Combination of in silico and cell culture strategies to predict biomaterial performance: Effects of sintering temperature on the biological properties of hydroxyapatite.

Advances in methodologies to evaluate biomaterials brought an explosive growth of data, ensuing computational challenges to better analyzing them and allowing for high-throughput profiling of biological systems cost-efficiently. In this sense, we have applied bioinformatics tools to better understand the biological effect of different sintering temperatures of hydroxyapatite (abbreviated HA; at 1100, 1150, and 1250°C) on osteoblast performance. To do, we have better analyzed an earlier deposited study, in which the access code is E-MTAB-7219, which the authors have explored different in silico tools on this purpose. In this study, differential gene expression analyses were performed using the gene set variation analysis (GSVA) algorithm from the transcriptomes respecting the thermal changes of HA, which were validated using exclusively in vitro strategies. Furthermore, in silico approaches elected biomarkers during cell behavior in response to different sintering temperatures of HA, and it was further validated using cell culture and qPCR technologies. Altogether, the combination of those strategies shows the capacity of sintered HA at 1250°C to present a better performance in organizing an adequate microenvironment favoring bone regeneration, angiogenesis and material resorption stimulus once it has promoted higher involvement of genes such as CDK2, CDK4 (biomarkers of cell proliferation), p15, Osterix gene (related with osteogenic differentiation), RANKL (related with osteoclastogenesis), VEGF gene (related with angiogenesis), and HIF1α (related with hypoxia microenvironment). Altogether, the combination of in silico and cell culture strategies shows the capacity of sintered HA at 1250°C in guaranteeing osteoblast differentiation and it can be related in organizing an adequate microenvironment favoring bone regeneration, angiogenesis, and material resorption stimulus.

Boosting Titanium Surfaces with Positive Charges: Newly Developed Cationic Coating Combines Anticorrosive and Bactericidal Properties for Implant Application.

Along with poor implant-bone integration, peri-implant diseases are the major causes of implant failure. Although such diseases are primarily triggered by biofilm accumulation, a complex inflammatory process in response to corrosive-related metallic ions/debris has also been recognized as a risk factor. In this regard, by boosting the titanium (Ti) surface with silane-based positive charges, cationic coatings have gained increasing attention due to their ability to kill pathogens and may be favorable for corrosion resistance. Nevertheless, the development of a cationic coating that combines such properties in addition to having a favorable topography for implant osseointegration is lacking. Because introducing hydroxyl (-OH) groups to Ti is essential to increase chemical bonds with silane, Ti pretreatment is of utmost importance to achieve such polarization. In this study, plasma electrolytic oxidation (PEO) was investigated as a new route to pretreat Ti with OH groups while providing favorable properties for implant application compared with traditional hydrothermal treatment (HT). To produce bactericidal and corrosion-resistant cationic coatings, after pretreatment with PEO or HT (Step 1), surface silanization was subsequently performed via immersion-based functionalization with 3-aminopropyltriethoxysilane (APTES) (Step 2). In the end, five groups were assessed: untreated Ti (Ti), HT, PEO, HT+APTES, and PEO+APTES. PEO created a porous surface with increased roughness and better mechanical and tribological properties compared with HT and Ti. The introduction of -OH groups by HT and PEO was confirmed by Fourier transform infrared spectroscopy and the increase in wettability producing superhydrophilic surfaces. After silanization, the surfaces were polarized to hydrophobic ones, and an increase in the amine functional group was observed by X-ray photoelectron spectroscopy, demonstrating a considerable amount of positive ions. Such protonation may explain the enhanced corrosion resistance and dead bacteria (Streptococcus aureus and Escherichia coli) found for PEO+APTES. All groups presented noncytotoxic properties with similar blood plasma protein adsorption capacity vs the Ti control. Our findings provide new insights into developing next-generation cationic coatings by suggesting that a tailorable porous and oxide coating produced by PEO has promise in designing enhanced cationic surfaces targeting biomedical and dental implant applications.

Copper source determines chemistry and topography of implant coatings to optimally couple cellular responses and antibacterial activity.

Implant-related infections at the early healing period are considered one of the main risk factors in implant failure. Designing coatings that control bacterial adhesion and have cell stimulatory behavior remains a challenging strategy for dental implants. Here, we used plasma electrolytic oxidation (PEO) to produce antimicrobial coatings on commercially pure titanium (cpTi) using bioactive elements (calcium and phosphorus) and different copper (Cu) sources: copper acetate (CuAc), copper sulfate (CuS), and copper oxide (CuO); coatings containing only Ca and P (CaP) served as controls. Cu sources drove differential physical and chemical surface features of PEO coatings, resulting in tailorable release kinetics with a sustained Cu ion release over 10 weeks. The antibacterial effects of Cu-containing coatings were roughness-dependent. CuAc coating exhibited optimal properties in terms of its hydrophilicity, pores density, and limited surface roughness, which provided the most robust antibacterial activity combined with appropriate responses of human primary stem cells and angiogenic cells. Our data indicate that Cu source selection largely determines the functionality of Cu-containing PEO coatings regarding their antibacterial efficacy and cytocompatibility.

Optimizing citric acid protocol to control implant-related infections: An in vitro and in situ study.

OBJECTIVE: The present study aimed to establish an optimized protocol for biofilm removal from titanium (Ti) surfaces using citric acid (CA) solutions. BACKGROUND: Biofilm accumulation is the main factor to trigger peri-implant infections and to increase the risk of treatment failures. Although CA has been suggested as the anti-infective agent with highest potential for biofilm removal on Ti, there is no consensus that CA could improve the anti-infective treatment and its effect. METHODS: Physical and chemical alterations, electrochemical behavior, cytotoxicity, and antimicrobial effect of CA on Ti discs were evaluated using four concentrations (1, 10, 20, and 40%) and two application methods (immersion and rubbing). Negative control using 0.9% NaCl was used in all experiments. To evaluate whether different application times can have similar response, polymicrobial biofilm (microcosm model) was formed on Ti and treated with CA for 1, 2, 4, and 8 min. An in situ study was conducted to verify whether the established protocol is equally effective in biofilms formed on machined and sandblasted, large-grit, and acid-etched (SLA) Ti surfaces. RESULTS: CA 40% induced significantly higher surface alterations observed by confocal images and profilometry. In general, rubbing protocol decreased the surface roughness and increased the wettability (p < 0.05), exhibiting better surface cleaning by biofilm removal. CA 10% presented no indirect cytotoxicity and, when applied by rubbing for 8 min, presented proper in vitro antibacterial action and potential corrosion inhibition. When CA 10% was rubbed on Ti surfaces for 4 min, it displayed optimum cleaning ability as 8 min, working equally to remove in situ biofilm on machined and SLA surfaces. CONCLUSIONS: The application of CA 10% by rubbing for at least 4 min demonstrated to be a promising protocol to eliminate biofilms formed in smooth and rougher surfaces, which could improve implant-related infection therapies.

Thin Film Deposition by Atmospheric Pressure Dielectric Barrier Discharges Containing Eugenol: Discharge and Coating Characterizations.

Eugenol (4-Allyl-2-methoxyphenol) is the main constituent of clove oil. In addition to being widely used as a condiment, it has been recognized as a powerful bactericide. Owing to that, Eugenol has been used in several applications including odontology and as a conservative for food products. Aiming at the development of natural bactericide coatings, in this work, using an atmospheric pressure plasma in a dielectric barrier discharge (DBD) reactor Eugenol was deposited on stainless steel substrate, with argon as a carrier gas. The discharge power supply was a transformer at 14.4 kV peak-to-peak voltage and 60 Hz frequency. Operating with a gas flow rate at 4 L/min, the active power was around 1.2 W. The maximum plasma electron temperature of the plasma with monomers was about 1.5 eV, estimated by visible emission spectroscopy using a local thermodynamic equilibrium approach. The study also comprehended the analysis of the film structure, aging, and thermal stability using infrared reflectance spectroscopy, and its thicknesses and roughness by profilometry. The thickness of the films was in the range of 1000 to 2400 nm with a roughness of up to 800 nm with good adhesion on the substrate. The FTIR result shows a stable coating with a chemical structure similar to that of the monomer. Aging analysis showed that the film does not degrade, even after exposing the film for 120 days in ambient air and for 1.0 h under a high thermal UV-lamp.

OsteoBLAST: Computational Routine of Global Molecular Analysis Applied to Biomaterials Development.

For bone purposes, surface modifications are a common trend in biomaterials research aiming to reduce the time necessary for osteointegration, culminating in faster recovery of patients. In this scenario, analysis of intracellular signaling pathways have emerged as an important and reliable strategy to predict biological responses from in vitro approaches. We have combined global analysis of intracellular protein phosphorylation, systems biology and bioinformatics into an early biomaterial analysis routine called OsteoBLAST. We employed the routine as follows: the PamChip tyrosine kinase assay was applied to mesenchymal stem cells grown on three distinct titanium surfaces: machined, dual acid-etched and nanoHA. Then, OsteoBLAST was able to identify the most reliable spots to further obtain the differential kinome profile and finally to allow a comparison among the different surfaces. Thereafter, NetworKIN, STRING, and Cytoscape were used to build and analyze a supramolecular protein-protein interaction network, and DAVID tools identified biological signatures in the differential kinome for each surface.

Synthesis of bioactive glass-based coating by plasma electrolytic oxidation: Untangling a new deposition pathway toward titanium implant surfaces.

HYPOTHESIS: Although bioactive glass (BG) particle coatings were previously developed by different methods, poor particle adhesion to surfaces and reduced biological effects because of glass crystallization have limited their biomedical applications. To overcome this problem, we have untangled, for the first time, plasma electrolytic oxidation (PEO) as a new pathway for the synthesis of bioactive glass-based coating (PEO-BG) on titanium (Ti) materials. EXPERIMENTS: Electrolyte solution with bioactive elements (Na2SiO3-5H2O, C4H6O4Ca, NaNO3, and C3H7Na2O6P) was used as a precursor source to obtain a 45S5 bioglass-like composition on a Ti surface by PEO. Subsequently, the PEO-BG coating was investigated with respect to its surface, mechanical, tribological, electrochemical, microbiological, and biological properties, compared with those of machined and sandblasted/acid-etched control surfaces. FINDINGS: PEO treatment produced a coating with complex surface topography, Ti crystalline phases, superhydrophilic status, chemical composition, and oxide layer similar to that of 45S5-BG (~45.0Si, 24.5 Ca, 24.5Na, 6.0P w/v%). PEO-BG enhanced Ti mechanical and tribological properties with higher corrosion resistance. Furthermore, PEO-BG had a positive influence in polymicrobial biofilms, by reducing pathogenic bacterial associated with biofilm-related infections. PEO-BG also showed higher adsorption of blood plasma proteins without cytotoxic effects on human cells, and thus may be considered a promising biocompatible approach for biomedical implants.

Atmospheric Pressure Plasma Chemical Vapor Deposition of Carvacrol Thin Films on Stainless Steel to Reduce the Formation of E. Coli and S. Aureus Biofilms.

In this paper, we have investigated the deposition of thin films from natural carvacrol extract using dielectric barrier discharge (DBD) plasma polymerization, aiming at the inhibition of bacteria adhesion and proliferation. The films deposited on stainless steel samples have been characterized by scanning electron microscopy, infrared reflectance-absorbance spectroscopy, profilometry, and contact angle measurements. Films with thicknesses ranging from 1.5 µm to 3.5 µm presented a chemical structure similar to that of carvacrol. While the formation of biofilm was observed on untreated samples, the coating completely inhibited the adhesion of E. coli and reduced the adhesion of S. aureus biofilm in more than 90%.

UV-photofunctionalization of a biomimetic coating for dental implants application.

Photofunctionalization mediated by ultraviolet (UV) rays changes the physico-chemical characteristics of titanium (Ti) and improves the biological activity of dental implants. However, the role of UV-mediated photofunctionalization of biofunctional Ti surfaces on the antimicrobial and photocatalytic activity remains unknown and was investigated in this study. Commercially pure titanium (cpTi) discs were divided into four groups: (1) machined samples without UV light application [cpTi UV-]; (2) plasma electrolytic oxidation (PEO) treated samples without UV light application [PEO UV-]; (3) machined samples with UV light application [cpTi UV+]; and (4) PEO-treated samples with UV light application [PEO UV+]. The surfaces were characterized according to their morphology, roughness, crystalline phase, chemical composition and wettability. The photocatalytic activity and proteins adsorption were measured. For the microbiological assay, Streptococcus sanguinis was grown on the disc surfaces for 1 h and 6 h, and the colony forming units and bacterial organization were evaluated. In addition, to confirm the non-cytotoxic effect of PEO UV +, human gingival fibroblast (HGF) cells were cultured in a monolayer onto each material surface and the cells viability and proliferation evaluated by a fluorescent cell staining method. PEO treatment increased the Ti surface roughness and wettability (p < 0.05). Photofunctionalization reduced the hydrocarbon concentration and enhanced human blood plasma proteins and albumin adsorption mainly for the PEO-treated surface (p < 0.05). PEO UV+ also maintained higher wettability values for a longer period and provided microbial reduction at 1 h of bacterial adhesion (p = 0.012 vs. PEO UV-). Photofunctionalization did not increase the photocatalytic activity of Ti (p > 0.05). Confocal microscopy analyses demonstrated that PEO UV+ had no cell damage effect on HGF cells growth even after 24 h of incubation. The photofunctionalization of a biofunctional PEO coating seems to be a promising alternative for dental implants as it increases blood plasma proteins adsorption, reduces initial bacterial adhesion and presents no cytotoxicity effect.

Targeting Pathogenic Biofilms: Newly Developed Superhydrophobic Coating Favors a Host-Compatible Microbial Profile on the Titanium Surface.

Polymicrobial infections are one of the most common reasons for inflammation of surrounding tissues and failure of implanted biomaterials. Because microorganism adhesion is the first step for biofilm formation, physical-chemical modifications of biomaterials have been proposed to reduce the initial microbial attachment. Thus, the use of superhydrophobic coatings has emerged because of their anti-biofilm properties. However, these coatings on the titanium (Ti) surface have been developed mainly by dual-step surface modification techniques and have not been tested using polymicrobial biofilms. Therefore, we developed a one-step superhydrophobic coating on the Ti surface by using a low-pressure plasma technology to create a biocompatible coating that reduces polymicrobial biofilm adhesion and formation. The superhydrophobic coating on Ti was created by the glow discharge plasma using Ar, O2, and hexamethyldisiloxane gases, and after full physical, chemical, and biological characterizations, we evaluated its properties regarding oral biofilm inhibition. The newly developed coating presented an increased surface roughness and, consequently, superhydrophobicity (contact angle over 150°) and enhanced corrosion resistance (p < 0.05) of the Ti surface. Furthermore, proteomic analysis showed a unique pattern of protein adsorption on the superhydrophobic coating without drastically changing the biologic processes mediated by proteins. Additionally, superhydrophobic treatment did not present a cytotoxic effect on fibroblasts or reduction of proliferation; however, it significantly reduced (≈8-fold change) polymicrobial adhesion (bacterial and fungal) and biofilm formation in vitro. Interestingly, superhydrophobic coating shifted the microbiological profile of biofilms formed in situ in the oral cavity, reducing by up to ≈7 fold pathogens associated with the peri-implant disease. Thus, this new superhydrophobic coating developed by a one-step glow discharge plasma technique is a promising biocompatible strategy to drastically reduce microbial adhesion and biofilm formation on Ti-based biomedical implants.

Role of the Plasma Activation Degree on Densification of Organosilicon Films.

The possibility of controlling the density of organosilicon films was investigated by tuning the plasma activation degree without providing extra energy to the structure, as usually reported in the literature. For this purpose, thin films were deposited in plasmas fed with hexamethyldisiloxane/Ar mixtures at a total pressure of 9.5 Pa. The power of the radiofrequency excitation signal, P, ranged from 50 to 300 W to alter the average energy of the plasma species while the electrical configuration was chosen to avoid direct ion bombardment of the growing films. In this way, it was possible to evaluate the effect of P on the film properties. Thickness and deposition rate were derived from profilometry data. X-ray energy dispersive and infrared spectroscopies were, respectively, applied to analyze the chemical composition and molecular structure of the layers. Surface topography and roughness were determined by atomic force microscopy while nanoindentation was used to evaluate the mechanical properties of the films. From electrochemical impedance spectroscopy the total resistance to the flow of electrolyte species was derived. The main alteration observed in the structure with changing P is related to the proportion of the methyl functional which remains connected to the Si backbone. Chain crosslinking and film density are affected by this structural modification induced by homogeneous and heterogeneous plasma reactions. The density increase resulted in a film with hardness comparable to that of the silica and more resistant to the permeation of oxidative species, but preserving the organosilicon nature of the structure.

Visible-Light-Induced Photocatalytic and Antibacterial Activity of TiO2 Codoped with Nitrogen and Bismuth: New Perspectives to Control Implant-Biofilm-Related Diseases.

Biofilm-associated diseases are one of the main causes of implant failure. Currently, the development of implant surface treatment goes beyond the osseointegration process and focuses on the creation of surfaces with antimicrobial action and with the possibility to be re-activated (i.e., light source activation). Titanium dioxide (TiO2), an excellent photocatalyst used for photocatalytic antibacterial applications, could be a great alternative, but its efficiency is limited to the ultraviolet (UV) range of the electromagnetic spectrum. Since UV radiation has carcinogenic potential, we created a functional TiO2 coating codoped with nitrogen and bismuth via the plasma electrolytic oxidation (PEO) of titanium to achieve an antibacterial effect under visible light with re-activation potential. A complex surface topography was demonstrated by scanning electron microscopy and three-dimensional confocal laser scanning microscopy. Additionally, PEO-treated surfaces showed greater hydrophilicity and albumin adsorption compared to control, untreated titanium. Bismuth incorporation shifted the band gap of TiO2 to the visible region and facilitated higher degradation of methyl orange (MO) in the dark, with a greater reduction in the concentration of MO after visible-light irradiation even after 72 h of aging. These results were consistent with the in vitro antibacterial effect, where samples with nitrogen and bismuth in their composition showed the greatest bacterial reduction after 24 h of dual-species biofilm formation ( Streptococcus sanguinis and Actinomyces naeslundii) in darkness with a superior effect at 30 min of visible-light irradiation. In addition, such a coating presents reusable photocatalytic potential and good biocompatibility by presenting a noncytotoxicity effect on human gingival fibroblast cells. Therefore, nitrogen and bismuth incorporation into TiO2 via PEO can be considered a promising alternative for dental implant application with antibacterial properties in darkness, with a stronger effect after visible-light application.

Proteome analysis of the salivary pellicle formed on titanium alloys containing niobium and zirconium.

The chemical composition of biomaterials can drive their biological responses; therefore, this in vitro study aimed to evaluate the proteomic profile of the salivary pellicle formed on titanium (Ti) alloys containing niobium (Nb) and zirconium (Zr). The experimental groups consisted of Ti35NbxZr (x = 5 and 10 wt%) alloys, and commercially pure titanium (cpTi); titanium aluminium vanadium (Ti6Al4V) alloys were used as controls. The physical and chemical characteristics of the Ti materials were analysed. The proteomic profile was evaluated by liquid chromatography coupled with tandem mass spectrometry. Bacterial adhesion (2 h) of mixed species (Streptococcus sanguinis and Actinomyces naeslundii) was investigated as colony-forming units (n = 6). This paper reports the finding that salivary pellicle composition can be modulated by the composition of the Ti material. The Ti35NbxZr group showed a significant ability to adsorb proteins from saliva, which can favour interactions with cells and compatibility with the body.

Characterisation of a new plasma-enhanced film to improve shear bond strength between zirconia and veneering ceramic.

The aim of this study was to develop and characterise a new plasma-enhanced chemical vapor deposition (PECVD) film for improving shear bond strength (SBS) between yttria-stabilised tetragonal zirconia (Y-TZP) and veneering ceramic. In total, 192 Y-TZP samples (13 × 5.4 × 5 mm) were divided into 6 groups: control - no treatment (C), airborne-particle abrasion with 27 µm aluminum oxide particles (Al27), 110 µm aluminum oxide particles (Al110), and 250 µm aluminum oxide particles (Al250), application of liner for zirconia (L) and the PECVD film application (P). The Y-TZP surface was characterised by means of Scanning Electronic Microscopy (SEM), Energy-dispersive Spectroscopy (EDS), atomic force microscopy (AFM), surface profilometry and surface-free energy (SFE). SBS between Y-TZP and veneering ceramic was tested before and after thermocycling (20,000 cycles of 5 and 55 °C), and failure mode was also evaluated. Data were analysed by ANOVA and Tukey's HSD test (α = 0.05). Data analysis showed that PECVD film had no effect on surface roughness of Y-TZP (p > 0.05 vs control), whilst the other groups presented higher roughness values (p < 0.05). All treatments increased SFE, except the Al27 group. The highest SBS was presented by the P group (p < 0.05), and values were similar to those of the Al27 group (p = 0.107). Mixed failures were prevalent in all groups, and premature failures were found only in Al groups after thermocycling. Whilst PECVD treatment did not affect Y-TZP surface roughness, high SBS between Y-TZP and the veneering layer was observed. Therefore, PECVD treatment is a promising alternative to improve the performance of bi-layer zirconia-based restorations.

Characterization of chemically treated Ti-Zr system alloys for dental implant application.

Materials and surfaces developed for dental implants need to withstand degradation processes that take place in the oral cavity. Therefore, the aim of the study was to develop and evaluate the topographical, mechanical, chemical, electrochemical and biological properties of Ti-xZr alloys (x = 5, 10, and 15 wt%) with two surface features (machined and double acid etched). Commercially pure titanium (cpTi) and Ti-6Al-4V alloy were used as controls. Surface characterization was performed using dispersive energy spectroscopy, X-ray diffraction, scanning electron microscopy, atomic force microscopy, profilometry and surface energy. The mechanical properties were assessed using Vickers microhardness, elastic modulus and stiffness. The electrochemical behavior analysis was conducted in a body fluid solution (pH 7.4). In addition, MC3T3-E1 cells were used to determine the impact of material and surface treatment on cell morphology by SEM analysis. Data were analyzed by two-way ANOVA and Bonferroni test (α = 0.05). Ti-Zr alloys showed lower surface roughness, elastic modulus and stiffness, as well as higher hardness and surface energy when compared to cpTi. Ti-Zr system increased the polarization resistance values and significantly decreased the capacitance, corrosion current density (icorr), and passivation current density (ipass) values. The acid treatment increased the resistance and corrosion potential of the oxide layer. SEM data analysis demonstrated that Ti-Zr alloys displayed normal cell attachment/spreading and slightly changed cell morphology in the double etched surface. In conclusion, Zr addition and surface treatment altered surface, mechanical, biological and electrochemical properties of Ti material.

Antibacterial photocatalytic activity of different crystalline TiO2 phases in oral multispecies biofilm.

OBJECTIVE: Titanium dioxide (TiO2) incorporation in biomaterials is a promising technology due to its photocatalytic and antibacterial activities. However, the antibacterial potential of different TiO2 crystalline structures on a multispecies oral biofilm remains unknown. We hypothesized that the different crystalline TiO2 phases present different photocatalytic and antibacterial activities. METHODS: Three crystalline TiO2 films were deposited by magnetron sputtering on commercially pure titanium (cpTi), in order to obtain four groups: (1) machined cpTi (control); (2) A-TiO2 (anatase); (3) M-TiO2 (mixture of anatase and rutile); (4) R-TiO2 (rutile). The morphology, crystalline phase, chemical composition, hardness, elastic modulus and surface free energy of the surfaces were evaluated. The photocatalytic potential was assessed by methylene blue degradation assay. The antibacterial activity was evaluated on relevant oral bacteria, by using a multispecies biofilm (Streptococcus sanguinis, Actinomyces naeslundii and Fusobacterium nucleatum) formed on the treated titanium surfaces (16.5h) followed by UV-A light exposure (1h) to generate reactive oxygen species production. RESULTS: All TiO2 films presented around 300nm thickness and improved the hardness and elastic modulus of cpTi surfaces (p<0.05). A-TiO2 and M-TiO2 films presented superior photocatalytic activity than R-TiO2 (p<0.05). M-TiO2 revealed the greatest antibacterial activity followed by A-TiO2 (≈99.9% and 99% of bacterial reduction, respectively) (p<0.001 vs. control). R-TiO2 had no antibacterial activity (p>0.05 vs. control). SIGNIFICANCE: This study brings new insights on the development of extra oral protocols for the photocatalytic activity of TiO2 in oral biofilm-associated disease. Anatase and mixture-TiO2 showed antibacterial activity on this oral bacterial biofilm, being promising surface coatings for dental implant components.

Development of binary and ternary titanium alloys for dental implants.

OBJECTIVE: The aim of this study was to develop binary and ternary titanium (Ti) alloys containing zirconium (Zr) and niobium (Nb) and to characterize them in terms of microstructural, mechanical, chemical, electrochemical, and biological properties. METHODS: The experimental alloys - (in wt%) Ti-5Zr, Ti-10Zr, Ti-35Nb-5Zr, and Ti-35Nb-10Zr - were fabricated from pure metals. Commercially pure titanium (cpTi) and Ti-6Al-4V were used as controls. Microstructural analysis was performed by means of X-ray diffraction and scanning electron microscopy. Vickers microhardness, elastic modulus, dispersive energy spectroscopy, X-ray excited photoelectron spectroscopy, atomic force microscopy, surface roughness, and surface free energy were evaluated. The electrochemical behavior analysis was conducted in a body fluid solution (pH 7.4). The albumin adsorption was measured by the bicinchoninic acid method. Data were evaluated through one-way ANOVA and the Tukey test (α=0.05). RESULTS: The alloying elements proved to modify the alloy microstructure and to enhance the mechanical properties, improving the hardness and decreasing the elastic modulus of the binary and ternary alloys, respectively. Ti-Zr alloys displayed greater electrochemical stability relative to that of controls, presenting higher polarization resistance and lower capacitance. The experimental alloys were not detrimental to albumin adsorption. SIGNIFICANCE: The experimental alloys are suitable options for dental implant manufacturing, particularly the binary system, which showed a better combination of mechanical and electrochemical properties without the presence of toxic elements.

Deciphering pyritization-kerogenization gradient for fish soft-tissue preservation.

Soft-tissue preservation provides palaeobiological information that is otherwise lost during fossilization. In Brazil, the Early Cretaceous Santana Formation contains fish with integument, muscles, connective tissues, and eyes that are still preserved. Our study revealed that soft-tissues were pyritized or kerogenized in different microfacies, which yielded distinct preservation fidelities. Indeed, new data provided the first record of pyritized vertebrate muscles and eyes. We propose that the different taphonomic pathways were controlled by distinct sedimentation rates in two different microfacies. Through this process, carcasses deposited in each of these microfacies underwent different residence times in sulphate-reduction and methanogenesis zones, thus yielding pyritized or kerogenized soft-tissues, and a similar process has previously been suggested in studies of a late Ediacaran lagerstätte.

Three-species biofilm model onto plasma-treated titanium implant surface.

In this study, titanium (Ti) was modified with biofunctional and novel surface by micro-arc oxidation (MAO) and glow discharge plasma (GDP) and we tested the development of a three-species periodontopatogenic biofilm onto the treated commercially-pure titanium (cpTi) surfaces. Machined and sandblasted surfaces were used as control group. Several techniques for surface characterizations and monoculture on bone tissue cells were performed. A multispecies biofilm composed of Streptococcus sanguinis, Actinomyces naeslundii and Fusobacterium nucleatum was developed onto cpTi discs for 16.5h (early biofilm) and 64.5h (mature biofilm). The number of viable microorganisms and the composition of the extracellular matrix (proteins and carbohydrates) were determined. The biofilm organization was analyzed by scanning electron microscopy (SEM) and Confocal laser scanning microscopy (CLSM). In addition, MC3T3-E1 cells were cultured on the Ti surfaces and cell proliferation (MTT) and morphology (SEM) were assessed. MAO treatment produced oxide films rich in calcium and phosphorus with a volcano appearance while GDP treatment produced silicon-based smooth thin-film. Plasma treatments were able to increase the wettability of cpTi (p<0.05). An increase of surface roughness (p<0.05) and formation of anatase and rutile structures was noted after MAO treatment. GDP had the greatest surface free energy (p<0.05) while maintaining the surface roughness compared to the machined control (p>0.05). Plasma treatment did not affect the viable microorganisms counts, but the counts of F. nucleatum was lower for MAO treatment at early biofilm phase. Biofilm extracellular matrix was similar among the groups, excepted for GDP that presented the lowest protein content. Moreover, cell proliferation was not significantly affected by the experimental, except for MAO at 6days that resulted in an increased cell proliferative. Together, these findings indicate that plasma treatments are a viable and promising technology to treat bone-integrated dental implants as the new surfaces displayed improved mechanical and biological properties with no increase in biofilm proliferation.