Biomedical subjects

C A Scotchford

Publications and source records attributed to C A Scotchford.

At least 19 recordsLinked to original sources

PubMed2006

The effect of anisotropic architecture on cell and tissue infiltration into tissue engineering scaffolds.

A common phenomenon in tissue engineering is rapid tissue formation on the outer edge of the scaffold which restricts cell penetration and nutrient exchange to the scaffold centre, resulting in a necrotic core. To address this problem, we generated scaffolds with both random and anisotropic open porous architectures to enhance cell and subsequent tissue infiltration throughout the scaffold for applications in bone and cartilage engineering. Hydroxyapatite (HA) and poly(D,L-lactic acid) (P(DL)LA) scaffolds with random open porosity were manufactured, using modified slip-casting and by supercritical fluid processing respectively, and subsequently characterised. An array of porous aligned channels (400 microm) was incorporated into both scaffold types and cell (human osteoblast sarcoma, for HA scaffolds; ovine meniscal fibrochondrocytes, for P(DL)LA scaffolds) and tissue infiltration into these modified scaffolds was assessed in vitro (cell penetration) and in vivo (tissue infiltration; HA scaffolds only). Scaffolds were shown to have an extensive random, open porous structure with an average porosity of 85%. Enhanced cell and tissue penetration was observed both in vitro and in vivo demonstrating that scaffold design alone can influence cell and tissue infiltration into the centre of tissue engineering scaffolds.

Animals↗

PubMed2005

The influence of dispersant concentration on the pore morphology of hydroxyapatite ceramics for bone tissue engineering.

There is a clinical need for synthetic scaffolds that will promote bone regeneration. Important factors include obtaining an optimal porosity and size of interconnecting windows whilst maintaining scaffold mechanical strength, enabling complete penetration of cells and nutrients throughout the scaffold, preventing the formation of necrotic tissue in the centre of the scaffold. To address this we investigated varying slip deflocculation in order to control the resulting porosity, pore size and interconnecting window size whilst maintaining mechanical strength. Hydroxyapatite (HA) porous ceramics were prepared using a modified slip casting process. Rheological measurements of the HA slips were used to identify deflocculation conditions which resulted in changes in the cell and window sizes of the resulting ceramics. Sintered ceramics were characterised by X-ray diffraction (XRD) and scanning electron microscopy (SEM). Pore and window size distribution was determined by SEM. XRD analysis confirmed that the crystal structure remained HA after the sintering process. SEM showed that HA porous ceramics presented a highly interconnected porous network with average pore sizes ranging from 391+/-39 to 495+/-25 microm. The average window size varied from 73+/-5 to 135+/-7 microm. Pore diameters obtained were controllable in the range 200-500 microm. Window sizes were in the range 30-250 microm. The use of dispersant concentration allows pore and window size to be modified whilst maintaining control over porosity demonstrated by a porosity of 85% for seven different dispersant concentrations. The advantage of this approach allows the correlation between the rheological conditions of the slip and the resultant sintered ceramic properties. In particular, optimising the ceramic strength by controlling the agglomeration during the casting process.

Bone Substitutes↗

PubMed2005

Preparation of poly(epsilon-caprolactone)/continuous bioglass fibre composite using monomer transfer moulding for bone implant.

Poly(epsilon-caprolactone) (PCL)/continuous bioglass fibre composite was prepared using the monomer transfer moulding technique coupled with a surface initiated polymerisation. The bioglass fibres were surface treated with an amine ended silane in order to initiate polymerisation of epsilon-caprolactone from the fibre surface. Surface initiated polymerisation significantly improved the Young's modulus and flexural strength and water resistance of the composite. Initial in vitro biocompatibility assessment suggests that amine ended silane treatment of bioglass fibres before their inclusion in the composite does not have a negative effect on the biological responses in terms of macrophage activation as measured by IL-1beta release and craniofacial osteoblast attachment.

Animals↗

PubMed2005

Thin-film theories for two-phase reactive flow models of active cell motion.

The aim of this paper is to develop a broadly-applicable and self-consistent thin-film biphasic modelling framework for the full crawling cycle of a single animal cell. A hierarchy of thin-film two-phase 'reactive flow' models is derived; between them these cover a wide range of biologically relevant parameter regimes. The mathematical properties and biological implications of the resulting systems of high-order nonlinear degenerate parabolic-elliptic evolution equations are investigated. Linear-stability arguments suggest the formation of highly localized regions of high or low network density associated with small irregular oscillations or 'ruffling' of the plasma membrane. Local analyses at the contact line identify the classes of admissible contact-line conditions, through which we study for the first time the effect on the cell-scale motion of the 'mesoscopic' contact-line physics, which consists of the chemical and mechanical mechanisms for protrusive and retractive force generation near the outer cell periphery. One of the formulations is used to develop a minimal model for cell body translocation over a thin pseudopod, which predicts that myosin-driven contraction is not essential for rapid translocation. An analytic prediction for the translocation speed is given in terms of the network viscosity and slip coefficient (a parameter measuring the adhesion strength), of the membrane tension and of the thicknesses of the pseudopod and actin cortex; this is in good agreement with the translocation speed of osteoblasts on biomaterial substrates commonly used for orthopaedic implants. Limitations of the modelling approach and directions for future work are outlined.

Actins↗

PubMed2004

Synthesis and degradation of sodium iron phosphate glasses and their in vitro cell response.

The degradation profiles of six sodium iron phosphate glass formulations have been investigated using simple dissolution trials in deionized water. The glasses were produced from the appropriate phosphate salts by melting at 1200 degrees C in 5% Au/95% Pt crucibles. Dissolution rates varied from 0.2 gcm(-2)h(-1) for the 1% Fe glass to essentially zero over the 6-week test period for the 15% Fe and 20% Fe glasses. The overall degradation rate was found to vary according to the approximate relation: rate = 1.3e(-0.79x) gcm(-2)h(-1), where x is the percentage iron content of the glass. Glasses with 10% or greater iron content were observed to maintain a constant density over the course of the tests and thus appeared to degrade from the surface and not the bulk. In vitro cell response tests were conducted on the glasses using macrophages and primary craniofacial osteoblasts. These tests were performed on the glasses with 10% or greater iron content because glasses with lower iron content degraded too quickly. Confocal microscopy revealed a rounded macrophage morphology and IL-1beta production was low, suggesting little macrophage activation. However, a significant level of peroxide production was observed. Osteoblasts were observed to attach to the glass surfaces and spread, exhibiting a similar cytosketetal organization to the cells on the Thermanox controls, with a high level of F-actin organization. On balance, the 15% Fe glass performed slightly better than the 20% Fe glass in these assays.

Animals↗

PubMed2004

The effect of production regime and crucible materials on the thermal properties of sodium phosphate glasses produced from salts.

Changes in the thermal properties of sodium phosphate glasses during melt production have been investigated using Pt/Au and fused alumina crucibles. Glasses were produced from NaH(2)PO(4) as a starting material, providing an intrinsic Na(2)O:P(2)O(5) ratio of 1:1 and giving an O/P = 3, that is, a metaphosphate. In Pt/Au crucibles, glass transition temperatures rose to a plateau value of 295 degrees C at a rate determined by melt temperature. No contamination of the glass by platinum or gold was detected or indicated in the results. E(a) for the reaction was found to be 66.4 kJ mol(-1). In fused alumina crucibles, glass transition temperatures rose to over 450 degrees C, with these values showing some convergence at higher furnace temperatures. Extensive erosion of the alumina crucibles was observed. The amount of alumina incorporation within the glasses correlated well with the rise in glass transition temperature up to a maximum of 15.5 mol % Al(2)O(3) content. Al(2)O(3) incorporation above this value caused a reduction in the value of the T(g).

Aluminum Oxide↗

PubMed2004

Controlled degradation and macrophage responses of a fluoride-treated polycaprolactone.

We have developed a new bone replacement material based on polycaprolactone (PCL), which can act as a suitable matrix for monomer transfer molding of degradable composites. A boron trifluoride catalyst with glycerol additive was used to produce PCL with a degradation rate that can be altered by treatment with fluoride ions. The effect of cations on the degradation of the polymer and macrophage cell responses are discussed. We found that treatment with fluoride ions reduced the degradation rate. No significant difference between these three fluorides was observed although a general trend was seen where KF-treated PCL appeared to degrade slower than NaF-treated PCL which was slower than NH(4)F-treated PCL. Variation in solubilities of the salts was observed where the K(+) cation had the highest solubility and the Na(+) cation had the lowest solubility, which suggests that NaF was able to degrade the polymer more efficiently than the other fluorides. No significant macrophage activation was observed after culture on the polymer surfaces as determined by peroxide and IL-1 beta release, whereas some activation occurred after culture in degradation products.

Animals↗

PubMed2004

Porous methacrylate scaffolds: supercritical fluid fabrication and in vitro chondrocyte responses.

This paper describes a method of foaming a polymer system comprising poly(ethyl methacrylate)/tetrahydrofurfuryl methacrylate (PEMA/THFMA), characterisation of the resulting porosity and use of the foam for chondrocyte culture. The potential for this polymer system to support cartilage repair has been investigated previously, both in vivo and in vitro. PEMA/THFMA foamed created using supercritical carbon dioxide were characterised using scanning electron microscopy, mercury intrusion porosimetry and helium pycnometry. Foams were found to be 82% porous with open porosities of 57%. The mean pore diameter was found to be 99+60 microm. Bovine chondrocytes seeded directly onto the surface of the foamed and unfoamed PEMA/THFMA demonstrated lower proliferation on the foamed material, greater retention of the rounded cell morphology and increased glycosaminoglycan synthesis. In conclusion, this study has shown that a porous PEMA/THFMA system can further enhance the ability of the material to support chondrocytes in vitro. However, further modifications in processing are necessary to determine optimum conditions for cartilage tissue formation.

Animals↗

PubMed2003

Long-term craniofacial osteoblast culture on a sodium phosphate and a calcium/sodium phosphate glass.

The aim of this study was to determine the characteristics of human craniofacial osteoblasts cultured on sodium phosphate glass and calcium-sodium phosphate glass in a long-term culture of up to 28 days. The characteristics studied were attachment, proliferation, alkaline phosphatase activity, collagen-1 production, and mineralization. A comparison of the degradation rate, measured by mass loss of the glasses, which are intended for use as a component of a novel degradable composite for craniofacial bone repair, was also performed. It was our hypothesis that the glass would be degradable with a change in degradation rate observed by calcium addition and support osteoblast proliferation and expression of the above characteristics. The inclusion of calcium into the reaction mixture significantly decreased the degradation rate, and it is suggested that the slower degradation is the result of pseudo crosslinking (ionic crosslinks rather than covalent bonding) of the polyphosphate chains by the calcium ions. Therefore, twice as many P-O bonds will need to be hydrolyzed for dissolution of the metal phosphate to occur, therefore greatly reducing the rate of hydrolysis. Osteoblasts were able to attach, spread, and proliferate in a manner comparable with the positive control, as shown by analysis of variance. Formation of a collagen-rich mineralized matrix was also observed. The results presented here suggest that a biocompatible soluble glass has been produced, which has potential to be included in a novel biodegradable craniofacial implant.

Calcium↗

PubMed2003

Seeding cells into needled felt scaffolds for tissue engineering applications.

Tissue engineering methods are under development that will enable the repair or replacement of a variety of tissues, including articular cartilage and bone. To engineer functional tissue it is necessary that scaffolds initially be seeded with a large number of cells distributed evenly throughout the scaffold structure. It previously has been shown that, compared to static seeding conditions, seeding scaffolds under dynamic conditions facilitates high seeding densities and even distributions of cells (Li et al., Biotechnology Progress 2001;17:935-944). The efficiency of seeding HOSTE85 cells and bovine chondrocytes into needled felt scaffolds following agitation at different speeds was determined. Seeding efficiency was determined using the Hoechst 33258 assay, and cell viability was assessed using the Alamar Blue trade mark assay. The distribution of cells within the scaffolds was imaged using scanning electron microscopy. It was found that the optimum seeding conditions varied for HOSTE85 cells and bovine chondrocytes, with different agitation speeds leading to different seeding efficiencies, cell viabilities, and distributions of cells within scaffolds. The optimum agitation speeds for seeding a high number of viable cells into scaffolds so that they were arranged evenly were 300 rpm for HOSTE85 cells and 200 rpm for bovine chondrocytes.

Animals↗

PubMed2003

Chemically patterned, metal-oxide-based surfaces produced by photolithographic techniques for studying protein- and cell-interactions. II: Protein adsorption and early cell interactions.

Protein adsorption and adhesion of primary human osteoblasts on chemically patterned, metal-oxide-based surfaces comprising combinations of titanium, aluminium, vanadium and niobium were investigated. Single metal samples with a homogeneous surface and bimetal samples with a surface pattern produced by photolithographic techniques were used. The physical and chemical properties of the samples have been extensively characterised and are presented in a companion paper. Here, we describe their properties in terms of cell responses during the initial 24h of cell culture. Regarding the cell number and activity there was no significant difference between any of the single metal surfaces. However the morphology of cells on vanadium surfaces became spindle-like. In contrast to the behaviour on single metal samples, cells exhibited a pronounced reaction on bimetallic surfaces that contained aluminium. Cells tended to stay away from aluminium, which was the least favoured metal in all two-metal combinations. An initial cell alignment relative to the pattern geometry was detectable after 2h and was fully developed after 18h of incubation. The organisation of f-actin and microtubules as well as the localisation of vinculin were all more pronounced on non-aluminium regions. We hypothesised that the differences in cell response could be associated with differences in the adsorption of serum proteins onto the various metal oxides. Protein adsorption experiments were performed using microscopy in conjunction with immunofluorescent stains. They indicated that both fibronectin and albumin adsorption were significantly greater on the non-aluminium regions, suggesting that differences in cellular response correlate with a modulation of the concentration of serum proteins on the surface.

Adsorption↗

PubMed2003

Craniofacial osteoblast responses to polycaprolactone produced using a novel boron polymerisation technique and potassium fluoride post-treatment.

There is no ideal material for craniofacial bone repair at present. The aim of this study was to test the biocompatibility of polycaprolactone (PCL) synthesised by a novel method allowing control of molecular weight and degradation rate, with regard to it being used as matrix for a biodegradable composite for craniofacial bone repair. Human primary craniofacial cells were used, isolated from paediatric skull after surgery. Cell responses were analysed using various assays and antibody staining. Cells attached and spread on the PCL in a similar manner to the Thermanox controls as shown by phalloidin staining of F-actin. Cells maintained the osteoblast phenotype as demonstrated by alkaline phosphatase assay and antibody staining throughout the time points studied, up to 28 days. Cells proliferated on the PCL as shown by a DNA assay. Collagen-1 staining showed extensive production of a collagen-1 containing extracellular matrix, which was also shown to be mineralised by alizarin red staining. Short-term (up to 48 h) attachment studies and long-term (up to 28 days) expression of markers of the osteoblast phenotype have been demonstrated on the PCL. This new method of synthesising PCL shows biocompatibility characteristics that give it potential to be used for craniofacial bone repair.

Absorbable Implants↗

PubMed2003

Properties of calcium phosphate coatings deposited and modified with lasers.

Physical, chemical and biological properties of calcium phosphate coatings fabricated by a pulse laser deposition method at room temperature (RT PLD) have been studied. In vitro evaluation of RT PLD coatings on bioresorbable polymers (Poly-epsilon-caprolactone and Poly-L-lactide) have been carried out. It was shown that both polymers support osteoblast growth, with increased cell activity, alkaline phosphatase activity and total protein content on those surfaces that have been coated. The advantages of RT PLD coatings in biomaterials surface optimization are discussed.

Journal Article↗

PubMed2002

Synthesis, degradation, and in vitro cell responses of sodium phosphate glasses for craniofacial bone repair.

This report outlines the initial synthesis, degradation, and short-term biocompatibility of sodium phosphate glasses, for use in the drawing of fibers and manufacture of biodegradable composites. Biocompatibility studies were performed using a macrophage cell line and primary human craniofacial osteoblasts. Sodium hydrogen phosphate and sodium dihydrogen phosphate glass synthesized for less than 1 h, resulted in a higher degradation rate than glass synthesized for 3 h or more (0.015 mg cm(-2) h(-1)). Glasses with high and low ratios of hydrogen phosphate to dihydrogen phosphate had very similar degradation rates. A condensation route for the formation of the glass should give rise to varying degradation rates with varying ratios of starting materials. It is suggested that the degradation rate of the glass is independent of the concentrations of the initial reagents and that ring-opening polymerization, which reaches an equilibrium state, occurs. Biocompatibility studies suggest minimal macrophage activation (low levels of peroxide and interleukin-1beta release and rounded morphology) and high osteoblast biocompatibility. The ultimate aim of our studies is to produce a biocompatible soluble phosphate glass that can be drawn into fibers for incorporation into a polycaprolactone matrix for craniofacial bone repair. This report demonstrates the successful production of a soluble glass, which is biocompatible with regard to osteoblasts and macrophages. Recent data from our laboratory have demonstrated successful fiber drawing and production of a novel polycaprolactone.

Absorbable Implants↗

PubMed2001

Osteoblast growth on titanium foils coated with hydroxyapatite by pulsed laser ablation.

Pulsed laser ablation is a new method for deposition of thin layers of hydroxyapatite (HA) on to biomaterial surfaces. In this paper, we report activity and morphology of osteoblasts grown on HA surfaces fabricated using different laser conditions. Two sets of films were deposited from dense HA targets, at three different laser fluences: 3, 6 and 9 Jcm(-2). One set of the surfaces was annealed at 575 degrees C to increase the crystallinity of the deposited films. Primary human osteoblasts were seeded onto the material surfaces and cytoskeletal actin organisation was examined using confocal laser scanning microscopy. The annealed surfaces supported greater cell attachment and more defined cytoskeletal actin organisation. Cell activity, measured using the alamar Blue assay, was also found to be significantly higher on the annealed samples. In addition, our results show distinct trends that correlate with the laser fluence used for deposition. The cell activity increases with increasing fluence. This pattern was repeated for alkaline phosphatase production by the cells. Differences in cell spreading were apparent which were correlated with the fluence used to deposit the HA. The optimum surface for initial attachment and spreading of osteoblasts was one of the HA films deposited using 9 J cm(-2) laser fluence and subsequently annealed at 575 degrees C.

Alkaline Phosphatase↗

PubMed1998

Growth of human osteoblast-like cells on alkanethiol on gold self-assembled monolayers: the effect of surface chemistry.

Primary human osteoblasts were cultured on self-assembled monolayers (SAMs) of alkylthiols on gold with carboxylic acid and methyl termini, and the kinetics of cell attachment and proliferation were measured. Over 90 min approximately twice as many cells attached to carboxylic-acid-terminated monolayers as attached to methyl-terminated monolayers. After 24 h the number of cells attached to carboxylic-acid-terminated monolayers was ten times that attached to the methyl-terminated monolayers. Cell morphology and cytoskeletal actin organization also were found to be different. Osteoblasts were cultured on SAMs that were patterned by photolithographic techniques. Cells attached almost exclusively to carboxylic-acid-functionalized areas of the patterned surfaces, leaving methyl-functionalized regions bare. The patterns strongly influenced the morphology of the attached cells. After 24 h, cells were observed to bridge between carboxylic-acid-terminated regions separated by 75 microns, methyl-terminated regions but not those separated by 150 microns methyl-terminated regions. After 6 days in culture osteoblasts formed multilayers on the carboxylic-acid-terminated regions of the pattern.

Actins↗

PubMed1997

Association of magnesium whitlockite crystals with lipid components of the extracellular matrix in human articular cartilage.

Several basic calcium phosphate mineral phases have been reported to be associated with osteoarthritis joint disease. Magnesium whitlockite crystal deposition has been reported in both osteoarthritic and normal human articular cartilage. Existing data suggest that likely prevailing conditions within cartilage would not support de novo whitlockite formation. It would appear, therefore, that additional factors must be extant at sites of crystal deposition. In this study normal articular cartilage specimens were examined for the presence and distribution of lipids relative to crystal deposition within the extracellular matrix. Specimens were examined using light and transmission electron microscopy (TEM), with standard processing protocols plus a malachite green-glutaraldehyde-osmium tetroxide (MGO) method, used to retain lipids normally removed from tissues during preparation for electron microscopy. Elemental maps of sections produced using this method were also made using X-ray microanalysis. Positive oil red O staining for lipid was clearly apparent immediately below and parallel to the articular surface of cartilage specimens using light microscopy. The extent and distribution of staining correlated well with the distribution of crystals, observed by TEM, in sections of tissue from adjacent sites of the same specimens. Using standard TEM, crystals were frequently observed scattered amongst intramatrical lipidic debris, particularly pericellularly, in areas of cell necrosis and amongst close packed tangential fibers between the articular surface and initial superficial zone chondrocytes. Cartilage specimens processed using the MGO method demonstrated electron dense features, not apparent using standard techniques, identified as lipid. Such extracellular lipid deposition varied with depth, with 100 nm globular bodies present in the superficial region, where colocalization of crystals and lipid were observed in about 10% of crystal observations. The association of lipid and crystal deposition is discussed in the context of phospholipid associated mineral formation and the potential role of such magnesium whitlockite deposition assessed.

Adult↗

PubMed1995

The isolation and characterization of magnesium whitlockite crystals from human articular cartilage.

A number of basic calcium phosphate crystals have been demonstrated in human articular tissues. The exact relationship between crystal deposition and disease remains obscure, although there is evidence supporting a rapid degenerative arthropathy within a specific set of patients. Limited reports of 'cuboid' calcium phosphate microcrystals in articular cartilage have been made over the last 10 years. In this study the occurrence of such crystals, not apparent by light microscopy, in human articular cartilage has been confirmed by transmission electron microscopy and X-ray microanalysis of tissue prepared by aqueous and anhydrous processing techniques. A crystal isolation technique involving collagenase digestion, centrifugation and sodium hypochlorite treatment was developed enabling crystal characterization by electron and X-ray diffraction. Crystals were identified as magnesium whitlockite; the first report of this mineral in articular cartilage. The presence of this mineral phase in normal and osteoarthritic articular cartilage is discussed with consideration given to physical conditions known to favor whitlockite formation and those extant in articular cartilage.

Calcium Phosphates↗