The effect of hydrogen peroxide concentration on metal ion release from dental casting alloys.

There are concerns that tooth bleaching agents may adversely affect dental materials. The aim of this study was to test the hypothesis that increasing concentrations of hydrogen peroxide (HP) are more effective than water at increasing metal ion release from two typical dental casting alloys during bleaching. Discs (n = 28 for each alloy) were prepared by casting and heat treated to simulate a typical porcelain-firing cycle. Discs (n = 7) of each alloy were immersed in either 0%, 3%, 10% or 30% (w/v) HP solutions for 24 h at 37 degrees C. Samples were taken for metal ion release determination using inductively coupled plasma-mass spectrometry and the data analysed using a two-way anova followed by a one-way anova. The surface roughness of each disc was measured using a Talysurf contact profilometer before and after bleaching and the data analysed using a paired t-test. With the exception of gold, the differences in metal ion concentration after treatment with 0% (control) and each of 3%, 10% and 30% HP (w/v) were statistically significant (P < 0.05). Metal ion release from the two alloys increased with increasing HP concentrations (over 3000% increase in Ni and 1400% increase in Pd ions were recorded when HP concentration increased from 0% to 30%). Surface roughness values of the samples before and after bleaching were not significantly different (P > 0.05) Exposure of the two dental casting alloys to HP solutions increased metal ion release of all the elements except gold.

Pre-clinical evaluation of novel mucoadhesive bilayer patches for local delivery of clobetasol-17-propionate to the oral mucosa.

Oral lichen planus (OLP) and recurrent aphthous stomatitis (RAS) are chronic inflammatory conditions often characterised by erosive and/or painful oral lesions that have a considerable impact on quality of life. Current treatment often necessitates the use of steroids in the form of mouthwashes, creams or ointments, but these are often ineffective due to inadequate drug contact times with the lesion. Here we evaluate the performance of novel mucoadhesive patches for targeted drug delivery. Electrospun polymeric mucoadhesive patches were produced and characterised for their physical properties and cytotoxicity before evaluation of residence time and acceptability in a human feasibility study. Clobetasol-17-propionate incorporated into the patches was released in a sustained manner in both tissue-engineered oral mucosa and ex vivo porcine mucosa. Clobetasol-17 propionate-loaded patches were further evaluated for residence time and drug release in an in vivo animal model and demonstrated prolonged adhesion and drug release at therapeutic-relevant doses and time points. These data show that electrospun patches are adherent to mucosal tissue without causing tissue damage, and can be successfully loaded with and release clinically active drugs. These patches hold great promise for the treatment of oral conditions such as OLP and RAS, and potentially many other oral lesions.

In vitro biocompatibility of fluorcanasite glass-ceramics for bone tissue repair.

Fluorcanasite glass-ceramics were produced by controlled two stage heat-treatment of as-cast glasses. These glasses were modified from stoichiometric fluorcanasite composition by either adding P(2)O(5) or altering the molar ratios of Na(2)O and CaO. Commercial bioactive 45S5 Bioglass(R) was also prepared in-house to evaluate the relative in vitro biocompatibility of fluorcanasite glass-ceramics. The scanning electron microscopy (SEM) images showed that cells had colonized the surfaces of fluorcanasite glass-ceramics to form a confluent sheet. Quantitative MTT assay results were in good agreement with the qualitative SEM observations. It was concluded that incorporation of excess calcium oxide or P(2)O(5) in stoichiometric glass composition improved in vitro biocompatibility. Controlled heat-treatment further improved the biological response of cultured bone cells to modified fluorcanasite glass-ceramics when compared with their parent glasses. Ion release and pH data suggested a strong correlation between solubility (in particular, Na ion release) and biocompatibility. Reduced solubility, Na ion release, and related pH effects appeared to be the principal mechanisms responsible for improvement in in vitro biocompatibility.

The effect of 24h non-stop hydrogen peroxide concentration on bovine enamel and dentine mineral content and microhardness.

OBJECTIVES: Tooth bleaching agents may adversely affect tooth structure. The aim of this study was to investigate the effect of hydrogen peroxide concentration on mineral loss and microhardness of bovine teeth. METHODS: Twenty-six freshly extracted intact bovine incisor teeth were stored in distilled water. Five teeth were sectioned and four samples (2 mm x 2 mm x 1.5 mm) each of enamel and dentine were obtained from each tooth. The samples of enamel and dentine were divided into four groups and immersed in either 0%, 3%, 10% or 30% (w/v) hydrogen peroxide solutions for 24h at 37 degrees C. Samples from the solutions were taken for ion release analysis using inductively coupled plasma mass spectrometry. The remaining 21 teeth were mounted in epoxy resin and the upper surface of the specimens were ground and polished to expose the enamel and dentine for microhardness measurements. These specimens were randomly divided into three equal groups and Vickers microhardness values were recorded on the enamel and dentine surfaces of each group before and after bleaching. RESULTS: The differences in ion release concentration after treatment with 0% (control) and each of 3%, 10% and 30% hydrogen peroxide (w/v) were statistically significant (p<0.025). The release of calcium and phosphorous ions increased with increasing hydrogen peroxide concentrations. A significant reduction (p<0.05) in Vickers microhardness values for enamel was recorded after bleaching. CONCLUSIONS: Ion release from both enamel and dentine increased with increasing hydrogen peroxide concentration. Microhardness of enamel decreased significantly with bleaching.

The effect of hydrogen peroxide concentration on metal ion release from dental amalgam.

OBJECTIVES: The aim of this study was to investigate the effect of hydrogen peroxide (HP) concentration on metal ion release from dental amalgam. METHODS: Dental amalgam discs (n=25) were prepared by packing amalgam into cylindrical plastic moulds (10 mm diameter and 2 mm height). The discs were divided into five equal groups and each group was immersed in 20 ml of either 0%, 1%, 3%, 10% or 30% HP solution for 24 h at 37 degrees C. Samples were taken for metal ion release determination (Hg, Ag, Sn and Cu) using inductively coupled plasma mass spectrometry (ICP-MS). The surface roughness of each disc was measured before and after bleaching. RESULTS: The differences in concentration of metal ions released after treatment with 0% (control) and each of 1%, 3%, 10% and 30% HP were statistically significant (p<0.05). Metal ion release for the elements (Hg, Ag, Sn and Cu) increased with exposure to increasing concentrations of HP. Surface roughness measurements of the samples before and after treatments with HP solutions were not significantly different (p>0.05). CONCLUSIONS: Exposure to HP bleaching agent was associated with increased metal ion released from dental amalgams compared to treatment with a control solution. Ion release was in proportion to the peroxide concentration tested, with the highest concentration associated with the greatest metal ion release for all elements investigated.

Alginate-based hydrogels functionalised at the nanoscale using layer-by-layer assembly for potential cartilage repair.

Injuries to articular cartilage are frequently difficult to repair, in part because of the poor regenerative capacity of this tissue. To date, no successful system for complete regeneration of the most challenging cartilage defects has been demonstrated. The aim of this work was to develop functionalised hydrogels at the nanoscale by Layer-by-Layer (LbL) assembly to promote cartilage healing. Hydrogels, based on sodium alginate (NaAlg) and gelatin (G), were prepared by an external gelation method consisting of CaCl2 diffusion and genipin addition for G crosslinking. Successively, hydrogels were coated with G to obtain a positive charge on the surface, then functionalised by LbL assembly to create 16 nanolayers, based on poly(styrene sulfonate)/poly(allyl amine) (PSS/PAH), including a specific peptide sequence (CTATVHL) and transforming growth factors ß1 (TGF-ß1). Physico-chemical properties were evaluated by XPS, ATR-FTIR and rheological analyses while in vitro cytocompatibility was studied using bovine articular chondrocytes (BAC). XPS spectra showed N1s and S2p peaks, indicating that PAH and PSS have been introduced with success. ATR-FTIR indicated the specific PAH and PSS absorption peaks. Finally, the biomolecule incorporation influenced positively the processes of BAC adhesion and proliferation, and glycosamynoglycan secretion. The functionalised alginate-based hydrogels described here are ideally suited to chondral regeneration in terms of their integrity, stability, and cytocompatibility.

Preparation and Antibacterial Properties of Silver-Doped Nanoscale Hydroxyapatite Pastes for Bone Repair and Augmentation.

The treatment of deep bone infections remains a significant challenge in orthopaedic and dental surgery. The relatively recent commercial manufacture of nanoscale hydroxyapatite has provided surgeons with an injectable biomaterial that promotes bone tissue regeneration, and with further modification it may be possible to incorporate antimicrobial properties into these devices. Silver-doped nanoscale hydroxyapatite pastes (0, 2, 5 and 10 mol.% silver) were prepared using a rapid mixing method. When the process was modified to prepare a 10 mol.% silver-doped material, silver phosphate was detected in addition to nanoscale hydroxyapatite. Thermal decomposition occurred more readily with greater silver content following calcination at 1000 °C for 2 h. Silver-doped nanoscale hydroxyapatite pastes showed antibacterial activity against Staphylococcus aureus and Pseudomonas aeruginosa in a dose dependent manner using both agar diffusion assays and suspension cultures. It was concluded that the enhanced antibacterial activity of the silver-doped pastes was due to the action of diffusible silver ions. Based on these results, silver-doped nanoscale hydroxyapatite pastes represent a highly promising new biomaterial system for the prevention and treatment of deep infections in bone tissue.

The relationship between particle morphology and rheological properties in injectable nano-hydroxyapatite bone graft substitutes.

Biomaterials composed of hydroxyapatite (HA) are currently used for the treatment of bone defects resulting from trauma or surgery. However, hydroxyapatite supplied in the form of a paste is considered a very convenient medical device compared to the materials where HA powder and liquid need to be mixed immediately prior to the bone treatment during surgery. In this study we have tested a series of hydroxyapatite (HA) pastes with varying microstructure and different rheological behaviour to evaluate their injectability and biocompatibility. The particle morphology and chemical composition were evaluated using HRTEM, XRD and FTIR. Two paste-types were compared, with the HA particles of both types being rod shaped with a range of sizes between 20 and 80nm while differing in the particle aspect ratio and the degree of roundness or sharpness. The pastes were composed of pure HA phase with low crystallinity. The rheological properties were evaluated and it was determined that the pastes behaved as shear-thinning, non-Newtonian liquids. The difference in viscosity and yield stress between the two pastes was investigated. Surprisingly, mixing of these pastes at different ratios did not alter viscosity in a linear manner, providing an opportunity to produce a specific viscosity by mixing the two materials with different characteristics. Biocompatibility studies suggested that there was no difference in vitro cell response to either paste for primary osteoblasts, bone marrow mesenchymal stromal cells, osteoblast-like cells, and fibroblast-like cells. This class of nanostructured biomaterial has significant potential for use as an injectable bone graft substitute where the properties may be tailored for different clinical indications.

The effect of carbamide peroxide treatment on metal ion release from dental amalgam.

OBJECTIVES: There is concern that hydrogen peroxide generated by tooth bleaching agents may cause enhanced metal ion release (including mercury) from dental amalgam following contact. The aim of this in vitro study was therefore to investigate the effect of a carbamide peroxide (CP) based tooth bleaching gel on metal ion release from dental amalgam. METHODS: Dental amalgam discs were prepared according to the manufacturers' instructions. These were treated with either a 10% carbamide peroxide (CP) gel or a 0% CP gel for 24h. Discs were carefully wiped with cotton wool before immersion in distilled water (20 ml) for 24h at 37 degrees C. Following immersion, water samples were taken for metal ion release determination (Ag, Cu, Hg and Sn) using inductively coupled plasma mass spectrometry methods. The specimens were further evaluated for surface changes using scanning electron microscopy (SEM) and Talysurf surface roughness measurements. RESULTS: The differences in concentration of metal ions released after treatment with the 10% CP gel and a placebo gel treatment were not statistically significant (p>0.05). For example, mercury release following treatment with the 10% CP gel and the 0% CP gel was found to be 1.17(0.5) and 0.57(0.1)microgcm(-2), respectively. Roughness measurements for samples treated with the 10% CP gel and 0% CP gel were 2.23(0.47) and 1.74(0.16)microm, respectively, again showing no significant difference between groups (p>0.05). SEM images of the amalgam surfaces showed no apparent differences between treatments. SIGNIFICANCE: Treatment with a 10% CP gel did not significantly enhance subsequent metal ion release from dental amalgams compared to a control gel, contradicting previously published studies.

Biocompatibility of glass-ionomer bone cements.

Glass-ionomer cements (GIC) have been extensively used in dentistry for over 30 years. Due to their excellent biocompatibility in dental applications GIC have been formulated for medical applications. The past decade has seen some impressive advances in the development of medical GICs, however these advances have been matched by serious critical problems. This review examines the properties of GICs, which can influence their behaviour in a biological environment. The progress made and the problems encountered in the development of these bone cements will also be addressed. The review will conclude with the research currently being employed to optimise the biocompatibility of these important biomaterials. There is little doubt that GICs compare favourably with alternative bone cements for specific applications, based on in vitro and in vivo studies. There is however, a degree of risk inherent in the use of any medical device or biomaterial. GICs must therefore be used carefully and in accordance with the instructions that are based on a significant body of research data.

In vitro biocompatibility of a novel Fe2O3 based glass ionomer cement.

INTRODUCTION: Since their invention in the late 1960s, glass ionomer cements (GICs) have been used extensively in dentistry but recently they have also been utilised in ear nose and throat (ENT) surgery. Unfortunately, Al3+, a component of conventional ionomer glasses, has been linked to poor bone mineralisation and neurotoxicity. OBJECTIVE: The aim of the research was to modify a commercial ionomer glass composition by substituting Al2O3 with Fe2O3. METHODS: Glasses with the following molar compositions were fabricated: 4.5SiO2*3M2O3*XP2O5*3CaO*2CaF2 (M = Al or Fe, X = 0-1.5). The glasses were characterised using X-ray fluorescence (XRF) and X-ray powder diffraction (XRD). Cements were prepared using a standard ratio of; 1 g of glass powder: 0.2 g of dried polyacrylic acid: 0.3 g of 10% tartaric acid solution. Cement formation was assessed using a Gilmore needle and in vitro biocompatibility was investigated for novel cement formulations. RESULTS: XRF revealed that the Fe2O3-based glasses had Al2O3 contamination from the crucibles and also had undergone substantial F- losses. XRD gave peaks that corresponded to magnetite Fe3O4 (JCPDS # 19-629) in all compositions. Apatite Ca5(PO4)3(OH,F) (JCPDS # 15-876) was identified in P2O5 containing glasses. It was possible to fabricate cements from all of the Fe2O3-based ionomer glasses. Good in vitro biocompatibility was observed for the Fe2O3-based cements. CONCLUSION: Ionomer glasses may be prepared by entirely replacing Al2O3 with Fe2O3. Cement setting times appeared to be related to P2O5 content. Fe2O3-based cements showed good in vitro biocompatibility.

Extracellular ATP and UTP stimulate cartilage proteoglycan and collagen accumulation in bovine articular chondrocyte pellet cultures.

Bovine articular chondrocytes were maintained in high density pellet cultures with and without serum and nucleotide triphosphates for different periods of time. Despite half-lives in culture of about 3 h, adenosine triphosphate and uridine triphosphate in the presence of serum increased sulphated glycosaminoglycan and collagen deposition above control levels. In the presence of serum a single dose of uridine triphosphate on the first day of culture was sufficient to induce significant increases in subsequent proteoglycan and collagen deposition. We conclude that both adenine triphosphate and uridine triphosphate are anabolic for articular chondrocytes, and that this effect on the chondrocyte is long-term.

Expression of type X collagen and matrix calcification in three-dimensional cultures of immortalized temperature-sensitive chondrocytes derived from adult human articular cartilage.

Chondrocytes isolated from normal adult human articular cartilage were infected with a retroviral vector encoding a temperature-sensitive mutant of the simian virus 40 large tumor antigen and a linked geneticin (G418)-resistance marker. G418-resistant colonies were then isolated, ring-cloned, and expanded in serum-containing media. Several immortalized chondrocyte cell lines were established from the clones that survived, some of which have been maintained in continuous culture for over 2 years. Despite serial subcultures and maintenance as monolayers, these cells retain expression of markers specific for cells of the lineage, namely type II collagen and aggrecan, detected immunocytochemically. We also examined the phenotype of three of these immortalized cell lines (designated HAC [human articular chondrocyte]) using a pellet culture system, and in this report, we present evidence that a prototype of these lines (HAC-F cells) expresses markers normally associated with hypertrophic chondrocytes. When HAC-F cells were cultivated in centrifuge tubes, for periods of up to 63 days, at 39 degrees C with mild and intermittent centrifugation they continued to express both lineage markers; total type II collagen/pellet remained stable, whereas there was a temporal decrease in cartilage-specific glycosaminoglycans content. In addition, in the presence of ascorbate but in the absence of a phosphate donor or inorganic phosphate supplement, the cells also begin to express a hypertrophic phenotype characterized by type X collagen synthesis and extensive mineralization of the extracellular matrix in late stage cultures. The mRNA encoding type X collagen was detected in the cell pellets by reverse transcriptase polymerase chain reaction as early as day 2, and anti-type X collagen immunoreactivity was subsequently localized in the matrix. The mineral was characterized by energy-dispersive X-ray microanalysis as containing calcium (Ca) and phosphorus (P) with a Ca:P peak height ratio close to that of mineralized bone tissue. The unexpected phenotype of this human chondrocyte cell line provides an interesting opportunity for studying chondrocyte maturation in vitro.

Glass-ionomers: bioactive implant materials.

Glass-ionomer cements (GICs) originally designed for use as dental materials have a number of advantages over acrylic bone cements. These include lack of exotherm during setting, absence of monomer and improved release of incorporated therapeutic agents; this has resulted in the development of GICs for biomedical applications. Major landmarks in this history are the formulation of defined-composition ionomer glasses and an improved understanding of the biological and material properties of GICs. Following implantation, GICs can form a stable integration with bone, and affect the growth and development of bone, both adjacent to their surface and systemically, through an ion release mechanism. The 'non-inert' nature of this group of materials is also demonstrated by their adverse effects on neural tissue. Successful clinical use of GICs, both as bone cements and as preformed implants for hard tissue replacement, have been reported in the fields of otologic surgery (Cochlear implant fixation, repair of the tympanic chain, eustation tube obliteration and as ear ossicles), and oral and reconstructive surgery. The use of GICs in situations where they will come into contact with nerves or neural tissue is contraindicated.

Role of exchanged ions in the integration of ionomeric (glass polyalkenoate) bone substitutes.

Ionomeric (glass polyalkenoate) implants are synthetic materials which can be used for repairing bone defects. It has been suggested that ions are leached from these implants during healing and that they influence cellular activity in the surrounding tissues. Morphological, immunohistochemical and microanalytical techniques were used to compare the osteogenic capacity of implants which eluted aluminium ions with implants which did not elute aluminium ions. The extracellular matrix molecules fibronectin and tenascin were located upon the surface of both implanted materials. Thick seams of lamellar bone were apposed to implants containing labile aluminium ions, but the bone was poorly mineralized. At the same time, transient increases were apparent in osteoblast activity on periosteal and endosteal surfaces and in chondrocyte activity in the growth plate and articular cartilages. In contrast, small amounts of mineralized lamellar bone were apposed to substituted implants (without aluminium) and the growth plate and articular cartilages remained normal in thickness and morphology. These results suggest that exchanged ions can influence the amount and quality of bone apposed to the implant. They also suggest that the effect of the ions depends upon their concentration and the state of differentiation of osteogenic cells.

Bone cell interactions with a granular glass-ionomer bone substitute material: in vivo and in vitro culture models.

The interface between bone and a synthetic bone substitute constructed from glass-ionomer cement (ionomeric microimplant) was studied in diffusion chambers implanted in a primate baboon model (Papio ursinus) and in in vitro primary bone organ cultures derived from neonate rat calvaria. In both models osteoblast-like cells colonized the surface of the implant producing a collagenous extracellular matrix. An electron-dense bonding zone similar to that reported for hydroxyapatite and titanium was seen in both models but was a more constant feature of the tissue/implant interface in calvarial culture.

Devitrification of ionomer glass and its effect on the in vitro biocompatibility of glass-ionomer cements.

The effects of devitrification of an ionomer glass with a molar composition 4.5SiO(2).3Al(2)O(3).1.5P(2)O(5).3CaO.2CaF(2) on cement formation and in vitro biocompatibility were investigated. Differential thermal analysis was used to study the phase evolution in the glass, and to determine the heat treatments for production of glass-ceramics. X-ray diffraction patterns from glass frit heat-treated at 750 degrees C for 2h contained peaks corresponding to apatite (JCPDS 15-876), whereas for samples heat-treated at 950 degrees C for 2h apatite and mullite (JCPDS 15-776) were the major phases detected. Transmission electron microscopy (TEM) confirmed that apatite and apatite-mullite phases were present after heat treatments at 750 degrees C and 950 degrees C respectively. Glass and glass-ceramics were ground to prepare <45microm powders and glass ionomer cements were produced using a ratio of 1g powder: 0.2g PAA: 0.3g 10% m/v tartaric acid solution in water. In vitro biocompatibility was evaluated using cultured rat osteosarcoma (ROS) cells. Scanning electron microscopy (SEM) showed that cells colonised the surfaces of cements prepared using untreated ionomer glass and glass crystallised to form apatite (750 degrees C/2h). However, quantitative evaluation using MTT and total protein assays indicated that more cell growth occurred in the presence of cements prepared using ionomer glasses crystallised to apatite than cements prepared using untreated glass. The least cell growth and respiratory activity was observed on cements made with crystallised glass containing both apatite and mullite. It was concluded that the controlled devitrification of ionomer glasses could be used to produce GIC bone cements with improved biocompatibility.

Synthetic implant surfaces. 1. The formation and characterization of sol-gel titania films.

Sol-gel has been used to prepare thin titania films. We have investigated the effects of dip rate, sintering temperature and time on the chemical composition of the films, their physical structure and thickness, and adherence to a silica substrate. Our aim has been to produce films that mimic as closely as possible the natural oxide layer that is found on titanium. These films are to be used as substrates in an in vitro model of osseointegration.

In vivo bone tissue response to a canasite glass-ceramic.

The aim of this study was to determine the biocompatibility and osteoconductive potential of a high-strength canasite glass ceramic. Glass-ceramic rods were produced using the lost-wax casting technique and implanted in the mid-shafts rabbit femurs. Implants were harvested at 4, 13 and 22 weeks and prepared for light and electron microscopy. Hydroxyapatite was used as a control material. Hydroxyapatite implants were surrounded by new mineralised bone tissue after 4 weeks of implantation. The amount of bone surrounding the implant increased slightly at 13 weeks. In contrast, canasite glass and glass ceramic implants were almost entirely surrounded by soft tissue during all the time periods. Close contact between bone and canasite glass-ceramic implant without the intervening fibrous tissue was observed in only a few regions. The canasite formulation evaluated was not osteoconductive and appeared to degrade in the biological environment. It was therefore concluded that the canasite formulation used was unsuitable for use as implant. Further work is required to improve the biocompatibility of these materials with bone tissue. It is possible that this could be achieved by reducing the solubility of the glass and glass ceramic.