RESUMO
The aim of this study was to evaluate the transdentinal cytotoxicity of experimental adhesive systems (EASs) with different hydrophilicity and dentin saturation solutions on odontoblast-like cells. One hundred 0.4-mm-thick dentin discs were mounted in in vitro pulp chambers and assigned to 10 groups. MDPC-23 cells were seeded onto the pulpal side of the discs, incubated for 48h. The EASs with increasing hydrophilicity (R1, R2, R3 and R4) were applied to the occlusal side after etching and saturation of etched dentin with water or ethanol. R0 (no adhesive) served as controls. R1 is a non-solvated hydrophobic blend, R2 is similar to a simplified etch-and-rinse adhesive system and R3 and R4 are similar to self-etching adhesives. After 24h, cell metabolism was evaluated by MTT assay (n=8 discs) and cell morphology was examined by SEM (n=2 discs). Type of cell death was identified by flow cytometry and the degree of monomer conversion (%DC) was determined by infrared spectroscopy (FTIR) after 10s or 20s of photoactivation. Data were analyzed by the Kruskal-Wallis and Mann-Whitney tests (α=0.05). Dentin saturation with ethanol resulted in higher necrotic cell death ratios for R2, R3 and R4 compared with water saturation, although R2 and R3 induced higher SDH production. Photoactivation for 20s significantly improved the %DC of all EASs compared with 10s. A significant positive correlation was observed between the degree of hydrophilicity and %DC. In conclusion, except for R1, dentin saturation with ethanol increased the cytotoxicity of EASs, as expressed by the induction of necrotic cell death.
Assuntos
Adesivos Dentinários/toxicidade , Dentina/efeitos dos fármacos , Etanol/farmacologia , Odontoblastos/efeitos dos fármacos , Solventes/farmacologia , Condicionamento Ácido do Dente/métodos , Animais , Bis-Fenol A-Glicidil Metacrilato/química , Bis-Fenol A-Glicidil Metacrilato/toxicidade , Técnicas de Cultura de Células , Morte Celular/efeitos dos fármacos , Linhagem Celular , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Colagem Dentária/métodos , Permeabilidade da Dentina/efeitos dos fármacos , Adesivos Dentinários/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Cura Luminosa de Adesivos Dentários , Metacrilatos/química , Metacrilatos/toxicidade , Camundongos , Necrose , Organofosfatos/química , Organofosfatos/toxicidade , Polietilenoglicóis/química , Polietilenoglicóis/toxicidade , Polimerização , Ácidos Polimetacrílicos/química , Ácidos Polimetacrílicos/toxicidade , Água/química , para-Aminobenzoatos/química , para-Aminobenzoatos/toxicidadeRESUMO
This study evaluated the cytotoxicity of experimental adhesive systems (EASs) on odontoblast-like cells. Paper discs (n = 132) were impregnated with 10 µL of each EAS-R1, R2, R3, R4, and R5 (in an ascending order of hydrophilicity), followed by photoactivation. R1 and R2 are nonsolvated hydrophobic blends, R3 represents a simplified etch-and-rinse adhesive system, and R4 and R5 represent simplified self-etch adhesive systems. Discs were immersed in Dulbecco's modified Eagle's medium for 24 h to obtain eluates applied on MDPC-23 cell cultures. No material was applied on discs used as control (R0). Cell viability [3-(4,5-dimethythiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay], total protein (TP) production, alkaline phosphatase (ALP) activity, type of cell death, and degree of monomer conversion Fourier transform infrared (%DC-FTIR) were evaluated. Data were analyzed by Kruskal-Wallis and Mann-Whitney tests (α = 0.05). Considering R0 (control) as having 100% of cell viability, R1, R2, R3, R4, and R5 reduced the metabolic activity of cells by 36.4, 3.1, 0.2, 21.5, and 65.7%, respectively, but only R1 and R5 differed from R0. Comparing with R0, lower TP production was observed for R1, R4, and R5, while ALP activity decreased for R1 and R5. Necrotic cell death was predominant for all EASs, but only R1, R4, and R5 differed from R0. Only R5 presented a different apoptotic cell death ratio from R0. R1 presented the lowest %DC (ca. 37%), whereas R4 and R5 presented the highest (ca. 56%). In conclusion, R2 and R3 were not toxic to the MDPC-23 cells, suggesting that the degree of hydrophilicity or %DC of the EASs alone were not responsible for their cytopathic effects.
Assuntos
Materiais Biocompatíveis/química , Adesão Celular/efeitos dos fármacos , Odontoblastos/citologia , Odontoblastos/efeitos dos fármacos , Fosfatase Alcalina/metabolismo , Animais , Apoptose , Morte Celular , Linhagem Celular , Sobrevivência Celular , Difusão , Citometria de Fluxo , Interações Hidrofóbicas e Hidrofílicas , Teste de Materiais , Camundongos , Necrose , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier , Sais de TetrazólioRESUMO
O objetivo geral desse trabalho, dividido em dois experimentos (capítulos 1 e 2), foi avaliar a citotoxicidade de sistemas adesivos experimentais (SAEs), com diferentes graus de hidrofilia, e do etanol como solução de solvatação da dentina, sobre células odontoblastóides. No capítulo 1, discos de papel filtro esterilizados foram impregnados com 10 uL de cada SAE (n=22): R1, R2, R3, R4 e R5 (em ordem crescente de hidrofilia), seguido de fotoativação. Os discos foram individualmente imersos em meio de cultura DMEM para obtenção de extratos (DMEM + componentes liberados dos SAEs), os quais foram posteriormente aplicados sobre células MDPC-23 em cultura. Discos não impregnados (R0) serviram como controle. Foram avaliados o metabolismo celular (teste de MTT), a expressão de proteína total (PT) e a atividade de fosfatase alcalina (FA), além do tipo de morte celular (citometria de fluxo) e grau de conversão monomérica (FTIR) dos SAEs. Os dados de cada variável resposta do estudo foram analisados por testes de Kruskal-Wallis e Mann-Whitney (α=0,05). Considerando R0 como 100%, foi observada redução do metabolismo celular de 36,4%, 3,1%, 0,2%, 21,5% e 65,7%, respectivamente para R1, R2, R3, R4 e R5. Apenas R1 e R5 diferiram estatisticamente do controle. Para PT, R1, R4 e R5 tiveram expressão estatisticamente inferior ao controle, enquanto que a atividade de FA foi significativamente reduzida por R1 e R5. Esses mesmos SAEs juntamente com R4 induziram as maiores porcentagens de morte...
The overall aim of this study, divided into two experiments, was to evaluate the cytotoxicity of experimental adhesive systems (EAS) with different hydrophilicity, and ethanol as a dentin solvation solution, on odontoblast-like cells. In the first experiment, sterilized filter paper discs were impregnated with 10 uL of each EAS: R1, R2, R3, R4 and R5 (in increasing rank of hydrophilicity), followed by light activation. The paper discs were individually immersed in DMEM culture medium for obtaining the extracts (DMEM + released components of EAS), which were applied on MDPC-23 cells in culture. Non-impregnated paper discs (R0) were used as control. Cell metabolism (MTT assay), total protein expression (TP) and alkaline phosphatase activity (ALP) were assessed, in addition to the type of cell death (flow cytometry) and the degree of monomer conversion (FTIR). Data for each response variable were submitted to Kruskal-Wallis and Mann-Whitney tests (α=0.05). Compared to the control (100%), cell metabolism was decreased by 36.4%, 3.1%, 0.2%, 21.5% and 65.7% for R1, R2, R3, R4 and R5, respectively. However, only R1 and R5 differed from the control. R1, R4 and R5 decreased the expression of TP compared to the control, whereas only R1 and R5 significantly reduced the activity of ALP. The later EAS plus R4 caused the highest percentages of cell death by necrosis. A higher percentage of monomer conversion was detected as a function of the hydrophilicity. According to the experimental conditions, it could be...