OBJECTIVES: To compare the effects of different sintering protocols on density, translucency, microstructure, crystallography, and biaxial flexural strength of different yttria concentrations of Tosoh zirconia materials (TZ-3YSB-E, TZ-3YB-E, Zpex, and Zpex Smile). MATERIALS AND METHODS: A total of 242 zirconia discs were manufactured from commercially available Tosoh zirconia powders. Zpex smile, Zpex, TZ-3YB-E, and TZ-3YSB-E were uniaxially dry pressed into cylindrical blocks using a 5/8-inch internal diameter steel die set under a hydraulic press load of 3,000 N (Carver Press). Blocks were partially sintered at 1000 °C and sectioned into discs with a thickness of 2 mm x 15 mm by an IsoMet 5000 section machine. The discs were fully sintered using three protocols: 1) regular sintering using Vita Zyrcomat T furnace at 1520 °C for 2 h (total cycle time of 7 h); 2) fast sintering using Sirona inFire HTC SPEED at 1540 °C for 30 min (total cycle time of ~1.5 h). 3) speed sintering using Sirona CEREC Speed Fire furnace at 1579 °C for 5 min (total cycle time of ~18 min). After sintering, the translucency was characterized by contrast ratio using a spectrophotometer (X-Rite Ci7600). The specimens were then subjected to one of the following post treatments: 1) no treatment (control), 2) low temperature degradation (LDT) using 120 °C steam for 1 day (LDT 1d), 3) low temperature degradation for 1 week (LDT 1wk), 4) cyclic loading fatigue for 50,000 cycles, and 5) cyclic loading fatigue for 100,000 cycles. The microstructure of the materials was characterized using a scanning electron microscope (SEM), and the crystal grain size was measured using NIH ImageJ. A ball-on-three-balls biaxial flexural strength was performed at a crosshead speed of 0.5 mm/min using an Instron 5566A. The crystallographic phases of Tosoh zirconia were evaluated by X-ray diffraction (XRD) utilizing a D2 Phaser desktop diffractometer and the phase fraction was determined by full-profile refinements using GSAS-II software. The statistics were analyzed using JMP Pro 15.0 for comparing translucency, flexural strength, and grain size of all specimens. Statistical significance was assigned to p-values less than 0.05. RESULTS: There was a significant difference in the density of zirconia powders, with Zpex Smile powder having the lowest density of 5.81g/cm3, whereas Zpex powder had the highest density of 5.93 g/cm3. Mean grain size for speed sintering Zpex Smile was significantly larger than fast and regular sintering protocols. Grain size of TZ-3YSBE for regular sintering was significantly larger than that of speed and fast sintering. Speed sintered zirconia showed a significantly higher contrast ratio than other sintering protocols. Fast sintering groups had significantly higher contrast ratios than regular sintering for Zpex and Zpex Smile. For regular and fast sintering, TZ-3YBE had the highest contrast ratio, and Zpex Smile had the lowest value. Overall, speed sintering resulted in the highest contrast ratio and regular sintering the lowest ratio. There was a statistically significant effect on crystal phases resulting from the zirconia powders and post sintering treatments. The results also indicated that Zpex Smile had a combination of cubic and tetragonal phases with a higher percentage of the cubic phase than other zirconia types. Speed sintered zirconia had significantly lower biaxial flexural strength values. However, there was no significant difference in biaxial flexural strength of TZ-3YB-E for different sintering protocols and post treatments. CONCLUSION: The sintering protocols significantly impacted zirconia grain size, crystallography, and biaxial flexural strength. Speed sintering provided lower translucency and flexural strength.

This study was designed to evaluate the cell attachment efficiency, cell proliferation, cytotoxicity and genotoxicity of different sizes and doses of silica on human dental pulp cell culture. Human dental pulp cells (HDPCs) derived from extracted third molars were cultured in growth media and supplemented with two sizes of silica particles (70 nm – 225 nm) and (1 µm – 5 µm) in different doses: (12.5 µg/mL, 50 µg/mL, 100 µg/mL and 1000 µg/mL) and media without silica particles as a control, for the time intervals of 16h, 24h, 7 and 14 days. Attachment efficiency and cell proliferation were measured by comparing optical density of crystal violet stained cells. Cytotoxicity was measured using water-soluble dye that changes its absorption spectra upon cellular reduction. Genotoxicity was measured using specific antibody-based detection. Cultures with Si-np50, Si-np100, Si--mp12.5, Si-mp50, Si-mp100 at 16h yielded significantly higher attachment efficiency compared to the other doses. Cultures with Si-np50 and Si-np100 showed increased attachment efficiency compared to cultures with their larger microparticles counterparts(P<0.05). Cultures with Si-mp12.5 displayed similar proliferation compared to the control at day 14. Higher doses showed significant reduction in proliferation compared to lower doses in all sizes of Si-np and Si-mp at day 14. All the silica nanoparticles groups showed lower proliferation when compared to their larger microparticles counterparts at 14 days (P<0.05). When comparing different doses in the cytotoxicity assay all the groups displayed cytotoxicity by day 14, except the Si-mp12.5 group. And when comparing different sizes, all the doses of Si-np showed higher levels of cytotoxicity when compared to Si-mp at day 14 (P<0.05). Cultures with 1000 µg/mL group displayed significantly higher genotoxicity in both sizes when compared to lower doses at day 1. When comparing both sizes to each other, nanoparticles showed a higher genotoxicity than microparticles (p < 0.05). In conclusion, the higher doses of silica nanoparticles significantly decreased proliferation and increased cytotoxicity and genotoxicity on normal HDPCs in a dose and size dependent manner. This is the first report to demonstrate the effect of dose and size of silica particles on the proliferation, cytotoxicity and genotoxicity of normal HDPCs.

The Regional Acceleratory Phenomenon (RAP), a post-injury transient bone remodeling phenomenon, is the foundation of most modern corticotomy-assisted orthodontics treatment. The piezoelectric knife (PIEZO) is an alternative to mechanical devices in conventional oral surgery procedures by rotary bur (BUR). It enables cutting bone with low ultrasonic frequency while protecting fragile anatomical structures. PIEZO may enhance RAP leading to cellular activities while BUR causes severe trauma in the medullary tissue resulting in excessive inflammation. Our previous PIEZO study demonstrated that the trans-cortical penetration (TCP) significantly activated biological responses by RAP more than the intra-cortical defect (ICD). PIEZO-TCP increases initial osteocyte apoptosis, osteoclast, and osteogenic activities. The cortical alveolar bone is a crucial structural element to support teeth or implant stability. Yet, there is no reliable metric for predicting the mechanical properties of the bone in this critical region. Recent studies suggest that micro-porosity assessed by deep learning from micro-CT images correlates with cortical bone’s elastic modulus and ultimate compressive strength by tissue mineral density. We hypothesized that cortical porosity might increase in the resorption phase and decrease in the formation phase associated with RAP. In this study, we used deep learning analysis to compare cortical micro-porosity from post-operative micro-CT images of PIEZO, BUR, and Control at day 7 and 14. Eighteen 9-week-old male Sprague-Dawley rats were randomly divided into three groups: PIEZO, BUR and Control with deep and shallow defects on the right and left tibias in test groups. 3D rendered micro-CT images, with approximately 1000 slices each, were analyzed for cortical micro-porosity with deep learning algorithms for multi-label segmentation. The deep learning model was trained to analyze the image and classify the pixels in one of these labels: background, cortical bone, reactive calcified tissue and cortical porosity. Cortical porosity was considered to be all void in the cortical bone, except for the notch or canal of the tibial nutrient artery. Regional cortical porosity was assessed using the full scan volume (approximately 6mm long). Local cortical porosity was measured and compared for 3 standardized local regions of interest (ROIs, 0.5, 1.0, 1.5mm from the defect edge). Results indicate that Piezo corticotomies have a significantly deeper impact on the RAP versus conventional rotary burs and that the deep learning process, a subset of machine learning that makes the computation of multi-layer neural networks, could be a very powerful new tool and an innovative approach to dental research.

INTRODUCTION: Lysine-specific demethylase 1 (LSD1) plays a crucial role in controlling cell homeostasis in health and disease. LSD1 is elevated in oral cancer and promotes metastasis and correlates with poor prognosis. The objective of this study is to determine the mechanism by which LSD1 plays a critical role in suppressing oral squamous cell carcinoma (OSCC) development. METHODS: The effect of knocking down LSD1 and the use of LSD1 inhibitor (SP2509) was investigated using 4-NQO mouse models in vivo. The mice were sacrificed, and their tongues were subjected to real-time PCR analysis, hematoxylin and eosin staining, pathological analysis and immunohistochemistry staining for different markers. The effect of SP2509 and different inhibitors on human squamous cell carcinoma (HSC3) was investigated. Cell proliferation assays were used to evaluate the effect of SP2509 alone and in combination with other inhibitors on HSC3 cells. RESULTS: The data showed that LSD1 knockout mice showed inhibition of tongue OSCC, downregulation of oncogenic genes. Immunostaining analysis confirmed the downregulation of PD-L1, YAP and other oncogenic factors in LSD1 knockout mice. CONCLUSION: Genetic deletion of LSD1 in Keratin14-Cre-Lsd1floxed mice provided a novel model to study epigenetic attenuation of HNSCC and provided a proof-of-concept that Lsd1 could be a specific target during tongue OSCC. Next, the pharmacological attenuation of Lsd1 with specific small molecule LSD1 inhibitor, SP,2509, inhibited pathological changes and provided evidence that SP2509 could be used for translational studies. Next, SP2509 was shown to be useful for immunotherapy and combination therapy applications. Overall, this study identified that LSD1 is a key regulator of OSCC. In addition, targeting LSD1 with SP2509 has an implication for immunotherapy, combination therapy, and future translational studies.

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OBJECTIVES: The objective of this study is to assess the effects of different surface treatments on flexural strength, surface roughness, and crystal structure of zirconia with different yttria concentrations on YZ ceramics are to be evaluated. Moreover, the effects yttria stabilization on the mechanical properties of four variants of highly translucent yttria-stabilized zirconia (VITA YZ ® Zahnfabrik) and porcelain-fused YZ. MATERIALS AND METHODS: Four different monolithic zirconia ceramics: VITA-Zahnfabrik VITA-YZ T, VITA-YZ HT, VITA-YZ ST, VITA-YZ XT, as well as porcelain-fused zirconia (PFZ) were used in this study. Disc specimens were prepared by core-drilling and sectioning the bulk discs, which were sintered according to the manufacturer’s recommendations. These sintered zirconia specimens were subsequently subjected to surface treatments with diamond discs of various fineness parameters such as 1μm, 50μm, 125μm, and without surface treatment as a control group. Surface roughness was evaluated at three different locations of five polished specimens from each group of the five different materials by using a contact profilometer Mitutoyo Surftest SJ-201 with a 2μm diamond stylus. A Bruker D2 Phaser diffractometer was used for X-ray diffraction (XRD) measurements. Crystallographic phase identification and analysis were performed through Rietveld refinements on XRD raw data using GSAS-II software implemented by Python. An Instron universal testing machine was used to test biaxial flexural strength of the materials, in which a ball-on-three-balls configuration was set up to perform biaxial flexural strength testing with a speed of 1 mm/min and a load cell capacity of 10 kN. The microstructures of materials were analyzed by using a SU6600 Hitachi field emission scanning electron microscope (FESEM). Descriptive statistics such as surface roughness, phase contents, grain size, and flexural strength were indicated as mean and standard deviation (SD). Statistical analysis was performed using multi-way ANOVA and linear regression model followed by the post hoc Tukey test. RESULTS: XRD analyses for all control, as well as treated specimens, indicate the presence of multiple phases of zirconia in each specimen, which include tetragonal, cubic, monoclinic, and rhombohedral crystalline phases. Some of the control samples have no detectable monoclinic phase of zirconia while all surface treated specimens have various amounts of monoclinic phases and rhombohedral phases. The presence of these phases has affected the flexural strength as well as the crack toughening characteristics. The YZ T samples have the highest flexural strength while YZ XT have the lowest flexural strength. The sample surface roughness depends on the type of treatment. The surface treatment affects each group of specimens in a different way depending upon its various pristine crystal phases. The surface treatment with different fineness parameters such as 1μm, 50μm, and 125μm was found to induce the stress-induced phase transformation in the surface layers of the specimens, which includes tetragonal to monoclinic phase transformation. This phase transformation seriously affects the flexural strength and crack propagation of zirconia due to the transformation toughening mechanism. The results indicated that grain size was not a crucial factor influencing either the transformation toughening mechanism or the tetragonal phase content in the control groups in each monolithic material system. In case of the porcelain fused zirconia (PFZ), Several factors can affect the flexural strength of the veneering materials. CONCLUSIONS: The higher yttria content over 4 mol% decreases the flexural strength of the ceramics studied. Moreover, the surface polishing treatments seem to enhance the flexural strength in all cases. The biaxial flexural strength of porcelain-fused zirconia (PFZ) was significantly lower than monolithic zirconia (YZ)

OBJECTIVE: Determine the impact of clinically used anti-resorption drug alendronate (ALN) on bone cells and the capacity of bone to regenerate, and to provide insights into the interplay between cancer stem cells and osteocytes/osteoblasts and mesenchymal stem cells using a three-dimensional (3D) live cancer-bone interactive model. METHODS: Co-cultures of live mouse neonatal calvarial bone and cancer cells, PC3 prostate and breast MDA-MB-231, in a roller tube model system in the absence and presence of alendronate (ALN) and osteoprotgrin (OPG). These models were used under conditions whereby the two naturally occurring bone remodeling stages were dissociated, viz., bone resorption and formation. The used media and calvarial bones were evaluated by chemical, biochemical, histological and quantitative histomorphometric analyses. RESULTS: These studies revealed that cancer cells are unaffected to BP treatment in the presence of bone and colonize live bone irrespective to the bone remodeling stage, hence, cancer-bone metastasis/interactions are though to be ‘‘in- dependent of bone remodeling stages’’. Under resorption conditions cancer cells induce differentiation of osteoclasts and bone resorption and inclusion of alendronate (ALN) inhibited cancer-induced bone resorption at the osteoclast differentiation level. However, alendronate treated bones were adversely impacted as demonstrated by their inability to respond to stimulation for new bone formation. These data indicated that the bone stem/progenitor cells appeared non-vital after alendronate exposure. Similarly, in the formation model system the new bone formation was also limited. Mass spectrometric analysis of the media from cancer-bone organ cultures in the absence and presence of ALN was consistent with the organ culture results. The mineral-bound ALN impacts the bone organs by limiting transformation of mesenchymal stem cells to osteoblasts and leads to diminish endosteal cell population and degenerated osteocytes within the mineralized bone matrix.