Engineering Precise Interconnected Porosity in ß-Tricalcium Phosphate (ß-TCP) Matrices by Means of Top-Down Digital Light Processing.

This study evaluated the biocompatibility and accuracy of 3D-printed ß-tricalcium phosphate (ß-TCP) pure ceramic scaffolds. A specific shaping process associating a digital light processing (DLP) 3D printer and a heat treatment was developed to produce pure ß-TCP scaffolds leaving no polymer binder residue. The ß-TCP was characterised using X-ray diffraction, infrared spectroscopy and the detection of pollutants. The open porosity of produced matrices and their resorption were studied by hydrostatic weighing and calcium release measures. The biocompatibility of the printed matrices was evaluated by mean of osteoblast cultures. Finally, macroporous cubic matrices were produced. They were scanned using a micro-Computed Tomography scanner (micro-CT scan) and compared to their numeric models. The results demonstrated that DLP 3D printing with heat treatment produces pure ß-TCP matrices with enhanced biocompatibility. They also demonstrated the printing accuracy of our technique, associating top-down DLP with the sintering of green parts. Thus, this production process is promising and will enable us to explore complex phosphocalcic matrices with a special focus on the development of a functional vascular network.

Effect of dimensional variations on the manufacturing process and the 3D shrinkage ratio of stereolithographic dental alumina ceramics.

The purpose of the present study was to evaluate the influence of height and length variations of alumina ceramics manufactured by stereolithography on deformations caused by the manufacturing process and on the 3D shrinkage ratio to control the final dimensions and improve the adaptation of stereolithographic ceramic dental prostheses. Two different U-shaped models were designed with variable heights or lengths. The specimens were manufactured by stereolithography and were scanned using a microtomographic device before and after the heat treatment. Dimensional variations were measured using inspection software. The number and surface of layers of alumina ceramic influenced the reliability of the stereolithography manufacturing but did not influence the 3D shrinkage ratio. The larger the layer surface, the larger the deformation of the ceramic. Dental ceramics manufactured by stereolithography with smallest layer surface are the most reliable. This helps in the selection of the build orientation.

Polydopamine treatment of chitosan nanofibers for the conception of osteoinductive scaffolds for bone reconstruction.

We report the influence of treatment time of electrospun chitosan nanofibers (CHT NFs) in dopamine hydrochloride bath (2 mg.mL-1 in 10 mM Tris buffer, pH 8.5) on the extent of the polydopamine (pDA) coating on NFs surface. The reaction was characterized by FTIR and SEM analysis and the cytocompatibility of the scaffolds toward MT3C3-E1 cells was assessed. Biomimetic deposition of hydroxyapatite (HA) in 1.5xSBF batch was investigated by SEM-EDS and XRD. Samples treated in dopamine bath during 2 h promoted the structural stability of NFs in PBS, provided optimal cytocompatibility and induced the in vitro biomineralization from 6 days in 1.5xSBF. The XRD and SEM-EDS investigations confirmed formation of spherical-shaped particles composed of apatitic phase. Finally, this study shows that these NFs-pDA scaffolds prepared in the optimal experimental conditions defined here are promising candidates for application as osteoinductive scaffolds for bone regeneration applied to orthopedic and dental applications.

Effect of build orientation on the manufacturing process and the properties of stereolithographic dental ceramics for crown frameworks.

STATEMENT OF PROBLEM: Stereolithography (SLA) ceramic crown frameworks are suitable for clinical use, but the impact of SLA build orientation has not been identified. PURPOSE: The purpose of this in vitro study was to investigate the effect of 3 build orientations on the physical and mechanical properties and the microstructure of SLA alumina dental ceramics. MATERIAL AND METHODS: The physical and mechanical properties and microstructures of 3 different oriented SLA alumina ceramics (ZX, ZY, and XY) were evaluated by visual observation, hydrostatic weighing (n=10/group), Weibull analyses (n=30/group), scanning electron microscopy, 3-point flexural strength (n=30/group), fracture toughness (indentation, single-edge-V-notched-beam) (n=4/group), and Vickers hardness (n=15/group) testing. The hydrostatic weighing, 3-point flexural strength, fracture toughness, and Vickers hardness testing data were statistically analyzed (α=.05). RESULTS: The minimum resting period of slurries between the polymerization of 2 layers was shorter for the ZY- and ZX-oriented specimens and increased with the layer surface. The density and Vickers hardness of the SLA-manufactured specimens were similar for all groups (P>.05). The 95% confidence intervals of the Weibull moduli of the ZX- and ZY-oriented specimens were higher than that of the XY-oriented specimens, with no overlap fraction. The ZY-oriented specimens displayed significantly higher 3-point flexural strength (P<.05) and fracture toughness as evaluated by the single-edge-V-notched-beam method than the ZX-oriented specimens (P<.05). They also displayed significantly higher 3-point flexural strength than the XY-oriented specimens (P<.05). The microstructural analysis showed that the texturing was heterogeneous and that the major axis of the large grains of alumina ran parallel to the orientation of the layers. CONCLUSIONS: The ZY orientation produced a reliable dental ceramic by SLA, with the shortest general manufacturing time and the highest mechanical strength when the layers were perpendicular to the test load surface.

A Localized Phage-Based Antimicrobial System: Effect of Alginate on Phage Desorption from ß-TCP Ceramic Bone Substitutes.

Tricalcium phosphate (TCP) is a prosthetic material commonly used as a bone substitute to repair osteoarticular diseases and injuries. In this type of bone reconstruction surgery, antibiotics remain the common preventive and therapeutic treatment for bacterial infection. Nevertheless, the emergence of multi-resistant strains requires complimentary or alternative treatments. Today, one of the promising alternative approaches is phage therapy. Phages are bacterial viruses that have several advantages over chemotherapy, such as the specificity of bacterial strain, the absence of side effects, and a rapid response. In this work, we studied the impact of alginate hydrogels for overlaying λvir-phage-loaded ß-TCP ceramic bone substitutes, delaying the phage desorption. The results show that the use of a 1% alginate-CaCl2 hydrogel overlapping the ß-TCP ceramic pellets leads to higher initial phage concentration on the material and extends the released time of phages to two weeks when compared with control pellets. These alginate-coated biomaterials also generate faster bacterial lysis kinetics and could therefore be a good practical prosthetic device for bone and joint surgeries by allowing local treatment of bacterial infections with phage therapy for a longer period of time.

3D printed bioceramic for phage therapy against bone nosocomial infections.

This study provides a new therapeutic response to postoperative joint and bone infections. Alone or in combination with antibiotics, phage therapy has many advantages, including accurate targeting of pathogenic bacteria. In addition, a decrease in harmful side effects can improve the healing process. Integrating the bacteriophage directly into the graft product will improve the antibacterial spread over the site of the surgery. The phage cocktail-filled ceramics are an innovative device for localized and curative phage therapy (in prosthetic replacement surgery, for example) in bone and joint surgery. Calcium phosphate-based ceramics were synthesized and shaped by stereolithography (3D) before loading by a phage cocktail to lyse a heterospecific bacterial population. In addition, the device makes possible the protection of osteoblastic cells against Staphylococcus aureus infection during their colonization on the ceramic material and prevents the formation of biofilm on the surface of biomaterials.

Stereolithography: A new method for processing dental ceramics by additive computer-aided manufacturing.

OBJECTIVES: The aim of this study was to compare the physical and mechanical properties of stereolithography (SLA)- manufactured alumina ceramics of different composition to those of subtractive- manufactured ceramics and to produce suitable dental crown frameworks. METHODS: The physical and mechanical properties of a control and six experimental SLA ceramics prepared from slurries with small (S) and large (L) particles (0.46±0.03 and 1.56±0.04µm, respectively) and three dry matter contents (70%, 75%, 80%) were evaluated by dynamic rheometry, hydrostatic weighing, three3-point flexural strength measurements, and Weibull analyses, and by the micrometrics measurement of shrinkage ratio before and after the heat treatments. RESULTS: S75 was the only small particle slurry with a significantly higher viscosity than L70. The viscosity of the S80 slurry made it impossible to take rheological measurements. The viscosities of the S75 and S80 slurries caused deformations in the printed layers during SLA manufacturing and were excluded from further consideration. SLA samples with low dry matter content had significantly lower and densityflexural strengths. Only SLA samples with a large particle size and high dry matter content (L75 and L80) were similar in density and flexural strength to the subtractive- manufactured samples. The 95% confidence intervals of the Weibull modulus of the L80 ceramic were higher (no overlap fraction) than those of the L75 ceramic and were similar to the control (overlap fraction). The Weibull characteristics of L80 ceramic were higher than those of L75 ceramic and the control. SLA can be used to process suitable crown frameworks but shows results in anisotropic shrinkage. SIGNIFICANCE: The hH High particle size and dry matter content of the L80 slurry allowed made it possible to produce a reliable ceramic by SLA manufacturing with an anisotropic shrinkage, and a density, and flexural strength similar to those of a subtractive-manufactured ceramic. SLA allowed made it possible to build up a dense 3D alumina crown framework with controlled shape. Further studies on the marginal adaptation and shrinkage model of alumina crown frameworks will be required to optimize the process.

Optimization and Preclinical Perception of an Artificial Simulator for Endodontic Training: A Preliminary Study.

The aim of this study was to evaluate the performance of ceramic, hybrid ceramic, and commercial plastic bloc root canal simulator (RCS) as preclinical training aids in the learning phase of endodontic treatments. A previously developed hydroxyapatite ceramic RCS was improved by adding epoxy resin to the ceramic matrix to more closely mimic the organic phase of dentin and to simulate the clinical situation as realistically as possible. The sintered hydroxyapatite ceramic RCS was vacuum infiltrated with epoxy resin, and the degree of infiltration was evaluated by methylene blue staining. The suitability of the resin-infiltrated ceramic simulator (CR) for preclinical endodontic training was compared to that of a non-infiltrated ceramic simulator (C) and a commercial epoxy bloc (P) using a cohort of 30 dental students at one dental school in France. The study was conducted in 2016. The students' perceptions following the required exercises using the CR, C, and P were scored using a questionnaire. The learning outcomes were also assessed by examining the canal preparations that the students performed on extracted teeth using a master cone try-in test. The vacuum process resulted in a good degree of resin infiltration into the ceramic. The questionnaire showed that the C and CR groups generally reported greater satisfaction, especially for radiographic visualizations, than the P group. The CR group had a higher score than the P group for tactile sensation. There was no significant difference among the three groups with respect to the canal preparations using extracted teeth. Resin infiltration improved the performance of the ceramic RCS, especially with respect to perception during root canal instrumentation. A larger scale student training investigation and an assessment by experienced endodontists are required to validate the model.

Influence of Root Canal Curvature on Wall Cleanliness in the Apical Third during Canal Preparation.

OBJECTIVE: The aim of this study was to evaluate the influence of root canal curvature (curved and straight root canals), prepared using reciprocating and rotary files, on wall cleanliness during root canal treatments. METHODS: Thirty curved and 30 straight human root canals were prepared using ProTaper (Dentsply), Pro- Taper Next (Dentsply) and Reciproc files (Dentsply) (n=20/group). The roots were split longitudinally and observed using a scanning electron microscope. Six micrographs were obtained at 1, 3 and 5 mm from the working length (WL). Two blinded observers scored the amount of debris. Mean debris scores were compared using a non-parametric the Mann-Whitney U test or Kruskal-Wallis test, and a Bonferroni correction was used for multiple comparisons. RESULTS: Considering all the shaping systems together, the debris scores were lower in curved root canals (P<0.05). Reciproc and ProTaper Next performed better than ProTaper in straight canals (P<0.05). No difference was found between ProTaper Next and Reciproc regardless of the canal curvature or distance from the WL. Considering all the shaping systems together, cleanliness increased when pulling away from the WL. CONCLUSION: The anatomical configuration of the root canal influences the quality of cleaning by shaping instruments regardless of the instrument kinematics during endodontic procedures. In every circumstance, the last millimetres of the apical third remain the most difficult area to clean.

Surrogate Outcome Measures of In Vitro Osteoclast Resorption of ß Tricalcium Phosphate.

Introduction of porosity to calcium phosphate scaffolds for bone repair has created a new challenge when measuring bioresorption in vitro, rendering traditional outcome measures redundant. The aim of this study is to identify a surrogate endpoint for use with 3D scaffolds. Murine RAW 264.7 cells are cultured on dense discs of ß-tricalcium phosphate in conditions to stimulate osteoclast (OC) formation. Multinucleated OCs are visible from day 6 with increases at days 8 and 10. Resorption pits are first observed at day 6 with much larger pits visible at days 8, 10, and 12. The concentration of calcium ions in the presence of cells is significantly higher than cell-free cultures at days 3 and 9. Using linear regression analysis, Ca ion release could account for 35.9% of any subsequent change in resorption area. The results suggest that Ca ion release is suitable to measure resorption of a beta-tricalcium phosphate ceramic substrate in vitro. This model could replace the more accepted resorption pit assay in circumstances where quantification of pits is not possible, e.g., when characterizing 3D tissue engineered bone scaffolds.

Influence of heat and ultrasonic treatments on the setting and maturation of a glass-ionomer cement.

PURPOSE: To compare the effect of different treatments (heat capsule, ultrasound, and dual treatments) on the setting kinetics and maturation properties of a conventional GIC (EQUIA, GC) to that of standard setting. METHODS: The optimal durations of the heat and ultrasonic treatments were determined by monitoring changes in the COO-/COOH ratio, surface hardness, and temperature within the samples. The influence of optimal treatments on the maturation properties of the GIC (microhardness, and 3-point flexural strength) were assessed using GIC samples incubated in artificial saliva for 24 hours, 1 month, and 3 months. RESULTS: The optimal durations of the heat and ultrasonic treatments for accelerating setting were 5 minutes and 35 seconds, respectively. The dual treatment using the optimal conditions of the individual treatments further enhanced the setting kinetics. A temperature peak (49°C) within the GIC was detected during setting. Only the dual treatment increased the mechanical properties of the GIC after 24 hours compared to the control, while no significant difference was observed after 1 and 3 months.

Comparison of the Weibull characteristics of hydroxyapatite and strontium doped hydroxyapatite.

The effects of two strontium (Sr) additions, 5% and 10% of the total calcium (Ca) content, on the phase assemblage and Weibull statistics of hydroxyapatite (HA) are investigated and compared to those of undoped HA. Sintering was carried out in the range of 900-1200 °C in steps of 1000 °C in a conventional furnace. Sr content had little effect on the mean particulate size. Decomposition of the HA phase occurred with Sr incorporation, while ß-TCP stabilization was shown to occur with 10% Sr additions. Porosity in both sets of doped samples was at a comparable level to porosity in the undoped HA samples, however the 5% Sr-HA samples displayed the greatest reduction in porosity with increasing temperature while the porosity of the 10% Sr-HA samples remain relatively constant over the full sintering temperature range. The undoped HA samples displayed the greatest Weibull strengths and the porosity was determined to be the major controlling factor. However, with the introduction of decompositional phases in the Sr-HA samples, the dependence of strength on porosity is reduced and the phase assemblage becomes the more dominant factor for Weibull strength. The Weibull modulus is relatively independent of the porosity in the undoped HA samples. The 5% Sr-HA samples experience a slight increase in Weibull modulus with porosity, indicating a possible relationship between the parameters. However the 10% Sr-HA samples show the highest Weibull modulus with a value of approximately 15 across all sintering temperatures. It is postulated that this is due to the increased amount of surface and lattice diffusion that these samples undergo, which effectively smooths out flaws in the microstructure, due to a saturation of Sr content occurring in grain boundary movement.

New antibacterial microporous CaP materials loaded with phages for prophylactic treatment in bone surgery.

Hydroxyapatite and beta-tricalcium phosphate (ß-TCP) are materials commonly used in bone repair. The most important problem occurring in bone repair surgery is bacterial infection which is usually overcome by treatment with antibiotics. Currently, emergence of multidrug resistant strains has led to development of alternative treatments such as phage therapy. Phages are bacterial viruses with several advantages over chemotherapy such as specificity of bacterial strain, no side effects and fast response. This study evaluates the possibility of loading hydroxyapatite and ß-tricalcium phosphate ceramics used as bone substitutes with phages and their antibacterial activity against Escherichia coli K12. The majority of phages were retained in dense and microporous HA and ß-TCP samples during at least 6 days suggesting the occurrence of strong interaction between phages and ceramics, which did not prevent bacterial attachment and lysis. This study has shown for the first time that phage loaded ceramics could be used in prophylactic treatments.

Molecular interactions between zoledronic acid and bone: An in vitro Raman microspectroscopic study.

The aim of this study was to investigate molecular interactions between a bisphosphonate (BP), zoledronic acid, and bone tissue by the use of Raman microspectroscopy. In this way, samples of hydroxyapatite (HA), as a bone model, and Wistar rat femurs were soaking in zoledronic acid solutions. Sample surfaces were studied by Environmental Scanning Electron Microscopy and Raman spectroscopy. The amount of zoledronic acid incorporated onto the samples and the inorganic phosphate released in solution were determined by (31)P NMR spectroscopy. Total carbonate content in solution was evaluated by inorganic carbon analyser. After impregnation new Raman bands with frequencies close to characteristic peaks of zoledronic acid (in particular phosphate moieties and imidazole ring of the R2 side-chain) were observed on both types of samples. Physico-chemical parameters of the bone were also significantly modified (P<0.0001). The mineral to organic ratio and the carbonate to phosphate ratio decreased and the crystallinity increased. Released inorganic phosphate and carbonate were detected in the solutions. The Raman shift of the bands corresponding to the phosphate groups and the imidazole ring of the BP highlight their implication in the binding to the mineral. The detection of released inorganic phosphate and carbonate in solution, the modifications of the mineral to phosphate ratio and the carbonate to phosphate ratio reveal that BP decrease the amount of inorganic phosphate and limit the dissolution of bone mineral. The increase of the crystallinity after BP binding shows a re-organisation of the lattice with a higher symmetry. Thus, it seems that zoledronic acid has an important contribution on the increase of crystallinity. The use of Raman spectrometry brings new and complementary information on the impact of zoledronic acid on bone composition at molecular level. Raman spectrometry could help to understand by which way BPs improve bone strength and decrease fracture risk.

Prolonged local antibiotics delivery from hydroxyapatite functionalised with cyclodextrin polymers.

Per-operative infection is a common complication for bone-graft surgery. Combining antiseptic agents with graft materials may offer a solution by increasing local drug concentration at target sites. Aiming to achieve a sustained local antibiotic (ATB) delivery for a widely applied bone substitute material - hydroxyapatite (HA), we attempted incorporating hydroxypropyl-beta-cyclodextrin polymer (polyHPbetaCD) into microporous HA via impregnating either in a CD monomers mixture solution or a pre-synthesized CD polymer solution, followed by thermal fixation processing. In such functionalised material (CD-HA), polyHPbetaCD could entrap ATBs and release them progressively. Infrared-spectroscopic analysis confirmed the presence of polyHPbetaCD in functionalised HA via both processing pathways; polyHPbetaCD functionalisation yields were quantitated by thermogravimetric analysis for optimising the processing regime. Ciprofloxacin (CFX) and vancomycin (VCM), commonly applied in orthopaedics, have been respectively loaded on CD-HA by dip-coating. For both ATBs, kinetic release test in phosphate buffered saline showed significantly increased initial-burst amount and prolonged release from CD-HA compared with those from non-functionalised HA. Encouragingly, ATBs loaded CD-HA also revealed a prolonged bacteriostatic activity against Staphylococcus aureus and progressively increased cytocompatibility to osteoblasts (MC3T3-E1). Overall, polyHPbetaCD functionalisation on HA could be an effective drug-delivery model for loading different drug molecules in prevention of infection.

The corrosion and biological behaviour of titanium alloys in the presence of human lymphoid cells and MC3T3-E1 osteoblasts.

Corrosion behaviour of biomedical alloys is generally determined in mineral electrolytes: unbuffered NaCl 0.9% (pH 7.4) or artificial saliva (pH 6.8). The assays with exclusive utilization of these electrolytes are of low relevance for the biological condition, to which the alloys will be exposed once implanted in the human organism. As an approach to the biological situation regarding the interaction of proteins, electrolytes and metals, we added the RPMI cell culture medium containing foetal calf serum as a biological electrolyte (pH 7.0). The analysis of corrosion behaviour was also performed in the presence of human lymphoid cells (CEM). The rest potential (Er) and the global polarization were determined on cp-Ti, micro-arc oxidized cp-Ti (MAO-Ti), four different Ti-alloys (Ti6Al4V, Ti12Zr, Ti(AlMoZr), Ti(NbTaZr)) and 316L stainless steel. The 316L exhibited an appropriate Er and a good passive current density (Ip), but a high corrosion potential (Ec) and a very low breakdown potential (Eb) in all electrolytes. All Ti-alloys exhibited a much better electrochemical behaviour: better Er and Ec and very high Eb. No significant differences of the above parameters existed between the Ti-alloys, except for Zr-containing alloys that showed better corrosion behaviour. A remarkable difference, however, was stated with respect to the electrolytes. NaCl 0.9% induced strong variations between the Ti-alloys. More homogeneous results were obtained with artificial saliva and RPMI medium, which induced a favourable Ec and an increased Ip. The presence of cells further decreased these values. The unbuffered NaCl solution seems to be less appropriate for the analysis of corrosion of metals. Additional in vitro biological assessments with CEM cell suspensions and MC3T3-E1 osteoblasts confirmed the advantages of the Ti(AlMoZr) and Ti(NbTaZr) alloys with an improved cell proliferation and vitality rate.

beta-TCP microporosity decreases the viability and osteoblast differentiation of human bone marrow stromal cells.

Association of osteoprogenitor cells to calcium phosphate ceramics is currently under intense investigation, for its considered ability to induce bone formation and therefore to allow the successful repair of large bone defects. However, if the first experimental and clinical studies provided promising results, lack of new bone formation has been reported in a large number of animal experiments. In this context and since it has been reported that in some conditions, calcium phosphate ceramic microstructure induces ectopic bone formation, we investigated the effects of ceramic microporosity on the behavior of osteoprogenitor cells for the development of hybrid materials. Human bone marrow stromal cells (BMSCs) were seeded on beta-tricalcium phosphate (TCP) ceramics with 0, 25, or 45% microporosity and cell adhesion, viability, and osteoblastic differentiation have been studied for three weeks. Cell counts, measurement of MTS conversion, and LDH activity indicated that microporosity decreased the viability of BMSCs in a time and rate-dependent manner. In addition, microporosity inhibited osteoblastic differentiation as compared with nonmicroporous ceramics, as revealed by decreased alkaline phosphatase activity and osteocalcin secretion. Results of this in vitro study therefore highlight a negative role for beta-TCP microporosity in the behavior of human osteoprogenitor cells.