomers affect cell viability and cytokine expression in human leukocytes stimulated with Porphyromonas gingivalis

Abstract Objectives: Dental composites release unreacted resin monomers into the oral environment, even after polymerization. Periodontal cells are, therefore, exposed to substances that potentially elicit the immune inflammatory response. The underlying molecular mechanisms associated with the interaction between resin monomers and human immune cells found in the gingival crevicular fluid are not fully understood yet. This study investigated the ability of bisphenol A-glycidyl methacrylate (BISGMA), urethane dimethacrylate (UDMA) and triethylene glycol dimethacrylate (TEGDMA) to induce apoptosis and cytokine release by human leukocytes stimulated with a periodontal pathogen. Methodology: Peripheral blood mononuclear cells (PBMC) from 16 healthy individuals were included in this study. To determine the toxicity, the PBMC were incubated for 20 hours, with monomers, for the analysis of cell viability using MTT assay. To evaluate cell death in the populations of monocytes and lymphocytes, they were exposed to sub-lethal doses of each monomer and of heat-inactivated Porphyromonas gingivalis (P. gingivalis) for 5 hours. Secretions of IL-1β, IL-6, IL-10 and TNF-α were determined by ELISA after 20 hours. Results: UDMA and TEGDMA induced apoptosis after a short-time exposure. Bacterial challenge induced significant production of IL-1β and TNF-α (p<0.05). TEGDMA reduced the bacterial-induced release of IL-1β and TNF-α, whereas UDMA reduced IL-1β release (p<0.05). These monomers did not affect IL-10 and IL-6 secretion. BISGMA did not significantly interfere in cytokine release. Conclusions: These results show that resin monomers are toxic to PBMC in a dose-dependent manner, and may influence the local immune inflammatory response and tissue damage mechanisms via regulation of bacterial-induced IL-1β and TNF-α secretion by PBMC.

Increased whitening efficacy and reduced cytotoxicity are achieved by the chemical activation of a highly concentrated hydrogen peroxide bleaching gel

Abstract Objective This study was designed for the chemical activation of a 35% hydrogen peroxide (H2O2) bleaching gel to increase its whitening effectiveness and reduce its toxicity. Methodology First, the bleaching gel - associated or not with ferrous sulfate (FS), manganese chloride (MC), peroxidase (PR), or catalase (CT) - was applied (3x 15 min) to enamel/dentin discs adapted to artificial pulp chambers. Then, odontoblast-like MDPC-23 cells were exposed for 1 h to the extracts (culture medium + components released from the product), for the assessment of viability (MTT assay) and oxidative stress (H2DCFDA). Residual H2O2 and bleaching effectiveness (DE) were also evaluated. Data were analyzed with one-way ANOVA complemented with Tukey's test (n=8. p<0.05). Results All chemically activated groups minimized MDPC-23 oxidative stress generation; however, significantly higher cell viability was detected for MC, PR, and CT than for plain 35% H2O2 gel. Nevertheless, FS, MC, PR, and CT reduced the amount of residual H2O2 and increased bleaching effectiveness. Conclusion Chemical activation of 35% H2O2 gel with MC, PR, and CT minimized residual H2O2 and pulp cell toxicity; but PR duplicated the whitening potential of the bleaching gel after a single 45-minute session.

The influence of Aloe vera with mesenchymal stem cells from dental pulp on bone regeneration: characterization and treatment of non-critical defects of the tibia in rats

Abstract Objective This study aimed to evaluate the inflammatory effect and bone formation in sterile surgical failures after implantation of a collagen sponge with mesenchymal stem cells from human dental pulp (hDPSCs) and Aloe vera. Material and Methods Rattus norvegicus (n=75) were divided into five experimental groups according to treatment: G1) control (blood clot); G2) Hemospon®; G3) Hemospon® in a culture medium enriched with 8% Aloe vera; G4) Hemospon® in a culture medium containing hDPSCs and G5) Hemospon® in a culture medium enriched with 8% Aloe vera and hDPSCs. On days 7, 15 and 30, the animals were euthanized, and the tibia was dissected for histological, immunohistochemistry and immunofluorescence analyses. The results were analyzed using nonparametric Kruskal-Wallis test and Dunn's post-test. Results On days 7 and 15, the groups with Aloe vera had less average acute inflammatory infiltrate compared to the control group and the group with Hemospon® (p<0.05). No statistically significant difference was found between the groups regarding bone formation at the three experimental points in time. Osteopontin expression corroborated the intensity of bone formation. Fluorescence microscopy revealed positive labeling with Q-Tracker® in hDPSCs before transplantation and tissue repair. Conclusion The results suggest that the combination of Hemospon®, Aloe vera and hDPSCs is a form of clinical treatment for the repair of non-critical bone defects that reduces the inflammatory cascade's effects.

Root canal dressings for revascularization influence in vitro mineralization of apical papilla cells

Abstract Endodontic revascularization is based on cell recruitment into the necrotic root canal of immature teeth after chemical disinfection. The clinical outcome depends on the ability of surviving cells from the apical tissue to differentiate and promote hard tissue deposition inside the dentinal walls. Objective To investigate the effect of calcium hydroxide (CH) and modified triple antibiotic paste (mTAP - ciprofloxacin, metronidazole and cefaclor) on the viability and mineralization potential of apical papilla cells (APC) in vitro . Material and Methods APC cultures were kept in contact with CH or mTAP (250-1000 µg/mL) for 5 days, after which cell viability was assessed using 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Next, APCs were subjected to CH or mTAP at 250 µg/mL for 5 days before inducing the differentiation assay. After 14 and 21 days, calcium deposition was assessed by the Alizarin Red S staining method, followed by elution and quantification using spectrophotometry. Data were analyzed using ANOVA followed by Tukey post hoc test. Results CH induced cell proliferation, whereas mTAP showed significant cytotoxicity at all concentrations tested. APC treated with CH demonstrated improved mineralization capacity at 14 days, while, for mTAP, significant reduction on the mineralization rate was observed for both experimental periods (14 and 21 days). Conclusion Our findings showed that CH induces cell proliferation and improves early mineralization, whereas mTAP was found cytotoxic and reduced the mineralization potential in vitro of APCs.

Effects of MTA and Brazilian propolis on the biological properties of dental pulp cells

Abstract: The aim of this study was to evaluate the effect of mineral trioxide aggregate (MTA) and Brazilian propolis on the cell viability, mineralization, anti-inflammatory ability, and migration of human dental pulp cells (hDPCs). The cell viability was evaluated with CCK-8 kit after 1, 5, 7, and 9 days. The deposition of calcified matrix and the expression of osteogenesis-related genes were evaluated by Alizarin Red staining and real-time PCR after incubation in osteogenic medium for 21 days. The expression of inflammation-related genes in cells was determined after exposure to 1 μg/mL LPS for 3 h. Finally, the numbers of cells that migrated through the permeable membranes were compared during 15 h. Propolis and MTA significantly increased the viability of hDPCscompared to the control group on days 7 and 9. In the propolis group, significant enhancement of osteogenic potential and suppressed expression of IL-1β and IL-6 was observed after LPS exposure compared to the MTA and control groups. The number of migration cells in the propolis group was similar to that of the control group, while MTA significantly promoted cell migration. Propolis showed comparable cell viability to that of MTA and exhibited significantly higher anti-inflammatory and mineralization promotion effects on hDPCs.

Addition of zirconium oxide to Biodentine increases radiopacity and does not alter its physicochemical and biological properties

Abstract Objectives: To evaluate the radiopacity of Biodentine (BD) and BD associated with 15% calcium tungstate (BDCaWO4) or zirconium oxide (BDZrO2), by using conventional and digital radiography systems, and their physicochemical and biological properties. Materials and Methods: Radiopacity was evaluated by taking radiographs of cement specimens (n=8) using occlusal film, photostimulable phosphor plates or digital sensors. Solubility, setting time, pH, cytocompatibility and osteogenic potential were also evaluated. Data were analyzed using one-way ANOVA and Tukey post-test or two-way ANOVA and Bonferroni post-test (α=0.05). Results: BD radiopacity was lower than 3 mm Al, while BD ZrO2 and BD CaWO4 radiopacity was higher than 3 mm Al in all radiography systems. The cements showed low solubility, except for BDCaWO4. All cements showed alkaline pH and setting time lower than 34 minutes. MTT and NR assays revealed that cements had greater or similar cytocompatibility in comparison with control. The ALP activity in all groups was similar or greater than the control. All cements induced greater production of mineralized nodules than control. Conclusions: Addition of 15% ZrO2 or CaWO4 was sufficient to increase the radiopacity of BD to values higher than 3 mm Al. BD associated with radiopacifiers showed suitable properties of setting time, pH and solubility, except for BDCaWO4, which showed the highest solubility. All cements had cytocompatibility and potential to induce mineralization in Saos-2 cells. The results showed that adding 15% ZrO2 increases the radiopacity of BD, allowing its radiography detection without altering its physicochemical and biological properties.

Cytotoxicity of intracanal dressings on apical papilla cells differ upon activation with E. faecalis LTA

Abstract Objective The aim of this study was to investigate the cytotoxic effects of modified triple antibiotic paste and an experimental composition using calcium hydroxide on lipoteichoic acid (LTA)-primed apical papilla cells (APC). Material and Methods Human APC were tested for in vitro cytotoxicity of modified Triple Antibiotic Paste (mTAP - Ciprofloxacin, Metronidazole and Cefaclor at 1:1:1) and of a paste of Ciprofloxacin, Metronidazole and Calcium hydroxide (CMC - 1:1:2) and modified CMC (mCMC - 2:2:1) by using MTT assay. The substances were reconstituted in DMEM at 1,000 µg/mL and » serially diluted before being kept in contact with cells for 1, 3, 5 and 7 days. Further, cells were primed with 1 µg/mL of Enterococcus faecalis LTA for 7 days prior to the viability test with 1,000 µg/mL of each substance. Statistical analysis was performed using one-way analysis of variance (ANOVA) and two-way ANOVA respectively followed by Tukey's post-test. Significance levels were set at p<0.05. Results In the first assay, the higher cytotoxic rates were reached by mTAP for all experimental periods. CMC was found toxic for APC at 5 and 7 days, whereas mCMC did not affect the cell viability. Only CMC and mCMC were able to induce some cellular proliferation. In the second assay, when considering the condition with medium only, LTA-primed cells significantly proliferated in comparison to LTA-untreated ones. At this context, mTAP and CMC showed similar cytotoxicity than the observed for LTA-untreated cells, while mCMC was shown cytotoxic at 7 days only for LTA-primed APC. Comparing the medications, mTAP was more cytotoxic than CMC and mCMC. Conclusion mTAP showed higher cytotoxicity than CMC and mCMC and the effect of topic antimicrobials might differ when tested against apical papilla cells under physiological or activated conditions.

Effect of ProRoot MTA® and Biodentine® on osteoclastic differentiation and activity of mouse bone marrow macrophages

Abstract Objectives This investigation aimed to assess the differentiation inhibitory effects of ProRoot MTA® (PMTA) and Biodentine® (BIOD) on osteoclasts originated from murine bone marrow macrophages (BMMs) and compare these effects with those of alendronate (ALD). Materials and Methods Mouse BMMs were cultured to differentiate into osteoclasts with macrophage colony-stimulating factor and receptor activator of NF-κB (RANKL), treated with lipopolysaccharide. After application with PMTA, BIOD, or ALD, cell toxicities were examined using WST-1 assay kit, and RANKL-induced osteoclast differentiation and activities were determined by resorption pit formation assay and tartrate-resistant acid phosphate (TRAP) staining. The mRNA levels of osteoclast activity-related genes were detected with quantitative real time polymerase chain reaction. Expressions of molecular signaling pathways were assessed by western blot. All data were statistically analyzed with one-way ANOVA and Tukey's post-hoc test (p<0.05). Results Mouse BMMs applied with PMTA, BIOD, or ALD showed highly reduced levels of TRAP-positive osteoclasts. The BIOD treated specimens suppressed mRNA expressions of cathepsin K, TRAP, and c-Fos. Nonetheless, it showed a lower effect than PMTA or ALD applications. Compared with ALD, PMTA and BIOD decreased RANKL-mediated phosphorylation of ERK1/2 and IκBα. Conclusions PMTA and BIOD showed the inhibitory effect on osteoclast differentiation and activities similar to that of ALD through IκB phosphorylation and suppression of ERK signaling pathways.