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Diverse applications of nanoparticles due to their unique properties has rapidly increased human exposure to numerous nanoparticles such as calcium hydroxide (Ca(OH) 2 ), calcium titanate (CaTiO 3 ), and yttrium oxide (Y 2 O 3 ) nanoparticles almost in all aspect of daily life. However, very limited data are available on the effect of these nanoparticles on genomic DNA integrity and inflammation induction in the gastric tissues. Hence, this study estimated the effect of Ca(OH) 2 , CaTiO 3, or/and Y 2 O 3 nanoparticles multiple oral administration on the genomic DNA damage and inflammation induction in the mice gastric tissues. A suspension containing 50 mg/kg b.w of Ca(OH) 2 , CaTiO 3, or Y 2 O 3 nanoparticles were given orally to male mice separately or together simultaneously three times a week for two consecutive weeks. Multiple oral administration of Ca(OH) 2 nanoparticles led to significant elevations in DNA damage induction and ROS generation, in contrast to the non-significant changes observed in the level of induced DNA damage and generated ROS after administration of CaTiO 3 or Y 2 O 3 nanoparticles separately or in combination with Ca(OH) 2 nanoparticles. Oral administration of Ca(OH) 2 nanoparticles alone also highly upregulated INOS and COX-2 genes expression and extremely decreased eNOS gene expression. However, high elevations in eNOS gene expression were detected after multiple administration of CaTiO 3 and Y 2 O 3 nanoparticles separately or together simultaneously with Ca(OH) 2 nanoparticles. Meanwhile, non-remarkable changes were noticed in the expression level of INOS and COX-2 genes after administration of CaTiO 3 and Y 2 O 3 nanoparticles separately or simultaneously together with Ca(OH) 2 nanoparticles. In conclusion: genomic DNA damage and inflammation induced by administration of Ca(OH) 2 nanoparticles alone at a dose of 50 mg/kg were mitigated by about 100% when CaTiO 3 and Y 2 O 3 nanoparticles were coadministered with Ca(OH) 2 nanoparticles until they reached the negative control level through altering the expression level of eNOS, INOS and COX-2 genes and scavenging gastric ROS. Therefore, further studies are recommended to investigate the toxicological properties of Ca(OH) 2 , CaTiO 3 and Y 2 O 3 nanoparticles and possibility of using CaTiO 3 and Y 2 O 3 nanoparticles to mitigate genotoxicity and inflammation induction by Ca(OH) 2 nanoparticles., (© 2023. The Author(s).)

Intensive uses of Calcium hydroxide (Ca(OH) 2 NPs), calcium titanate (CaTiO 3 NPs) and yttrium oxide (Y 2 O 3 NPs) nanoparticles increase their environmental release and human exposure separately or together through contaminated air, water and food. However, too limited data are available on their genotoxicity. Therefore, this study explored the effect of Ca(OH) 2 NPs, CaTiO 3 NPs or/and Y 2 O 3 NPs administration on the genotoxicityand oxidative stress induction in mice hepatic tissue. Mice were orally administered Ca(OH) 2 NPs, CaTiO 3 NPs and Y 2 O 3 NPs separately or simultaneously together at a dose level of 50 mg/kg b.w. for two successive weeks (3 days per week). Marked induction of DNA damage noticed after oral administration of Ca(OH) 2 NPs or CaTiO 3 NPs alone together with high Ca(OH) 2 NPs induced reactive oxygen species (ROS) generation and a slight CaTiO 3 NPs induced ROS production were highly decreased after simultaneous coadministration of administration of Y 2 O 3 NPs with Ca(OH) 2 NPs and CaTiO 3 NPs up to the negative control level. Oral administration of Y 2 O 3 NPs alone also did not cause observable changes in the genomic DNA integrity and the ROS generation level compared to the negative control levels. Similarly, significant elevations in P53 gene expression and high reductions in Kras and HSP-70 genes expression were observed only after administration of Ca(OH) 2 NPs alone, while, remarkable increases in the Kras and HSP-70 genes expression and non-significant changes in p53 gene expression were noticed after administration of CaTiO 3 NPs and Y 2 O 3 NPs separately or simultaneously together with Ca(OH) 2 NPs. Conclusion: Ca(OH) 2 NPs exhibited the highest genotoxic effect through oxidative stress induction and disruption of apoptotic (p53 and Kras) and protective (HSP-70) genes expression. Slight DNA damage was noticed after CaTiO 3 NPs administration. However, administration of Y 2 O 3 NPs alone was non-genotoxic and coadministration of Y 2 O 3 NPs with Ca(OH) 2 NPs and CaTiO 3 NPs restored genomic DNA integrity and normal expression of apoptotic p53 and protective HSP-70 genes disrupted by Ca(OH) 2 NPs and CaTiO 3 NPs. Thus co-administration of Y 2 O 3 NPs with Ca(OH) 2 NPs and CaTiO 3 NPs is recommended to counter Ca(OH) 2 NPs and CaTiO 3 NPs induced genotoxicity and oxidative stress., (© 2023. The Author(s).)

Objective: To evaluate the solubility, pH, antimicrobial action, and cytotoxicity of ambroxol hydrochloride (AMB), N-acetylcysteine (NAC), and calcium hydroxide (CH) pastes for use as intracanal medications., Methods: Solubility was determined by micro-CT, based on the paste volume remaining after immersion in water for 7 days. pH was measured by immersing acrylic tubes containing the pastes in ultrapure water and then measuring pH after 3 hours, 3 days, and 7 days. Antimicrobial action against Enterococcus faecalis was assessed based on the percentage of living cells, using the live/dead staining method under confocal microscopy. Cytotoxicity was assessed based on the cell viability of L929 fibroblast-like cells after 6, 24, and 48 hours. Cytotoxicity data were compared using the ANOVA and Tukey tests, and the antimicrobial data were compared using the Kruskal-Wallis and Dunn tests. The significance level used was 5% (α=0.05)., Results: The solubility values for all the study groups were significantly different (P<0.05), where the highest values were for NAC, followed by AMB, and then CH. Likewise, the pH levels were all significantly different (P<0.05), where NAC and AMB levels were acidic, and CH levels were alkaline. The antimicrobial action of AMB was significantly higher than that of CH (P<0.05), and that of NAC was also higher than that of CH, albeit not significantly. AMB and NAC were more cytotoxic than CH, and higher dilutions of CH promoted higher cell viability levels than lower dilutions of the same paste (P<0.05)., Conclusion: The NAC and AMB pastes were more soluble and cytotoxic than the CH paste and had acidic pH levels. The AMB paste displayed the highest antimicrobial action against Enterococcus faecalis biofilm.

Objective: This study aimed to assess the cytotoxicity and genotoxicity of triple antibiotic paste, double antibiotic paste and calcium hydroxide medicaments on human stem cells of the apical papilla., Methods: In this experimental study, stem cells were isolated from the apical papilla and cultured. They are treated with different concentrations 0.1, 0.5, 1, 10 and 100 mg/ml of medicaments for 24, 48 and 72 hours. The cytotoxicity and genotoxicity of the medicaments were determined using methyl thiazolyl tetrazolium assay and Comet test, respectively., Results: Results showed all tested concentrations of the calcium hydroxide had no significant effect on stem cells at any time point. Triple antibiotic paste showed cytotoxicity in 10 and 100 mg/mL concentrations at all-time points and in 1, 10 and 100 mg/ml concentrations at 72 hours. In addition, its genotoxicity was significantly higher than that of other groups (P<0.05). Double antibiotic paste showed cytotoxic effects only in 100 mg/ml concentration at 24 hours and 10 and 100 mg/ml concentrations at 48 and 72 hours. And also, its genotoxicity in these concentrations was significantly higher than that of control and calcium hydroxide groups (P<0.05)., Conclusion: In contrast to calcium hydroxide, triple antibiotic paste and double antibiotic paste, especially in their higher concentrations, induced cytotoxicity and genotoxicity on human stem cells of the apical papilla.

With the high increases in the uses of calcium hydroxide in various applications due its distinctive properties, human exposure has increased to normal- and nano-calcium hydroxide. However, its impact on the DNA integrity, expression of inflammatory cytokines, and induction of oxidative stress has not been clearly studied. Therefore, here we estimate the induction of DNA damage, inflammation, and oxidative stress in mice orally administrated a single dose (100 mg/kg) of normal- or nano-sized calcium hydroxide for 24 hour. Comet, Diphenylamine and laddered DNA fragmentation assays were done to assess DNA damage induction. Acute oral administration of normal- or nano-calcium hydroxide particles disrupted the DNA integrity, caused generation of ROS and also concurrent increases in both the nitric oxide concentration and inducible nitric oxide synthase gene expression in a reverse proportional to the calcium hydroxide particles' size. Increases in the concentration of calcium ions as well as alterations in the expression level of p53 and proinflammatory cytokines were also observed in calcium hydroxide administrated groups. Moreover, administration of normal- or nano-calcium hydroxide particles suspension elevated the level of malondialdehyde and decreased both the glutathione peroxidase activity and the reduced glutathione level, as well as caused tissue injuries (e.g. renal tube degeneration, congested blood vessels, atrophied lymphoid follicles, interstitial inflammatory reaction, and hyalinosis of myocardial muscles). Thus, we conclude that calcium hydroxide acutely orally administrated in its ordinary or nano-particulate form causes DNA damage induction by generating free radicals and altering the expression levels of p53 gene and proinflammatory cytokines.

The aims of this study were to evaluate the physical and chemical properties, cytotoxicity and dentinal tubule penetration of a new calcium silicate-based root canal dressing. For pH and calcium ion release evaluation (1, 24, 72 and 168 h) were used a pH meter and colorimetric spectrophotometer, respectively. Radiopacity evaluation followed the ISO 6876:2012. Cytotoxicity was evaluated by the percentage of cell viability using MTT assay. Illustrative images of dentinal tubule penetration were obtained using confocal laser scanning microscopy (CLSM). Data from pH and calcium ion release were statistically analyzed by two-way analysis of variance and Tukey test. Radiopacity was analyzed using the Student t-test. The statistical tests for cytotoxicity results were the one-way analysis of variance and Tukey test. Both materials showed alkaline pH in all experimental times. The pH values for calcium hydroxide paste were higher than bioceramic paste at 1, 24, and 72 h (p<0.05). The calcium ion release of bioceramic was lower than the calcium hydroxide paste only at 24 h (p<0.05). The bioceramic was more radiopaque than the calcium hydroxide paste (p<0.05). Bioceramic paste presented a dose and time-dependent cytotoxic effect after MTT assay. CLSM images showed absence of tubule penetration for both pastes. The new calcium silicate-based canal dressing presented alkaline pH, high calcium release, and acceptable radiopacity. Bio C Temp showed a dose and time-dependent cytotoxic and absence of dentinal tubule penetration.

The aim of this study was to evaluate the degree of conversion, cytotoxicity, solubility and pH of photopolymerizable calciumbased cements submitted to preheating. The degree of conversion was analyzed by Fourier transform infrared, cytotoxicity by the MTT test and solubility through loss of mass. The data were subjected to statistical tests (ANOVA / Tukey's, p<0.05). The photopolymerizable materials showed a low degree of conversion, regardless of preheating. All materials caused a reduction in cell viability at 24 hours and 7 days, with the Dycal (control) being more cytotoxic. Heat had a positive effect on Biocal at 7 days. Dycal is the most soluble material. Heat had no effect on the solubility or pH of the polymerizable materials. It is concluded that photopolymerizable calcium-based cements have a low degree of conversion and are soluble, which results in mild to moderate cytotoxicity., Competing Interests: none, (Sociedad Argentina de Investigación Odontológica.)

Introduction: A successful outcome of root canal therapy relies on effective disinfection of the root canal system, including the use of intracanal medicaments, which vary in their bactericidal and cytotoxic properties. Assessing the benefits and risks associated with the use of these medicaments is of extreme importance, especially in regenerative endodontic procedures, because residual stem cells may be harmed. In this study, we tested the cytotoxicity and genotoxicity of a novel agent, 2-hydroxyisocaproic acid (HICA), and compared its properties with those of a well-established medicament, calcium hydroxide., Methods: Human periodontal ligament fibroblasts were exposed to varying concentrations of HICA (1, 2.5, 5, 10, 20, and 40 mg/mL) for 24 hours, and a dose-response curve was generated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay. Immunofluorescence for 2 markers of DNA double-strand breaks, phosphorylated γH2AX and 53BP1, was used to establish the genotoxicity of HICA at various half maximal effective concentration (EC50) fractions. Cytotoxicity and genotoxicity of HICA and calcium hydroxide at 1 mg/mL were compared at 24 and 48 hours using the same methods., Results: At 10 mg/mL and higher, HICA was significantly more cytotoxic and genotoxic than the control (P < .05 and P < .0001, respectively). Calcium hydroxide at 1 mg/mL was more cytotoxic than HICA at 1 mg/mL at 24 and 48 hours (P < .05 for both), whereas no difference in the accumulated DNA damage was observed., Conclusions: HICA is not cytotoxic and genotoxic at concentrations <10 mg/mL. At the concentration of 1 mg/mL, HICA is significantly less cytotoxic than calcium hydroxide., (Copyright © 2019 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Nowadays, there is increasing interest in using calcium hydroxide (Ca(OH) 2 ) nanoparticles rather than normal-sized Ca(OH) 2 because of its higher antimicrobial activities. However, the genotoxicity of Ca(OH) 2 nanoparticles has not been well investigated. Therefore, this study was performed to estimate the possible genomic instability and mitochondrial DNA damage induction by normal and nano-sized Ca(OH) 2 particles in mice. Oral administration of normal or nano-sized Ca(OH) 2 particles induced DNA breakages and apoptosis causing genomic instability as a result of increased Calcium content, ROS and MDA levels and decreased SOD and Gpx activities reversely proportional to Ca(OH) 2 particles size in the liver, brain and bone marrow tissues. However, decreases in mitochondrial membrane potential concurrently with downregulated expression of POLG, POLG1 and TFAM genes were only observed in the brain and bone marrow tissues confirmed mitochondrial DNA damage. In contrast, the expressions of POLG, POLG1 and TFAM genes have been improved and mitochondrial membrane potential has not unchanged in liver tissues. Conclusion: single oral administration of normal or nano-sized Ca(OH) 2 particles induced genomic instability through ROS generation that exhausted the antioxidant defense system in the liver, brain and bone marrow tissues but impaired the mitochondrial DNA only in the brain and bone marrow tissues., (Copyright © 2019 Elsevier B.V. All rights reserved.)

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 ¼ 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.

This study evaluated the cytotoxicity and biocompatibility of a new bioceramic endodontic sealer (i.e., Sealer Plus BC) in comparison with those of MTA Fillapex and AH Plus. L929 fibroblasts were cultured and Alamar Blue was used to evaluate cell viability of diluted extracts (1:50, 1:100, and 1:200) from each sealer at 24 h. Polyethylene tubes that were filled with material or empty (as a control) were implanted in the subcutaneous tissue of rats. The rats were killed after 7 and 30 d (n = 8), and the tubes were removed for histological analysis. Parametric data was analyzed using a one-way ANOVA test, and nonparametric data was analyzed via the Kruskal-Wallis test followed by the Dunn test (p < 0.05). A reduction in cell viability was observed in the extracts that were more diluted for Sealer Plus BC when compared to that of Control and AH Plus (p < 0.05). However, the 1:50 dilution of the Sealer Plus BC was similar to that of the Control (p > 0.05). Conversely, more diluted extracts of MTA Fillapex (1:200) and AH Plus (1:100 and 1:200) were similar to the Control (p > 0.05). Histological analysis performed at 7 d did not indicate any significant difference between tissue response for all materials, and the fibrous capsule was thick (p > 0.05). At 30 d, Sealer Plus BC was similar to the Control (p > 0.05) and MTA Fillapex and AH Plus exhibited greater inflammation than the Control (p < 0.05). The fibrous capsule was thin for the Control and for most specimens of Sealer Plus BC and AH Plus. Thus, Sealer Plus BC is biocompatible when compared to MTA Fillapex and AH Plus, and it is less cytotoxic when less-diluted extracts are used.

Objectives: The aim of this study was to evaluate the subcutaneous response induced by Roeko Guttaflow2 (RG), Sealapex Xpress (SX), AH Plus (AHP) sealers., Methods: 100 BALB/c mice received implants in the subcutaneous tissue with the tested materials (10 animals per period for each evaluated sealer) and were evaluated after different experimental periods (7, 21 and 63 days), in each animal was placed a tube, the control group was an empty tube. Histological analysis evaluated semi-quantitatively the inflammatory infiltration, collagen fiber formation and tissue thickness. In addition, immunohistochemistry was performed for interleukin-6 (IL-6). Data were statistically analyzed (α = 0.05)., Results: RG promoted a greater collagen fiber formation at 7 days and 63 days compared to the CG (p = 0.004) and AHP (p = 0.005) respectively, while at 21 days, the SX promoted a greater reaction (p = 0.021). For the tissue thickness, there was a greater reaction at 7 days with CG (p = 0.0156) and with RG at 63 days (p = 0.03). Regarding the inflammatory infiltrate, there was no difference at 7 days and 63 days (p = 0.5; p = 0.27), while at 21 days, a statistically difference was found between SX, CG (p = 0.04) and RG (p = 0.027). In addition, the presence of IL-6 was observed in almost all groups, with a more intense marking at 7days., Significance: All cements evaluated presented a satisfactory tissue response, however, RG was the one that presented a more satisfactory tissue response., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Aim: To evaluate and compare the cytotoxic effects of different types of root canal., Materials and Methods: The sealers were eluted with culture medium for 1 hour, 7 days, and 14 days. Cell viability was estimated by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay and trypan blue exclusion method on human periodontal ligament (PDL) fibroblast cells. Sealers used are mineral trioxide aggregate (MTA)-based sealer (MTA Fillapex, Angelus), calcium hydroxide-based sealer (Apexit Plus, Ivoclar Vivadent), resin-based sealer (AH Plus, Dentsply), and zinc oxide eugenol-based sealer (Tubli Seal, SybronEndo)., Results: The order of cytotoxicity through MTT assay, at the end of the second week, was observed as MTA Fillapex> Tubli Seal> Apexit Plus > AH Plus. The percentage cell viability obtained after trypan blue exclusion method decreased in the order of Apexit Plus> Tubli Seal> AH Plus> MTA Fillapex, which was similar to the reported cytotoxicity from the MTT assay after 1 hour., Conclusion: Each type of sealer showed moderate-to-severe cytotoxic response when compared with the control. The MTA Fillapex was found to be the most cytotoxic sealer. Use of resin-based material as a root canal sealer may result in a more favorable response to PDL fibroblasts., Clinical Significance: Having knowledge of the cytotoxicity of various sealers will help in increasing patient's comfort.

This study aims to evaluate, in vitro, the effect of Aloe vera associated with endodontic medication, with or without laser photobiomodulation (FTL) irradiation in FP6 human pulp fibroblasts. The materials were divided into eight groups: CTR - control; CL - FTL alone; AA - Aloe vera with distilled water; AL - Aloe vera with distilled water and FTL; HA - calcium hydroxide P.A. with distilled water; HL - calcium hydroxide P.A. with distilled water and FTL; HAA - calcium hydroxide P.A. with Aloe vera and distilled water; HAL - calcium hydroxide P.A. with Aloe vera, distilled water, and FTL. The cytotoxicity was evaluated by MTT assay at 24, 48, and 72h and the genotoxicity by micronucleus test assay. This study was performed in triplicate. Data obtained in both tests were statistically analyzed by ANOVA and Tukey's tests (p≤0.05). Group AA presented high genotoxicity and low cytotoxicity. After 24, 48, and 72h, the group HAA significantly reduced the cell viability. Interaction with FTL showed slightly increase cell viability after 24 and 48h in groups CL and HL (p<0.001), despite the high genotoxicity in group CL and low genotoxicity in group HL. Group AL showed higher cell survival rate at 72h (p<0.05) and high genotoxicity (p<0.001). It was concluded that Aloe vera allowed higher cell viability in human pulp fibroblasts in the presence of calcium hydroxide or with FTL separately, but genotoxicity increased in these associations., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Introduction: Root canal sealers exhibit varying degrees of cytotoxicity to periapical tissues. This in turn results in inflammation, delayed wound healing, and even bone resorption. This study aimed to explore the effect of the addition of an antioxidant like pachymic acid on the cytotoxicity of 4 root canal sealers, namely, Tubliseal (Kerr, Romulus, MI), a zinc oxide eugenol-based sealer; AH Plus (Dentsply De Trey GmbH, Konstanz, Germany), an epoxy resin-based sealer; Sealapex (Kerr), a calcium hydroxide-based sealer; and EndoREZ (Ultradent Products, South Jordan, UT), a methacrylate resin-based sealer., Methods: Sealers mixed according to the manufacturers' instructions formed the experimental groups. Subgroups were determined based on the absence (subgroup A) or addition (subgroup B) of pachymic acid. The experimental sealers were added to L929 mouse fibroblast cells immediately after mixing. Cell viability was evaluated by methylthiazoletetrazolium assay after 24 hours. Data were analyzed using 1-way analysis of variance. Intergroup comparisons were performed using the Kruskal-Wallis test, and intragroup comparisons were done using independent t and post hoc tests., Results: All 4 sealers were cytotoxic but to varying degrees. In both the subgroups, Sealapex exhibited the lowest cytotoxicity followed by AH Plus, Tubliseal, and EndoREZ (P < .05). The addition of pachymic acid reduced the cytotoxicity of all the sealers except that of EndoREZ (P > .05)., Conclusions: Calcium hydroxide-based Sealapex showed the least cytotoxicity compared with the other sealers. Pachymic acid could be a viable therapeutic agent to overcome the potential adverse effects associated with root canal sealers., (Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Pulp therapy is the last resort for preserving deciduous teeth. However, the genotoxic and cytotoxic effects of many products used in this therapy are not well established. The aim of this study was to use the micronucleus test on bone marrow from mice to evaluate the genotoxic and cytotoxic effects of four filling pastes: zinc oxide, calcium hydroxide P.A., mineral trioxide aggregate and an iodoform paste (iodoform + camphorated + paramonochlorophenol + rifamycin + prednisolone). Male Swiss mice were divided into 4 groups of 10 animals, each exposed to one of the pastes, and were subdivided according to the dilutions tested: 1/10, 1/50, 1/500 and 1/1000 administered intraperitoneally (0.1ml/10g of weight). Cyclophosphamide was the positive control. The negative controls were dimethylsulfoxide and buffered saline solution. Five animals were killed 24h and five 48h after the treatment. The material was processed in accordance with Schmid (1976) and micronuclei were counted in 1000 polychromatic erythrocytes (PCE), under an optical microscope in a blinded test. Cytotoxicity was evaluated using the PCE/normochromatic erythrocyte (NCE) ratio in 200 erythrocytes. The micronucleus analysis results were evaluated using the conditional test for comparing proportions in situations of rare events. Analysis of variance and Tukey's test were used to evaluate the PCE/NCE ratio. There was significantly greater occurrence of micronuclei in the animals treated with iodoform paste at all the dilutions tested, at both sacrifice times. Greater occurrence of micronuclei was observed among the animals treated with zinc oxide and sacrificed 48h after the treatment, at the dilutions 1:50; 1:500 and 1:1000. Calcium hydroxide P.A. and mineral trioxide aggregate did not present any genotoxic or cytotoxic effects. The genotoxicity and cytotoxicity of zinc oxide and iodoform paste revealed here constitute an initial step towards their contraindication, but additional studies will be necessary in order to securely establish the risks involved in their use., (© The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Works of art are constantly under physical, chemical and biological degradation, so constant restoration is required. Consolidation is an important step in restoration, and traditional approaches and materials have already shown their limitations. To solve these problems, new nanoparticle-based consolidants were developed. No information on their toxicity is yet available. In this work, we focused our attention on potential risks posed by three commercially available nanoparticle-based consolidants: silica (SiO2 NPs), silanized silica (silanized SiO2 NPs) and calcium hydroxide (nanolime) nanoparticle dispersions. Occupational exposure impact was tested on three in vitro models mimicking inhalation, dermal contact and systemic routes. While no toxic effects were observed for nanolime and silanized SiO2 NPs, bare SiO2 NPs showed a dose- and time-dependent damage in all considered models. Corrosion test on EpiSkin® revealed no viability reduction. Works of art degradation is partially due to microorganism activity. Consolidant antibacterial activity was evaluated on three representative bacterial strains. Silica NPs-based consolidants showed effect on specific bacterial groups, while no specificity was observed with nanolime. In conclusion, silanized SiO2 NPs-based consolidant emerged as the safest and bacteriologically active product. The different biological impact of bare and silanized SiO2 NPs highlights the importance of safe-by-design approach in developing nanoparticle-containing products., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Introduction: This study aimed to investigate the effect of BioAggregate, a calcium silicate-based nanoparticulate bioceramic, on the regulation of receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclast differentiation and bone resorption in vitro, as well as to delineate the underlying molecular mechanism. The performance of BioAggregate was compared with that of ProRoot mineral trioxide aggregate (MTA)., Methods: Cells of a murine macrophage cell line RAW 264.7 were treated with various concentrations of BioAggregate and MTA extracts. Cytotoxicity of material extracts was evaluated with Cell Counting Kit-8 assay. RANKL-induced osteoclast differentiation and function were assessed with tartrate-resistant acid phosphatase staining, F-actin staining, and lacunar resorption pits assay. The mRNA expression associated with osteoclast function was detected with quantitative real-time polymerase chain reaction. Related molecular signaling pathways were investigated with Western blot and immunofluorescence., Results: BioAggregate extracts dose-dependently inhibited RANKL-induced osteoclast formation and resorption capacity without evident cytotoxicity. RAW 264.7 cells exposed to BioAggregate extracts also presented a decrease in RANKL-stimulated mRNA expression of osteoclast-related genes and transcription factors. Moreover, cells treated with BioAggregate extracts exhibited attenuated TRAF6 expression, suppressed mitogen-activated protein kinase signaling, and decreased nuclear translocation of NFATc1 and c-Fos in the presence of RANKL. Comparable effects were induced by MTA., Conclusions: BioAggregate and MTA exhibit comparable inhibitory effect on osteoclast differentiation and function in vitro, and our findings provide valuable insights into the mechanism of bioceramic-mediated anti-osteoclastogenic activity., (Copyright © 2015 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Purpose: To describe presentation, treatment, and follow-up after unilateral alkaline injuries to the eye in four dogs., Material and Method: The case notes of four patients that suffered from alkaline injuries to the eye were included in this series., Results: Acute clinical signs included blepharospasm and edema of the eyelids, chemosis and conjunctival hyperemia, conjunctival ischemia, destruction of the corneal epithelium, a whitish haze of the corneal stroma, mild corneal edema, and uveitis. Two patients showed depigmentation of the eyelids. Presumed endothelial cell damage resulted in severe corneal edema in two dogs. Long-term complications included phthisis bulbi, scarring of the eyelids and damage to the meibomian glands, symblepharon formation, conjunctivalization of the cornea, corneal vascularization, pigmentation, and fibrosis. Persisting corneal edema was seen in the dogs with presumed endothelial cell damage. One dog developed a mild bullous keratopathy with superficial corneal ulcerations 4½ years after the injury and had a reduced anterior chamber depth on ultrasound., Conclusion: The damage to the ocular structures described here mainly affects the ocular surface. One patient presumably suffered an injury to the ciliary body epithelium resulting in a phthisical globe. Chronic corneal edema, conjunctivalization, and scarring can result in permanent visual impairment. Healing of the ocular surface can take weeks and is associated with a dramatic vascular response. However, a severely vascularized cornea has the potential to clear and allow a good visual outcome long term. Ongoing discomfort was only seen in one case with persistent corneal edema and a secondary bullous keratopathy., (© 2014 American College of Veterinary Ophthalmologists.)

Aim: The purpose of this in vitro study was to evaluate the effects of intracanal medicaments commonly used in endodontic regeneration on the survival of human dental pulp cells (DPCs)., Methods: DPCs were cultured and exposed to either no medicament treatment or low concentrations (0.3-5 mg ml(-1) ) of calcium hydroxide [Ca(OH)2 ], triple antibiotic paste (TAP), or double antibiotic paste (DAP) for 3 days. After that, toxicity to the DPCs was determined by lactate dehydrogenase activity assays (LDH) and cell proliferation was measured by colorimetric assays (WST-1). Two-way anova followed by Fisher's protected least significant differences was used for statistical analyses (α = 0.05)., Results: The group-by-concentration interactions were significant for the LDH and WST-1 assays (P < 0.0001). For the LDH assays, only the highest tested concentration (5 mg ml(-1) ) of Ca(OH)2 and TAP caused significant toxicity to the DPCs compared with the untreated control, while four tested concentrations of DAP (0.5, 1, 2.5, and 5 mg ml(-1) ) caused significant toxicity to the DPCs compared with the untreated control. For the WST-1 assays, the highest concentrations that did not negatively affect the proliferation rate of DAP, TAP and Ca(OH)2 were 0.3, 2, and 2.5 mg ml(-1) , respectively., Conclusion: The low concentrations of intracanal medicaments tested in this study were not cytotoxic in cultured cells. However, these concentrations are much lower than the concentrations that have been advocated in endodontic regeneration. Furthermore, the negative effects of TAP on DPCs were detected at lower concentrations by using the WST-1 assays than by measuring the LDH release., (© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: The objective of this study was to compare the cytotoxicity, inflammatory response, osteogenic effect, and the signaling mechanism of these biologic activities of 4 calcium compound-based root canal sealers (ie, Sealapex [Sybron Kerr, WA], apatite root sealer [ARS; Dentsply Sankin, Tokyo, Japan], MTA Fillapex [Angelus Indústria de Produtos Odontológicos S/A, Londrina, PR, Brazil], and iRoot SP [Innovative BioCreamix Inc, Vancouver, Canada]) in human periodontal ligament cells., Methods: Cytotoxicity was assessed using the 3-(4,5-dimethylthiazolyl-2-yl)-2,5-diphenyltetrazolium bromide assay. Levels of inflammatory mediators were measured by enzyme-linked immunosorbent assay, reverse-transcription polymerase chain reaction, and Western blot analysis. Osteogenic potential was evaluated by alkaline phosphatase activity, alizarin red staining, and marker genes by reverse-transcription polymerase chain reaction. The signal transduction pathways were examined by Western blotting., Results: None of the sealers were cytotoxic. ARS, MTA Fillapex, and iRoot SP induced a lower expression of proinflammatory mediators than Sealapex. All sealers increased ALP activity and the formation of mineralized nodules and up-regulated the expression of osteoblastic marker messenger RNA. ARS, MTA Fillapex, and iRoot SP showed superior osteogenic potential compared with Sealapex. The expression and/or activation of integrin receptors and downstream signaling molecules, including focal adhesion kinase, paxillin, Akt, mitogen-activated protein kinase, and nuclear factor κB, was induced by ARS, MTA Fillapex, and iRoot SP treatment but not by Sealapex treatment., Conclusions: We show for the first time that ARS, MTA Fillapex, and iRoot SP induce a lower expression of inflammatory mediators and enhance osteoblastic differentiation of PDLCs via the integrin-mediated signaling pathway compared with Sealapex., (Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Aim: To investigate and compare the systemic toxic effect of DiaRoot BioAggregate and grey ProRoot Mineral trioxide aggregate (MTA) on the liver and kidney after 7 and 30 days., Methodology: Forty-two white albino rats were divided into two main groups. Group (1), considered the control group (n = 18), was further divided into two subgroups. The negative control subgroup (n = 6) received no treatment. The empty tube subgroup (n = 12) received empty sterile Teflon tubes. In Group (2), considered the experimental group (n = 24), the rats were divided equally into two subgroups. One subgroup received MTA, whilst the other received BioAggregate. The materials in the Teflon tubes were implanted subcutaneously in the dorsal side of the rats. Blood samples were taken to investigate the change of kidney and liver functions on day 7 and day 30. The liver and kidney organs were subjected to histopathological examination and calculation of the number of inflammatory cells. Data analysis was performed using one-way anova with post hoc multiple comparisons with the Tukey's test. Student's t-test was used to compare the changes in liver and kidney functions amongst the groups., Results: On day 7, a significantly more severe inflammatory reaction was observed in both experimental subgroups compared with the control (P < 0.05); the severity decreased after 30 days. The kidney functions were not affected after 7 days but had subsequently increased after 30 days (P < 0.001). Liver functions increased after 7 days and had decreased in the BioAggregate subgroup after 30 days, whilst in the MTA subgroup, a continuous increase in the level of liver function was observed., Conclusions: Mineral trioxide aggregate had adverse effects on the liver and kidney that were significantly more severe than BioAggregate but with no permanent damage., (© 2013 International Endodontic Journal. Published by John Wiley & Sons Ltd.)

Introduction: Calcium hydroxide (Ca[OH]2) microcapsules were synthesized for use in controlled release. The aim of this study was to evaluate the cytotoxicity, antibacterial properties, and influence on gene expression of bone-related markers of 2 different formulas of Ca(OH)2 microcapsules., Methods: Two formulas of Ca(OH)2 microcapsules (A and B) were evaluated, and pure Ca(OH)2 powder was used as a positive control. The shell material of formula A was pure EC, and the PLA/EC blend of 1:1 was used as the shell material for formula B. The MG63 cells/Cell Counting Kit-8 (Dojindo, Kumamoto, Japan) were used to evaluate the cytotoxicity, and the colony-forming units of Enterococcus faecalis were monitored for the antibacterial effect. The relative messenger RNA expression of collagen I and osteocalcin was determined by real-time polymerase chain reaction., Results: Both formulas of the Ca(OH)2 microcapsules showed no cytotoxicity in MG63 cells; however, the Ca(OH)2 positive control did exhibit cytotoxicity. The antibacterial effect of the 2 microcapsule formulas lasted longer than the positive control, and formula A lasted longer than formula B. For both Ca(OH)2 microcapsule formulas, the relative messenger RNA expression of collagen I and osteocalcin was prolonged and up-regulated. The time effects of the influence on messenger RNA expression of collagen I and osteocalcin were different between the 2 microcapsule formulas., Conclusions: Ca(OH)2 microcapsules had prolonged antibacterial activity and prolonged the up-regulation of bone-related markers with reduced cytotoxicity., (Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Objectives: The purpose of this study was to evaluate and compare the cytotoxic effects of ProRoot MTA and DiaRoot BA, a bioceramic nanoparticulate cement, on subcutaneous rat tissue., Study Design: Fifty Sprouge Dawley rats were used in this study. Polyethylene tubes filled with ProRoot MTA and DiaRoot BioAggregate, along with a control group of empty, were implanted into dorsal connective tissue of rats for 7, 15, 30, 60, and 90 days. After estimated time intervals the rats were sacrificed. The specimens were fixed, stained with hematoxylin and eosin, and then evaluated under a light microscope for inflammatory reactions and mineralization., Results: All groups evoked a severe to moderate chronic inflammatory reaction at 7 and 15 days, which decreased with time. Both the MTA and BioAggregate groups showed similar inflammatory reactions, except at 90 days when MTA showed statistically significant greater inflammation (p>0.05). The MTA group showed foreign body reaction at all times. Compared to BioAggregate, MTA showed significantly more foreign body reaction at 60 and 90 days (p<0.0001). After 30 days foreign body reaction of BioAggregate decreased significantly. Both MTA and BioAggregate groups showed similar necrosis at 7 and 15 days (p=0.094 and p=0.186 respectively). No necrosis was observed after 15 days. Similarly there was no fibrosis after 30 days for both MTA and BioAggregate groups (p>0.05)., Conclusions: Since DiaRoot BioAggregate showed significantly better results than MTA, we can conclude that it is more biocompatible. However, further studies are required to confirm this result.

A 9-year-old boy presented with unilateral, total limbal stem cell deficiency (LSCD) complicated by the presence of a large fibro-vascular ocular surface mass lesion secondary to accidental lime injury. The pathological tissue covering the cornea was excised and simple limbal epithelial transplantation (SLET) was performed using autologous limbal tissue from the fellow eye. Histopathology of the excised ocular surface tissue revealed exuberant granulation tissue interspersed with retained calcium particles. At 6 weeks postoperatively, a focal recurrence of LSCD with symblepharon and forniceal shortening was noted superiorly. This was successfully managed by performing conjunctival autografting along with supplemental SLET. The unaided vision had improved from light perception at presentation to 20/40 at 6 months postoperatively. The fornices were deep and the corneal surface was avascular, epithelised and stable. This case demonstrates the efficacy of SLET in a child with severe ocular burns, highlighting the role of supplementary procedures customised to treat focal recurrences of LSCD.

Aim: The present study evaluated the inflammatory/irritant potential of propolis in comparison with commonly used intracanal irrigants such as chlorhexidine and calcium hydroxide, with normal saline solution as control using an animal (Wistar rats) model., Method: 2% Evans blue was intravenously injected into the lateral caudal vein. 0.1 ml each of the test solutions was intradermally injected into the experimental sites designed on their shaved backs. The animals were then sacrificed after 1 1/2 and 3 hours respectively. Each piece of skin containing the injected solution was excised, immersed in 4 ml formamide and incubated at 45 degrees C for 72 hours. After filtration with glass wool, optical density(OD) was measured using a spectro-photometer and analyzed statistically., Results: At 620 nm irrespective of time, the mean optical density with Calcium Hydroxide was found to be maximum (0.197 +/- 0.095) while that with DMSO Propolis was found to be minimum (0.070 +/- 0.016). Both at 90 min and 180 min, the mean optical density with Calcium Hydroxide was found to be maximum., Conclusions: On short-term evaluation, maximum inflammation was seen with calcium hydroxide followed by chlorhexidine and DMSO extract of propolis. Minimum inflammation was seen with sterile physiologic saline. With progress of time, maximum inflammation was seen with calcium hydroxide followed by chlorhexidine and DMSO extract of propolis which was non-significant.

Objectives: The aim of the present study was to investigate the effects of root canal sealers on the cytotoxicity of 3T3 fibroblasts during a period of 5 weeks., Material and Methods: Fibroblasts (3T3, 1×105 cells per well) were incubated with elutes of fresh specimens from eight root canal sealers (AH Plus, Epiphany, Endomethasone N, EndoREZ, MTA Fillapex, Pulp Canal Sealer EWT, RoekoSeal and Sealapex) and with elutes of the same specimens for 5 succeeding weeks after immersing in simulated body fluid. The cytotoxicity of all root canal sealers was determined using the MTT assay. Data were analyzed using ANOVA and Tukey's test., Results: RoekoSeal was the only sealer that did not show any cytotoxic effects (p<0.05). All the other tested sealers exhibited severe toxicity initially (week 0). MTA Fillapex remained moderately cytotoxic after the end of experimental period. Toxicity of the other tested sealers decreased gradually over time. The evaluated root canal sealers presented varying degrees of cytotoxicity, mainly in fresh mode., Conclusions: RoekoSeal had no cytotoxic effect both freshly mixed and in the other tested time points. MTA Fillapex was associated with significantly less cell viability when compared to the other tested root canal sealers.

Aim: To determine whether cells pre-stressed by known cytotoxic or inflammatory agents are more susceptible to the deleterious effects of a calcium hydroxide formulation used in pulp capping., Methodology: Adult human dermal fibroblasts were treated for 48 h with 0.001% chlorhexidine, 0.2% ethanol, 5 μg mL(-1) Escherichia coli lipopolysaccharide (LPS) or 0.05 mmol L(-1) nicotine. Cells were subsequently treated with the soluble materials extracted from Dycal pellets for an additional 24 h. Controls included cells cultured in medium only and cells exposed to Dycal only. Cytotoxicity was measured using colorimetric MTT, WST and secreted lactate dehydrogenase assays. In addition, mitotic activity was evaluated using a colorimetric histone H3 phosphorylation assay. Data were statistically analysed using anova with Tukey's multiple comparison post-test and significance at P ≤ 0.05., Results: For all assays, measured values for cells treated with chlorhexidine, ethanol, LPS or nicotine plus the soluble materials extracted from Dycal pellets were significantly lower compared to control (P < 0.05) for all comparisons between experimental conditions. However, between treatments and for comparisons of treatments with Dycal, there were no differences observed for any assay., Conclusions: Calcium hydroxide in a formulation used in dental clinical procedures is highly cytotoxic to cultured cells, as evidenced by several cellular assays. However, other known toxic agents, including chlorhexidine, ethanol, bacterial LPS and nicotine, do not appear to function synergistically to increase the deleterious cellular effects of the calcium hydroxide in an in vitro model of cytotoxicity., (© 2012 International Endodontic Journal.)

Introduction: Regenerative endodontic procedures are an alternative treatment for immature teeth with necrotic pulps. Typically, intracanal medicaments such as triple antibiotic paste (TAP) or double antibiotic paste (DAP) and calcium hydroxide (Ca[OH](2)) are used for disinfection. However, their effect on human stem cells of the apical papilla (SCAPs) is unknown. We hypothesized that intracanal medicaments at high concentrations are toxic to SCAPs. To test this hypothesis, a cell culture assay was used., Methods: Briefly, SCAPs were cultured and subjected to either no drug treatment or various concentrations including TAP, DAP, modified TAP (ciprofloxacin, metronidazole and cefaclor), Augmentin (Champs Pharmacy, San Antonio, TX), or Ca(OH)(2). Viable stem cells counts were obtained using an automated method of detecting trypan blue dye at 3 days after treatment., Results: All 4 antibiotics significantly reduced SCAP survival in a concentration-dependent fashion. Interestingly, Ca(OH)(2) was conducive with SCAP survival at all concentrations., Conclusions: Collectively, our data show that high concentrations of antibiotics have a detrimental effect on SCAP survival, whereas lower concentrations as well as Ca(OH)(2) at all tested concentrations are conducive with SCAP survival and proliferation. These studies highlight the clinically important point that intracanal medicaments must be used at concentrations that are bactericidal while having minimal effects on stem cell viability., (Copyright © 2012 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

The aim of this work was to evaluate the effects of different times of extraction on the cytotoxicity of six representatives of different root canal sealer groups-Real Seal SE, AH Plus, GuttaFlow, Sealapex, Roth 801, and ThermaSeal Plus-with human gingival fibroblasts. The materials were prepared according to manufacturers' specifications, and were incubated in culture medium (DMEM) at 37ºC for 1, 7, 14, 21, and 28 days, with daily washing, to simulate periodontal ligament clearance. Human fibroblasts were exposed to the final extracts at 24 hours, and cell viability was determined by MTT assay, with exposure to unconditioned DMEM as a negative control. Statistical analysis comparing cytotoxicities at each exposure time was performed by ANOVA with Scheffé adjustment for multiple comparisons at a 95% confidence level. Results indicated that GuttaFlow was significantly less cytotoxic than all other sealers (p < 0.05) at 1 day of extraction. After 7 days of extraction, cell viability for GuttaFlow was significantly increased as compared with that of all groups except sealer AH Plus. At day 14, cytotoxicity of Sealapex was significantly higher than that of all other sealers (p < 0.05). At days 21 and 28, there were no significant differences in cytotoxicity among sealer groups. All materials presented some level of cytotoxicity to fibroblasts, while GuttaFlow was the least cytotoxic sealer tested. However, the cytotoxicity of all materials seemed to decrease similarly in a time-dependent manner.

Aim: To investigate the effects of root repair materials on the cytotoxicity and gelatinolytic activity of matrix metalloproteinases (MMPs) in 3T3 fibroblasts., Methodology: Fibroblasts (3T3, 3 × 10(5) cells per well) were incubated with elutes of calcium hydroxide (Biodinâmica, Ibiporã, PR, Brazil), EndoBinder (Binderware, São Carlos, SP, Brazil) and mineral trioxide aggregate (MTA) (Angelus, Londrina, PR, Brazil) for 24 h. The cytotoxicity of all root repair materials was determined using the MTT assay. Supernatants of cell cultures incubated with materials were collected after 24 h to determine the levels of MMP-2 and MMP-9 gelatinolytic activity by gelatin zymography. Data were analysed using anova and Tukey's test., Results: Cells secreted MMP-2 after 24 h with calcium hydroxide inducing significantly greater MMP-2 expression in relation to the control and the other root repair materials (P < 0.05). The cytotoxicity results revealed that there was no significant difference in the cell viability of MTA, EndoBinder and the control group. However, there was a significantly reduced cell viability of 3T3 fibroblasts in association with calcium hydroxide (P < 0.05)., Conclusions: Calcium hydroxide was associated with significantly less cell viability when compared with EndoBinder and MTA. All materials had gelatinolytic activity for MMP-2 with calcium hydroxide being associated with the greatest activity., (© 2012 International Endodontic Journal.)

Introduction: The purpose of this in vitro study was to compare the cytotoxicity of white mineral trioxide aggregate cement (AMTA, MTA-Angelus), Brasseler Endosequence Root Repair Putty (ERRM), Dycal, and Ultra-blend Plus (UBP) by using human dermal fibroblasts and a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay., Methods: Cultured adult human dermal fibroblasts were exposed to multiple concentrations of material elutes. The test material samples were immersed and incubated in the culture medium for 2, 5, or 8 days at 37°C. The cytotoxic effects were recorded by using an MTT-based colorimetric assay. Positive and negative controls were used. The results were statistically examined by one-way analysis of variance and Tukey post tests., Results: The cell viability of cultures exposed to all dilutions of AMTA, ERRM, and UBP was statistically similar to the negative control at 2 and 5 days. Only the Dycal-exposed specimens exhibited a statistically significant increase in cytotoxicity at the 2 initial evaluation periods. After exposure to the 8-day elutes, the respective percentage of cell survivability was 91% (Brasseler), 88% (MTA-Angelus), 76% (Ultra-blend Plus), and 37% (Dycal)., Conclusions: From the data in this in vitro study, AMTA, ERRM, and UBP had statistically similar adult human dermal fibroblast cytotoxicity levels. Relative to the negative control, only Dycal was shown to have a statistically significant cytotoxic effect to adult human dermal fibroblasts at all tested intervals., (Copyright © 2012 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Objective: To investigate the cytotoxicity of calcium hydroxide (CH) and composite resin on fibroblasts derived from human embryo body fibroblasts-H9 (EBf-H9), human dental pulp cells (hDPCs) and immortalized fibroblasts L929; and to evaluate the use of EBf-H9 as a cellular model for cytotoxicity screening of dental materials., Methods: The EBf-H9 cells were derived from human embryonic stem cells (H9) via outgrowth of embryonic body (EB); hDPCs were isolated from healthy dental pulp, and identified by immunochemical staining. Cell Counting Kit-8 (CCK-8) assay was applied to analyze the cytotoxicity of CH and composite resin with serial concentrations on the 3 kinds of cells., Results: Following 24 h and 48 h (or 72 h) post-treatment of CH and composite resin, the viability of L929 cells was significantly lower than that of EBf-H9 and hDPCs (P<0.05), and there was no significant difference between the last two groups (P>0.05)., Conclusion: Immortalized fibroblasts L929 cells exhibited different response to CH and composite resin compared with EBf-H9 and hDPCs, and the last two cell types were similar to each other. This study indicated that fibroblasts derived from human embryonic stem cells were a potential cellular model instead of traditional immortalized murine cell line for cytotoxicity screening assay.

Aim:   To compare the cytotoxicity of four endodontic sealers (Sealapex, Pulp Canal Sealer EWT, Real Seal and MTA Fillapex) either 1 or 7 days after mixing, when assessed through a multiparametric analysis employing human primary cells closely related to periapical tissues., Methodology:   Extracts of each sealer were prepared following 24-h exposure to culture media, at either 24 h or 7 days after mixing. Primary human osteoblasts were exposed to extracts for 24 h, at 37 °C with 5% CO(2) , and cell viability was evaluated by a multiparametric assay assessing sequentially, on the same cells, mitochondrial activity (XTT), membrane integrity (neutral red test) and total cell density (crystal violet dye exclusion test). Results from each test and experimental time were compared by 2-way analysis of variance (anova)., Results:   All endodontic sealers had strong cytotoxicity 24 h after mixing, according to all parameters evaluated. At a longer setting period (7 days), viability for Sealapex was significantly increased (P < 0.05) and Pulp Canal Sealer achieved levels of cytocompatibility similar to the control group. The anova indicated a general correlation between the cytotoxicity of the materials and the time after mixing, with some level of dependence on the cell viability assay employed., Conclusions:   All materials had high cytotoxic levels for human primary cells, mostly on a time-dependent basis, as shown by three different cell viability tests., (© 2011 International Endodontic Journal.)

Purpose: To determine whether exposure to alkaline chemicals results in predictable changes in corneal protein profile. To determine whether protein profile changes are indicative of severity and duration of alkali exposure., Methods: Enucleated bovine and porcine (n = 59 each) eyes were used for exposure to sodium, ammonium, and calcium hydroxide, respectively. Eyes were subjected to fluorescein staining, 5-bromo-2'-deoxy-uridine (BrdU) labeling. Excised cornea was subjected to protein extraction, spectrophotometric determination of protein amount, dynamic light scattering and SDS-PAGE profiling, mass spectrometric protein identification, and iTRAQ-labeled quantification. Select identified proteins were subjected to Western blot and immunohistochemical analyses., Results: Alkali exposure resulted in lower protein extractability from corneal tissue. Elevated aggregate formation was found with strong alkali exposure (sodium hydroxide>ammonium, calcium hydroxide), even with a short duration of exposure compared with controls. The protein yield after exposure varied as a function of postexposure time. Protein profiles changed because of alkali exposure. Concentration and strength of the alkali affected the profile change significantly. Mass spectrometry identified 15 proteins from different bands with relative quantification. Plexin D1 was identified for the first time in the cornea at a protein level that was further confirmed by Western blot and immunohistochemical analyses., Conclusions: Exposure to alkaline chemicals results in predictable and reproducible changes in corneal protein profile. Stronger alkali, longer durations, or both, of exposure resulted in lower yields and significant protein profile changes compared with controls.

Aim: To evaluate the antibacterial and cytotoxic effects of Liquorice as a root canal medicament and to compare its action to the commonly used root canal medicament calcium hydroxide Ca(OH)(2)., Methodology: The antibacterial effect of Liquorice and Ca(OH)(2) either separately or in combination was investigated against Enterococcus faecalis. Agar-well diffusion methods, broth microdilution tests and biofilm susceptibility assays were used to determine the antibacterial activity. Human periodontal ligament fibroblast tissue culture was used to assess the cytotoxicity of the preparations under investigation., Results: Liquorice extract either by itself or in combination with Ca(OH)(2) had a significant inhibitory effect against Enterococcus faecalis compared with that of Ca(OH)(2) alone. The use of Liquorice extract followed by Liquorice/Ca(OH)(2) mixture retained significantly more viable periodontal ligament cells than Ca(OH)(2) , which had a strong lethal effect on the cells., Conclusion: Liquorice extract either separately or as Liquorice/Ca(OH)(2) mixture had a potent bactericidal effect against Enterococcus faecalis and retained compatibility with fibroblasts in tissue culture compared to the commonly used root canal medicament Ca(OH)(2)., (© 2010 International Endodontic Journal.)

Aim: To investigate the cytotoxicity of bioaggregate (BA; Innovative Bioceramix, Vancouver, BC, Canada) to human periodontal ligament (PDL) fibroblasts and its effect on differentiation of human PDL fibroblasts and to compare its performance to that of mineral trioxide aggregate (MTA; Dentsply, Tulsa, OK, USA)., Methodology: Cytotoxicity was assessed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay on days 1, 2, and 3 after incubation with BA or MTA. The influence of BA and MTA on differentiation of human PDL fibroblasts on days 3, 5, and 7 was evaluated by gene expression of alkaline phosphatase (ALP) and collagen type I (COLI) via quantitative real-time polymerase chain reaction (PCR). The data were analysed by one-way analysis of variance followed by Tukey's test., Results: Cell numbers in the BA group were similar to that of the control group throughout the culture period, whereas MTA suppressed the proliferation of fibroblasts. ALP expression was significantly increased in the BA group on day 7, whilst it was enhanced by MTA on day 3. Gene expression of COLI was induced by both BA and MTA compared to the control group., Conclusions: Bioaggregate was nontoxic to human PDL fibroblasts and appeared to induce the differentiation of human PDL fibroblasts., (© 2010 International Endodontic Journal.)

Introduction: The objective was to determine the effects of growth factor treatment on dental pulp cell sensitivity to toxicity of 2 composite restoration materials, Flow Line and Durafill VS, and a calcium hydroxide pulp capping material, Dycal., Methods: Toxicity of the dental materials to cultures of primary dental pulp cells was determined by the MTT metabolism assay. The ability of 6 different growth factors to influence the toxicity was tested., Results: A 24-hour exposure to either Flow Line or Durafill VS caused approximately 40% cell death, whereas Dycal exposure caused approximately 80% cell death. The toxicity of Flow Line and Durafill VS was mediated by oxidative stress. Four of the growth factors tested (bone morphogenetic protein [BMP]-2, BMP-7, epidermal growth factor [EGF], and transforming growth factor [TGF]-beta) decreased the basal MTT values while making the cells resistant to Flow Line and Durafill VS toxicity except BMP-2, which made the cells more sensitive to Flow Line. Treatment with fibroblast growth factor-2 caused no change in basal MTT metabolism, prevented the toxicity of Durafill VS, but increased the toxicity of Flow Line. Treatment with insulin-like growth factor-I (IGF-I) increased basal MTT metabolism and made the cells resistant to Flow Line and Durafill VS toxicity. None of the growth factors made the cells resistant to Dycal toxicity., Conclusions: The results indicated that growth factors can be used to alter the sensitivity of dental pulp cells to commonly used restoration materials. The growth factors BMP-7, EGF, TGF-beta, and IGF-I provided the best profile of effects, making the cells resistant to both Flow Line and Durafill VS toxicity., (Copyright 2010 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.)

Purpose: The purpose of this study was to determine the cytotoxic effects of five different primary tooth root canal filling materials on L929 permanent cell line with MTT assay., Methods: Kri 1 paste (iodoform), Diapex (iodoform+Ca(OH)(2)), Metapaste (Ca(OH)(2) with distilled water), Dentalis (iodoform+ZOE+Ca(OH)(2)) and Kalsin (Ca(OH)(2) with glycerin) were used in this study. Tested materials were in contact for 24, 48 and 72 hours with L929 cells. At the end of the test periods, MTT test solutions were added to the plates and incubated for 3 hours at 37 degrees C. Then optic densities were read using UV visible spectrophotometer. All assays were repeated three times to ensure reproducibility. The obtained data were analyzed statistically by one-way analysis of variance and Dunnett T3 post hoc test (P<0.05)., Results: All tested materials were found cytotoxic on L929 cell line. It was found that Kri 1 paste group showed the highest survival rates., Conclusions: We concluded that the use of Kri 1 paste as a root canal filling material is a better option than other medications in primary teeth. Further research is necessary to determine the effect of root canal filling materials on vital tissues.

Introduction: The aim of this study was to evaluate the usefulness of a new product called Mineral Trioxide Aggregate (MTA) used in direct and indirect pulp capping. The results of pulp treatment using therapeutic MTA with the results after application of calcium hydroxide were compared. Moreover, the aim of the study was to determine the toxicity of the investigated materials in relation to human fibroblast cultures derived from dental pulp., Material and Methods: The study included 97 patients, male and female in the age 8-56 years. A total of 135 vital pulp therapy treatments were performed. Depending on the material and methods, the patients were assigned to four groups., Results: In the carried out study 88.2% of the results were positive after a direct coverage of the pulp using MTA and 86.7% of positive results were after the application of calcium hydroxide. In the case of indirect use of both the MTA and Ca(OH)2 100% positive results were obtained. Biocompatibility test preparations showed significantly lower toxicity of MTA compared with Ca(OH)2., Conclusions: On the basis of clinical examination Mineral trioxide Aggregate proved to be as effective as calcium hydroxide in the direct and indirect pulp capping. It has been shown that the deeper damage to the pulp, of the kind during its injury, worse the prognosis compared to an exposure in the pulp. Mineral Trioxide Aggregate proved to be highly statistically significantly less toxic compared to Ca(OH)2.

Objective: The objective of this study was to determine whether the addition of 5% calcium hydroxide to AH Plus sealer improves its biocompatibility in subcutaneous connective tissue of rats., Study Design: Thirty female rats distributed into 3 groups of 10 animals each received subcutaneous dorsal implants of silicone tubes filled with AH Plus (Group 1), AH Plus containing 5% (wt/wt) calcium hydroxide (Group 2), or no material (Group 3: control). The animals were killed after 14 days and the subcutaneous tissue containing the tubes was removed and processed for histological analysis. Biocompatibility was assessed by evaluating the inflammatory response to the implants qualitatively and quantitatively. Data were analyzed statistically by the Wilcoxon and Kruskal-Wallis tests (alpha = 0.05)., Results: The area adjacent to the implant was characterized by nonspecific chronic inflammation and was qualitatively similar in the 3 groups. Animals implanted with AH Plus/calcium hydroxide showed significantly less intense inflammatory response when compared with the animals implanted with AH Plus alone., Conclusion: The addition of calcium hydroxide to AH Plus root canal sealer improved its histopathological behavior within 14 days of analysis, producing less severe inflammatory response and less cytotoxicity when implanted in the subcutaneous tissue of rats., (Copyright 2010. Published by Mosby, Inc.)

Objectives: To examine the cytotoxic effects of MTA and Ca(OH)2 on 3T3 fibroblasts at different time intervals., Methods: Confluent cells were cultured with Ca(OH)2 and MTA in six-well plates. Wells with only fibroblasts served as controls. Cell number and viability were determined after 24 and 48 hours and 7 days of incubation. For cell viability, trypane blue exclusion assay was used. A Leitz inverted microscope was used to observe the morphological behavior of cells. The proliferation of cells was evaluated by BrdU assay. The results were analyzed with Dunn's multiple comparison, Friedman, and Kruskal-Wallis tests., Results: No difference was seen in morphology of cells for either test material. Cells treated with MTA and Ca(OH)2 were reduced in number after 24 and 48 hours, respectively. A statistically significant difference was found in number of viable cells between test groups at 48 hours of incubation. The results of BrdU assay revealed low percentages of capable cells incorporating BrdU., Conclusions: The cytotoxic effects of the test materials-MTA and Ca(OH)2-on 3T3 cell line were evaluated as cytostatic for 24 and 48 hours, respectively. But this effect was reversible because the incubated cells showed normal cell proliferation at 48 hours and 7 days, respectively; MTA showed a significantly shorter cytotoxic effect on the cells.

Objective: The aim of this investigation was to evaluate the cytotoxicity of two brands of root canal sealers, epoxy-resin based and zinc oxide-eugenol based, and one commercial calcium hydroxide paste on a monocyte cell line THP-1., Material and Methods: Undiluted (crude extract) and diluted extracts to 10%, 1%, 0.1%, 0.01%, 0.001% and 0.0001% of the sealers were tested for cytotoxicity to THP-1 cells using the trypan blue assay. Extracts were obtained according to ISO standard. Data were analyzed statistically by the Kruskal-Wallis and Mann-Whitney tests at 5% significance level., Results: Crude extract of AH Plus and Fill Canal killed approximately 90% of THP-1 cells versus 36% of THP-1 cells killed by L&C crude extract (p<0.05). Ten-fold dilutions of L&C, Fill Canal and AH Plus killed 24, 35 and 61% of THP-1 cells (p<0.05), respectively. Dilutions lesser than 1% caused minimal cell death as compared to the control groups (p>0.05), except for L&C 1% extract., Conclusions: The results revealed that the L&C paste crude extract was less cytotoxic to THP-1 cells than AH Plus or Fill Canal crude extracts.

The aim of this review was to consider laboratory experiments and clinical studies of calcium hydroxide-based root canal sealers. An extensive search of the endodontic literature was made to identify publications related to calcium hydroxide-based root canal sealers. The articles were assessed for the outcome of laboratory and clinical studies on their biological properties and physical characteristics. Comparative studies with other sealers were also considered. Several studies were evaluated covering different properties of calcium hydroxide-based sealers including physical properties, biocompatibility, leakage, adhesion, solubility, antibacterial properties, and periapical healing effect. Calcium hydroxide-based root canal sealers have a variety of physical and biological properties. Comparative studies reveal their mild cytotoxicity, but their antibacterial effects are variable. Further research is required to establish the tissue healing properties of calcium hydroxide in root canal sealers.

Objective: The main goal of this work is to compare the In vitro toxicity of MTA with other primary teeth pulpotomy agents., Study Design: The In vitro toxicity of MTA, calcium hydroxide, ferric sulphate solution, diluted formocresol and Buckley's formocresol were tested using MTT and Neutral Red Uptake cell viability assays. The results for MTA were compared to those obtained for the other substances using ANOVA and Tukey statistical tests (p < 0.05)., Results: MTA had the lower in vitro toxicity and Buckley's formocresol, the higher, with statically significant difference., Conclusion: Among the primary teeth pulpotomy agents tested, MTA showed the lower In vitro toxicity, standing as the most promising substitute to formocresol.

Aim: The aim of this study was to evaluate the cytotoxicity and genotoxicity of the new castor oil bean cement (COB) material in comparison to commonly used pulp capping materials., Methodology: Specimens of COB, calcium hydroxide (Hydro C), and mineral trioxide aggregate (white and gray MTA) were extracted in culture medium (91.6 mm(2) sample surface mL(-1)). Transfected human pulp cells (tHPCs) were exposed to dilutions of the extracts for 1 h, and the generation of reactive oxygen species (ROS) was determined by flow cytometry (FACS) using H(2)DCF-DA as a dye. Survival of tHPCs was measured photometrically using a crystal violet assay after a 24-h exposure period. Genotoxicity as indicated by the formation of micronuclei in V79 cells, and the modification of the normal cell cycle by extracts of the materials was analysed by FACS., Results: Clear cytotoxic effects were detected only with extracts of Hydro C under the current experimental conditions. The two MTA preparations induced an insignificant reduction in the number of cells. In contrast, the extracts of COB slightly induced cell proliferation. Extracts of Hydro C caused a twofold increase in ROS production, whilst the other tested materials were ineffective. An increase in the number of micronuclei was not detected with any material tested; Hydro C slightly increased the number of cells in G1 and G2., Conclusions: The COB and the two MTA preparations did not negatively influence cell survival or ROS production and may thus be further considered for pulp capping studies.

Objective: Our aim was to analyze the effect of laser phototherapy on the secretory activity of macrophages activated by interferon-gamma (IFN-gamma) and lipopolysaccharide (LPS), and stimulated by substances leached from an epoxy resin-based sealer (AH-Plus) and a calcium hydroxide-based sealer (Sealapex)., Background Data: Laser phototherapy can modulate the inflammatory process, improving wound healing. This type of therapy could be useful for modulating postoperative symptoms seen after endodontic treatment., Materials and Methods: Cytotoxicity was indirectly assessed by measuring mitochondrial activity. Macrophages were stimulated by the leached substances or not (controls), and the groups were then irradiated or not. The secretion of pro-inflammatory cytokines (TNF-alpha and MMP-1) was analyzed using ELISA. Two irradiations at 6-h intervals were done with an As-Ga-Al diode laser (780 nm, 70 mW, spot size 4.0 mm(2), 3 J/cm(2), for 1.5 sec) in contact mode., Results: The sealers were non-cytotoxic to macrophages. The production of TNF-alpha was significantly decreased by laser phototherapy, regardless of experimental group. The level of secretion of MMP-1 was similar in all groups., Conclusion: Based on the conditions of this study we concluded that in activated macrophages, laser phototherapy impairs the secretion of the pro-inflammatory cytokine TNF-alpha, but has no influence on MMP-1 secretion.

The aim of this study was to evaluate the rat subcutaneous tissue response to implanted polyethylene tubes filled with Endo-CPM-Sealer (Portland Cement Modified Sealer) (EGEO S.R.L., Buenos Aires, Argentina) compared with Sealapex (SybronEndo, Glendora, CA) and Angelus MTA (Angelus, Londrina, Brazil). These materials were placed in polyethylene and dentin tubes and implanted into dorsal connective tissue of Wistar rats for 7, 15, 30, 60, and 90 days. The specimens were prepared to be stained with hematoxylin and eosin or Von Kossa or not stained for polarized light. Qualitative and quantitative evaluations of the reaction were performed. Both materials caused mild to moderate reactions at 7 days that decreased with time. The response was similar to the control on the 30th day with Endo-CPM-Sealer and Angelus MTA and on the 60th day with Sealapex. Mineralization and granulations birefringent to the polarized light were observed with all materials. It was possible to conclude that Endo-CPM-Sealer was biocompatible and stimulated mineralization.

Suspensions of micro- or nanoparticulate SiO(2)-Na(2)O-CaO-P(2)O(5) bioactive glasses could potentially be used as dressings in traumatized front teeth with open apices as an alternative to Ca(OH)(2). These materials have a disinfecting capacity similar to Ca(OH)(2), but bear the advantage of bioactivity. However, because bioactive glasses initially act as alkaline biocides just as Ca(OH)(2) does, they may also negatively affect mechanical dentin properties over time. This was assessed in the current study using standardized human root dentin bars. Specimens were immersed in 1:20 (wt vol(-1)) suspensions of nanometric bioactive glass 45S5 or calcium hydroxide for 1, 10, or 30 days. Control specimens were immersed in pure saline for 30 days (n = 20 per group). Subsequently, modulus of elasticity (E) and flexural strength (FS) of the specimens were determined. Results were compared between groups using one-way anova and Scheffé's post-hoc test. Ca(OH)(2) caused a significant (P < 0.001) 35% drop in mean flexural strength values compared to the control treatment after 10 days. No further change was observed between 10 days and 30 days. Bioactive glass caused a 20% drop in mean flexural strength as compared to the control after 10 days. However, this difference did not reach statistical significance (P > 0.05). No effects of either material on dentin modulus of elasticity values were observed. It was concluded that the calcium hydroxide suspension affected the dentin more than the bioactive glass counterpart; however, the effect was self-limiting and probably restricted to superficial dentin layers, as suggested by the mere decrease in flexural strength but not in modulus of elasticity values.