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Objective: The aim of this study was to evaluate the effect of an adhesive loaded with 0.2 % copper (Cu) and 5 % zinc oxide (ZnO) nanoparticles (Nps) on its adhesive properties and enzymatic activity at the hybrid layer ex vivo in a randomized clinical model., Methods: Fifteen patients participated in this study, and a total of 30 third molars were used. Occlusal cavities (4 × 4 × 2 mm) were made in each tooth, and randomly divided into 2 groups: (i) Experimental group: commercial adhesive loaded with 0.2wt % CuNps and 5wt % ZnONps; and (ii) Control Group: non-loaded commercial adhesive. Teeth were restored with resin composite. Thirty days later, extractions were performed. Extracted teeth were longitudinally sectioned. Nps in powder were characterized by field emission scanning electron microscope (FE-SEM) and energy dispersive X-ray (EDX) analysis. Microtensile bond strength (μTBS), degree of conversion (DC), and nanoleakeage (NL) tests were executed. In situ zymography (Zym) was performed to evaluate the gelatinolytic activity at the hybrid layer. Student's t-test (α = 0.05) was applied for all tests., Results: μTBS and DC did not show significant differences (p > 0.05) between both groups. However, NL and gelatinolytic activity at the hybrid layer showed significant values (p < 0.05) for experimental group in comparison with control group., Conclusion: The addition of 0.2 % CuNps and 5 % ZnONps to a universal adhesive decreases NL and gelatinolytic activity at the hybrid layer, without jeopardizing its adhesive properties., Significance: This randomized clinical trial with ex vivo analysis demonstrate that a commercial adhesive modified with 0.2wt % Cu and 5wt % ZnO Nps that does not affect its adhesive properties, reducing gelatinolytic activity and nanoleakage at the hybrid layer, which should contribute to an improvement of long term bonding-dentine clinical performance., Competing Interests: Declaration of competing interest The authors declare no competing financial interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Objective: This cross-sectional study evaluated the bone thickness on mini-implants insertion site, the factors that influence the digital planning of MARPE appliance (miniscrew-assisted rapid palatal expansion), and its different designs., Methods: A total of 135 plannings were assessed regarding the size of the expander screw used, the positioning and the type of the mini-implant rings, and their location in relation to the teeth. Bone thickness measurements were assessed in the region of the mini-implants' trajectory. Differences between the sexes was verified using the ANOVA test (5% significance)., Results: 73 cases were planned with 4 mini-implants and 62 cases, with 6 mini-implants. In 90% of cases, teeth #16 and #26 were used as supports, and the most used expander screw was 13mm (64.1% of cases). The anterior mini-implants of conventional MARPE showed more pronounced insertion in bone in males (5.9 ± 2mm; p= 0.025). The extra mini-implants (anterior region) were inserted with greater bone thickness in males (11.1 ± 2.3mm) compared to females (9.9 ± 1.8mm; p=0.041). A greater bone thickness was observed in males (10.1 ± 2.1 mm) when using mini-implants in the paramedian region., Conclusion: Additional rings allow more pronounced bone insertion. Male patients had greater bone thickness, which may be related to greater difficulty in opening the sutures. The alveolar process region seems to be a satisfactory site for mini-implants to those patients with reduced bone thickness in the paramedian posterior region. MARPE appliance must be customized for each patient, due to bone thickness and anatomical variations.

Objective: To analyze the effect of hookah and cigarettes on the oral mucosa of smokers through the use of exfoliative cytology., Study Design: Smear samples were collected by exfoliative cytology from the tongue of 33 hookah smokers, 22 cigarette smokers, and 30 non-smokers. The selected analyses include micronuclei (MN), metanuclear anomalies, epithelial maturation, and cytomorphology (nuclear area [NA], cytoplasmic area [CA], and NA/CA ratio)., Results: The largest differences observed for MN and metanuclear anomalies were between cigarette smokers and the control group (notably 1 MN P = .04; total cells with MN P = .039; total MN P = .042; karyorrhexis and binucleation, P = .0001). The hookah group, compared with the control group, showed the greatest differences for karyolysis (P = .0023), binucleation (P = .0003), and broken egg (P = .008). Significant differences were found between the smokers and the control groups regarding changes in the superficial cell without nucleus, perinuclear halo, vacuolization, color change, mucus, and keratohyalin granules. There was a significant increase in the NA and NA/CA ratio in the smoker groups., Conclusion: This study showed that a combined analysis of exfoliative cytology associated with other diagnostic methods is a useful tool for studying oral carcinogenesis. Hookah and cigarettes showed similar effects in terms of displaying substantial cytogenetic and cytotoxic damage., Competing Interests: Disclosure None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: this study evaluated dentin microtensile bond strength (µTBS) and failure modes (at 24 h and one year), bonding interface regarding hybridization, surface morphology regarding demineralization, in situ metalloproteinase (MMP) activity, and antibacterial effect of three dentin etchants compared to 35% phosphoric acid (PA)., Materials and Methods: The Adper Single Bond 2 adhesive (3 M Oral Care) was applied on moist dentin etched with PA (control) or on air-dried dentin etched with 3% aluminum nitrate + 2% oxalic acid (AN), 6.8% ferric oxalate + 10% citric acid (FO), or 10% citric acid (CA). The µTBS test used 40 human teeth (n = 10). Failure modes and surface morphology were analyzed by scanning electron microscopy (n = 3), while bonding interface morphology and MMP activity were evaluated by laser scanning confocal microscopy (n = 3). Antibacterial activity was evaluated against S. Mutans biofilm by means of viable cells count (CFU/mL)., Results: PA presented the highest bond strengths regardless of aging time. PA, AN, and CA showed stable bond strengths after one year of storage. Adhesive and mixed failures were predominant in all groups. Thin hybrid layers with short resin tags were observed for the experimental etchants. The AN-based etchant was able to inhibit MMP activity. All tested etchants presented antibacterial activity against S. Mutans biofilm., Significance: This study suggests different dentin etchants capable of inhibiting MMP activity while also acting as cavity disinfectants., Competing Interests: Declaration of Competing Interest The authors declare that no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Evaluate the effects of incorporating silica-coated silver nanoparticles (Ag@SiO2 NPs) into odontological clinic resin materials., Material and Methods: Silver nanoparticles coated with silicon dioxide were added to the experimental resin matrix at 1, 3, and 5wt%. Degree of conversion (DC), optical properties (total transmittance and color change), and microstructural analysis were evaluated. Materials were tested for silver ion release, cytotoxicity in dental pulp fibroblasts, Streptococcus mutans biofilm growth by Colony-Forming Unit (CFU) and confocal laser scanning microscopy (CLSM)., Results: Groups had a similar DC, despite significant differences observed in transmittance and color change analysis for all groups with NPs. Silver ion release values were below the detection limit after 72h for all groups, and NPs incorporation did not show a statistical difference from the control on pulp fibroblasts assay. After 72h, the CFU count was significantly reduced by 74% from 3wt% of Ag@SiO2NPs. CLSM evaluation on S. mutans colonies showed a dose-dependent decrease in the emitted fluorescence., Conclusions: The application of Ag@SiO2 NPs in a resinous matrix, demonstrates a significant reduction of S. mutans CFU in oral biofilm, at concentrations from 3wt%, without an increase in cytotoxicity. The reduced transmittance values did not affect the DC, although a significant color change was perceived in all concentrations. Key words: Nanoparticles, Silver Compounds, Composite Dental Resin, Anti-Bacterial Agent, Optical Imaging., Competing Interests: Conflicts of interest None declared., (Copyright: © 2024 Medicina Oral S.L.)

In recent years, alternative pulpal therapies targeting dentinogenesis signaling pathways using different peptides have been investigated. The aim of this study was to verify the effectiveness of poly(aspartic acid), pAsp, in dentin regeneration using an animal model., Methods: Mechanical pulp exposure was performed in the upper molars of 56 Wistar rats, randomly divided as follows (n = 14): control (no treatment); MTA group-pulp capping with mineral trioxide aggregate (MTA Angelus); pAsp group-application of 20 μL of pAsp solution (25 mg·mL -1 ); MTA+pAsp group-application of MTA mixed with pAsp (5:1 by mass). Animals were euthanized after 7 or 21 days. Histological sections were submitted to hematoxylin-eosin and Brown and Brenn staining and immunohistochemical analysis for osteopontin (OPN) and dentin matrix protein 1 (DMP 1)., Results: At 7 days, an acute inflammatory infiltrate and the presence of disorganized mineralized tissue were observed in all groups. At 21 days, the quality and thickness of the reparative dentin in treated groups were superior to the control, and bacterial contamination was observed in two MTA-pAsp specimens. While all treated groups showed intense immunostaining for OPN at 21 days, only the pAsp group expressed DMP 1, indicating the presence of fully differentiated odontoblast-like cells., Conclusion: Poly(aspartic) acid promoted dentin regeneration in rat molars in the absence of an additional calcium source and may be an alternative to MTA as a pulp-capping agent.

Objectives: Evaluate molecularly the role of P 11 -4 self-assembly peptide in dentin remineralization and its interaction with collagen I., Methods: The calcium-responsive P 11 -4 peptide was analyzed by intrinsic fluorescence emission spectrum, circular dichroism spectrum (CD), and atomic force microscope (AFM). Differential light scattering was used to monitor the nucleation growth rate of calcium phosphate nanocrystals in the absence or in the presence of P 11 -4. AFM was used to analyze the radial size (nm) of calcium phosphate nanocrystals formed in the absence or in the presence of P 11 -4, as well as to verify the spatial structure of P 11 -4 in the absence or in the presence of Ca 2+ ., Results: The interaction of Ca 2+ with the P 11 -4 (K D = 0.58 ± 0.06 mM) promotes the formation of β-sheet antiparallel structure, leads to its precipitation in saturated solutions of Ca/P = 1.67 and induces the formation of parallel large fibrils (0.6 - 1.5 µm). P 11 -4 organized the HAP nucleation by reducing both the growth rate and size variability of nanocrystals, analyzed by the F test (p < 0.0001, N = 30). P 11 -4 interacts (K D = 0.75 ± 0.06 μM) with the KGHRGFSGL motif present at the C-terminal collagen telopeptide domain. P 11 -4 also increased the amount of HAP and collagen in the MDPC-23 cells., Significance: The presented data propose a mechanism that will help future clinical and/or basic research to better understand a molecule able to inhibit structural collagen loss and help the impaired tissue to remineralize., (Copyright © 2023 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.)

The plasma jet transfer technique relies on a conductive wire at floating potential, which, upon entering in contact with a primary discharge, is capable of igniting a small plasma plume at the distal end of a long flexible plastic tube. In this work, two different long tube configurations were employed for the surface modification of polypropylene (PP) samples using argon as the working gas. One of the jet configurations has a thin copper (Cu) wire, which was installed inside the long tube. In the other configuration, the floating electrode is a metallic mesh placed between two plastic tubes in a coaxial arrangement. In the first case, the tip of the Cu wire is in direct contact with the working gas at the plasma outlet, whereas, in the second, the inner plastic tube provides an additional dielectric barrier that prevents the conductor from being in contact with the gas. Water contact angle (WCA) measurements on treated PP samples revealed that different surface modification radial profiles are formed when the distance ( d ) between the plasma outlet and target is changed. Moreover, it was found that the highest WCA reduction does not always occur at the point where the plasma impinges the surface of the material, especially when the d value is small. Through X-ray photoelectron spectroscopy (XPS) analysis, it was confirmed that the WCA values are directly linked to the oxygen-functional groups formed on the PP surfaces after the plasma treatment. An analysis of the WCA measurements along the surface, as well as their temporal evolution, together with the XPS data, suggest that, when the treatment is performed at small d values, the plasma jet removes some functional groups at the point where the plasma hits the surface, thus leading to peculiar WCA profiles.

Mitochondria have pivotal roles in cellular physiology including energy metabolism, reactive oxygen species production, Ca 2+ homeostasis, and apoptosis. Altered mitochondrial morphology and function is a common feature of cancer cells and the regulation of mitochondrial homeostasis has been identified as a key to the response to chemotherapeutic agents in human leukemias. Here, we explore the mechanistic aspects of cytotoxicity produced by thioridazine (TR), an antipsychotic drug that has been investigated for its anticancer potential in human leukemia cellular models. TR exerts selective cytotoxicity against human leukemia cells in vitro. A PCR array provided a general view of the expression of genes involved in cell death pathways. TR immediately produced a pulse of cytosolic Ca 2+ , followed by mitochondrial uptake, resulting in mitochondrial permeabilization, caspase 9/3 activation, endoplasmic reticulum stress, and apoptosis. Ca 2+ chelators, thiol reducer dithiothreitol, or CHOP knockdown prevented TR-induced cell death. TR also exhibited potent cytotoxicity against BCL-2/BCL-xL-overexpressing leukemia cells. Additionally, previous studies have shown that TR exhibits potent antitumor activity in vivo in different solid tumor models. These findings show that TR induces a Ca 2+ -mediated apoptosis with involvement of mitochondrial permeabilization and ER stress in leukemia and it emphasizes the pharmacological potential of TR as an adjuvant in antitumor chemotherapy.

To evaluate the flexural strength (FS) and flexural modulus (FM) of a commercial 3Y-TZ0P ceramic after artificial aging and either without or with two application times of non-thermal plasma treatments (NTP). In addition, changes in crystalline phase transformation and surface nano-topography after NTP application, during different aging periods, were evaluated. Ninety 3Y-TZP bars (45x4x3 mm) were made for FS and FM testing, and assigned to nine groups (n=10): no NTP/no aging (Control); no NTP/4h aging; no NTP/30h aging; 10s NTP/no aging; 10s NTP/4h aging; 10s NTP/30h aging; 60s NTP/no aging; 60s NTP/4h aging and 60s NTP/30h aging. Artificial accelerated aging was simulated using an autoclave (134º C at 2 bar) for up to 30h. FS and FM were assessed using a universal testing machine and data analyzed using a ANOVA and Tukey test (α=0.05). The volume change in zirconia monoclinic phase (MPV) was evaluated using X-ray diffraction and surface nano-topography was assessed using atomic force microscopy (baseline until 30h-aging). NTP application did not influence the FS and FM of zirconia. Compared to the Control (no NTP/no aging), the FS of zirconia samples treated for 30 hours in autoclave ("no NTP/30h aging" group) increased. Artificial aging for 30 hours significantly increased the FM of zirconia, regardless of NTP application. MPV tended to increase following the increase in aging time, which might result in the surface irregularities observed at 30h-aging. NTP did not alter the zirconia properties tested, but 30h-aging can change the zirconia FS, FM and MPV.

The tissue engineering of dental oral tissue is tackling significant advances and the use of stem cells promises to boost the therapeutical approaches of regenerative dentistry. Despite advances in this field, the literature is still scarce regarding the modulatory effect of laser photobiomodulation (PBM) on genes related to inflammation and osteogenesis in Postnatal Human Dental Pulp Stem cells (DPSCs). This study pointedly investigated the effect of PBM treatment in proliferation, growth and differentiation factors, mineralization, and extracellular matrix remodeling genes in DPSCs. Freshly extracted human third molars were used as a source for DPSCs isolation. The isolated DPSCs were stimulated to an inflammatory state, using a lipopolysaccharide (LPS) model, and then subjected or not to laser PBM. Each experiment was statistically evaluated according to the sample distribution. A total of 85 genes related to inflammation and osteogenesis were evaluated regarding their expression by RT-PCR. Laser PBM therapy has shown to modulate several genes expression in DPSCs. PBM suppressed the expression of inflammatory gene TNF and RANKL and downregulated the gene expression for VDR and proteolytic enzymes cathepsin K, MMP-8 and MMP-9. Modulation of gene expression for proteinase-activated receptors (PARs) following PBM varied among different PARs. As expected, PBM blocked the odontoblastic differentiation of DPSCs when subjected to LPS model. Conversely, PBM has preserved the odontogenic potential of DPSCs by increasing the expression of TWIST-1/RUNEX-2/ALP signaling axis. PBM therapy notably played a role in the DPSCs genes expression that mediate inflammation process and tissue mineralization. The present data opens a new perspective for PBM therapy in mineralized dental tissue physiology., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Rocha, Alvarez, Pelosine, Carrilho, Tersariol and Nascimento.)

Objective: To evaluate the influence of surface treatment on roughness (SA), topography, and shear bond strength (SBS) of computer-aided designer and manufacture (CAD/CAM) zirconia-reinforced lithium silicate (ZLS) and feldspathic (FEL) glass-ceramics., Materials and Methods: FEL and ZLS specimens were submitted to 5% or 10% hydrofluoric acid (HF) or self-etching ceramic primer (MEP) and different application times (20, 40, and 60 s), or to sandblasting (Control, 20 s). Resin cement cylinders were bonded to the specimens and tested in shear (n = 10) after 24 h and 16-months of water storage. SA and topography were evaluated by atomic force (AFM, n = 10) and scanning electron microscopy. Data were analyzed by ANOVA and Bonferroni test (α = 0.05)., Results: Sandblasting promoted the highest SA for ZLS, but 10% HF (40, 60 s) promoted higher SBS at 16 months. 10% HF produced the highest SA for FEL, but sandblasting and 5% HF (20 s) maintained SBS after 16 months, without differences from 10% HF (20 s) (p > 0.05). Overall, MEP produced lower SA and SBS among groups (p < 0.05). HF displayed greater morphological changes on FEL., Conclusion: 10% HF (40 s) provided better results for ZLS, while 5% or 10% HF (20 s) was suitable for FEL., Clinical Significance: Surface treatments influenced SA, topography, and SBS of materials. HF etching is the surface treatment of choice for both CAD/CAM glass-ceramics., (© 2021 Wiley Periodicals LLC.)

Background/objective: Until 2010, adults underwent surgical treatment for maxillary expansion; however, with the advent of micro-implant-assisted rapid maxillary expansion (MARME), the availability of less invasive treatment options has increased. Nevertheless, individuals with severe transverse maxillary deficiency do not benefit from this therapy. This has aroused interest in creating a new device that allows the benefit of maxillary expansion for these individuals. The aim of this study was to evaluate the efficacy of three MARME models according to tension points, force distribution, and areas of concentration in the craniofacial complex when transverse forces are applied using finite element analysis., Materials and Methods: Digital modeling of the three MARME models was performed. Model A comprised five components: one body screw expander and four adjustable arms with rings for mini-implant insertion. These arms have an individualized height adjustment that allows MARME positioning according to the patient's palatal anatomy, thereby preventing body screw expander collision with the lateral mucosa in severe cases of maxillary deficiency. Model B was a maxillary expander with screw rings joined to the body, and model C was similar to model B, except that model C had open rings for the insertion of the mini-implants. Through the MEF (Ansys software), the stresses, distribution, and area of concentration of the stresses were evaluated when transverse forces of 7.85 N were applied., Results: The three models maintained the following pattern: model C presented weak stress peaks with limited distribution and lower concentration area, model B obtained median stress peaks with better distribution when compared to that of model C, and model A showed better stress distribution and larger concentration area. In model A, tensions were located in the lateral lamina of the pterygoid process, which is an important site for maxillary expansion. The limitation of the present study was that it did not include the periodontal tissues and muscles in the finite element method evaluation., Conclusions: Model A showed the best stress distribution conditions. In cases of severe atresia, model A seems to be an excellent option.

Objectives: To determine the impact of treating caries-affected dentin (CAD) with: 0.2% sodium fluoride (NaF), casein phosphopeptide-amorphous calcium phosphate (CPP-ACP/MI Paste™) or peptide P 11 -4 (Curodont™ Repair) on the longevity of resin/CAD interface at storage times of 24 -h, 6- and 18-month., Methods: 255 caries-free third molars were used, and CAD was produced by a biological method. The teeth were randomly distributed into: G1- Sound dentin (SD); G2- CAD; G3- CAD + 0.2% NaF (CAD/NaF); G4- CAD + CPP-ACP (CAD/ACP); G5- CAD + Curodont™ Repair (CAD/P 11 -4). The Filtek Z350 composite resin block was bonded to dentin using Adper™ Single 2 (4 mm/height). Resin/dentin blocks were stored in a solution of Simulated Body Fluid at 37 °C, pressures were modified to simulate natural pulpal pressures. Specimens were investigated by microtensile bond strength (μTBS) (n = 8), Scanning Electron Microscopy (to assess the failure mode) (n = 8), nanoinfiltration (to assess the interface sealing) (n = 3), in situ zymography (to assess the gelatinolytic activity) (n = 3) and micro-computed microtomography (μ-CT) (to assess the mineralization) (n = 3). Data from μTBS, μ-CT and, nanoinfiltration and hybrid layer formation/degradation were submitted to two-way ANOVA and Tukey tests, and failure patterns and in situ zymography to Kruskal-Wallis and Dunn tests (α = 5%)., Results: The highest mineral density change by μ-CT, smallest silver nitrate infiltration and proteolytic activity in the adhesive layer were obtained significantly for the groups SD, CAD/ACP and CAD/P 11 -4, with most mixed fractures at 18-month (p < 0.001). CAD/NaF showed significantly similar values to CAD, CAD and CAD/NaF which presented a high percentage of adhesive fracture (p < 0.001) at all time periods., Significance: Treating caries-affected dentin with remineralizing agents CPP-ACP and Curodont™ Repair, has the potential to be a clinically relevant treatment protocol to increase the longevity of adhesive restorations., (Copyright © 2021 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.)

Objective: In professional sports activities, the search for increased performance is constant. Electrophysical agents, including photobiomodulation (PBM), have been used in the sports context to accelerate postworkout recovery, prevent injuries, and even to improve performance. This study aims to investigate the effects of infrared laser (904 nm) on skeletal muscle gene expression of performance-related proteins of rats submitted to a chronic resistance training protocol. Materials and methods: Male Wistar rats ( n  = 40), weighing ±300 g were divided into four groups: sedentary control (CT, n  = 10); irradiated control (CTL, n  = 10); exercised not irradiated (EX, n  = 10); exercised irradiated (EXL, n  = 10). To assess the performance, the maximum carrying test was adapted and applied 72 h prior the training and 72 h after the last exercise session. The vertical weight climbing protocol was adapted for resistance training 3 × per week with 48 h interval between each session: first week adaptation, second week 25% of body weight (BW), third week 50% BW, fourth week 75% BW, and fifth week 100% BW. Animals were irradiated before exercise on hind paws 50 sec each, with infrared laser 904 nm 5 days per week, during 4 weeks, 9 J per leg in a total of 18 J energy per day. Results: The EXL performed more climbing (7.1 ± 0.91) compared to EX (4.4 ± 0.63). PBM promoted increased expression of lactate dehydrogenase enzyme, mammalian target of rapamycin protein, and androgen receptor ( p  < 0.05) but not the myosin heavy chain ( p  = 0.43). Conclusions: PBM therapy increases the expression of performance-related muscle mass gain genes besides improving the resistance training performance.

Objective: Cysteine proteases are lysosomal enzymes that, under specific circumstances, may be secreted into the extracellular space and participate in protein turnover. This study investigated the involvement of cathepsin B in the gelatinolytic activity of mature dentin matrices at neutral pH., Design: Human dentin fragments were made into powder and enzymes were extracted using guanidine-HCl/EDTA. Host-derived dentin proteases (cathepsin B, MMP-2 and MMP-9) were identified by immunoblotting, and their activities were evaluated spectrofluorimetrically using fluorogenic substrates. Proteases activities were monitored by measuring the rate of hydrolysis of substrates in the presence/absence of MMP- or cysteine cathepsin inhibitors, at neutral pH (7.4). Mass spectroscopy was used to determine the substrates' cleavage points. Reverse zymography was performed to examine the gelatinolytic activity of cathepsin B., Results: Western-blots of dentin extracts yielded strong bands at 95, 72 and 30 kDa, corresponding respectively to MMP-9, MMP-2 and Cathepsin B. Greater fluorogenic substrates hydrolysis occurred in the absence of MMP and cysteine cathepsin inhibitors than in their presence. Cathepsin B exhibited significant gelatinolytic activity., Conclusions: Together with MMP-2 and MMP-9, cathepsin B also account for the host-derived gelatinolytic activity and matrix turnover of mature dentin at physiological, neutral pH., Competing Interests: Declaration of Competing Interest We, undersigned authors of the manuscript titled “Insights into Cathepsin-B Activity in Mature Dentin Matrix”, declare no conflict of interest of any nature and that the present manuscript has been submitted solely to the Achieves Oral Biology and is not under consideration for publication in any other journal. This paper has been compiled with the knowledge, input and approval of all the named authors., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Purpose: Verify the influences of physical activity level, nutritional status and screen habits on the prevalence of back pain in Brazilian students., Methods: The sample consisted of 577 schoolchildren (female = 274; male = 303) aged between 10 and 16 years old, regularly enrolled in the 6 th grade of elementary school living in the metropolitan area of the Alto Tietê of the state of São Paulo, Brazil. The prevalence, intensity and frequency of pain was verified with the Back Pain Assessment Instrument. The usual practice of physical activity was verified with the Physical Activity Questionnaire for Older Children/Adolescent. Nutritional status was analyzed using Body Mass Index. Screen habits were obtained through a previously structured questionnaire., Results: The Chi-square test indicated that pain complaint and its prevalence in the cervical region are significantly higher in females (p < 0.05). The multiple logistic regression test revealed that watching television influences the prevalence of cervical pain and that the use of more than one screen increases the occurrence of low back pain in male students (p < 0.05)., Conclusion: Female students were the most affected by back pain complain, especially in the cervical region. However, factors associated with the prevalence of back pain were found only in males., Competing Interests: Conflict of interest: All authors declare no conflicts of interest in this paper., (© 2020 the Author(s), licensee AIMS Press.)

Objectives: This study evaluated the cell viability and expression of different major genes involved in mineralization in odontoblast-like cells exposed to sodium trimetaphosphate (STMP). It was also investigated the influence of STMP on the rate of calcium phosphate crystal growth, its anti-proteolytic action against the enzymatic degradation of type I collagen, the binding mechanism of STMP to collagen fibrils, and the potential mechanism to induce collagen stabilization., Methods: Immortalized rat odontoblast MDPC-23 cells were cultured. Cell viability was assessed by trypan blue staining, and the changes in gene expression balance induced by STMP were assessed by quantitative reverse transcription (qRT) PCR assays. Crystalline particle formation was monitored by light-scattering detectors to estimate pH variation and the radial size of the crystalline particles as a function of reaction time (pH 7.4, 25°C) in the presence of STMP in supersaturated calcium phosphate solution (Ca/P=1.67). Images were obtained under atomic force microscopy (AFM) to measure the particle size in the presence of STMP. A three-point bending test was used to obtain the elastic modulus of fully demineralized dentin beams after immersion in STMP solution. The binding mechanism of STMP to collagen fibrils and potential stabilization mechanism was assessed with circular dichroism spectrometry (CD). The data were analyzed statistically (α=0.05)., Results: STMP had no significant influence on the cell viability and gene expression of the MDPC-23 cells. STMP greatly increased the rate of crystal growth, significantly increasing the average radial crystal size. AFM corroborated the significant increase of STPM-treated crystal size. Mineralized collagen I fibrils exhibited less collagenase degradation with lower STMP concentration. CD analysis demonstrated changes in the conformational stability after STMP binding to type I collagen., Significance: The increased resistance of collagen against the proteolytic activity of collagenases appears to be related to the conformational change induced by STMP binding in collagen I and the STMP capacity for promoting biomimetic mineralization in type I collagen fibrils., (Copyright © 2020 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.)

Background: Adipose tissue is the main energy storage tissue in the body. Its catabolic and anabolic responses depend on several factors, such as nutritional status, metabolic profile, and hormonal signaling. There are few studies addressing the effects of laser photobiomodulation (PBM) on adipose tissue and results are controversial. Objective: Our purpose was to investigate the metabolic effects of PBM on adipose tissue from Wistar rats supplemented or not with caffeine. Materials and methods: Wistar rats were divided into four groups: control (CTL), laser-treated [CTL (L)], caffeine (CAF), and caffeine+PBM [CAF (L)]. Blood was extracted for quantification of triglyceride and cholesterol levels and white adipose tissues were collected for analysis. We evaluated gene expression in the adipose tissue for the leptin receptor, lipase-sensitive hormone, tumor necrosis factor alpha, and beta adrenergic receptor. Results: We demonstrated that the low-level laser irradiation was able to increase the feed intake of the animals and the relative mass of the adipose tissue in the CTL (L) group compared with CTL. Laser treatment also increases serum triglycerides [CTL = 46.99 ± 5.87; CTL (L) = 57.46 ± 14.38; CAF = 43.98 ± 5.17; and CAF (L) = 56.9 ± 6.12; p  = 0.007] and total cholesterol (CTL = 70.62 ± 6.80; CTL (L) = 79.41 ± 13.07; CAF = 71.01 ± 5.52; and CAF (L) = 79.23 ± 6.881; p  = 0.003). Conclusions: Laser PBM decreased gene expression of the studied genes in the adipose tissue, indicating that PBM is able to block the catabolic responses of this tissue. Interestingly, the CAF (L) and CAF animals presented the same CLT (L) phenotype, however, without increasing the feed intake and the relative weight of the adipose tissue. The description of these phenomena opens a new perspective for the study of the action of low-level laser in adipose tissue.

Objectives:  In this study, a collagen-rich biomembrane obtained from porcine -intestinal submucosa for application in guided bone regeneration was developed and characterized. Then, its biological and mechanical properties were compared with that of commercial products ( GenDerm [Baumer], Lumina-Coat [Critéria], Surgitime PTFE [Bionnovation], and Surgidry Dental F [Technodry])., Materials and Methods:  The biomembrane was extracted from porcine intestinal submucosa. Scanning electron microscopy, spectroscopic dispersive energy, glycosaminoglycan quantification, and confocal microscopy by intrinsic fluorescence were used to evaluate the collagen structural patterns of the biomembrane. Mechanical tensile and deformation tests were also performed., Statistical Analysis:   The results of the methods used for experimental membrane characterizations were compared with that obtained by the commercial membranes and statistically analyzed (significance of 5%)., Results:  The collagen-rich biomembrane developed also exhibited a more organized, less porous collagen fibril network, with the presence of glycosaminoglycans. The experimental biomembrane exhibited mechanical properties, tensile strength, and deformation behavior with improved average stress/strain when compared with other commercial membranes tested. Benefits also include a structured, flexible, and -bioresorbable characteristics scaffold., Conclusions:  The experimental collagen-rich membrane developed presents physical-chemical, molecular, and mechanical characteristics similar to or better than that of the commercial products tested, possibly allowing it to actively participating in the process of bone neoformation., Competing Interests: None declared., (Dental Investigation Society.)

Objective: Clinical issues have been raised about problems related to cytotoxic effects caused when applying self-adhesive cement. It was hypothesized that byproducts eluted from self-adhesive cements modulate oxidative stress response, the gene expression of signaling pathways of inflammatory process/transcriptional activators, and the expression and activity of interstitial collagenases, and modify the phenotypic characteristics of cellular proliferation and mineral deposition in odontoblastic-like cells., Methods: Cements (MaxCem Elite [MAX] and RelyX U200 [U200)]) were mixed, dispensed into moulds, and photoactivated according to the manufacturers' instructions. Immortalized rat odontoblast-like cells (MDPC-23) were cultured and exposed to polymerized specimens of cements for 4 h. Reactive oxidative specimen production and quantification of gene expression were evaluated. Cell proliferation assay and alizarin red staining were also performed to evaluate the disturbance induced by the cements on cellular proliferation and mineralization., Results: Despite their cytotoxic effects, both self-adhesive cements influenced the metabolism in the odontoblast cells on different scales. MAX induced significantly higher oxidative stress in odontoblast cells than U200. Gene expression varied as a function of exposure to self-adhesive cements; MAX induced the expression of pro-inflammatory cytokines such as TNF-α, whereas U200 downregulated, virtually depleted TNF-α expression, also inducing overexpression of the transcriptional factor Runx2. Overexpression of heme oxygenase-1 (HO-1) and thioredoxin reductase 1 (TRXR1) occurred after exposure to both cements, antioxidant genes that are downstream of Keap1-Nrf2-ARE system. MAX significantly induced the overexpression of collagenase MMP-1, and U200 induced the expression of gelatinase MMP-2. MAX significantly inhibited cell proliferation whereas U200 significantly activated cell proliferation. Alizarin red staining revealed significantly decreased mineral deposition especially when exposed to MAX., Significance: These results support the hypothesis that byproducts of different self-adhesive cements play important roles in the highly orchestrated process which ultimately affect the cellular proliferation and the mineral deposition in odontoblastic-like cells, possibly delaying the reparative dentin formation after cementation of indirect restorations, especially on recently exposed dentin preparations., (Copyright © 2019 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.)

The major goal in restorative dentistry is to develop a true regenerative approach that fully recovers hydroxyapatite crystals within the caries lesion. Recently, a rationally designed self-assembling peptide P 11 -4 (Ace-QQRFEWEFEQQ-NH 2 ) has been developed to enhance remineralization on initial caries lesions, yet its applicability on dentin tissues remains unclear. Thus, the present study investigated the interaction of P 11 -4 with the organic dentin components as well as the effect of P 11 -4 on the proteolytic activity, mechanical properties of the bonding interface, and nanoleakage evaluation to artificial caries-affected dentin. Surface plasmon resonance and atomic force microscopy indicated that P 11 -4 binds to collagen type I fibers, increasing their width from 214 ± 4 nm to 308 ± 5 nm ( P < 0.0001). P 11 -4 also increased the resistance of collagen type I fibers against the proteolytic activity of collagenases. The immediate treatment of artificial caries-affected dentin with P 11 -4 enhanced the microtensile bonding strength of the bonding interface ( P < 0.0001), reaching values close to sound dentin and decreasing the proteolytic activity at the hybrid layer; however, such effects decreased after 6 mo of water storage ( P < 0.05). In conclusion, P 11 -4 interacts with collagen type I, increasing the resistance of collagen fibers to proteolysis, and improves stability of the hybrid layer formed by artificial caries-affected dentin.

Background: The lipid metabolism is essential for maintaining the body's energy responses. Laser photobiomodulation triggers many important cellular effects, but these effects on lipid metabolism are not well described. In this study, we analyzed the laser photobiomodulation in the hormone-sensitive lipase (HSL) activity, a key enzyme in the triglycerides (TAG) hydrolysis in adipose tissue 3T3-L1. Methods: Cells were submitted to the differentiation protocol in adipose cells, irradiated with 1, 2, and 3J with laser (904 nm-60 mw-laser diode) and incubated for 4 h after irradiation. Results: The response of laser photobiomodulation was able to trigger an inhibition of HSL activity (control = 0.057 ± 0.0008; 1J = 0.050 ± 0.0003; 2J = 0.0477 ± 0.002; 3J = 0.051 ± 0.002; p  = 0.0003 against the control), but no modulation was observed in TAG levels into the medium (control = 26.5856 ± 0.52; 1J = 26.5856 ± 0.52; 2J = 27.2372 ± 1.41; 3J = 25.9991 ± 0.1303; p  = 0.18). Conclusions: This is the first study of HSL activity modulation with laser radiation, suggesting that photobiomodulation can influence adipose tissue metabolism and open a new field of study.

This study investigated the effect of application of non-thermal atmospheric plasma (NTAP) on the topography and composition of the dentin surface, as well as the microtensile bond strength (μTBS) of a universal adhesive to NTAP-treated dentin. Exposed flat dentin surfaces from human third molars were either treated with NTAP for 10 and 30 s or untreated (control). The dentin-surface topography and chemical composition were characterized by atomic force microscopy (n = 3) and Raman confocal spectroscopy (n = 5), respectively. The μTBS (n = 8) of Scotchbond Universal to dentin was determined after storage for 24 h and 1 yr, either by direct water exposure or under simulated pulpal pressure. In-situ zymography was used to evaluate the influence of NTAP on the dentin-enzymatic activity. Non-thermal atmospheric plasma produced no remarkable topographical or chemical alterations at the dentin surface; only the amount of phosphate decreased following 10 s of treatment with NTAP. After 1 yr of direct water exposure, the μTBS of NTAP-treated specimens did not differ statistically significantly from that of untreated controls, whereas simulated pulpal pressure-aging resulted in a significantly higher μTBS for NTAP-treated dentin. The dentin-enzymatic activity appeared to be treatment-dependent, but the untreated controls showed more intense fluorescence within the hybrid layer. Scotchbond Universal maintained its μTBS strength after 1 yr of direct water exposure and simulated pulpal pressure, although remarkable statistical differences between treatments were observed depending on the aging condition., (© 2017 Eur J Oral Sci.)

Objectives: To evaluate the cytotoxic effects of exposing odontoblast cells to a variety of commercial self-adhesive cements polymerized using different activation modes., Methods: Five cements: MaxCem Elite (MAX), Bifix SE (BSE), G-Cem LinkAce (GCE), Clearfil SA Luting (CAS), and RelyX U200 (U200) were mixed, dispensed into molds, and distributed in groups, according to polymerization protocols: immediate photoactivation; delayed photoactivation (10min self-curing plus light-activation); and chemical activation (no light exposure). Immortalized rat odontoblast cells (MDPC-23) were cultured. Cell viability was assessed by Trypan Blue staining and total cell death was assessed by annexin V-APC/7-AAD double staining and flow cytometry. Volatilized compounds from polymerized specimens of cements were evaluated by gas chromatography/mass spectrometry (GC-MS). Data was analyzed with 2-way ANOVA/Tukey tests (α=0.05)., Results: Exposure to all of the cements tested significantly reduced the cell viability, irrespective of the activation protocol (p<0.05). The least harmful cements were CSA and U200. Total death of cells significantly increased when exposed to BSE, GCE, and MAX, especially when chemically activated (p<0.05). Characteristic apoptotic cells increased after exposure to cements, mainly for MAX, regardless of the activation mode. Chemical activation of MAX also induced necrosis. Moreover, GCE and MAX exhibited higher percentages of late apoptotic/dead cells. Chromatograms revealed 28 compounds released from the cements tested, some of them with known carcinogenic effects. Selection of self-adhesive cements and polymerization protocols affect the cytotoxicity and cell viability of odontoblastic cells., Clinical Significance: Despite the simplified cementation protocol, care is needed when cementing indirect restorations with self-adhesive cements, especially on recently exposed dentin. This category of material may cause differential cytotoxic effects and should be considered when selecting a cement. This is particularly true in clinical cases of light attenuation, where the polymerization depends on chemical activation, inducing higher cytotoxic damages when using some of the cements tested., (Copyright © 2017 The Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.)

Protease-activated receptors (PARs) are G protein-coupled receptors, which are activated by proteolytical cleavage of the amino-terminus and act as sensors for extracellular proteases. We hypothesized that PAR-1 and PAR-2 can be modulated by inflammatory stimulus in human dental pulp cells. PAR-1 and PAR-2 gene expression in human pulp tissue and MDPC-23 cells were analyzed by quantitative polymerase chain reaction. Monoclonal PAR-1 and PAR-2 antibodies were used to investigate the cellular expression of these receptors using Western blot, flow cytometry, and confocal microscopy in MDPC-23 cells. Immunofluorescence assays of human intact and carious teeth were performed to assess the presence of PAR-1 and PAR-2 in the dentin-pulp complex. The results show for the first time that human odontoblasts and MDPC-23 cells constitutively express PAR-1 and PAR-2. PAR-2 activation increased significantly the messenger RNA expression of matrix metalloproteinase (MMP)-2, MMP-9, MMP-13, and MMP-14 in MDPC-23 cells ( P < 0.05), while the expression of these enzymes decreased significantly in the PAR-1 agonist group ( P < 0.05). The high-performance liquid chromatography and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry analysis showed the presence of MMP-13 activity cleaving PAR-1 at specific, noncanonical site TLDPRS 42 ↓F 43 LL in human dental pulp tissues. Also, we detected a presence of a trypsin-like activity cleaving PAR-2 at canonical site SKGR 20 ↓S 21 LIGRL in pulp tissues. Confocal microscopy analysis of human dentin-pulp complex showed intense positive staining of PAR-1 and PAR-2 in the odontoblast processes in dentinal tubules of carious teeth compared to intact ones. The present results support the hypothesis of activation of the upregulated PAR-1 and PAR-2 by endogenous proteases abundant during the inflammatory response in dentin-pulp complex.

It has been hypothesized that cysteine cathepsins (CTs) along with matrix metalloproteases (MMPs) may work in conjunction in the proteolysis of mature dentin matrix. The aim of this study was to verify simultaneously the distribution and presence of cathepsins B (CT-B) and K (CT-K) in partially demineralized dentin; and further to evaluate the activity of CTs and MMPs in the same tissue. The distribution of CT-B and CT-K in sound human dentin was assessed by immunohistochemistry. A double-immunolabeling technique was used to identify, at once, the occurrence of those enzymes in dentin. Activities of CTs and MMPs in dentin extracts were evaluated spectrofluorometrically. In addition, in situ gelatinolytic activity of dentin was assayed by zymography. The results revealed the distribution of CT-B and CT-K along the dentin organic matrix and also indicated co-occurrence of MMPs and CTs in that tissue. The enzyme kinetics studies showed proteolytic activity in dentin extracts for both classes of proteases. Furthermore, it was observed that, at least for sound human dentin matrices, the activity of MMPs seems to be predominant over the CTs one., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Purpose: To investigate the bonding efficacy of a multimode adhesive to plasma-treated and -untreated (control) dentin using a mini-interfacial fracture toughness (mini-iFT) test., Materials and Methods: Twenty human molars were used in a split-tooth design (n = 10). The adhesive Scotchbond Universal (SBU; 3M ESPE) was applied in etch-and-rinse (E&R) and self-etch (SE) modes. Mid-coronal dentin was exposed and covered with a standardized smear layer ground to 320 grit. One half of each dentin surface received 15 s of non-thermal atmospheric plasma (NTAP), while the other half was covered with a metallic barrier and kept untreated. Following the E&R mode, dentin was plasma treated immediately after phosphoric acid etching. SBU and a resin-based composite were applied to dentin following the manufacturer's instructions. Six mini-iFT specimens were prepared per tooth (1.5 x 2.0 x 16 to 18 mm), and a single notch was prepared at the adhesive-dentin interface using a 150-μm diamond blade under water cooling. Half of the mini-iFT specimens were immediately loaded until failure in a 4-point bending test, while the other half were first stored in distilled water for 6 months. After testing, the exact dimensions of the notch were measured with a measuring optical microscope, from which ΚIc was determined., Results: Three-way ANOVA revealed higher mini-iFT for SBU applied in E&R than SE mode for both storage times, irrespective of NTAP treatment., Conclusion: Overall, mini-iFT did not decrease for any of the experimental groups upon 6-month aging, while plasma treatment did not show a direct beneficial effect on mini-iFT of SBU applied in either E&R or SE mode.

Objectives: To evaluate whether the concentration of phosphoric acid (PA) has an effect on the proteolytic activity of sound human demineralized dentin. It is hypothesized that the activity of matrix-bound and extracted enzymes depends on the PA concentration used to demineralize dentin., Methods: One-gram aliquots of mid-coronal human dentin powder were demineralized with 1wt%, 10wt% and 37wt% PA. Concentrations of released calcium were measured for each set of demineralization. Extracted MMP-2 was immunologically identified by western blot and its activity was determined by conventional gelatin zymography. Analysis of released hydroxyproline (HYP) and in situ zymography were performed to evaluate the activity of insoluble, bound-matrix enzymes., Results: The amount of released calcium from dentin powder treated with 37wt% PA was significantly higher (p≤0.05) than that obtained by dentin demineralization with 10wt% and 1wt% PA. Expression and activity of endogenous enzymes, extracted from or bound to dentin matrix, were detected for all samples regardless of the PA concentration. However, the expression and activity of extracted MMP-2 were significantly higher when dentin was treated with 10wt% PA (p<0.05), followed by 1wt% and 37wt% PA. Similarly, the highest concentration of released HYP (i.e. meaning higher percentage of collagen degradation) and the highest activity in in situ zymography were observed when dentin samples were treated with 10wt% PA (p<0.05)., Conclusions: It was confirmed that PA does not denature endogenous enzymes of dentin matrices, but it may somehow modulate the expression and activity of these enzymes in a concentration-dependent manner., Clinical Significance: Endogenous proteases have been identified and suggested to be responsible for the digestion of dentin matrix when activated by the acidic components of dental adhesives. Proteolytic activity of dentinal MMPs showed to be dependent on phosphoric acid concentration. The clinically-used concentration (37%) does not inhibit MMPs activity, but slows it., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Proteolytic enzymes mediate the activation or inactivation of many physiologic and pathologic processes. The PHEX gene (Phosphate-regulating gene with homologies to endopeptidase on the X chromosome) encodes a metallopeptidase, which is mutated in patients with a prevalent form (1:20,000) of inherited rickets-X-linked hypophosphatemia (XLH). XLH shows growth retardation, hypophosphatemia, osteomalacia, and defective renal phosphate reabsorption and metabolism of vitamin D. Most PHEX studies have focused on bone, and recently we identified osteopontin (OPN) as the first protein substrate for PHEX, demonstrating in the murine model of XLH (Hyp mice) an increase in OPN that contributes to the osteomalacia. Besides its role in bone mineralization, OPN is expressed in many tissues, and therein has different functions. In tumor biology, OPN is known to be associated with metastasis. Here, we extend our PHEX-OPN studies to investigate PHEX expression in a squamous cell carcinoma (SCC) cell line and its possible involvement in modulating OPN function. Real-time PCR showed PHEX-OPN co-expression in SCC cells, with sequencing of the 22 exons showing no mutation of the PHEX gene. Although recombinant PHEX hydrolyze SCC-OPN fragments, unlike in bone cells, SCC-PHEX protein was not predominantly at the plasma membrane. Enzymatic activity assays, FACs and immunoblotting analyses demonstrated that membrane PHEX is degraded by cysteine proteases and the decreased PHEX activity could contribute to inappropriate OPN regulation. These results highlight for the first time PHEX in tumor biology., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Human H-kininogen (120 kDa) plays a role in many pathophysiological processes and interacts with the cell surface through protein receptors and proteoglycans, which mediate H-kininogen endocytosis. In the present work we demonstrate that H-kininogen containing bradykinin domain is internalized and different endogenous kininogenases are present in CHO-K1 cells. We used CHO-K1 (wild type) and CHO-745 (mutant deficient in proteoglycans biosynthesis) cell lines. H-kininogen endocytosis was studied using confocal microscopy, and its hydrolysis by cell lysate fraction was determined by immunoblotting. Bradykinin release was also measured by radioimmunoassay. H-kininogen interaction with the cell surface of CHO-745 cells resulted in bradykinin release by serine proteases. In CHO-K1 cells, which produce heparan and chondroitin sulfate proteoglycans, internalization of H-kininogen through its bradykinin domain can occur on lipid raft domains/caveolae. Nevertheless bradykinin-free H-kininogen was not internalized by CHO-K1 cells. The H-kininogen present in acidic endosomal vesicles in CHO-K1 was approximately 10-fold higher than the levels in CHO-745. CHO-K1 lysate fractions were assayed at pH 5.5 and intact H-kininogen was totally hydrolyzed into a 62 kDa fragment. By contrast, at an assay pH 7.4, the remained fragments were 115 kDa, 83 kDa, 62 kDa and 48 kDa in size. The antipain-Sepharose chromatography separated endogenous kininogenases from CHO-K1 lysate fraction. No difference was detected in the assays at pH 5.5 or 7.4, but the proteins in the fraction bound to the resin released bradykinin from H-kininogen. However, the proteins in the unbound fraction cleaved intact H-kininogen at other sites but did not release bradykinin. H-kininogen can interact with extravascular cells, and is internalized dependent on its bradykinin domain and cell surface proteoglycans. After internalization, H-kininogen is proteolytically processed by intracellular kininogenases. The present data also demonstrates that serine or cysteine proteases in lipid raft domains/caveolae on the CHO cell can hydrolyze H-kininogen, thus releasing kinins.

Objective: Dentin matrices release ICTP and CTX fragments during collagen degradation. ICTP fragments are known to be produced by MMPs. CTX fragments are thought to come from cathepsin K activity. The purpose of this study was to determine if quaternary methacrylates (QAMs) can inhibit matrix MMPs and cathepsins., Methods: Dentin beams were demineralizated, and dried to constant weight. Beams were incubated with rh-cathepsin B, K, L or S for 24h at pH 7.4 to identify which cathepsins release CTX at neutral pH. Beams were dipped in ATA, an antimicrobial QAM to determine if it can inhibit dentin matrix proteases. Other beams were dipped in another QAM (MDPB) to determine if it produced similar inhibition of dentin proteases., Results: Only beams incubated with cathepsin K lost more dry mass than the controls and released CTX. Dentin beams dipped in ATA and incubated for 1 week at pH 7.4, showed a concentration-dependent reduction in weight-loss. There was no change in ICTP release from control values, meaning that ATA did not inhibit MMPs. Media concentrations of CTX fell significantly at 15wt% ATA indicating that ATA inhibits capthesins. Beams dipped in increasing concentrations of MDPB lost progressively less mass, showing that MDPB is a protease-inhibitor. ICTP released from controls or beams exposed to low concentrations were the same, while 5 or 10% MDPB significantly lowered ICTP production. CTX levels were strongly inhibited by 2.5-10% MDPB, indicating that MDPB is a potent inhibitor of both MMPs and cathepsin K., Significance: CTX seems to be released from dentin matrix only by cathepsin K. MMPs and cathepsin K and B may all contribute to matrix degradation., (Copyright © 2014 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.)

The search for new compounds that induce p53-independent apoptosis is the focus of many studies in cancer biology because these compounds could be more specific and would overcome chemotherapy resistance. In this study, we evaluated the in vitro antitumour activity of a Biphosphinic Palladacycle Complex (BPC) and extended preclinical studies to an in vivo model. Saos-2 cells, a p53-null human osteosarcoma drug-resistant cell line, were treated with BPC in the presence or absence of a cathepsin B inhibitor and a calcium chelator (CA074 and BAPTA-AM, respectively), and several parameters related to apoptosis were evaluated. Preclinical studies were performed with mice that were intravenously inoculated with murine melanoma B16F10-Nex2 cells and treated intraperitoneally (i.p.) with BPC (8 mg/kg/day) for ten consecutive days, when lung metastatic nodules were counted. In vitro data show that BPC induces cell death in Saos-2 cells mainly by apoptosis, which was accompanied by the effector caspase-3 activation. These events are most likely related to Bax translocation and increased cytosolic calcium mobilisation, mainly from intracellular compartments. Lysosomal Membrane Permeabilisation (LMP) was also observed after 12 h of BPC exposure. Interestingly, BAPTA-AM and CA074 significantly decreased BPC cytotoxicity, suggesting that both calcium and cathepsin B are required for BPC antitumour activity. In vivo studies demonstrated that BPC protects mice against murine metastatic melanoma. In conclusion, BPC complex is an effective anticancer compound against metastatic murine melanoma. This complex is cytotoxic to the drug-resistant osteosarcoma Saos-2 human tumour cells by inducing apoptosis triggered by calcium signalling and a lysosomal-dependent pathway., (Copyright © 2014 Elsevier Masson SAS. All rights reserved.)

Introduction: The aim of this work was to evaluate the role of human plasma prekallikrein assembly and processing in cells and to determine whether proteoglycans, along with high molecular weight kininogen (H-kininogen), influence this interaction., Methods: We used the endothelial cell line ECV304 and the epithelial cell lines CHO-K1 (wild type) and CHO-745 (deficient in proteoglycans). Prekallikrein endocytosis was studied using confocal microscopy, and prekallikrein cleavage/activation was determined by immunoblotting using an antibody directed to the prekallikrein sequence C364TTKTSTR371 and an antibody directed to the entire H-kininogen molecule., Results: At 37°C, prekallikrein endocytosis was assessed in the absence and presence of exogenously applied H-kininogen and found to be 1,418.4±0.010 and 1,070.3±0.001 pixels/cell, respectively, for ECV304 and 1,319.1±0.003 and 631.3±0.001 pixels/cell, respectively, for CHO-K1. No prekallikrein internalization was observed in CHO-745 in either condition. Prekallikrein colocalized with LysoTracker in the absence and presence of exogenous H-kininogen at levels of 76.0% and 88.5%, respectively, for ECV304 and at levels of 40.7% and 57.0%, respectively, for CHO-K1. After assembly on the cell surface, a plasma kallikrein fragment of 53 kDa was predominant in the incubation buffer of all the cell lines studied, indicating specific proteolysis; plasma kallikrein fragments of 48-44 kDa and 34-32 kDa were also detected in the incubation buffer, indicating non-specific cleavage. Bradykinin free H-kininogen internalization was not detected in CHO-K1 or CHO-745 cells at 37°C., Conclusion: The prekallikrein interaction with the cell surface is temperature-dependent and independent of exogenously applied H-kininogen, which results in prekallikrein endocytosis promoted by proteoglycans. Prekallikrein proteolysis/activation is influenced by H-kininogen/glycosaminoglycans assembly and controls plasma kallikrein activity.