Mineral deposition promoted by resin-based sealants with different calcium phosphate additions.

The aim of this study was to evaluate the influence of different calcium phosphates (CaPs) on the physical, biological, and remineralizing properties of experimental resin-based sealants (RBSs). Triethylene-glycol dimethacrylate (90wt%) and bisphenol A-glycidyl methacrylate (10wt%) were used to produce resin-based sealants. Hydroxyapatite (SHAp), α-tricalcium phosphate (Sα-TCP) and octacalcium phosphate (SOCP) were added to the sealants in a 10wt% concentration. One group without CaPs was used as the control group (SCG). The degree of conversion (DC) was assessed with Fourier-transformed infrared spectroscopy, whereas cytotoxicity was tested with the HaCaT keratinocyte cell line. The ultimate tensile strength (UTS) was used to assess the mechanical strength of the experimental RBSs. Sealed enamel was used for colorimetric assay. Mineral deposition was assessed with Raman spectroscopy after 7, 14, and 28 days of sample immersion in artificial saliva. Scanning electron microscopy was used to analyze the surface morphology after 28 days of immersion. The addition of 10wt% of fillers significantly reduced the DC of sealants. SOCP groups showed reduced cell viability. Higher UTS was found for Sα-TCP and SHAp. The color analysis showed that SGC and demineralized teeth presented higher mismatches with the sound tissue. Mineral deposition was observed for SHAp and Sα-TCP after 7 days, with increased phosphate content and mineral deposits for SHAp after 28 days. RBS with the addition of 10% HAp promoted increased mineralization in vitro after 28 days, and did not affect cell viability, DC, mechanical properties, or RBS color in the enamel.

Dental Sealants Part 4: Bisphenol A: What dentists should know.

BACKGROUND: Bisphenol A (BPA) is a synthetic chemical resin used worldwide to produce plastic products. It is also a component of the bisphenol A diglycidylether methacrylate (Bis-GMA), which is a monomer found in dental resin-based materials (including resin-based dental sealants, RBSs). The controversy about its possible toxicity begins around the early '30s. Even if the amount of BPA released by dental sealants is well below the limit proposed by the U.S. Environmental Protection Agency and the European Food Safety Authority, we can reduce the risk of exposure, particularly for children, following precautionary measures.

Investigation of the chemical profile and cytotoxicity evaluation of organic components eluted from pit and fissure sealants.

The aim of this study was to identify organic components eluted from five resin dental sealants using gas chromatography and mass spectrometry (GC/MS) after 1-day and 40-days storage and the effect of sealants on cell survival of cultured fibroblasts. Five resin materials were studied: BeautiSealant (SHOFU), Clinpro (3M/ESPE), Conseal F (SDI), Grandio Seal (VOCO) and Helioseal Clear (Ivoclar/Vivadent). The organic monomers detected were butylated hydroxytoluene (BHT), bis-phenol-A (BPA), camphoroquinone (CQ), diethylenglycoldimethacrylate (DEGDMA), 4N, N-dimethylaminobenzoic acid butylethoxyester (DMABEE), hydroxyethylmethcrylate (HEMA), hydroquinone monomethylether (MEHQ), triethylene glycol dimethacrylate (TEGDMA), tetrabutylammonium tetrafluoroborate (TBATFB), triphenylstibane (TPSb). The main monomer detected was TEGDMA, whereas BHT and DEGDMA were detected at lower concentrations. Higher monomer concentrations were detected after 40 days storage. The eluting chemical profiles of the tested materials differ qualitative and quantitative. For cytotoxicity evaluation, NIH/3T3 cells were exposed to eluates of sealants and cell viability was assessed by a quantitative technique at two observation periods. Decreased cell viability was observed. The cytotoxicity and the release of monomers from dental materials examined depends on the type of material and the observation time point. Resin-based dental materials have raised public concerns regarding possible adverse biological effects, thus it is essential to evaluate possible side effects for human health.

Comparative Evaluation of Cytotoxicity and Genotoxicity of Two Bioceramic Sealers on Fibroblast Cell Line: An in vitro Study.

AIM: The purpose of this study was to evaluate and compare the cytotoxicity and genotoxicity of two bioceramic root canal sealers: EndoSequence BC and iRoot SP with zinc oxide eugenol sealers on fibroblast cell line. MATERIALS AND METHODS: The sealers tested were zinc oxide eugenol, EndoSequence BC, and iRoot SP. Each material was mixed according to the manufacturer's instructions and mounted into sterile polyethylene color-coded rings, for cytotoxicity and genotoxicity evaluation. After 48 hours, the set materials were transferred to previously marked wells and cytotoxicity evaluation to L929 murine fibroblast cells was done by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The percentages of viable cells were then calculated and values were statistically analyzed by Kruskal-Wallis test. The evaluation of genotoxicity of the materials to L929 murine fibroblast cells was carried out by Comet assay. To quantify deoxyribonucleic acid (DNA) damage, the following comet parameters were evaluated in the assay using Comet scoring software: tail length, tail moment, and Olive moment. The values were statistically analyzed using Kruskal-Wallis test with a significance value set to p < 0.05. RESULTS: The results of the study showed that both cytotoxicity and genotoxicity evaluation by MTT assay and Comet assay can be done on L929 murine fibroblast cell line. Among the three tested materials, zinc oxide eugenol showed maximum cytotoxicity to the cells (30.64% viable cells), followed by EndoSequence BC (71.33% viable cells) and iRoot SP (75.11% viable cells). The evaluation of DNA damage by genotoxicity assessment showed iRoot SP to be least genotoxic followed closely by EndoSequence BC. Zinc oxide eugenol was genotoxic and induced more DNA damage on the fibroblast cell line studied. The statistical analyses for both the assays were nonsignificant. CONCLUSION: All the three tested sealers showed varying degrees of cytotoxicity and genotoxicity while using fibro-blast cell line. Zinc oxide eugenol was most toxic in both the assays and iRoot SP showed least toxicity, followed closely by EndoSequence BC.

DNA double-strand breaks caused by new and contemporary endodontic sealers.

AIM: To investigate the cytotoxicity and genotoxicity of a new silicate-based BioRoot RCS® sealer in comparison with contemporary sealers. METHODOLOGY: A periodontal ligament cell line using lentiviral gene transfer of human telomerase reverse transcriptase (hTERT) was used and exposed to subtoxic concentrations of 24-h eluates from two epoxy resin-based (AH Plus Jet® and Acroseal® ), four various methacrylate-based endodontic sealers (EndoREZ® , RealSeal® , RealSeal SE® and Hybrid Root SEAL® ) and three silicate-based sealers (BioRoot RCS® , iRootSP® and MTA Fillapex® ). The XTT-based cell viability assay was used for cytotoxicity screening of materials. The γ-H2AX assay was used for genotoxicity screening. In the γ-H2AX immunofluorescence assay, PDL-hTERT cells were exposed to eluates of the substances for 6 h and DNA double-strand breaks (DSB) were detected microscopically. Induced foci represented DSBs, which can induce ATM-dependent phosphorylation of the histone H2AX. The statistical significance of the differences between the experimental groups was compared using the Student's t-test (P < 0.05). RESULTS: The cytotoxicity of the 24-h eluates could be ranked in the following order: Hybrid Root SEAL® >RealSeal® >Acroseal® >RealSeal SE® ≥ AH Plus Jet® > EndoREZ® >MTA Fillapex® > iRoot SP® >BioRoot RCS® . In negative controls (cells which received medium only) 4.08 ± 0.53 DSB foci (mean ± SEM) whilst in positive controls 10.76 ± 4.05 DSB foci/cell were found. BioRoot RCS® and RealSeal SE® exhibited significant differences in foci formation at 1/3 EC50 compared with their 1/10 EC50 concentration (P < 0.05). Both concentrations (1/10 and 1/3 of EC50) of AH Plus Jet® , Acroseal® , RealSeal® and MTA Fillapex® sealers were not significantly different when compared with the medium control (P < 0.05). CONCLUSIONS: New BioRoot RCS® was not toxic whilst Hybrid Root SEAL® demonstrated more toxicity and DNA double-strand breaks when compared with other resin- and silicate-based root canal sealers.

In vitro studies on the cytotoxic potential of surface sealants.

OBJECTIVE: The objective of this in vitro study was an initial screening of the cytotoxic potential of widely used smooth enamel surface sealants. MATERIALS AND METHODS: A total of 20 products were allocated to four groups based on their chemical composition: (1) filled resin-based sealants, (2) unfilled resin-based sealants, (3) a resin-modified, glass ionomer-based sealant, and (4) silicone-based sealants. All materials were applied to human enamel slices both in accordance with manufacturers' instructions and in additional experiments applying 50% undercuring and 50% overcuring. An agar overlay assay was then used to test the specimens following ISO 10933. The cytotoxic potential of each material was interpreted based on a reaction index that summarized the decolorization and lysis scores obtained. RESULTS: The cytotoxic potential decreased as follows: unfilled resin-based sealants > filled resin-based sealants > resin-modified, glass ionomer-based sealant > silicone-based sealants. In 75% of the resin-based products, deliberate undercuring was associated with more extensive decolorization zones, leading to higher rates of cytotoxic potential in two of those products. Overcuring, by contrast, was associated with a tendency for smaller decolorization zones in 50% of the resin-based products. CONCLUSION: Surface sealants derived from resin monomers exhibited cytotoxic potential in the agar overlay assay. There is also evidence of a possible association with curing, as undercuring can increase the cytotoxic potential, whereas normal curing (as per manufacturers' instructions) or overcuring may help minimize such effects. More research into the biological implications of these materials is needed, especially with regard to their potential impact on the adjacent gingiva.

Influence of light-curing mode on the cytotoxicity of resin-based surface sealants.

BACKGROUND: Surface sealants have been successfully used in the prevention of erosive tooth wear. However, when multiple tooth surfaces should be sealed, the light-curing procedure is very time-consuming. Therefore, the aim of this study was to investigate whether reduced light-curing time (while maintaining similar energy density) has an influence on resin-based surface sealant cytotoxicity. METHODS: Bovine dentine discs were treated as follows: group 1: untreated, groups 2-5: Seal&Protect and groups 6-9: experimental sealer. Groups 2 and 6 were light-cured (VALO LED light-curing device) for 40 s (1000 mW/cm2), groups 3 and 7 for 10 s (1000 mW/cm2), groups 4 and 8 for 7 s (1400 mW/cm2) and groups 5 and 9 for 3 s (3200 mW/cm2). Later, materials were extracted in culture medium for 24 h, and released lactate dehydrogenase (LDH) activity as a measure of cytotoxicity was determined photometrically after cells (dental pulp cells and gingival fibroblasts) were exposed to the extracts for 24 h. Three independent experiments, for both sample preparation and cytotoxicity testing, were performed. RESULTS: Overall, lowest cytotoxicity was observed for the unsealed control group. No significant influence of light-curing settings on the cytotoxicity was observed (p = 0.537 and 0.838 for pulp cells and gingival fibroblasts, respectively). No significant difference in the cytotoxicity of the two sealants was observed after light-curing with same light-curing settings (group 2 vs. 6, 3 vs. 7, 4 vs. 8 and 5 vs. 9: p > 0.05, respectively). CONCLUSIONS: Shortening the light-curing time, while maintaining constant energy density, resulted in no higher cytotoxicity of the investigated sealants.

Quantification of elutable substances from methacrylate based sealers and their cytotoxicity effect on with human gingival fibroblasts.

OBJECTIVES: Previous studies have shown that resin composites may cause persistent inflammation of oral or pulpal tissues as well as cell death through eluted substances. The aim of this study was to investigate the leaching of ingredients from commercial dental fissure sealers as well as their cytotoxic effects on human gingival fibroblast (HGF). METHODS: The sealers tested were: Helioseal(®) F, Helioseal(®) Refill, Fissurit(®) F, Grandio(®) Seal, Ultraseal XT(®) plus and Delton(®) FS. Ten discs of each sealer were respectively immersed in methanol or water and incubated at 37°C. The eluates were analysed by gas chromatography/mass spectrometry at day 1, 3 and 7. In the XTT-test, eight discs of each fissure sealer were immersed into medium. The eluates of the respective sealer were mixed and used undiluted and diluted with medium. HGF were incubated with the dilutions at 37°C for 24h. Then XTT-salt was added and the XTT-formazan formation was quantified. RESULTS: In eluates from polymerized sealers, comonomers (triethylene glycol dimethacrylate (TEGDMA)) and additives were found (e.g. camphorquinone (CQ), butylated hydroxytoluene, triphenylstibane). 7 d after the beginning of the experiments the highest amount of TEGDMA was found in the aqueous eluate from Grandio(®) Seal (9944.31 (2250.56) µmol/l). The most cytotoxic eluate found in the XTT-test was from Fissurit(®) F (EC50 value at 27.13 (7.04)%; (mean(SD)). SIGNIFICANCE: Because of the use of sealers in preventative dental medicine it should be taken into account that substances like TEGDMA or CQ, that are often causing allergic reactions, are elutable. Before using the sealers patients should be asked for allergic reactions to these substances.

Is there a risk of harm or toxicity in the placement of pit and fissure sealant materials? A systematic review.

BACKGROUND: Recently, there has been increased interest in the in vivo release of dental sealant components, such as bisphenol A (BPA), which has the potential to bind the estrogen receptors of relevant cells at subtoxic concentrations in vitro, impairing the development, health and reproductive systems of wildlife. The purpose of this systematic review was to investigate whether the placement of pit and fissure sealant materials causes toxicity, and thus harms patients. METHODS: The literature search (from the earliest record up to March 2007) for relevant articles was done with Ovid MEDLINE, CINAHL and other bibliographic databases. RESULTS: A total of 377 articles were identified by the literature search; relevance was determined by examining the title and abstract of the articles. Eleven original studies met the inclusion criteria. These articles were read in full and scored independently by 2 reviewers. RECOMMENDATIONS: The evidence suggests that patients are not at risk for exposure to BPA from the use of dental sealants. To reduce the potential, if any, for BPA toxicity from sealants, dental providers should use a mild abrasive, such as pumice, either on a cotton applicator or in a prophy cup; have older children and adolescents gargle with tepid water for 30 seconds; or wash the sealant surface for 30 seconds with an air-water syringe while suctioning fluids and debris from a child"s mouth.

On the diagnosis and management of neurocutaneous syndrome, a toxicity disorder from dental sealants.

Neurocutaneous syndrome, a newly discovered toxicity disorder, is characterized by neurological sensations, pain, depleted energy, and memory loss as well as itchy cutaneous lesions that may invite various opportunistic infections. Components in the calcium hydroxide dental sealants Dycal, Life and Sealapex have been identified as sources of the observed symptoms. Sulfonamide and neurological toxicity issues are discussed, and three case histories are presented. Additional notes on zinc oxide, Fynal, IRM, and Sultan U/P sealers are also included. Diagnostic and management protocols at the Parasitology Center, Inc., are proposed.

Root canal sealers induce cytotoxicity and necrosis.

There are three types of the root canal sealers commonly used in clinical applications. They are calcium hydroxide base (Sealapex), zinc oxide-eugenol base (Canals), and epoxy-resin base (AH Plus). Elutable substances and degradation products from root canal sealers may gain access to periodontal tissue in a number of ways. The purpose of the present study was to evaluate the biologic effects of the root canal sealers on human oral cancer cell line (OC2). The tetrazolium bromide (MTT) assay was to evaluate the cell's survival rate. The DNA electrophoresis was used to evaluate the OC2 cell's DNA damage. The results demonstrated that the above root canal sealers' survival rates are in dose-dependent increase (p < 0.05). The toxicity of fresh mix group is higher than that of the mixed after 24h group. DNA fragmentation assay of sealer treated OC2 cells shows a smear layer pattern on the electrophoresis gel. There is no DNA damage found. The toxicity that regulated the cell death is not by the apoptic change of cells.

Estrogenicity of fissure sealants and adhesive resins determined by reporter gene assay.

It is controversial whether the dental resinous materials containing 2,2-bis[4-(2-hydroxy-3-methacryloyloxypropoxy)phenyl]propane (Bis-GMA), which is synthesized from the estrogenic compound bisphenol A (BPA), include unreacted BPA and/or can mimic the effects of natural steroid hormones. In the present study, the estrogenic activities of 3 fissure sealants and 5 adhesive resins, which were all unpolymerized, were determined by means of a reporter gene assay, and the relevance of the components to the estrogenicity was investigated. Two commercially available sealants were confirmed to have estrogenic activity, although none of the tested materials contained BPA. In contrast, hydrophobic monomer bisphenol A dimethacrylate (BPA-DMA), which is also estrogenic, was found to be included in these estrogenic sealants in an amount greater than the minimum concentration to show estrogenicity. This suggests that the estrogenicity of the two proprietary sealants was associated with BPA-DMA rather than with BPA.

Variability of cytotoxicity and leaching of substances from four light-curing pit and fissure sealants.

It was the aim of our study to investigate the composition and cytotoxicity of aqueous extracts of four light-curing pit and fissure sealants. Water extracts were analyzed by gas chromatography/mass spectrometry (GC/MS), and relative quantities of identified compounds were compared by means of an internal caffeine standard [%CF]. Cytotoxic effects due to medium extracts were determined by means of permanent 3T3 fibroblasts. All light-curing pit and fissure sealants segregated different ingredients into water, such as co-monomers (mainly ethylene glycol compounds) and initiating substances (e.g., camphorquinone). Bisphenol-A, however, which is easily detected by GC/MS, was not found in any of the analyzed eluates. The extracts of three sealants inhibited monolayer growth only moderately whereas the eluate of one product inhibited cell proliferation significantly. In the extracts of this sealant high quantities [%CF] of the co-monomer TEGDMA were detected. Our results indicate that light-curing pit and fissure sealants release substances into aqueous media that may induce cytotoxic effects. However, no concerns about potential estrogenic effects of Bisphenol-A are supported by our results.

BIS-GMA--based resins in dentistry: are they safe?

BACKGROUND: The authors critically surveyed research dealing with the release of resin components from dental composites and the potential of these agents to mimic or disrupt estrogenic cell responses. TYPES OF STUDIES REVIEWED: The studies reviewed included those on synthetic methods used to make bisphenol A glycidyl methacrylate, or BIS-GMA, and the biological effects of this resin in cell culture and animals. The estrogenic effect of bisphenol A was targeted because bisphenol A is present as an impurity in some resins (BIS-GMA) and as a degradation product from other resins (bisphenol A dimethacrylate, or BIS-DMA). RESULTS: The outcomes of this review revealed that short-term administration of BIS-GMA and/or bisphenol A in animals or cell cultures can induce changes in estrogen-sensitive organs or cells. However, considering the dosages and routes of administration and the modest response of estrogen-sensitive target organs, the authors conclude that the short-term risk of estrogenic effects from treatments using bisphenol A-based resins is insignificant. Long-term effects need to be investigated further. CLINICAL IMPLICATIONS: Commonly used dental resins should not be of concern to the general public; however, pharmacological evaluation of dental materials is needed to ensure biologically safe and therapeutically effective substances.

Toxicity testing of sealants: a tissue implant study.

The toxicity of pit and fisure sealants implanted into the subcutaneous tissues of guinea pigs was tested using the protocol for toxicity testing derived from the ADA/ANSI Document No. 41, 1982. The materials tested were Delton autopolymerized and photopolymerized, and Concise White autopolymerized and photopolymerized. After two weeks, reactive fibrosis and mild to moderately severe foreign body reactions were noted. After 12 weeks, only thin fibrous walls infiltrated occasionally by small numbers of chronic inflammatory cells were observed. The results of this investigation appear to indicate that following subcutaneous implantation of a pit and fissure sealant, a foreign body reaction will most likely take place during the first two weeks, but will be resolved by 12 weeks. Furthermore, than an initial reactive fibrosis will give way to a thin fibrosis wall by 12 weeks, and the initial inflammatory response will subside. It can also be stated that in this study, Delton and Concise White sealant materials produced similar tissue reactions, and that there were few differences between materials which were autopolymerized or photopolymerized.