Synthesis and characterization of a hybrid (chitosan-g-glycidyl methacrylate)-xanthan hydrogel.

This work reports the synthesis and characterization of a new material obtained by mixing the hybrid natural-synthetic chitosan-g-glycidyl methacrylate (CTS-g-GMA) biopolymer and xanthan gum (X). All materials were characterized by infrared spectroscopy (FTIR), X-ray diffraction, and thermal analysis (DSC and TGA) and the results were contrasted with those of the precursor materials. The swelling index of the hydrogels decreases when the GMA mass percentage increases. The X-ray diffraction patterns show that the hybrid hydrogels are amorphous in contrast to chitosan (CTS), which is semi-crystalline. FTIR analysis confirms the existence of physical interactions among constituents. Rheological properties, η, G', and G", were determined as a function of flow allowing one to conclude that (CTS-g-GMA)-X behaves as physical hydrogel. Additionally, we report viability of fibroblasts when cultured onto the synthesized hydrogels. This study shows that these hydrogels support cell viability and have potential for use in biomedical engineering applications.

Gel-cast glass-ceramic tissue scaffolds of controlled architecture produced via stereolithography of moulds.

Two glass-ceramic scaffolds with a simple cubic structure of 500 µm square ligaments and square channels of width 400 or 600 µm have been fabricated by gel-casting into moulds produced by stereolithography, followed by mould removal, polymer burnout and sintering. The scaffolds have crushing strengths of 41 ± 14 and 17 ± 5 Mpa, respectively. Using a method of assembling discrete slices of scaffold, we are able to study cell behaviour within a scaffold by disassembly. Both scaffold structures were seeded with primary human osteoblasts and these penetrate, adhere, spread and proliferate on the scaffold structure. The larger channel diameter scaffold shows a greater cell population (despite its smaller surface area) and more pronounced production of ECM components (collagen and mineralization) with increased time in culture. Studies of sectioned scaffolds show that cell density and ECM production decrease with depth and that the difference between the two scaffold architectures is maintained.

The Chemical and Physical Properties of Poly(ε-caprolactone) Scaffolds Functionalised with Poly(vinyl phosphonic acid-co-acrylic acid).

There is a clinical need for a synthetic alternative to bone graft substitute (BGS) derived from demineralised bone matrix. We report the electrospinning of Poly(ε-caprolactone) (PCL) to form a 3-dimensional scaffold for use as a synthetic BGS. Additionally, we have used Poly(vinyl phosphonic acid-co-acrylic acid) (PVPA) to improve bone formation. Fibres were formed using a 10% w/v PCL/acetone solution. Infrared spectroscopy confirmed that the electrospinning process had no effect on the functional groups present in the resulting structure. The electrospun scaffolds were coated with PVPA (PCL/PVPA), and characterised. The stability of the PVPA coating after immersion in culture medium was assessed over 21 days. There was rapid release of the coating until day 2, after which the coating became stable. The wettability of the PCL scaffolds improved significantly, from 123.3 ± 10.8° to 43.3 ± 1.2° after functionalisation with PVPA. The compressive strength of the PCL/PVPA scaffolds (72 MPa) was significantly higher to that of the PCL scaffold (14 MPa), and an intermediate between trabecular and cortical bone (7 MPa and 170 MPa, resp.). The study has demonstrated that the PCL/PVPA scaffold has the desired chemical and biomechanical characteristics required for a material designed to be used as a BGS.

Aligned electrospun polymer fibres for skeletal muscle regeneration.

Skeletal muscle repair is often overlooked in surgical procedures and in serious burn victims. Creating a tissue-engineered skeletal muscle would not only provide a grafting material for these clinical situations, but could also be used as a valuable true-to-life research tool into diseases affecting muscle tissue. Electrospinning of the elastomer PLGA produced aligned fibres that had the correct topology to provide contact guidance for myoblast elongation and alignment. In addition, the electrospun scaffold required no surface modifications or incorporation of biologic material for adhesion, elongation, and differentiation of C2C12 murine myoblasts.

Three-dimensional cell culture of chondrocytes on modified di-phenylalanine scaffolds.

The design of self-assembled peptide-based structures for three-dimensional cell culture and tissue repair has been a key objective in biomaterials science for decades. In search of the simplest possible peptide system that can self-assemble, we discovered that combinations of di-peptides that are modified with aromatic stacking ligands could form nanometre-sized fibres when exposed to physiological conditions. For example, we demonstrated that a number of Fmoc (fluoren-9-ylmethyloxycarbonyl) modified di- and tri-peptides form highly ordered hydrogels via hydrogen-bonding and pi-pi interactions from the fluorenyl rings. These highly hydrated gels allowed for cell proliferation of chondrocytes in three dimensions [Jayawarna, Ali, Jowitt, Miller, Saiani, Gough and Ulijn (2006) Adv. Mater. 18, 611-614]. We demonstrated that fibrous architecture and physical properties of the resulting materials were dictated by the nature of the amino acid building blocks. Here, we report the self-assembly process of three di-phenylalanine analogues, Fmoc-Phe-Phe-OH, Nap (naphthalene)-Phe-Phe-OH and Cbz (benzyloxycarbonyl)-Phe-Phe-OH, to compare and contrast the self-assembly properties and cell culture conditions attributable to their protecting group difference. Fibre morphology analysis of the three structures using cryo-SEM (scanning electron microscopy) and TEM (transmission electron microscopy) suggested fibrous structures with dramatically varying fibril dimensions, depending on the aromatic ligand used. CD and FTIR (Fourier-transform IR) data confirmed beta-sheet arrangements in all three samples in the gel state. The ability of these three new hydrogels to support cell proliferation of chondrocytes was confirmed for all three materials.

Osteoblast responses to tape-cast and sintered bioactive glass ceramics.

The advantage of tape-cast bioactive glasses lies in the manufacturing procedure, which allows the build-up of layers and, therefore, the production of complex shapes. This, therefore, has applications to tissue engineering, where specific shapes are required such as repair of craniofacial defects. The bioactivity of tape-cast discs sintered at temperatures ranging from 800 degrees C to 1000 degrees C and for 3 or 6 h was analyzed by FTIR. Tape-cast discs were used to culture primary human osteoblasts, and cell attachment, cell death, collagen production, nodule formation, and mineralization were studied. These responses were dependent upon Si and Na release profiles of the tape-cast discs, and development of the hydroxyapatite layer.

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.

Osteoblast attachment and mineralized nodule formation on rough and smooth 45S5 bioactive glass monoliths.

Human primary osteoblast responses to smooth and roughened bioactive glass of 45S5 (Bioglass trade mark ) composition (46.1% SiO(2), 26.9% CaO, 2.6% P(2)O(5), 24.4% Na(2)O) were analysed in vitro. The smooth and rough surfaces had R(a) values and peak to valley distances of 0.04, 4.397, 2.027, and 21.328 microm, respectively. Cell attachment and morphology was observed using phalloidin staining of the actin cytoskeleton and revealed significant differences between smooth and rough surfaces. Cells that were spiky in appearance on the rough compared to the smooth surface formed an organized actin matrix much later on the rough surface. Scanning electron microscopy revealed many cell filipodia extending from more rounded cell bodies on the rough surface. A significantly greater number of nodules on the rough surface was observed, and these were shown to mineralize when supplemented with beta-glycerophosphate and dexamethasone. Raman spectroscopy confirmed the presence of hydroxyapatite in the mineralized cultures showing a definite peak at 964 cm(-1). FTIR analysis showed hydroxyapatite formation occurred more rapidly on the rough surface. This study demonstrates that although initial cell morphology was less advanced on the roughened surface, the cells were able to form mineralized nodules in greater numbers. This may have implications to bone tissue engineering using bioactive glasses.

In vitro evaluation of novel bioactive composites based on Bioglass-filled polylactide foams for bone tissue engineering scaffolds.

Highly porous poly(DL-lactic acid) (PDLLA) foams and Bioglass-filled PDLLA composite foams were characterized and evaluated in vitro as bone tissue engineering scaffolds. The hypothesis was that the combination of PDLLA with Bioglass in a porous structure would result in a bioresorbable and bioactive composite, capable of supporting osteoblast adhesion, spreading and viability. Composite and unfilled foams were incubated in simulated body fluid (SBF) at 37 degrees C to study the in vitro degradation of the polymer and to detect hydroxyapatite (HA) formation, which is a measure of the materials' in vitro bioactivity. HA was detected on all the composite samples after incubation in SBF for just 3 days. After 28 days immersion the foams filled with 40 wt % Bioglass developed a continuous layer of HA. The formation of HA for the 5 wt % Bioglass-filled foams was localized to the Bioglass particles. Cell culture studies using a commercially available (ECACC) human osteosarcoma cell line (MG-63) were conducted to assess the biocompatibility of the foams and cell attachment to the porous substrates. The osteoblast cell infiltration study showed that the cells were able to migrate through the porous network and colonize the deeper regions within the foam, indicating that the composition of the foams and the pore structures are able to support osteoblast attachment, spreading, and viability. Rapid formation of HA on the composites and the attachment of MG-63 cells within the porous network of the composite foams confirms the high in vitro bioactivity and biocompatibility of these materials and their potential to be used as scaffolds in bone tissue engineering and repair.

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.

In vitro bioactivity of S520 glass fibers and initial assessment of osteoblast attachment.

Bioactive glass fibers are attractive materials for use as tissue-engineering scaffolds and as the reinforcing phase for resorbable bioactive composites. The bioactivity of S520 glass fibers (52.0 mol % SiO(2), 20.9 Na(2)O, 7.1 K(2)O, 18.0 CaO, and 2.0 P(2)O(5)) was evaluated in two media, simulated body fluid (SBF) and Dulbecco's modified Eagle's medium (DMEM), for up to 20 days at 37 degrees C. Hydroxyapatite formation was observed on S520 fiber surfaces after 5 h in SBF. After a 20-day immersion, a continuous hydroxyapatite layer was present on the surface of samples immersed in SBF as well as on those samples immersed in DMEM [fiber surface area to solution volume ratio (SA:V) of 0.10 cm(2)/mL]. Backscattered electron imaging and EDS analysis revealed that the hydroxyapatite layer formation was more extensive for samples immersed in SBF. Decreasing the SA:V ratio to 0.05 cm(2)/mL decreased the time required to form a continuous hydroxyapatite surface layer. ICP was used to reveal Si, Ca, and P release profiles in DMEM after the 1st h (15.1, 83.8, and 29.7 ppm, respectively) were similar to those concentrations previously determined to stimulate gene expression in osteoblasts in vitro (16.5, 83.3, and 30.4 ppm, respectively). The tensile strength of the 20-microm diameter fibers was 925 +/- 424 MPa. Primary human osteoblast attachment to the fiber surface was studied by using SEM, and mineralization was studied by using alizarin red staining. Osteoblast dorsal ruffles, cell projections, and lamellipodia were observed, and by 7 days, cells had proliferated to form monolayer areas as shown by SEM. At 14 days, nodule formation was observed, and these nodules stained positive for alizarin red, demonstrating Ca deposition and, therefore mineralization.

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.

Scaffolds and biomaterials for tissue engineering: a review of clinical applications.

Tissue engineering is a multidisciplinary area of research aimed at regeneration of tissues and restoration of organ function. This is achieved through implantation of cells/tissues grown outside the body or by stimulating cells to grow into an implanted matrix. In this short review, we discuss the use of biomaterials, in the form of scaffolds, for tissue engineering and review clinical applications to otorhinolaryngology-head and neck surgery.

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.

Novel bioresorbable and bioactive composites based on bioactive glass and polylactide foams for bone tissue engineering.

Bioresorbable and bioactive tissue engineering scaffolds based on bioactive glass (45S5 Bioglass(R)) particles and macroporous poly(DL-lactide) (PDLLA) foams were fabricated. A slurry dipping technique in conjunction with pretreatment in ethanol was used to achieve reproducible and well adhering bioactive glass coatings of uniform thickness on the internal and external surfaces of the foams. In vitro studies in simulated body fluid (SBF) demonstrated rapid hydroxyapatite (HA) formation on the surface of the composites, indicating their bioactivity. For comparison, composite foams containing Bioglass(R) particles as filler for the polymer matrix (in concentration of up to 40 wt %) were prepared by freeze-drying, enabling homogenous glass particle distribution in the polymer matrix. The formation of HA on the composite surfaces after immersion in phosphate buffer saline (PBS) was investigated to confirm the bioactivity of the composites. Human osteoblasts (HOBs) were seeded onto as-fabricated PDLLA foams and onto PDLLA foams coated with Bioglass(R) particles to determine early cell attachment and spreading. Cells were observed to attach and spread on all surfaces after the first 90 min in culture. The results of this study indicate that the fabricated composite materials have potential as scaffolds for guided bone regeneration.

Osteoblast cell death on methacrylate polymers involves apoptosis.

The success of an implant depends on the implant-tissue interface. There are many causes of implant failure, one of which is tissue necrosis. The aim of this in vitro study was to determine whether cell death of primary human osteoblasts (implant site specific cells) occurred by apoptosis (a form of programmed cell death) on two methacrylate polymers. Cells were cultured on poly(ethyl methacrylate)/tetrahydrofurfuryl methacrylate and poly(methyl methacrylate in the form of 13-mm discs, in conditioned medium containing leachable monomer and in the presence of various concentrations of monomer itself in the culture medium. It was found that monomer and leached monomer caused apoptosis of human osteoblast cells in this system. Tetrahydrofurfuryl methacrylate monomer was found to be more toxic than currently used monomer methylmethacrylate. Preincubation of polymers in serum containing medium was found to increase the biocompatibility of the polymers. High levels of apoptosis occurred on polymer used directly after polymerization. Apoptosis levels were decreased after polymer was incubated at 60 degrees C overnight or for 3 days. Apoptosis therefore may occur in cells at the implant site in vivo.

Are emergency department patients at risk for herb-drug interactions?

OBJECTIVES: To determine the prevalence of herbal and/or dietary supplement use and identify patients at risk for herb-drug interactions. METHODS: A convenience sample of 944 patients were surveyed to determine the prevalence and types of supplements used. Patients with heart disease, diabetes, psychiatric disorders, and/or hypertension were assessed for potential interactions. RESULTS: One hundred thirty-five (14.3%) patients reported regular use. Of these, 79.3% were taking supplements concurrently with prescription medications, and 80.0% were administered medication(s) in the emergency department. Cardiac: 19.8% (n = 33) reported regular use, with four potential interactions. Hypertension: 20.3% (n = 54) reported regular use, with two potential interactions. Diabetes: 15.9% (n = 20) reported regular use, with no known interactions. Psychiatric: 15.9% (n = 10) reported regular use, with one potential interaction. CONCLUSIONS: Six patients were identified at risk for seven known herb-drug interactions. The prevalence of undisclosed herbal supplement use and lack of research on these supplements suggest that more patients may be at risk.

Exertional rhabdomyolysis in a body builder abusing anabolic androgenic steroids.

Rhabdomyolysis, or acute skeletal muscle destruction, may be accompanied by myoglobinaemia, myoglobinuria, and an elevated serum creatine kinase level. This disorder has many potential causes. In this article, the authors describe a case of rhabdomyolysis occurring after vigorous weight lifting by a man who was supplementing his weight-training programme with the intake of anabolic androgenic steroids dispensed to him by a colleague.

Complications of a retrograde intubation in a trauma patient.

The authors report the case of an elder woman involved in a motor vehicle collision (MVC) requiring emergent intubation using the technique of retrograde intubation (RI). Since RI is a blind technique, potential complications arising from its use are numerous and may result in increased morbidity and mortality. Such was the case of this RI that involved incorrect placement of the endotracheal tube (ETT), resulting in suboptimal ventilation and increased morbidity. Additionally, this case illustrates how the failure to detect this error in multiple settings (ambulance, helicopter, emergency department) led to unnecessary and potentially deleterious procedures and significant delay in providing the basics of trauma care, oxygenation and ventilation. Although theoretical complications of RI have been addressed in the past, there have been very few published reports of actual complications. The emergency physician must be aware of difficult airways, options available to establish alternative airways, and methods to confirm appropriate placement of the ETT. The authors also discuss the indications, procedures, and complications involved in performing an RI.