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OBJECTIVES: To investigate the cytotoxicity on human dental pulp cells (HDPCs) and Streptococcus mutans (S.mutans) biofilm formation on universal resin composite cements (UCs). METHODS: Three UCs (RelyX Universal, 3 M Oral Care - RXU; Panavia SA Cement Universal, Kuraray Noritake - PSAU; SoloCem, Coltene - SCM) and one 'gold-standard' multi-step cement (Panavia V5, Kuraray Noritake - PV5) were used following two polymerization protocols (light-cured - LC; self-cured - SC). Cytotoxicity (MTT) tests were performed after 1, 3 and 7 days of direct contact. Carboxy-2',7'-dichlorodihydrofluorescein diacetate was used to detect the release of reactive oxygen species (ROS), and interleukin 6 (IL-6) expression was analyzed by IL-6 proquantum high sensitivity immunoassay. S. mutans biofilms were grown on UCs samples in a bioreactor for 24 h, then adherent viable biomass was assessed using MTT assay. For microbiological procedures, half of UCs samples underwent accelerated aging. Data were statistically analyzed (α = 0.05). RESULTS: The highest cytotoxicity was observed for PSAU SC, RXU SC, and PV5 SC at day 1, then for SC RXU after 3 days, and SC PSAU, LC PV5 and SCM after 1-week (p < 0.05). There was no increase in IL-6 expression after 1 day, while it increased depending on the group at 3 and 7 days. The highest ROS expression after 12 h was recorded for PSAU SC, PV5 SC and PV5 LC. Biofilm formation was as follows: RXU > > PSAU = PV5 > SCM, while light-curing systematically decreased biofilm formation (≈-33 %). Aging leveled out differences between UCs and between polymerization protocols. SIGNIFICANCE: The choice of cement brand, rather than category, and polymerization protocol influence cell viability and microbiological behavior. Light-curing is beneficial for reducing the harmful pulpal effect that UCs may possess.

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OBJECTIVES: to assess the impact of universal adhesives, cured with single-peak and polywave LEDs, on the metabolic activity and cytokine release of human dental pulp stem cells (hDPSCs). In addition, analyze the degree of conversion (DC) of the adhesives cured with the different LEDs. METHODS: Discs (5 mm diameter, 1 mm thick) were prepared using three universal adhesives: Single Bond Universal (SBU, 3 M ESPE), Optibond Universal (OBU, Kerr), and Zipbond Universal (ZBU, SDI). These discs were cured for 40 s using a single-peak (DeepCure, 3 M ESPE) or a polywave light-emmiting diode (LED) curing unit (Valo Grand, Ultradent). After 24 h, the specimens were placed in 24-well culture plates, each containing 1 mL of culture medium for 24 h. hDPSCs (1.8 ×104) were seeded in 96-well plates and allowed to grow for 24 h. Subsequently, the cells were exposed to the extracts (culture medium containing eluates from the adhesive discs) for an additional 24 h. Cells not exposed to the extracts were used as a control group. The mitochondrial metabolism was assessed using the MTT assay and the cytokine release evaluated through MAGPIX. The degree of conversion of the adhesives was analyzed using FTIR (n = 5). The results were analyzed by ANOVA two-way and Tukey's test. RESULTS: OBU and ZBU eluates caused a statistically significant reduction in mitochondrial metabolism, regardless of the LED used, indicating their cytotoxicity. In contrast, SBU did not significantly affect the MTT results, resembling the control group. A higher release of cytokines IL-1, IL-6, IL-10, and TNF-α were found in association to ZBU. SBU, on the other hand, increased the release of IL-8. OBU did not influenced the cytokine release. SBU presented the higher DC, while OBU and ZBU had similar DC, lower than SBU. SIGNIFICANCE: In conclusion, universal adhesives exhibit toxicity towards hDPSCs, but the extent of toxicity varies depending on the adhesive material. ZBU was associated with increased cytokine release, particularly pro-inflammatory mediators, from hDPSCs. The different LEDs did not influenced the cytotoxicity of the evaluated adhesives.

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OBJECTIVES: Although the introduction of self-adhesive composites in restorative dentistry is very promising, the innovation of new materials also presents challenges and unknowns. Therefore, the aim of this study was to investigate the cytotoxicity of four different self-adhesive composites (SAC) in vitro and to compare them with resin-modified glass ionomer cements (RM-GIC), a more established group of materials. METHODS: Samples of the following materials were prepared according to ISO 7405/10993-12 and eluted in cell culture medium for 24 h at 37 °C: Vertise Flow, Fusio Liquid Dentin, Constic, Surefil One, Photac Fil and Fuji II LC. Primary human pulp cells were obtained from extracted wisdom teeth and cultured for 24 h with the extracts in serial dilutions. Cell viability was evaluated by MTT assay, membrane disruption was quantified by LDH assay and apoptosis was assessed by flow cytometry after annexin/PI staining. RESULTS: Two SAC (Constic and Vertise Flow) and one RM-GIC (Photac Fil) significantly reduced cell viability by more than 30% compared to the untreated control (p < 0.001). Disruptive cell morphological changes were observed and the cells showed signs of late apoptosis and necrosis in flow cytometry. Membrane disruption was not observed with any of the investigated materials. CONCLUSION: Toxic effects occurred independently of the substance group and need to be considered in the development of materials with regard to clinical implications. CLINICAL SIGNIFICANCE: SAC have many beneficial qualities, however, the cytotoxic effects of certain products should be considered when applied in close proximity to the dental pulp, as is often required.

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STATEMENT OF PROBLEM: Based upon ethical questions and because of the difficulty of obtaining intact human teeth, researchers have used bovine teeth to assess the physical and mechanical properties of different dental materials. However, data from transdentinal cytotoxicity tests showing that the bovine dentin barrier is similar to the human dentin barrier is lacking. PURPOSE: The purpose of this in vitro study was to evaluate whether the bovine dentin barrier produces similar results to those obtained when the human dentin barrier is used to assess the transdentinal cytotoxicity of resin luting cements. MATERIAL AND METHODS: The number and diameter of dentinal tubules present in the human dentin barrier and bovine dentin barrier were evaluated and assessed with a t test (α=.05). After inserting the standardized dentin barriers into artificial pulp chambers, murine dental papilla-derived cells (MDPC-23) were seeded on the pulpal surface of the specimens, and the luting cements were applied to their occlusal surfaces. Then, the following groups were established for both human and bovine dentin barriers: no treatment (negative control); Single Bond Universal; RelyX Luting 2; RelyX U200; and RelyX Ultimate. After 24 hours, the viability (alamarBlue) and morphology (scanning electron microscopy) of the cells were evaluated with a 2-way analysis of variance and the Tukey honest significance test (α=.05). RESULTS: Dentinal tubules with larger diameters were observed in bovine dentin (P<.05), but the number of tubules was similar (P>.05). A reduction in viability and notable changes in the morphology of MDPC-23 cells occurred in the Single Bond Universal and RelyX Luting 2 groups in comparison with the negative control (P<.05). The RelyX U200 and RelyX Ultimate groups were statistically similar to the negative control (P>.05). No difference was found in cytotoxicity when the same luting cement was applied on human or bovine dentin barriers (P>.05). CONCLUSIONS: For transdentinal cytotoxicity tests of resin luting cements, the bovine dentin barrier proved similar results to the human dentin barrier.

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The aim was to develop dual-cured resin cements containing Sr-bioactive glass nanoparticles (Sr-BGNPs; 5 or 10 wt%) and monocalcium phosphate monohydrate (MCPM; 3 or 6 wt%). Effects of additives on degree of monomer conversion (DC), biaxial flexural strength/modulus, shear bond strength (SBS), mass/volume change, color stability, ion release, and cytotoxicity were examined. Controls included material without reactive fillers and Panavia SA Plus (PV). Experimental cements showed higher DC than PV regardless of light activation (p<0.05). Mean SBS and color stability were comparable between experimental cements and PV. Cell viability upon the exposure to sample extracts of experimental cements was 80%-92%. High additive concentrations led to lower strength and modulus than PV (p<0.05). The additives increased mass change, reduced color stability, and promoted ion release. The experimental resin cements demonstrated acceptable mechanical/chemical properties and cytotoxicity. The additives reduced the strength but provided ion release, a desirable action to prevent recurrent caries.

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AIM: This study evaluated cytotoxicity and antioxidant gene expression of resin cements on human gingival fibroblasts (hGF). MATERIALS AND METHOD: RelyX Ultimate™(RXU), Variolink™II(VLII), and RelyXU200™(RXU200) resin cements were incubated with culture medium for 24 h to obtain eluates. Then, the eluates were applied over hGF to assess cell viability at 24 h, 48 h, and 72 h and antioxidant gene expression at 24 h. hGF cultures non-exposed to the eluates were used as Control. Data were submitted to ANOVA and Bonferroni tests (α≤0.05). RESULTS: RXU and RXU200 reduced the number of viable cells in 24 h. Longer exposure to cement extracts caused cell death. Gene expression showed peroxiredoxin 1 (PRDX1) induction by all resin cement types, and superoxide dismutase 1 (SOD1) induction by RXU200 and VLII. Moreover, RXU200 induced not only PRDX1 and SOD1, but also glutathione peroxidase 1 (GPX1), catalase (CAT), and glutathione synthetase (GSS). CONCLUSIONS: All resin cements showed toxicity, and induced antioxidant genes in hGF. Antioxidant gene induction is at least partly associated with cytotoxicity of tested cements to oxidative stress experience.

OBJETIVO: O objetivo deste estudo foi avaliar a toxicidade dos cimentos resinosos Rely X Ultimate 2, Rely X U200 e Variolink II, bem como sua influência na expressão de genes antioxidantes em fibroblastos gengivais humanos. Materiais e Método: Corpos de prova de cada cimento foram colocados em meio de cultura por 24 h e os extratos correspondentes foram aplicados aos fibroblastos. A viabilidade celular foi avaliada após 24, 48 e 72 h de exposição pelo ensaio de exclusão do azul de tripano e MTT. A expressão gênica foi avaliada por PCR quantitativo após 24 h de exposição aos extratos. Estes parâmetros foram comparados aos das células não expostas aos cimentos. Os dados foram submetidos ao teste ANOVA, seguido pelo pós-teste de Bonferroni (a≤0.05). RESULTADOS: Os resultados demonstraram que todos os cimentos promoveram redução do número de células viáveis e da atividade mitocondrial nos períodos de 48 e de 72 h (p< 0,01), sendo que o Variolink II apresentou o menor efeito e os cimentos Rely X Ultimate e Rely X U200 promoveram similarmente os maiores efeitos. A análise de expressão gênica evidenciou influência significativa em todos os cimentos avaliados sobre os níveis de transcritos de PRDX1, SOD1, GPX1 e GSS (p> 0,05), com um aumento considerável no Rely X U200. Conclusão: A indução de genes antioxidantes está, pelo menos em parte, associada à citotoxicidade dos cimentos testados para a experiência de estresse oxidativo.

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The main aim of this study was to perform an integrative review on the toxic effects of resin-matrix cements and their products in contact with fibroblasts or mesenchymal cells. A bibliographic search was performed on PubMed using the following search terms: "cytotoxicity" AND "fibroblast" OR "epithelial" OR "mesenchymal" AND "polymerization" OR "degree of conversion" OR "methacrylate" OR "monomer" AND "resin cement" OR "resin-based cement". The initial search in the available database yielded a total of 277 articles of which 21 articles were included in this review. A decrease in the viability of mouse fibroblasts ranged between 13 and 15% that was recorded for different resin-matrix cements after light curing exposure for 20 s. The viability of human fibroblasts was recorded at 83.11% after light curing for 20 s that increased up to 90.9% after light curing exposure for 40 s. Most of the studies linked the highest toxicity levels when the cells were in contact with Bis-GMA followed by UDMA, TEGDMA and HEMA. Resin-matrix cements cause a cytotoxic reaction when in contact with fibroblasts or mesenchymal cells due to the release of monomers from the polymeric matrix. The amount of monomers released from the resin matrix and their cytotoxicity depends on the polymerization parameters.

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OBJECTIVES: The present study aimed to compare the in vitro cytocompatibility of two etch-and-rinse (Adper Scothbond, Optibond) and two self-etch (Clearfill SE Bond and Single Bond Universal) dental adhesives through a dentin-barrier model with human pulp fibroblasts. METHODS: Human fibroblasts were placed on a plastic device containing 500µm human dentin discs treated with each adhesive or without treatment (control). Other groups were directly exposed to media conditioned with adhesive samples according to ISO 10993-5:2009. After 24h exposure, cell viability was assessed by XTT, and released inflammatory mediators were detected with a multiparametric immunoassay. RESULTS: The standardized test without barrier indicated both etch-and-rinse adhesives and self-etch as cytotoxic, promoting viabilities under 70% of the control group (p<0.05). The dentin-barrier model identified increased cell viability for self-etch adhesives, with Clearfill SE Bond identified as non-cytotoxic. The immunoassay evidenced high rates of cytokines by cells exposed to the conditioned media of Adper Scotchbond, Optibond S, and Single Bond Universal. CONCLUSIONS: The use of a dentin-barrier in vitro model detected a better biocompatibility for self-etching adhesives and, in the case of Clearfill SE Bond, with a reversion from cytotoxic to biocompatible when compared to the indirect standardized test. CLINICAL SIGNIFICANCE: The use of a dentin-barrier in vitro model was able to detect a better biocompatibility for self-etching adhesives when compared to the indirect standardized test and presents itself as a predictive in vitro method for assessing the cytotoxicity of dental restorative materials that may simulate the clinical condition more accurately.

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OBJECTIVES: To investigate the transdentinal cytotoxicity (TC), degree of conversion (DC), and micro shear bond strength (µSBS) to dentin of light-cured resin cements (LCRCs) photoactivated directly or through a ceramic veneer( ± CV). MATERIALS AND METHODS: The TC was assessed using human dentin discs adapted into artificial pulp chambers. Odontoblast-like cells were seeded on the pulp surface of the discs, then three LCRCs( ± CV) were applied on their etched and hybridized occlusal surface (n = 8/group). The adhesive systems of each LCRCs and sterile phosphate-buffered saline were used as positive and negative controls, respectively. After 24 h, the viability and morphology of cells adhered on discs were assessed. The extracts (culture medium + components of the materials diffused through the discs) were applied on the MDPC-23 to evaluate their viability, adhesion/spreading (A/S), alkaline phosphatase activity (ALP), and mineralized nodule formation (MN). LCRCs( ± CV) specimens were evaluated concerning the DC and µSBS to dentin. Data were analyzed by one-, two-, or three-way ANOVA/Dunnett, Sidak, and Games-Howell tests (α = 5%). RESULTS: All LCRCs( ± CV) reduced cell viability, A/S, ALP, MN, and DC. Except for µSBS, the intensity of reduction was dependent on the LCRC used. LCRCs+CV resulted in lower DC and µSBS but did not increase the TC. SIGNIFICANCE: Besides the presence of CV between the light source and LCRCs reduces the degree of conversion and bond strength to dentin, these materials cause variable level of transdentinal toxicity to pulp cells. Thus, the composition and curing protocols of LCRCs should be revisited and reinforced to prevent mechanical and biological drawbacks.

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OBJECTIVES: A three-dimensional (3D) dentin/pulp tissue analogue, resembling the human natural tissue has been engineered in an in vitro setup, aiming to assess the cytocompatibility of resin-based dental restorative cements. METHODS: Stem Cells from Apical Papilla (SCAP) and Human Umbilical Vein Endothelial Cells (HUVEC) were embedded in Collagen-I/Fibrin hydrogels at 1:3 ratio within 24-well plates. Hanging culture inserts were placed over the hydrogels, housing an odontoblast-like cell layer and a human treated-dentin barrier. Shear modulus of the hydrogels at 3.5 and 5 mg/ml was evaluated by dynamic mechanical analysis. Eluates of two resin-based cements, a dual-cure- (Breeze™, Pentron: Cement-1/C1), and a self-adhesive cement (SpeedCEMplus™, Ivoclar-Vivadent: Cement-2/C2) were applied into the dentin/pulp tissue analogue after pre-stimulation with LPS. Cytocompatibility was assessed by MTT assay, live/dead staining and real-time PCR analysis. RESULTS: Both hydrogel concentrations showed similar shear moduli to the natural pulp until day (D) 7, while the 5 mg/ml-hydrogel substantially increased stiffness by D14. Both cements caused no significant toxicity to the dentin/pulp tissue analogue. C1 induced stimulation (p < 0.01) of cell viability (158 ± 3%, 72 h), while pre-stimulation with LPS attenuated this effect. C2 (±LPS) caused minor reduction of viability (15-20%, 24 h) that recovered at 72 h for the LPS+ group. Both cements caused upregulation of VEGF, ANGP-1, and downregulation of the respective receptors VEGFR-2 and Tie-1. SIGNIFICANCE: Both resin-based cements showed good cytocompatibility and triggered angiogenic response within the dentin/pulp tissue analogue, indicating initiation of pulp repair responses to the released xenobiotics.

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STATEMENT OF PROBLEM: Dental cements that release monomers that negatively impact adjacent oral soft tissues may adversely affect clinical outcomes. However, in vitro studies evaluating the cytotoxic and genotoxic potential of substances released from dental cements are lacking. PURPOSE: The purpose of this in vitro study was to define and compare the cytotoxicity and genotoxicity of the eluates of a self-adhesive resin cement (RelyX Unicem 2 Automix) autopolymerized and light polymerized with 2 other types of luting cements: a glass ionomer cement (Ketac Cem Easymix) and a resin-modified glass ionomer cement (Ketac Cem Plus). MATERIAL AND METHODS: The eluates were prepared, and 3T3 mouse fibroblast cells were exposed for 24 hours to serial eluate dilutions of the 3 types of cement. Cytotoxicity was determined by using a cell viability assessment through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and crystal violet assays. Genotoxic effects were determined by using the cytokinesis-block micronucleus assay. RESULTS: Cell viability was higher in the presence of the glass ionomer cement eluate than of the resin-modified glass ionomer cement and resin cement eluates. A pronounced decrease in viability was found when the cells were exposed to undiluted samples of resin-modified glass ionomer cement (around 50%) or resin cement (around 80% to 90%). No significant difference in cell viability was found between autopolymerized and light-polymerized resin cements. All cements induced a dose-dependent response of mononucleated cell formation. However, only the resin cements showed double strand breaks significant differences in the deoxyribonucleic acid (DNA) molecules against the basal DNA lesions that occurred spontaneously. CONCLUSIONS: The glass ionomer cement was not found to be cytotoxic or genotoxic, whereas the eluates derived from the resin-modified glass ionomer cement and resin cement, independently of the polymerization method, were cytotoxic in fibroblast cells. Maximum cytotoxicity was observed in the presence of resin cement, which also showed genotoxicity, independently of being light polymerized.

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This study evaluated the cytotoxicity of methacrylate-based resins containing dimethyl sulfoxide (DMSO). DMSO was incorporated into hydrophobic (R2) and hydrophilic (R5) resins at weight concentrations of 0, 0.01, 0.1, 1, 5, or 10 w/w %. Resin discs (n = 10/group) were prepared. Human gingival fibroblasts (HGF-1) were exposed to resin eluates for 24 h. Furthermore, dentin barrier test was performed using 3-D cultures of odontoblast-like cells (SV40 transfected pulp derived cells) with dentin slices of 400 µm thickness (n = 8). After acid etching of dentin, DMSO-modified resins were applied into the cavity part of the device and light-cured for 20 s. Cell viability (%) was assessed by MTT and analyzed spectrometrically. Data were analyzed by ANOVA and Tukey test (α = 0.05). Resin eluates showed statistically significantly lower % cell viability for all neat and DMSO-modified resins than seen for the negative control. Moreover, DMSO-R5 eluates resulted in significantly lower % cell viability than DMSO-R2 emulates. The dentin barrier test showed that DMSO-R2 did not result in significantly lower % cell viability, whereas incorporation of 1-10 w/w % DMSO into R5 resulted in significantly lower % of cell viability. Incorporating DMSO into hydrophilic self-etching resins may increase cytotoxicity. The biocompatibility is not influenced by the addition of DMSO into hydrophobic resin.

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OBJECTIVES: The biocompatibility of methacrylate-based adhesives is a topic that is intensively discussed in dentistry. Since only limited evidence concerning the cyto- and genotoxicity of orthodontic adhesives is available, the aim of this study was to measure the genotoxic potential of seven orthodontic methacrylate-based adhesives. MATERIALS AND METHODS: The XTT assay was utilized to determine the cytotoxicity of Assure Plus, Assure Bonding Resin, ExciTE F, OptiBond Solo Plus, Scotchbond Universal Adhesive, Transbond MIP, and Transbond XT after an incubation period of 24 h on human gingival fibroblasts. We also performed the γH2AX assay to explore the genotoxic potential of the adhesives within cytotoxic dose ranges after an incubation period of 6 h. RESULTS: The XTT assay showed a concentration-dependent reduction in cell viability. The decrease in cellular viability was in the same dose range most significant for Assure Plus, rendering it the adhesive material with the highest cytotoxicity. Employing the γH2AX assay, a concentration-dependent increase in H2AX phosphorylation was detected, indicating induction of DNA damage. CONCLUSIONS: For most products, a linear correlation between the material concentration and γH2AX foci was observed. The most severe effect on γH2AX focus induction was found for Transbond MIP, which was the only adhesive in the test group containing the co-initiator diphenyliodonium hexafluorophosphate (DPIHP). CLINICAL RELEVANCE: The data indicate that orthodontic adhesives, notably Transbond MIP, bear a genotoxic potential. Since the study was performed with in vitro cultivated cells, a direct translation of the findings to in vivo exposure conditions should be considered with great diligence.

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In this study, the cytotoxicity of different combinations of contemporary resin-based restoratives (adhesives, composites, luting agents) against human keratinocytes (HaCaT) was evaluated under two conditions, whether materials were applied to dentin or not. Adhesives (3-step etch-and-rinse/3ER: OptiBond FL; 2-step self-etch/2SE Clearfil SE Bond; Single Bond Universal/UNI), composites (conventional composite resin/CCR: Filtek Z350XT; flowable/FCR: Filtek Z350XT Flow; self-adhesive composite resin/SACR: Dyad Flow), and luting agents (conventional luting agent/CLA: Variolink-II; self-adhesive luting agent/SLA: RelyXU200) were combined according to their clinical use. Eluates from polymerized specimens applied to dentin were placed in contact with cells grown for 1 and 7 d. The controls were defined by cells without material contact. Cell viability was determined using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)] assay. C=C conversion was investigated using Fourier-transform infrared spectroscopy. After 1 d of incubation, when dentin was not present, 2SE yielded the highest cell viability, whereas 3ER, UNI, and SACR showed higher cell viability in the presence of dentin. After 7 d, when dentin was absent, 2SE and CLA achieved significantly higher cell viability. The presence of dentin resulted in a drastically higher cell viability for all materials, except 2SE and CLA. UNI had the lowest C=C conversion. The presence of dentin was a significant factor, which resulted in higher cell viability than what was seen for the material specimens per se. All materials resulted in a lower viability of HaCaT than what was seen under the no-material control conditions, with effects mainly limited to the first 24 h.

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OBJECTIVE: The objective of this work is to compare cellular toxicity in vitro of two resins for orthodontic use: an auto-polymerizable composite and a photo-polymerizable composite. MATERIALS AND METHODS: Samples were obtained by joining a couple of steel orthodontic brackets by using auto-polymerizing or photo-polymerizing resin. We used a halogen lamp, a mini LED lamp and a fast LED lamp used for orthodontics cure for 40 seconds. The 3T3 Swiss cellular line of fibroblasts was used. The samples obtained were used to determine the cellular toxicity in vitro using the Neutral Red Up-take (NRU) and the 3-(4,5-dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide (MTT) assay. RESULTS: Toxicity of the extract appraised at a low level at MTT and NRU assays. There were statistically relevant differences between the toxicity induced by the auto-polymerizing material and the toxicity induced by the photo-polymerizing composite material, polymerized with the blue-light lamp (p < 0.001) and with the mini LED lamp (p < 0.05). CONCLUSIONS: From the data collected in this study, we can conclude that both resins show a low level of cytotoxicity that, in the case of photochemical polymerizing resin, depends on the characteristics of the lamp.

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The aim of the present study was to evaluate the in vitro cytotoxicity of self-adhesive dual-cured resin cement (SADRC) polymerized beneath three different cusp inclinations of zirconia with different light curing time. A commercial SADRC (Multilink Speed) was polymerized beneath zirconia (ZrO2) with three different cusp inclinations (0°, 20°, and 30°) for 20 s or 40 s. After being stored in light-proof box for 24 h, the ZrO2-SADRC specimens were immersed in DMEM for 72 h and then we got the extract solution, cultured the human gingival fibroblasts (HGF, 8 × 103 per well) with 100% or 50% concentrations of the extract solution for 24 h, 72 h, and 120 h, respectively, and evaluated cytotoxicity of the polymerized SADRC with CCK-8 assay in optical density (OD) values, relative growth rates (RGR), and cytotoxicity grades. Statistical analysis was conducted using a two-way ANOVA followed by post hoc Student-Newman-Keuls test. The OD values varied from 0.8930 to 3.2920, the RGR varied from 33.93% to 98.68%, and the cytotoxicity grades varied from 0 to 2. There was significant difference in the OD values among the different cusp inclinations of zirconia (P < 0.001), and there was significant difference in the OD values between the different light curing times in some situations (P < 0.05). The cusp inclination of zirconia affects the in vitro cytotoxicity of SADRC. Prolonging the light curing time from 20 s to 40 s can reduce the in vitro cytotoxicity of SADRC when the cusp inclination of zirconia is smaller than 20°.

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PURPOSE: To qualitatively and quantitatively compare the cytotoxic potentials of five different one-step self-etching adhesives: Prime&Bond One-Select (PB-OS), Optibond All-in-One (OB-AIO), G-Bond (GB), Clearfil Universal Bond (CUB), Single Bond Universal (SBU). MATERIALS AND METHODS: During the first stage of the study, the cytotoxic activities of the test materials were evaluated qualitatively using the direct contact method. In this method, the test materials were placed directly into a monkey kidney epithelial cell culture medium. Reaction zones which occurred in the culture medium were evaluated, in addition to the density and changes in the morphology of the cells. During the second stage, the cytotoxic potential of four different dilutions (1%, 0.1%, 0.01%, 0.001%) of the test materials on L929 rat fibroblast cells was quantitatively evaluated at three different time periods (24 h, 48 h, 72 h) with the MTT tetrazolium-based assay. RESULTS: In the first stage, a zone exceeding 1 cm was observed around or below SBU, CUB, GB and OB-AIO. In PB-OS, the zone borders were approximately 1 cm. In the second stage after the MTT assay, CUB was the most cytotoxic after 24 h, GB and SBU after 48 h, and OB-AIO after 72 h. CONCLUSION: All adhesives tested showed different degrees of cytotoxicity, which statistically significantly increased with dose. Changes were seen related to time.

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The aim of this study was to evaluate the amount of leached residual monomers from self-adhesive resin cements and evaluate their toxicity in-vitro. A total of 60 disk-shaped specimens (5 mm in diameter and 0.5 mm in thickness) were prepared from each cement (RelyX U200, SpeedCEM, G-Cem) (n=20). Specimens were immersed in artificial saliva and the amount of released monomers [urethane dimethacrylate (UDMA) and triethyleneglycol dimethacrylate (TEGDMA)] was identified. Then, the cytotoxicity and genotoxicity effect on cells were evaluated using the defined amounts of released monomers from cements. The highest monomer release was detected in G-Cem (p<0.05). The highest cytotoxicity value was identified from SpeedCEM (p<0.01) and the highest genotoxicity values were calculated from RelyX U200 (p<0.05). Released UDMA and TEGDMA from self-adhesive resin cements induced cytotoxicity and genotoxicity effect on cells.

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OBJECTIVE: To compare cytotoxicity of extracts derived from commonly used luting cements: Hoffmann's Zinc Phosphate (ZPC), GC Fuji Plus Resin Modified Glass Ionomer (RMGIC) and 3M ESPE RelyX Unicem Resin Cement (RC) on primary human gingival fibroblasts (HGFs). DESIGN: HGFs were exposed to different concentrations of the ZPC, RMGIC and RC extracts. The cytotoxicity was assessed with the PrestoBlue Cell Viability Reagent and viable cells were counted by a haemocytometer using the trypan blue exclusion test. In order to determine the primary mechanism of the cell death induced by extracts from different luting cements, the real-time monitoring of caspase-3/-7 activity and membrane integrity of cells was employed. RESULTS: The extracts from the RMGIC and ZPC decreased the metabolic activity and numbers of viable cells. Unexpectedly, the extracts from the RC evoked only small effects on the metabolic activity of HGFs with a decreasing number of viable cells in a dose-and time-dependent manner. The live cell imaging revealed that the apoptosis was the primary mechanism of a cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death through a necrotic and caspase-independent pathway. CONCLUSIONS: The apoptosis was the primary mechanism of the cell death induced by the extracts derived from the RMGIC, whereas the extracts from the RC and ZPC induced a cell death via a necrotic pathway. We suggest that metabolic assays commonly used to assess the cytotoxicity of luting cements should be validated by alternative methods.

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OBJECTIVES: To evaluate the cytotoxic effects of exposing odontoblast cells to a variety of commercial self-adhesive cements polymerized using different activation modes. METHODS: Five cements: MaxCem Elite (MAX), Bifix SE (BSE), G-Cem LinkAce (GCE), Clearfil SA Luting (CAS), and RelyX U200 (U200) were mixed, dispensed into molds, and distributed in groups, according to polymerization protocols: immediate photoactivation; delayed photoactivation (10min self-curing plus light-activation); and chemical activation (no light exposure). Immortalized rat odontoblast cells (MDPC-23) were cultured. Cell viability was assessed by Trypan Blue staining and total cell death was assessed by annexin V-APC/7-AAD double staining and flow cytometry. Volatilized compounds from polymerized specimens of cements were evaluated by gas chromatography/mass spectrometry (GC-MS). Data was analyzed with 2-way ANOVA/Tukey tests (α=0.05). RESULTS: Exposure to all of the cements tested significantly reduced the cell viability, irrespective of the activation protocol (p<0.05). The least harmful cements were CSA and U200. Total death of cells significantly increased when exposed to BSE, GCE, and MAX, especially when chemically activated (p<0.05). Characteristic apoptotic cells increased after exposure to cements, mainly for MAX, regardless of the activation mode. Chemical activation of MAX also induced necrosis. Moreover, GCE and MAX exhibited higher percentages of late apoptotic/dead cells. Chromatograms revealed 28 compounds released from the cements tested, some of them with known carcinogenic effects. Selection of self-adhesive cements and polymerization protocols affect the cytotoxicity and cell viability of odontoblastic cells. CLINICAL SIGNIFICANCE: Despite the simplified cementation protocol, care is needed when cementing indirect restorations with self-adhesive cements, especially on recently exposed dentin. This category of material may cause differential cytotoxic effects and should be considered when selecting a cement. This is particularly true in clinical cases of light attenuation, where the polymerization depends on chemical activation, inducing higher cytotoxic damages when using some of the cements tested.

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