Biosynthesis of vitamin B3 and NAD+: incubating HepG2 cells with the alkaloid myosmine.

BACKGROUND: In the kynurenine pathway, it is reported that the essential amino acid tryptophan forms nicotinic acid (NA, vitamin B3) in biological systems. This pathway is part of the de novo pathway to perform nicotinamide adenine dinucleotide (NAD+) biosynthesis. Additionally, biosynthesis of NAD+ via the Preiss-Handler pathway involves NA and its analogue nicotinamide, both designated as niacin. Previous attempts were successful in converting myosmine (MYO) by organic synthesis to NA, and the assumption was that the alkaloid MYO, which is taken in from food, can be converted into NA by biological oxidation. RESULT: Incubation of HepG2 cells with MYO yielded NA. Moreover, a significant increase of NAD+ compared with the control has been found. CONCLUSION: Hence, MYO could be assumed to be the hitherto unknown origin of an alternative NA biosynthesis additionally influencing NAD+ biosynthesis positively. This novel MYO pathway may open new perspectives to improve knowledge and relevance of NA and NAD+ biosynthesis and bioactivation in cells and, moreover, in food staples, food, and diet. © 2024 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Salivary and gingival CXCL8 correlation with periodontal status, periodontal pathogens, and smoking.

OBJECTIVES: Neutrophil granulocytes have been proposed to play a major role in the mediation of periodontitis-associated tissue destruction. Their recruitment and activation are regulated by the chemokine CXCL8. This study aimed to delineate the dependency of CXCL8 expression in gingival crevicular fluid (GCF) and saliva on periodontal status, bacterial infection, and smoking, in patients with periodontitis. METHODS: The study cohort comprised 279 subjects with untreated periodontitis. Probing pocket depth (PPD), gingival recession, bleeding on probing (BOP), plaque index, and bone loss were evaluated. CXCL8 was determined in saliva and GCF using flow cytometry. RESULTS: Considering the entire study sample, CXCL8 levels were correlated with the mean PPD (ρ = 0.131; p = 0.029), severity of periodontitis (ρ = 0.121; p = 0.043), BOP (ρ = 0.204; p = 0.001), and smoking (ρ = -0.219; p < 0.0001) in GCF; and, in whole saliva, with mean PPD (ρ = 0.154; p = 0.010) severity of periodontitis (ρ = 0.140; p = 0.020), gender (ρ = 0.178; p = 0.003), and smoking (ρ = -0.156; p = 0.010). Subgroup analysis among non-smokers revealed significantly higher amounts of CXCL8 in GCF (p = 0.012) and saliva (p = 0.026) comparing subjects with mean PPD ≤3mm and >3mm. CONCLUSION: The current study revealed a strong dependency of CXCL8 expression in GCF on the severity and activity of periodontal disease. Smoking causes a significant reduction in CXCL8 expression in saliva and GCF.

Contact allergy to the Dexcom G6 glucose monitoring system-Role of 2,2'-methylenebis(6-tert-butyl-4-methylphenol) monoacrylate in the new adhesive.

BACKGROUND: Skin reactions to the glucose monitoring systems Dexcom G5 and G6 have been rare. In 2019, the components of the adhesive were exchanged for better skin fixation. Since then, more and more patients experienced severe skin reactions. A few months ago, 2,2'-methylenebis(6-tert-butyl-4-methylphenol) monoacrylate (MBPA) was identified as a new component in the adhesive of the G6 model. Furthermore, it was suspected that isobornyl acrylate (IBOA) was also a component of the exchanged adhesive. OBJECTIVES: Our objective was to investigate if MBPA plays a major role in the increasing skin problems of patients without a history of IBOA-sensitization. Furthermore, our aim was to examine whether IBOA is contained in the newer model adhesive and may also contribute to allergic contact dermatitis (ACD). PATIENTS AND METHODS: Five patients with a newly occurred ACD caused by the glucose monitoring system Dexcom G6 were investigated. Patch testing including MBPA in three different concentrations, as well as IBOA, were performed. Gas chromatography-mass spectrometry of the newer system Dexcom G6 was carried out. RESULTS: All patients were shown to be sensitized to MBPA, while MBPA 0,5% showed the strongest reaction. On the other hand, IBOA was tested negative. CONCLUSION: In our study group, MBPA was observed to be the triggering allergen of the recently changed adhesive.

Filtration efficiency of surgical and FFP3 masks against composite dust.

The aim of this study was to investigate the protection efficiency of two types of face masks against composite dust and to characterize the particles that penetrated through the masks. Composite dust was created by grinding a commercial nano-filled composite in a plexiglass box without using water cooling or high vacuum evacuation, in order to obtain a worst-case exposure. Dust particles were collected using a personal inhalable aerosol sampler (IOM) fixed inside a custom-made phantom head. Surgical and filtering facepiece (FFP3) masks were tested, and the situation without a mask served as control. The IOM sampler contained a cassette with two filters to collect large inhalable (4-100 µm) and respirable dust particles (<4 µm). The amount of particles was determined gravimetrically by weighing filters before and after composite grinding, and further characterized by electron microscopy. Particle collection for both inhalable and respirable dust was the highest when no mask was used, and the lowest with the use of a FFP3 mask. Different sizes and shapes of particles were observed, with the largest particles (>1 µm) being seen when no mask was applied, whereas only nanoparticles could be detected when either type of face mask was applied. Even though FFP3 masks showed a higher filtration efficacy than surgical masks of the inhalable dust fraction, penetration of a small respirable particle fraction was inevitable for both masks.

Actovegin® reduces PMA-induced inflammation on human cells.

PURPOSE: The effect of Actovegin® was investigated on PMA- and LPS-induced human peripheral blood mononuclear cells (PBMCs). METHODS: PBMCs (1 × 106 cells/ml) from five blood donors (2 f, 3 m; 45-55 years) were grown in medium and exposed to Actovegin® in the presence or absence of PMA or LPS. Supernatants were collected to assess the concentration of cytokines (TNF-α, IL-1beta, IL-6 and IL-10). The reactive oxygen species (ROS) were assessed by a ROS-GloTM H2O2 assay. RESULTS: Stimulation of cells by PMA or LPS (without Actovegin®) significantly increased the secretion of IL-1beta, IL-6, IL-10 and TNF-α from PBMCs, compared to controls. Pre-treatment of cells with Actovegin® (1, 5, 25, 125 µg/ml) plus PMA significantly decreased the secretion of IL-1beta from PBMCs, compared to controls (PMA without Actovegin®). In contrast, addition of Actovegin® (1, 5, 25, 125 and 250 µg/ml) plus LPS did not alter the IL-1beta production, compared to controls (LPS without Actovegin®). TNF-α, IL-6 and IL-10 do not contribute to the reduction of inflammatory reactions with Actovegin®. CONCLUSIONS: Actovegin® can reduce the PMA-induced IL-1beta release and the ROS production from PBMCs. These findings may help to explain the clinically known positive effects of Actovegin® on athletic injuries with inflammatory responses (e.g., muscle injuries, tendinopathies).

The implanted glucose monitoring system Eversense: An alternative for diabetes patients with isobornyl acrylate allergy.

BACKGROUND: Some patients with diabetes develop skin reactions when using systems for continuous glucose monitoring (CGM) or insulin pumps. Regular usage and long wearing periods lead not only to skin irritation, but also to allergic contact dermatitis. It has been shown that allergens such as isobornyl acrylate (IBOA) are present in the plastic housing and also in the adhesives of medical devices used for diabetes treatment. OBJECTIVES: To evaluate the IBOA content of all parts of a newly introduced, implanted CGM system (Eversense) to check whether this can be an alternative for IBOA-sensitized patients. METHODS: The IBOA content of the implanted sensor itself (n = 3), the transmitter (n = 3), and two different types of adhesive (white adhesive [n = 4] and clear adhesive [n = 4]) was measured by gas chromatography/mass spectrometry. RESULTS: No IBOA was found in any part of this CGM system. CONCLUSIONS: Patients with an IBOA allergy may be able to use this implanted CGM system.

The Dexcom glucose monitoring system-An isobornyl acrylate-free alternative for diabetic patients.

BACKGROUND: Glucose monitoring systems, for example, Freestyle Libre (Abott) and Dexcom (Nintamed), are increasingly being used instead of conventional blood sugar measurement. However, many patients have experienced adverse skin reactions such as severe allergic contact dermatitis (ACD). Finally, in August 2017, the culprit allergen in Freestyle Libre, isobornyl acrylate (IBOA), was identified. OBJECTIVES: After patients have developed ACD, it is recommended that they no longer use their glucose monitoring systems. Thus, it is important to find an alternative IBOA-free device. PATIENTS AND METHODS: Five patients presented with ACD caused by Freestyle Libre. Each was patch tested with allergens from the baseline series and from a plastics and glues series, and additionally with IBOA 0.1% pet. Gas chromatography-mass spectrometry (GC/MS) of the Freestyle Libre sensor and the Dexcom sensor was performed. The Dexcom sensor remained on the skin of all patients for at least 2 days. RESULTS: All patients were sensitized to IBOA. GC/MS showed the presence of IBOA in the Freestyle Libre sensor, whereas the Dexcom sensor was IBOA-free. None of the patients had skin reactions to the Dexcom sensor. CONCLUSIONS: Patients with Freestyle Libre and IBOA allergy may use the Dexcom sensor as an alternative for glucose monitoring.

Comprehensive Analytics of Actovegin® and Its Effect on Muscle Cells.

The ingredients of Actovegin® were analyzed and its effects on the muscle cell proliferation were investigated. C2C12 myoblasts were cultured in medium. Actovegin® was added in five different concentrations (1, 5, 25, 125, and 250 µg) to the differentiation medium. The formations of proliferation factor Ki67 and myosin heavy chains were measured by immunofluorescence. The first primary antibody was anti-Ki67 and anti-Mf20. Cells were washed and treated with the second fluorochrome. Thirty-one Actovegin® ingredients were found to contain significantly higher concentrations and twenty-nine ingredients were found to contain significantly lower concentrations, compared to the mean ranges as described in the literature for the normal physiological concentrations in human adult serum/plasma. A significant increase in the formation of Ki67 was observed in Actovegin® groups, compared to controls. The mean area of myotubes was significantly increased in Actovegin® groups. A significant decrease in the number of myotubes was observed. An increased myotube size (fusion) was observed. The intensity of Mf20 was significantly increased in Actovegin® groups. It could be demonstrated that Actovegin® contains many physiological substances in significantly higher and some in lower concentrations compared to human adult serum. Furthermore, it could be shown that Actovegin® improves muscle cell proliferation.

Effect of various light curing times on the elution of composite components.

OBJECTIVES: Polymerization of resin-based composites (RBCs) is incomplete. The aim of the present study was to determine whether a longer curing time than recommended by the manufacturer influences the amount of released composite components of RBCs. MATERIALS AND METHODS: The composites Clearfil AP-X and els extra low shrinkage were polymerized for six different curing times: 4, 10, 20, 40, 100, and 200 s. Light curing time recommended by the manufacturer for both composites is 20 s. Subsequently, samples were eluted in methanol and water for 1, 3, and 7 days and analyzed by gas chromatography/mass spectrometry (GC/MS). RESULTS: For Clearfil AP-X ethylene glycol dimethacrylate (EGDMA), diethylene glycol dimethacrylate (DEGDMA), triethylene glycol dimethacrylate (TEGDMA), 2-hydroxy-4-methoxybenzophenone (HMBP), camphorquinone (CQ) and 2,6-di-tert-butyl-4-methylphenol (BHT) were detected in methanol. In the aqueous eluate, only TEGDMA was detected. In els extra low shrinkage, HMBP, BHT, and CQ were detected in methanol. CONCLUSION: Increasing the curing time compared to recommendation of the manufacturer reduces the release of most composite components. This could result in less exposure to human due to these substances. CLINICAL RELEVANCE: Methacrylates are classified as potential allergens. An increasing number of dentists and patients show allergic reaction to methacrylates. Therefore, a reduced elution of composite components is an advantage.

Leaching components and initial biocompatibility of novel bioactive restorative materials.

OBJECTIVES: Bioactive restorative materials were developed on the premise that direct restorations should not only serve the purpose of reconstructing dental hard tissue defects but also exhibit biological features that prevent secondary caries development, without having adverse effects on the host cells. This study focuses on assessing the in vitro biocompatibility of two novel bioactive restorative materials. METHODS: Specimens of the bioactive restorative materials, Cention Forte (CF) and ACTIVA BioACTIVE RESTORATIVE (AB), a glass ionomer cement/glass hybrid (EQUIA Forte HT, EF) and an established nanohybrid composite (Venus Diamond, VD) were produced and finished. The specimens were eluted in water and methanol and the resulting eluates were analyzed via gas chromatography-mass spectrometry. hGF-1 cells were exposed to eluates prepared in cell culture medium. Cellular ATP levels, oxidized glutathione concentration, caspase-3/7 activity and the inflammatory response (IL-6 and PGE2 levels) were determined. Microscopic images were taken to examine the cell morphology. RESULTS: Methyl methacrylate and 2-Hydroxyethyl methacrylate were the main monomers detected in CF and AB eluates. All materials inhibited cell proliferation and led to significantly reduced ATP-levels. The cells exhibited a healthy morphology in the presence of CF and AB. Cells exposed to VD showed increased oxidized glutathione levels. Only EF led to enhanced caspase-3/7 activity. CF and AB caused IL-6 levels to increase, while EF and AB led to enhanced PGE2 levels. SIGNIFICANCE: CF and AB are promising materials from a biological point of view and seem to have improved bioactive properties compared to glass ionomer cements.

Intranuclear cell uptake and toxicity of titanium dioxide and zirconia particles as well as bacterial adhesion on dental titanium- and zirconia-implants.

OBJECTIVE: Previous studies have shown that particles can be released from dental titanium (Ti)- and zirconia (ZrO2)-implants. Titanium dioxide (TiO2)- and ZrO2-particles were compared regarding their toxicity and intranuclear cell uptake as well as the adhesion of various anaerobic bacteria on Ti- and ZrO2-implants. METHODS: Cyto- and genotoxicity of TiO2-microparticles (TiO2-MPs) and TiO2-nanoparticles (TiO2-NPs) in periodontal ligament (PDL)-hTERT cells were determined with XTT test and DNA damage with comet assay. Particle sizes of TiO2- and ZrO2-particles were measured with scanning electron microscope. Intranuclear uptake in PDL-hTERT cells was determined with laser scanning confocal microscopy. Adhesions of relevant anaerobic mouth bacteria Porphyromonas gingivalis, Prevotella intermedia and Aggregatibacter actinomycetemcomitans on Ti- and ZrO2-implants were investigated by cultivation and counting bacterial colonies. RESULTS: Particle size measurements revealed that 99% of the TiO2-NPs had a size below 100 nm and 88% of the TiO2-MPs sizes were between 50 and 200 nm. Following EC50 values were found for particles (mg/l): 92 (TiO2-MPs) and 15 (TiO2-NPs). A significant increase in olive tail moment (OTM) was found for TiO2-NPs at a concentration of 1/10 EC50. TiO2- and ZrO2-NPs had a higher intranuclear cell uptake efficiency, compared to corresponding TiO2- and ZrO2-MPs. All investigated particles could be detected in cell nucleus. Adhesion of all investigated bacterial species was significantly higher on Ti-implants, compared to ZrO2-implants. CONCLUSION: Ti usually develops an oxide layer (TiO2). Particles released from Ti-implants should be TiO2-particles or Ti-particles coated with a TiO2-layer. Toxicity of released Ti-particles depends on their oxidation state and on their size (NP or MP). Particularly, NPs were more cyto- and genotoxic compared to the corresponding MPs. TiO2- and ZrO2-NPs showed a significant increase in the intranuclear cell uptake ratio at higher exposure concentration, compared to lower concentrations and consequently might lead to a higher potential of DNA damage. Adhesion of bacteria to ZrO2-implants is reduced, compared to Ti-implants. Therefore, ZrO2-implants might contribute to reduced biological complications (e.g. periimplantitis).

Disrupted biomineralization in zebra mussels after exposure to bisphenol-A: Potential implications for molar-incisor hypomineralization.

OBJECTIVE: The aetiology of molar-incisor hypomineralization (MIH) is currently unclear. A major hurdle in MIH research is the lack of adequate model systems. The study investigated the feasibility of zebra mussel (Dreissena polymorpha) as a novel model to screen potential MIH-related factors. METHODS: In four experiments with overall 46 groups (n = 7 mussels/group), six groups per experiment were incubated with 100 mg/l calcein (mineralization marker) solution for 96 h to evaluate the dynamics of shell biomineralization, another six groups with tap water only (controls). Then zebra mussels with and without calcein pre-incubation were exposed to cadmium sulfate hydrate (3CdSO4•8H2O) (positive control; 0, 0.01, 0.1, 1, 10 and 100 mg/l), possible aetiological factors of MIH including bisphenol-A (BPA; 0, 0.02, 0.2, 2, 20 and 200 mg/l) and erythromycin (0, 0.1, 1, 10, 100 and 1000 mg/l) as mineralization "disruptors", and doxycycline (0, 0.1, 1, 10, 100 and 1000 mg/l) for 96 h, respectively. After two weeks, the mussels were sacrificed and shells were embedded in methylmethacrylate for fluorescence intensity analysis. RESULTS: Mortality rate was 100% after 20, 200 mg/l BPA and 10, 100 mg/l 3CdSO4•8H2O exposure. Thereby, the median lethal concentration (96 h-LC50) of BPA was 6.3 mg/l (95% CI, 1.3-34.4 mg/l), and that of cadmium was 3.1 mg/l (95% CI, 0.7-10.5 mg/l). Notably, calcein fluorescence in shells significantly decreased (p < 0.05) after 2 mg/l BPA and 1 mg/l 3CdSO4•8H2O exposure. SIGNIFICANCE: These findings suggest that BPA may disrupt biomineralization. Biomineralization in zebra mussels seems to be an effective model for investigating potential MIH-related factors.

Elution behavior of a 3D-printed, milled and conventional resin-based occlusal splint material.

OBJECTIVE: The elution of unpolymerized (co-)monomers and additives from methacrylic resin-based materials like polymethyl methacrylate (PMMA) can cause adverse side effects, such as mutagenicity, teratogenicity, genotoxicity, cytotoxicity and estrogenic activity. The aim of this study was to quantify the release and the cytotoxicity of residual (co-)monomers and additives from PMMA-based splint materials under consideration of real splint sizes. Three different materials used for additive (3D printing), subtractive (milling) and conventional (powder and liquid) manufacturing were examined. METHODS: The splint materials SHERAprint-ortho plus (additive), SHERAeco-disc PM20 (subtractive) and SHERAORTHOMER (conventional) were analysed. 16 (n = 4) sample discs of each material (6 mm diameter and 2 mm height) were polished on the circular and one cross-section area and then eluted in both distilled water and methanol. The discs were incubated at 37 °C for 24 h or 72 h and subsequently analysed by gas chromatography/mass spectrometry (GC/MS) for specifying and quantifying released compounds. XTT-based cell viability assays with human gingival fibroblasts (HGFs) were performed for Tetrahydrofurfuryl methacrylate (THFMA), 1,4-Butylene glycol dimethacrylate (BDDMA) and Tripropylenglycol diacrylate (TPGDA). In order to project the disc size to actual splint sizes in a worst-case scenario, lower and upper jaw occlusal splints were designed and volumes and surfaces were measured. RESULTS: For SHERAeco-disc PM20 and for SHERAORTHOMER no elution was determined in water. SHERAprint-ortho plus eluted the highest THFMA concentration of 7.47 µmol/l ±2,77 µmol/l after 72 h in water. Six (co-)monomers and five additives were detected in the methanol eluates of all three materials tested. The XTT-based cell viability assays resulted in a EC50 of 3006 ± 408 µmol/l for THFMA, 2569.5 ± 308 µmol/l for BDDMA and 596.7 ± 88 µmol/l for TPGDA. SIGNIFICANCE: With the solvent methanol, released components from the investigated splint materials exceeded cytotoxic concentrations in HGFs calculated for a worst-case scenario in splint size. In the water eluates only the methacrylate THFMA could be determined from SHERAprint-ortho plus in concentrations below cytotoxic levels in HGFs.

Monomer release from surgical guide resins manufactured with different 3D printing devices.

OBJECTIVES: The aim of this study was to assess the post curing monomer release of resins applicable for 3D printing of surgical implant guides in dependency of printing technique and storing media using high performance liquidchromatography. MATERIAL AND METHODS: Specimens of Nextdent SG, Freeprint Splint, Fotodent Guide, 3Delta Guide, and V-print SG (n = 4) were additively manufactured with the corresponding DLP/SLA printing devices (Rapidshape D20II, Form2, Solflex350). Postprocessing was done according to the manufacturer's specifications. Subsequently, samples were eluted in methanol and water for 3 days and analyzed with gas chromatography/mass spectrometry (GC/MS). RESULTS: A total of twelve different substances released from the tested resin materials. The highest eluted concentration for MMA in methanol was 20.27 ± 8.60 µg/mL followed by 12.66 ± 3.38 µg/mL of HPMA. HEMA was found at concentration of 11.17 ± 2.43 µg/mL in methanol and 1.15 ± 0.11 µg/mL in water. TPGDA and TEGDMA reached maximum concentration in methanol of 4.29 ± 0.54 µg/mL and 5.07 ± 0.93 µg/mL and in water of 0.79 ± 0.19 µg/mL and 0.36 ± 0.14 µg/mL, respectively. Significant difference was found for the material Nextdent SG manufactured on SLA and DLP printing device for THFMA (p = 0.041), TEGDMA (p = 0.026), TPGDA (p = 0.05) and EGDMA (p = 0.06). The amount of monomers released into water did not reach the detection threshold for V-print SG. SIGNIFICANCE: The study revealed significant influence of the printing technique and resin material on the elution of monomers. The elution in methanol and water was significantly different. While the relative amount of eluted monomers from 3D printed guides is comparable to conventional direct composites and below toxic relevant concentrations, the absolute amount of monomer can rise in a clinic situation due to the size of the guides.

Titanium and zirconium release from titanium- and zirconia implants in mini pig maxillae and their toxicity in vitro.

OBJECTIVE: Titanium (Ti)- and Zirconia (ZrO2)-implants in mini pig maxillae were compared with respect to Ti/zirconium (Zr) release into the surrounding bone tissues, the resulting short term tissue responses and the potential toxicity. METHODS: Ti/Zr release from Ti- and ZrO2-implants in mini pig maxillae was determined with inductively coupled plasma optical emission spectrometry (ICP-OES) and inductively coupled plasma mass spectrometry (ICP-MS). The spatial distribution of Ti and Zr in maxilla tissues near the implant surface was assessed with laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS). A histological analysis was performed to investigate the tissue responses after 12 weeks of implantation. The cytotoxicity and DNA damage of Ti particles and ZrO2 particles were studied with XTT and Comet assay. RESULTS: The mean Ti content in the bone adjacent to Ti-implants was 1.67 mg/kg-bone weight. The highest Ti content detected was 2.17 mg/kg-bone weight. The mean Zr content in the bone adjected to ZrO2-implants was 0.59 mg/kg-bone weight. The highest Zr content was 0.75 mg/kg-bone weight. The spatial distribution of the Ti and Zr in bone showed mainly a higher intensity of Ti and Zr close to the screw thread outer tip rather. Histological analysis indicated that near both implant-types signs of bone marrow fibrosis were present. EC50 of commercially available ZrO2-nanoparticles (NPs, <100 nm) and ZrO2-microparticles (MPs, <5 µm) was 13.96 mg/ml and 80.99 mg/ml, respectively. ZrO2-NPs and ZrO2-MPs can induce DNA damage at 70 µg/ml and 810 µg/ml, respectively. SIGNIFICANCE: After 12-weeks of implantation, increased concentrations of Ti and Zr can be detected in bone/tissues near Ti- and ZrO2-implants in mini pig maxillae. Ti content released from Ti-implants is two times higher than the Zr content released from ZrO2-implants. ZrO2-NPs showed lower cytotoxicity and DNA damage compared to results reported for Ti-NPs in human cells.

Effect of eluates from zirconia-modified glass ionomer cements on DNA double-stranded breaks in human gingival fibroblast cells.

OBJECTIVE: To formulate novel glass ionomer cements (GICs) containing zirconia (nanoparticles (NPs) and micro-particles (MPs)) and investigate the genotoxic effect of their eluates on DNA double-strand breaks of human gingival fibroblasts (HGFs) in vitro using a γ-H2AX fluorescent assay. METHODS: GIC (control, C), 10%ZrO2NPsGIC (T1) and 10%ZrO2MPsGIC (T2) were prepared per the manufacturer's instructions (hand-mixed, P/L=3.4:1w/w%). Dulbecco's modified Eagle's medium (DMEM) was used as the culture medium for HGFs and for eluate preparation. Eluates were collected from all specimens (n=5/g, 5×2mm) after 24h and used for XTT to obtain the EC50 using Graph Pad Prism4. A γ-H2AX immunofluorescence assay was performed to detect DSBs in HGFs. The mean foci per cells and percentage of free foci cells were statistically compared (one-way ANOVA with Tamhane's post hoc and Chi-square respectively) (p<0.05). RESULTS: (1) EC50 ranged from 31 to 36%. 5% and 20% eluate concentrations were selected for the genotoxicity test. (2) Cells exposed to eluates from T1 had lower mean foci per cell than cells in T2 and C eluates (p<0.05). Only cells in T1 at 5% had lower mean foci cell than medium (p<0.05). (3) T1 and C at both concentration showed a higher, but not significant, percentage of free foci cells than negative control (medium). At 20% eluate concentration T2 had a lower percentage of free foci cells than C (p<0.05). SIGNIFICANCE: Nano-zirconia GIC and micro-zirconia GIC were formulated. GIC and both zirconia modified GICs had no genotoxic effect on HGFs in vitro. Further studies related to physical properties should be performed to determine the future clinical applications for these novel nanomaterials.