Corrosive and cytotoxic properties of compact specimens and microparticles of Ni-Cr dental alloy.

PURPOSE: Nickel-chromium (Ni-Cr) dental alloys have been widely used in prosthodontic practice, but there is a permanent concern about their biocompatibility due to the release of metal ions. This is especially important when Ni-Cr metal microparticles are incorporated into gingival tissue during prosthodontic procedures. Therefore, the aim of this study was to examine and compare the corrosion and cytotoxic properties of compact specimens and microparticles of Ni-Cr dental alloy. MATERIALS AND METHODS: Ni-Cr alloy, Remanium CSe bars (4 mm diameter), were made by the standard casting method and then cut into 0.5-mm-thick disks. Metal particles were obtained by scraping the bars using a diamond instrument for crown preparation. The microstructure was observed by an optical microscope. Quantitative determination and morphological and dimensional characterization of metal particles were carried out by a scanning electron microscope and Leica Application Suite software for image analysis. Corrosion was studied by conditioning the alloy specimens in the RPMI 1640 medium, containing 10% fetal calf serum in an incubator with 5% CO2 for 72 hours at 37°C. Inductively coupled plasma-optical emission spectrometry was used to assess metal ion release. The cytotoxity of conditioning medium (CM) was investigated on L929 cells using an MTT test. One-way ANOVA was used for statistical analysis. RESULTS: After casting, the microstructure of the Remanium CSe compact specimen composed of Ni, Cr, Mo, Si, Fe, Al, and Co had a typical dendritic structure. Alloy microparticles had an irregular shape with a wide size range: from less than 1 µm to more than 100 µm. The release of metal ions, especially Ni and Mo from microparticles, was significantly higher, compared to the compact alloy specimen. The CM prepared from compact alloy was not cytotoxic at any tested dilutions, whereas CM from alloy microparticles showed dose-dependent cytotoxicity (90% CM and 45% CM versus control; p < 0.005). CONCLUSION: Ni-Cr microparticles showed less corrosion resistance and lower biocompatibility than compact alloy. This could affect health on long-term exposure, especially in sensitized individuals.

Biocompatibility effects of indirect exposure of base-metal dental casting alloys to a human-derived three-dimensional oral mucosal model.

OBJECTIVES: The study employed a three-dimensional (3D) human-derived oral mucosal model to assess the biocompatibility of base-metal dental casting alloys ubiquitous in fixed prosthodontic and orthodontic dentistry. METHODS: Oral mucosal models were generated using primary human oral keratinocyte and gingival fibroblast cells seeded onto human de-epidermidised dermal scaffolds. Nickel-chromium (Ni-Cr) and cobalt-chromium (Co-Cr) base-metal alloy immersion solutions were exposed to oral mucosal models for increasing time periods (2-72h). Analysis methodologies (histology, viable cell counts, oxidative stress, cytokine expression and toxicity) were performed following exposure. RESULTS: Ni-based alloy immersion solutions elicited significantly decreased cell viability (P<0.0004) with increased oxidative stress (P<0.0053), inflammatory cytokine expression (P<0.0077) and cellular toxicity levels (P<0.0001) compared with the controls. However, the Ni-free Co-Cr-based alloy immersion solutions did not elicit adverse oxidative stress (P>0.4755) or cellular toxicity (P<0.2339) responses compared with controls. CONCLUSIONS: Although the multiple analyses highlighted Ni-Cr base-metal alloy immersion solutions elicited significantly detrimental effects to the oral mucosal models, it was possible to distinguish between Ni-Cr alloys using the approach employed. The study employed a 3D human-derived full-thickness differentiated oral mucosal model suitable for biocompatibility assessment of base-metal dental casting alloys through discriminatory experimental parameters. CLINICAL SIGNIFICANCE: Increasing incidences of Ni hypersensitivity in the general population warrants serious consideration from dental practitioners and patients alike where fixed prosthodontic/orthodontic dental treatments are the treatment modality involved. The novel and analytical oral mucosal model has the potential to significantly contribute to the advancement of reproducible dental medical device and dental material appraisals.

Influence of S. mutans on base-metal dental casting alloy toxicity.

We have highlighted that exposure of base-metal dental casting alloys to the acidogenic bacterium Streptococcus mutans significantly increases cellular toxicity following exposure to immortalized human TR146 oral keratinocytes. With Inductively Coupled Plasma-Mass Spectrometry (ICP-MS), S. mutans-treated nickel-based (Ni-based) and cobalt-chromium-based (Co-Cr-based) dental casting alloys were shown to leach elevated levels of metal ions compared with untreated dental casting alloys. We targeted several biological parameters: cell morphology, viable cell counts, cell metabolic activity, cell toxicity, and inflammatory cytokine expression. S. mutans-treated dental casting alloys disrupted cell morphology, elicited significantly decreased viable cell counts (p < 0.0001) and cell metabolic activity (p < 0.0001), and significantly increased cell toxicity (p < 0.0001) and inflammatory cytokine expression (p < 0.0001). S. mutans-treated Ni-based dental casting alloys induced elevated levels of cellular toxicity compared with S. mutans-treated Co-Cr-based dental casting alloys. While our findings indicated that the exacerbated release of metal ions from S. mutans-treated base-metal dental casting alloys was the likely result of the pH reduction during S. mutans growth, the exact nature of mechanisms leading to accelerated dissolution of alloy-discs is not yet fully understood. Given the predominance of S. mutans oral carriage and the exacerbated cytotoxicity observed in TR146 cells following exposure to S. mutans-treated base-metal dental casting alloys, the implications for the long-term stability of base-metal dental restorations in the oral cavity are a cause for concern.

In vivo assessment of DNA damage induced in oral mucosa cells by fixed and removable metal prosthodontic appliances.

Given long-term effect on oral tissues due to contact with dental appliances, the biocompatibility studies of casting alloys are of great importance. It has been previously documented that metal dental appliances, due to corrosion, might induce genotoxic and mutagenic effects in cells. Therefore, the aim of presented study was to examine the genotoxicity of two dental casting alloys (Co-Cr-Mo and Ni-Cr) commonly used in fixed and removable prosthodontic appliances that are in contact with the oral epithelium for 5 years or more. For that purpose, 55 age-matched subjects were included in the study; 30 wearers of prosthodontic appliances and 25 controls. Buccal cells of oral mucosa were collected and processed for further analysis. The cell viability has been assessed by trypan blue exclusion test, while genotoxic effect of metal ions on DNA in oral mucosa cells was studied by use of alkaline comet assay. Results have shown significantly higher comet assay parameters (tail length and percentage DNA in the tail) in the group wearing metal appliances. Both subjects with Co-Cr-Mo alloy and Ni-Cr alloy showed significantly higher comet assay parameters when compared with controls. It has been confirmed that metal ions released by the two base metal dental casting alloys examined in this study, might be responsible for DNA damage of oral mucosa cells. Therefore, the results of this study emphasize the importance of the in vivo evaluation of dental materials with respect to their genotoxicity, which is of major importance to ensure long-term biocompatibility.

Development of a discriminatory biocompatibility testing model for non-precious dental casting alloys.

OBJECTIVES: To develop an enhanced, reproducible and discriminatory biocompatibility testing model for non-precious dental casting alloys, prepared to a clinically relevant surface finishing condition, using TR146 oral keratinocyte cells. METHODS: Comparative biocompatibility was determined following direct and indirect exposure of TR146 cells to two nickel-chromium (Ni-Cr) and a cobalt-chromium (Co-Cr) alloy-discs. The surface roughness of the discs was determined using a contact stylus profilometer and the elemental ion release by inductively coupled plasma mass spectrometry (ICP-MS). Subsequent biocompatibility analysis included cell morphology, cell density measurements with Trypan blue exclusion assay, inflammatory cytokine expression with ELISAs, cellular metabolic activity using XTT and cellular toxicity using lactate dehydrogenase (LDH) release assay. RESULTS: TR146 cell morphology was altered following direct and indirect exposure to the Ni-Cr alloys but not the Co-Cr alloy. Significant reductions (all P<0.001) in viable cell density measurements, cellular metabolic activity, significant increases inflammatory cytokine expression and cellular toxicity were observed when TR146 cells were exposed to the Ni-Cr alloys. Significant decreases in cell density measurements, cellular metabolic activity, significant increases inflammatory cytokine expression and cellular toxicity for the Ni-Cr d.Sign(®)15 alloy compared with d.Sign(®)10 alloy were identifiable (all P<0.001). Cellular toxicity was attributed to nickel ion release levels in solution detected by ICP-MS analysis. DISCUSSION: Nickel ions from the Ni-Cr alloys permeated the epithelial cells and activated a proinflammatory response, namely IL-1a, IL-8 and PGE2 expression. Further evidence of nickel ioninduced cell death was supported by the decreased biocompatibility of the highest nickel ion releasing alloy (d.Sign(®)15 compared with d.Sign(®)10) and the increased biocompatibility of the Co-Cr (d.Sign(®)30) alloy where nickel ions were absent.

Effect of temperature variation on the cytotoxicity of cast dental alloys and commercially pure titanium.

UNLABELLED: Cell culture system has been used to evaluate alloy cytotoxicity under different environments, testing the extracts, but the effect of temperature variation on the cytotoxicity of dental alloys has not been analyzed. OBJECTIVE: The aim of the present study was to investigate if temperature variation could affect dental alloy cytotoxicity, testing alloy extracts in an epithelial cell culture system. MATERIAL AND METHODS: Discs of Ni-Cr, Co-Cr-Mo, Ni-Cr-Ti, Ti-6Al-4V and commercially pure titanium (cp Ti) were cast by arc melting, under argon atmosphere, injected by vacuum-pressure. Discs were immersed in artificial saliva and subjected to different temperatures: 37 degrees C and thermocycling (37 degrees C/5 degrees C/37 degrees C/55 degrees C/37 degrees C). After thermocycling, extracts were put in a subconfluent culture during 6 h, and the number of cells and their viability were used to evaluate cytotoxicity in these temperatures. For each alloy, data from temperature conditions were compared by Student's t-test (alpha=0.05). RESULTS: The cytotoxicity tests with alloy/metal extracts showed that Ni-Cr, Co-Cr-Mo, Ti-6Al-4V and cp Ti extracts (p>0.05) did not affect cell number or cell viability, while Ni-Cr-Ti (p<0.05) extract decreased cell number and viability when the alloy was subjected to thermocycling. CONCLUSION: Within the limitations of the present study, the Ni-Cr-Ti alloy had cell number and viability decreased when subjected to temperature variation, while the other alloys/metal extracts did not show these results.

Effect of temperature variation on the cytotoxicity of cast dental alloys and commercially pure titanium

Cell culture system has been used to evaluate alloy cytotoxicity under different environments, testing the extracts, but the effect of temperature variation on the cytotoxicity of dental alloys has not been analyzed. OBJECTIVE: The aim of the present study was to investigate if temperature variation could affect dental alloy cytotoxicity, testing alloy extracts in an epithelial cell culture system. MATERIAL AND METHODS: Discs of Ni-Cr, Co-Cr-Mo, Ni-Cr-Ti, Ti-6Al-4V and commercially pure titanium (cp Ti) were cast by arc melting, under argon atmosphere, injected by vacuum-pressure. Discs were immersed in artificial saliva and subjected to different temperatures: 37ºC and thermocycling (37ºC/5ºC/37ºC/55ºC/37ºC). After thermocycling, extracts were put in a subconfluent culture during 6 h, and the number of cells and their viability were used to evaluate cytotoxicity in these temperatures. For each alloy, data from temperature conditions were compared by Student's t-test (α=0.05). RESULTS: The cytotoxicity tests with alloy/metal extracts showed that Ni-Cr, Co-Cr-Mo, Ti-6Al-4V and cp Ti extracts (p>0.05) did not affect cell number or cell viability, while Ni-Cr-Ti (p<0.05) extract decreased cell number and viability when the alloy was subjected to thermocycling. CONCLUSION: Within the limitations of the present study, the Ni-Cr-Ti alloy had cell number and viability decreased when subjected to temperature variation, while the other alloys/metal extracts did not show these results.

An investigation of the cytotoxic effects of dental casting alloys.

PURPOSE: This study investigated the cytotoxicity of a high-noble alloy (Bioherador N) and six commercially available base-metal alloys, including four Ni-Cr alloys (Remanium CS, Heranium NA, Wiron 99, CB Soft), one Co-Cr alloy (Wirobond C), and one Cu-based alloy (Thermobond). MATERIALS AND METHODS: Ten specimens from each alloy were prepared in the form of disks, which were placed in 24-well tissue culture plates together with a suspension containing Balb/C 3T3 fibroblasts (5 x 10(5) cells/mL). After 3 days of incubation at 37 degrees C, cell viability was determined by the MTT method. RESULTS: Variations in cytotoxicity of the alloys were observed and related to their composition. One-way ANOVA showed statistically significant differences in cytotoxicity of the alloys (P < .001). Tukey's multiple comparisons (alpha = .05) revealed that Bioherador N was significantly less cytotoxic than all the other alloys. Thermobond was the most cytotoxic, followed by CB Soft, and both of these alloys were significantly more cytotoxic than all the others. CONCLUSION: The cytotoxicity of casting alloys tested in this study was markedly affected by their composition. Differences were found in the cytotoxicity of alloys classified within the same category. The presence of Cu in the composition of the alloy adversely affected cell viability.

Cytotoxicity of dental casting alloys after conditioning in distilled water.

PURPOSE: This study investigated the cytotoxicity of various types of dental casting alloys after they had been conditioned in distilled water. MATERIALS AND METHODS: The casting alloys investigated included one high-noble alloy (Bioherador N) and six base-metal alloys, including four Ni-Cr alloys (Remanium CS, Heranium NA, Wiron 99, CB Soft), one Co-Cr alloy (Wirobond C), and one Cu-based alloy (Thermobond). Ten disks from each alloy were conditioned in distilled water at 37 degrees C for either 72 or 168 hours. The cytotoxicity of the alloys was then tested on Balb/C 3T3 fibroblasts, which were exposed to the alloys for 3 days at 37 degrees C. Cell viability was determined by the MTT method. The data were analyzed by ANOVA, and follow-up comparison between the groups was carried out using Tukey and t tests. RESULTS: ANOVA revealed a significant effect of alloy type and conditioning time (P < .001). Bioherador N was significantly less toxic than all the other alloys in the 72-hour conditioned group. After 168 hours of conditioning, its cytotoxicity was not different (P > .05) from that of Remanium CS, Wiron 99, and Wirobond C. Thermobond and CB Soft were significantly more toxic than the other alloys at both conditioning times. CONCLUSION: Conditioning of base-metal alloys, other than those containing Cu, for 168 hours in distilled water makes their cytotoxicity levels comparable to that of the high-noble alloy.

Assessment of dental material degradation product toxicity using a bioluminescent bacterial assay.

OBJECTIVES: This study examined dental material degradation product toxicity using the Microtox bacterial bioluminescence assay as well as the effects on toxicity of selective leaching, chelation with protein, the physical form of the products, and synergistic/antagonistic interactions among released ions. METHODS: Polarization was used to produce ionically dissolved (ID) and precipitated corrosion products from Litecast B alloy specimens, which were then chemically analyzed to determine their composition and to identify metal valence states. Corrosion product toxicity, as well as that of the individual alloying elements, alone and in the presence of mucin, was analyzed using Microtox. A mathematical approach identified synergistic/antagonistic interactions and determined element contribution to product toxicity. The mechanism by which the Microtox test bacterium interacts with solid products was explored. The toxicity of methyl methacrylate (MMA) monomer was also examined. RESULTS: Precipitated corrosion products were found to be more toxic than ID products. The metals in the precipitate have been shown to be available to the test bacterium. Be and Ni were the most toxic elements in the products and contributed significantly to their toxicity. Synergistic and slightly antagonistic interactions were observed in the ID and precipitated products, respectively. Mucin decreased toxicity of all elements except Be. MMA monomer toxicity was found to be low compared to metal toxicity. SIGNIFICANCE: Microtox is useful for evaluating dental degradation product biocompatibility and has significant promise for use in other types of studies, such as determining the effectiveness of antimicrobial treatments.

Effect of toothbrushing on the toxicity of casting alloys.

STATEMENT OF PROBLEM: The biological properties of casting alloys have been assessed largely under passive conditions. The effect of common intraoral stresses such as brushing, toothpastes, and low pH on alloy toxicity are not known. PURPOSE: This study assessed the toxicity of 5 types of casting alloys commonly used in prosthodontics after toothbrushing, brushing in an acidic environment, or brushing with toothpaste. These toxicities were compared with those observed without any brushing. MATERIAL AND METHODS: Au-Pt, Au-Pd, Pd-Cu-Ga, Ni-Cr-Be, and Ni-Cr (no Be) alloys were brushed for 48 hours in a toothbrushing machine at 90 strokes/minute and 200g force. Alloys were brushed with either saline at pH 7, saline at pH 4 (acidified with sodium lactate), or saline with 1:7 (wt/wt) toothpaste. After the brushing regimen, the cytotoxicity of the alloys was assessed in a standard in vitro test. Cytotoxicities of the alloys after different brushing treatments were compared with unbrushed (control) specimens. Analysis of variance (ANOVA) and Tukey multiple comparison intervals (alpha=.05) were used to identify significant differences among brushing conditions. RESULTS: Brushing at pH 7 significantly increased the toxicity of the Pd-Cu-Ga alloy (15% to 20% over unbrushed specimens). Brushing at pH 4 increased the toxicity of the Au-Pt and Au-Pd alloys by 30% and the Pd-Cu-Ga alloy by >40%. The Ni-based alloys were not affected by acid. After being brushed with toothpaste, both Ni-based alloys were significantly more toxic, but Ni-Cr-Be was the worst, increasing more than 60% in toxicity over the controls. The toxicity of the Au-Pd alloy also increased significantly (15%). CONCLUSION: Brushing dental casting alloys may increase their cytotoxicity in vitro, but the increase depends heavily on the alloy type and brushing condition.

Reaction of fibroblasts to various dental casting alloys.

The cytotoxicity of a series of dental casting alloys in the as-cast and polished condition was determined with cell culture techniques involving phase contrast microscopy to examine cell morphology and the succinic dehydrogenase histochemical reaction to measure any ring of inhibition of Balb/c 3T3 cellular respiration around alloys. Crown and bridge casting alloys and a nickel- and a cobalt-base alloy were biocompatible in the polished condition, but less so in the as-cast condition. The only two exceptions were casting alloys containing 50-60 wt% Cu. Porcelain-fused-to-metal alloys were biocompatible in either the as-cast or polished condition. This direct contact method appeared satisfactory for evaluating biocompatibility of dental casting alloys, especially since these materials are in contact with gingival tissues.

In vitro cytotoxicity of Ag-Pd-Cu-based casting alloys.

The cytotoxicity and its correlation to alloy composition, structure, corrosion, as well as galvanic coupling was studied with 12 Ag-Pd-Cu-type alloys, one conventional type III gold alloy and pure Ag, Cu, and Pd. The agar overlay cell culture technique was used. Single phase binary CuPd alloys were only slightly cytotoxic below a Cu content of 30 wt%. The tested multiphase alloys were all toxic, but no correlation between toxicity and Cu content could be observed. Solid solution annealing increased the cytotoxicity of a multiphase alloy. Exposure of a single phase alloy to an artificial saliva for 1 week prior to the test decreased its cytotoxicity significantly. Galvanic coupling of the alloys through an outer copper wire decreased their cytotoxicity.