Mechanical-physicochemical properties and biocompatibility of catechin-incorporated adhesive resins

Abstract Several anti-proteolytic dentin therapies are being exhaustively studied in an attempt to reduce dentin bond degradation and improve clinical performance and longevity of adhesive restorations. Objectives This study assessed the effect of epigallocatechin-3-gallate (EGCG) on long-term bond strength when incorporated into adhesives. Material and Methods Adhesive systems were formulated with EGCG concentrations of 0 wt%: (no EGCG; control); 0.5 wt% EGCG; 1.0 wt% EGCG, and 1.5 wt% EGCG. Flexural strength (FS), modulus of elasticity (ME), modulus of resilience (MR), compressive strength (CS), degree of conversion (DC), polymerization shrinkage (PS), percentage of water sorption (%WS), percentage of water solubility (%WL) and cytotoxicity properties were tested. Dentin microtensile bond strength (µTBS) was evaluated after 24 h and again after 6 months of water storage. The adhesive interface was analyzed using scanning electron microscopy (SEM). Results No significant differences were found among the groups in terms of FS, ME, MR, CS and PS. EGCG-doped adhesives increased the DC relative to the control group. EGCG concentrations of 1.0 wt% and 0.5 wt% decreased the WS of adhesives. WL decreased in all cases in which EGCG was added to adhesives, regardless of the concentration. EGCG concentrations of 1.0 wt% and 0.5 wt% reduced cytotoxicity. EGCG concentrations of 1.0 wt% and 0.5 wt% preserved µTBS after 6 months of storage, while 1.5 wt% EGCG significantly decreased µTBS. SEM: the integrity of the hybrid layer was maintained in the 0.5 wt% and 1.0 wt% EGCG groups. Conclusion EGCG concentrations of 1.0 wt% and 0.5 wt% showed better biological and mechanical performance, preserved bond strength and adhesive interface, and reduced cytotoxicity.

Cytotoxicity of Universal, Self-Etching and Etch-and-Rinse Adhesive Systems According to the Polymerization Time

This in vitro study evaluated in fibroblast cultures the direct cytotoxicity of universal, self-etching and etch-and-rinse adhesive systems according to the polymerization time. Paper discs were impregnated with adhesives and light-cured (10, 20 or 40 s). The discs were then immersed in culture medium to obtain the eluates for the experimental groups (A1-Single Bond 2; A2-Scotchbond Multi-purpose; A3-Clearfil SE Bond; A4 Scotchbond Universal). As a negative control, paper discs were immersed in culture medium only. After 24 h or 7 days, the eluate obtained was applied on fibroblast culture. Cell viability, cell morphology, membrane damage and the presence of residual monomers were evaluated by MTT assay, SEM, flow cytometry and high-performance liquid chromatography (HPLC), respectively. Data were analyzed by Kruskal-Wallis and Mann-Whitney tests (=0.05). All adhesive systems significantly reduced 33-51% cell metabolism when compared to the negative control, regardless of polymerization time, storage period and adhesive system. Moreover, the adhesives caused intense morphological alterations and cell membrane damage. Toxicity was directly related to the presence of residual monomers in the eluates. Residual monomers and additional components are capable of reducing mitochondrial activity, causing morphological alterations and disruption of the cell membrane in fibroblasts, regardless of the polymerization time. This study highlights that despite the more complex composition of the universal adhesive system, its biological response was not more toxic when compared with other systems, even when the shortest polymerization time was tested in cell culture.

O presente estudo in vitro avaliou a citotoxicidade direta dos sistemas adesivos convencionais, autocondicionantes e universais de acordo com o tempo de polimerização em cultura de fibroblastos. Discos de papel foram impregnados com adesivos e fotoativados (10, 20 e 40 s). Os discos foram posteriormente imersos em meio de cultura para obtenção dos eluatos dos grupos experimentais (A1-Single Bond 2; A2-Scotchbond Multi-purpose; A3-Clearfil SE Bond; A4 Scotchbond Universal). Para o controle negativo, os discos de papel foram imersos somente em meio de cultura. Após 24 h ou 7 dias, o eluato obtido foi aplicado na cultura de fibroblastos. O metabolismo celular, morfologia, dano de membrana e presença de monômeros residuais foram avaliados por teste de MTT, MEV, citometria de fluxo e HPLC, respectivamente. Os dados foram analisados estatisticamente por Kruskal-Wallis e Mann-Whitney. Todos os sistemas adesivos reduziram significativamente o metabolismo celular em 33 a 51% quando comparados ao grupo controle, independente do tempo de polimerização, período de armazenamento e tipo de sistema adesivo. O eluato do adesivos causou ainda intensas alterações morfológicas e danos à membrana celular. A toxicidade foi diretamente relacionada à presença de monômeros residuais nos eluatos experimentais. Monômeros residuais e componentes adicionais dos sistemas adesivos foram capazes de reduzir a atividade mitocondrial, causar alterações morfológicas e danos à membrana citoplasmática de fibroblastos, independente do tempo de polimerização. Esse estudo evidencia que apesar de uma composição mais complexa do sistema adesivo universal, sua resposta biológica não apresentou maior toxicidade quando comparado aos demais sistemas, mesmo no menor tempo de polimerização quando testados em cultura celular.

The cytotoxicity of methacryloxylethyl cetyl ammonium chloride, a cationic antibacterial monomer, is related to oxidative stress and the intrinsic mitochondrial apoptotic pathway

Antibacterial monomers incorporated in dentin bonding systems may have toxic effects on the pulp. Thus, the cytotoxicity of antibacterial monomers and its underlying mechanisms must be elucidated to improve the safety of antibacterial monomer application. The influence of an antibacterial monomer, methacryloxylethyl cetyl ammonium chloride (DMAE-CB), on the vitality of L929 mouse fibroblasts was tested using MTT assay. Cell cycle progression was studied using flow cytometry. Production of intracellular reactive oxygen species (ROS) after DMAE-CB treatment was measured using 2,7-dichlorodihydrofluorescein diacetate staining and flow cytometry analysis. Loss of mitochondrial membrane potential, disturbance of Bcl-2 and Bax expression, as well as release of cytochrome C were also measured using flow cytometry analysis or Western blot to explore the possible involvement of the mitochondrial-related apoptotic pathway. DMAE-CB elicited cell death in a dose-dependent manner and more than 50 percent of cells were killed after treatment with 30 µM of the monomer. Both necrosis and apoptosis were observed. DMAE-CB also induced G1- and G2-phase arrest. Increased levels of intracellular ROS were observed after 1 h and this overproduction was further enhanced by 6-h treatment with the monomer. DMAE-CB may cause apoptosis by disturbing the expression of Bcl-2 and Bax, reducing the mitochondrial potential and inducing release of cytochrome C. Taken together, these findings suggest that the toxicity of the antibacterial monomer DMAE-CB is associated with ROS production, mitochondrial dysfunction, cell cycle disturbance, and cell apoptosis/necrosis.

Biocompatibility of orthodontic adhesives in rat subcutaneous tissue

ABSTRACT OBJECTIVE: The objective of the present study was to verify the hypothesis that no difference in biocompatibility exists between different orthodontic adhesives. MATERIAL AND METHODS: Thirty male Wistar rats were used in this study and divided into five groups (n=6): Group 1 (control, distilled water), Group 2 (Concise), Group 3 (Xeno III), Group 4 (Transbond XT), and Group 5 (Transbond plus Self-Etching Primer). Two cavities were performed in the subcutaneous dorsum of each animal to place a polyvinyl sponge soaked with 2 drops of the respective adhesive in each surgical loci. Two animals of each group were sacrificed after 7, 15, and 30 days, and their tissues were analyzed by using an optical microscope. RESULTS: At day 7, Groups 3 (Transbond XT) and 4 (Xeno III) showed intense mono- and polymorphonuclear inflammatory infiltrate with no differences between them, whereas Groups 1 (control) and 2 (Concise) showed moderate mononuclear inflammatory infiltrate. At day 15, severe inflammation was observed in Group 3 (Transbond XT) compared to other groups. At day 30, the same group showed a more expressive mononuclear inflammatory infiltrate compared to other groups. CONCLUSION: Among the orthodontic adhesive analyzed, it may be concluded that Transbond XT exhibited the worst biocompatibility. However, one cannot interpret the specificity of the data generated in vivo animal models as a human response.

Citotoxidade de sistemas adesivos contemporâneos: testes in vitro sobre cultura de macrófagos murinos primários / Cytotoxicity of contemporary adhesive systems: in vitro tests on cultured primary murine macrophages

Este estudo, apresentado na forma de três artigos científicos, teve como objetivo geral avaliar o potencial de citotoxidade dos adesivos Adper Single Bond Plus (SB), Clearfil SE Bond (CF) e Xeno V (XE), observando-se a produção de óxido nítrico (NO) e taxa de sobrevivência celular (MTT assay) de macrófagos alveolares de ratos Wistar.

The main goal of this study, presented as three papers, was to evaluate the potencytotoxicity of Adper Plus Single Bond (SB), Clearfil SE Bond (CF), and Xeno V (XE) adhesives, by observing the production of nitric oxide (NO) and rate of cell survival (MTT assay) of aveolar macrophages from rats...

In vitro potential cytotoxicity of an adhesive system to alveolar macrophages

The purpose of this study was to evaluate the potential cytotoxicity of Adper Single Bond 2 (SB) simplified etch-and-rinse adhesive system in alveolar macrophage cultures, as a function of the post-polymerization time and duration of immersion in the culture medium for preparation of extracts, by observing the levels of nitric oxide (NO) release and cell survival rate (MTT assay). Wistar rat alveolar macrophages were exposed to 200 μL of extracts obtained from 24- or 72-h immersion of adhesive samples in culture medium (RPMI), immediately or 24 h after polymerization. Fresh RPMI and E. coli lipopolysaccharides were used as negative and positive controls, respectively. The cells were placed in a humidified incubator for 24 h. The results were analyzed by the Student's-t test (α=5 percent). The amount of NO produced and viable cells were significantly different (p<0.05) between the experimental and the control groups, showing that, irrespective of the post-polymerization time and duration of immersion in the culture medium, the adhesive system caused intense cytotoxicity to the macrophages. The cytotoxic effects were not statistically different (p<0.05) among the experimental groups. In conclusion, chemical components released from SB in aqueous environment were highly toxic to cell culture and thus an inflammatory pulpal response should be considered during the clinical application of dental adhesives.

O objetivo deste estudo foi avaliar o potencial de citotoxicidade do sistema adesivo Adper Single Bond 2 (SB), em função dos tempos pós-polimerização e imersão no meio de cultura para preparação dos extratos, observando-se os níveis de liberação de óxido nítrico (NO) e taxa de sobrevivência celular (MTT assay). Macrófagos alveolares de ratos Wistar foram expostos a 200 μL de extratos obtidos a partir da imersão de amostras do adesivo em meio de cultura (RPMI), imediatamente ou 24 h após sua polimerização, onde permaneceram durante 24 ou 72 h. RPMI puro e lipopolissacarídeos de E. coli foram utilizados como controles negativo e positivo, respectivamente. As células foram levadas à incubadora umidificada por 24 h. Os resultados foram submetidos ao teste "t" de Student (α=5 por cento). As quantidades de NO produzido e células sobreviventes foram significativamente diferentes entre os grupos experimentais e grupos controle, mostrando que, independente do tempo pós-polimerização e tempo de elaboração dos extratos, o sistema adesivo causou uma intensa citotoxicidade sobre os macrófagos. Os efeitos citotóxicos não foram estatisticamente diferentes entre os grupos experimentais. Componentes químicos do SB liberados em meio aquoso podem ser altamente citotóxicos para as células em cultura e, portanto, uma resposta inflamatória pulpar deve ser considerada durante a aplicação clínica de adesivos dentinários.

Avaliaçäo comparativa da toxidade do adesivo dentinário All Bond 2 e do cimento de óxido de Zinco e Eugenol, quando implantados no tecido conjuntivo subcutâneo de ratos / Comparative analysis of toxicity of dentin bonding agent All Bond 2 and Zinc Oxide and Eugenol implanted in subcutaneous tissue of rats

Com o objetivo de dar início ao estudo da biocompatibilidade do adesivo dentinário, All Bond 2 (Grupo experimental) quando comparado com o óxido de Zinco e Eugenol (Grupo OZE - controle), estes materiais foram implantados no tecido conjuntivo dorsal de 20 ratos. Decorridos os períodos de 7, 15, 30 e 60 dias, biópsias foram realizadas, sendo que após processamento laboratorial de rotina, os cortes histológicos de 6 *m de espessura foram corados com H/E. Como resultado, aos sete dias, o All Bond 2 promoveu intensa reaçäo inflamatória com predomínio de células mononucleares, moderada necrose de contato e degradaçäo de fibras colágenas, sendo que o cone capsular formado na área principal de análise foi amplo. Com o decorrer dos períodos houve tendência à regressäo do quadro inicial, mas a presença intensa de macrófagos e células gigantes ocorreu em alguns casos, até o último período de análise. No grupo controle, os achados histopatológicos foram menos intensos nos períodos iniciais de avaliaçäo, sendo que aos 30 e 60 dias, o tecido junto à abertura tubular, área principal de análise havia passado pelo processo de reparaçäo tecidual. Assim, apesar do adesivo dentinário All Bond 2 ter sido de maneira geral mais irritante do que o OZE, de acordo com as normas de ISO, foi possível concluir que ainda é um material aceitável quando implantado no tecido conjuntivo subcutâneo de ratos