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Articles by Alexander Bismarck in JoVE

 JoVE Bioengineering

Ex vivo Mimetismo de normal e anormal Hematopoese Humanos


JoVE 3654 4/10/2012

1Department of Chemical Engineering and Chemical Technology, South Kensington campus, Imperial College London, 2Department of Hematology, Northwick Park & St. Mark's campus, Imperial College London

Um sistema de cultura de 3D para a hematopoiese é descrito usando sangue do cordão umbilical humano e células de medula óssea leucémicas. O método baseia-se na utilização de um poliuretano poroso sintético andaime revestido com proteínas da matriz extracelular. Este andaime é adaptável para acomodar uma grande variedade de células.

Other articles by Alexander Bismarck on PubMed

Wetting Behavior of Flax Fibers As Reinforcement for Polypropylene

The wetting behavior of several flax (cellulose as reference) and polypropylene fibers is characterized by measuring the wetting rates (penetration velocities) of a series of liquids using the capillary rise technique. This present paper aims to provide a deeper understanding of the complex nature of natural fibers and their surface properties. The fiber surface tensions are estimated from plots of the normalized wetting rate as a function of the surface tension of the liquids assuming, in analogy to Zisman's method, that the maximum of the normalized wetting rate corresponds to the solid surface tension. The estimated surface tensions of the investigated flax fibers indicate that all the fibers are quite "hydrophobic." The method used to separate the fibers from the rest of the plants has a large influence on the estimated fiber surface tensions. In the case of polypropylene (PP) fibers, the estimated surface tension corresponds well with literature data. Grafting small amounts of maleic acid anhydrite (MAH) onto the PP surfaces will not affect the wetting behavior and, therefore, the surface tension, whereas grafting larger amounts (10 wt%) of MAH causes the polymer surface tension to increase significantly. Additional pH-dependent zeta-potential measurements show that even the "pure" PP-fibers contain acidic surface functions, possibly due to further processing at elevated temperatures (thermal degradation or other aging processes).

Towards a Methodology for the Effective Surface Modification of Porous Polymer Scaffolds

A novel low-pressure radio-frequency plasma treatment protocol was developed to achieve the effective through-thickness surface modification of large porous poly (D,L-lactide) (PDLLA) polymer scaffolds using air or water: ammonia plasma treatments. Polymer films were modified as controls. Scanning electron micrographs and maximum bubble point measurements demonstrated that the PDLLA foams have the high porosity, void fraction and interconnected pores required for use as tissue engineering scaffolds. The polymer surface of the virgin polymer does contain acidic functional groups but is hydrophobic. Following exposure to air or water: ammonia plasma, an increased number of polar functional groups and improved wetting behaviour, i.e. hydrophilicity, of wet surfaces was detected. The number of polar surface functional groups increased (hence the decrease in water contact angles) with increasing exposure time to plasma. The change in surface composition and wettablility of wet polymer constructs was characterised by zeta potential and contact angle measurements. The hydrophobic recovery of the treated PDLLA polymer surfaces was also studied. Storage of the treated polymer constructs in ambient air caused an appreciable hydrophobic recovery, whereas in water only partial hydrophobic recovery occurred. However, in both cases the initial surface characteristics decay as function of time.

Anisotropic Surface Energetics and Wettability of Macroscopic Form I Paracetamol Crystals

Advancing (theta(A)) and receding (theta(R)) contact angles were measured with several probe liquids on the external facets (201), (001), (011), and (110) of macroscopic form I paracetamol crystals as well as the cleaved (internal) facet (010). For the external crystal facets, dispersive surface energies gamma(d) calculated from the contact angles were found to be similar (34 +/- 1 mJ/m(2)), while the polar components varied significantly. Cleaving the crystals exposed a more apolar (010) surface with very different surface properties, including gamma(d) = 45 +/- 1 mJ/m(2). The relative surface polarity (gamma(p)/gamma) of the facets in decreasing order was (001) > (011) > (201) > (110) > (010), which agreed with the fraction of exposed polar hydroxyl groups as determined from C and O 1s X-ray photoelectron spectroscopy (XPS) spectra, and could be correlated with the number of non-hydrogen-bonded hydroxyl groups per unit area present for each crystal facet, based on the known crystal structures. In conclusion, all facets of form I paracetamol crystals examined exhibited anisotropic wetting behavior and surface energetics that correlated to the presence of surface hydroxyl groups.

Nondestructive Technique for the Characterization of the Pore Size Distribution of Soft Porous Constructs for Tissue Engineering

Polymer scaffolds tailored for tissue engineering applications possessing the desired pore structure require reproducible fabrication techniques. Nondestructive, quantitative methods for pore characterization are required to determine the pore size and its distribution. In this study, a promising alternative to traditional pore size characterization techniques is presented. We introduce a quantitative, nondestructive and inexpensive method to determine the pore size distribution of large soft porous solids based on the on the displacement of a liquid, that spreads without limits though a porous medium, by nitrogen. The capillary pressure is measured and related to the pore sizes as well as the pore size distribution of the narrowest bottlenecks of the largest interconnected pores in a porous medium. The measured pore diameters correspond to the narrowest bottleneck of the largest pores connecting the bottom with the top surface of a given porous solid. The applicability and reproducibility of the breakthrough technique is demonstrated on two polyurethane foams, manufactured using the thermally induced phase separation (TIPS) process, with almost identical overall porosity (60-70%) but very different pore morphology. By selecting different quenching temperatures to induce polymer phase separation, the pore structure could be regulated while maintaining the overall porosity. Depending on the quenching temperature, the foams exhibited either longitudinally oriented tubular macropores interconnected with micropores or independent macropores connected to adjacent pores via openings in the pore walls. The pore size and its distribution obtained by the breakthrough test were in excellent agreement to conventional characterization techniques, such as scanning electron microscopy combined with image analysis, BET technique, and mercury intrusion porosimetry. This technique is suitable for the characterization of the micro- and macropore structure of soft porous solids intended for tissue engineering applications. The method is sensitive for the smallest bottlenecks of the largest continuous pores throughout the scaffold that contributes to fluid flow.

Anisotropic Surface Chemistry of Crystalline Pharmaceutical Solids

The purpose of this study was to establish the link between the wetting behavior of crystalline pharmaceutical solids and the localized surface chemistry. A range of conventional wetting techniques were evaluated and compared with a novel experimental approach: sessile drop contact angle measurements on the individual facets of macroscopic (>1 cm) single crystals. Conventional measurement techniques for determining surface energetics such as capillary rise and sessile drops on powder compacts were found not to provide reliable results. When the macroscopic crystal approach was used, major differences for advancing contact angles, theta(a), of water were observed-as low as 16 degrees on facet (001) and as high as 68 degrees on facet (010) of form I paracetamol. theta(a) trends were in excellent agreement with X-ray photoelectron spectroscopy surface composition and known crystallographic structures, suggesting a direct relationship to the local surface chemistry. Inverse gas chromatography (IGC) was further used to probe the surface properties of milled and unmilled samples, as a function of particle size. IGC experiments confirmed that milling exposes the weakest attachment energy facet, with increasing dominance as particle size is reduced. The weakest attachment energy facet was also found to exhibit the highest theta(a) for water and to be the most hydrophobic facet. This anisotropic wetting behavior was established for a range of crystalline systems: paracetamol polymorphs, aspirin, and ibuprofen racemates. theta(a) was found to be very sensitive to the local surface chemistry. It is proposed that the hydrophilicity/hydrophobicity of facets reflects the presence of functional groups at surfaces to form hydrogen bonds with external molecules.

Anisotropic Surface Chemistry of Aspirin Crystals

The wettability of the (001), (100), and (011) crystallographic facets of macroscopic aspirin crystals has been experimentally investigated using a sessile drop contact angle (theta) method. theta for a nonpolar liquid was very similar for all three facets, though significant theta differences were observed for three polar probe liquids. The observed hydrophobicity of the (001) and (100) facets is ascribed to a reduced hydrogen bonding potential at these surfaces, whilst the observed hydrophilicity of facet (011) may be attributed to presence of surface carboxylic functionalities as confirmed by X-ray photoelectron spectroscopy (XPS). The dispersive component of the surface free energy (gamma(s)(d)) was similar for all three facets (35 +/- 2 mJ/m2). The total surface energy, gammas varied between 46 and 60 mJ/m2 due to significant variations in the polar/acid-base components of gamma for all facets. Surface polarity as determined by gamma measurements and XPS data were in good agreement, linking the variations in wettability to the concentration of oxygen containing surface functional groups. In conclusion, the wettability and the surface energy of a crystalline organic solid, such as aspirin, was found to be anisotropic and facet dependant, and in this case, related to the presence of surface carboxylic functionalities.

Removal of Oxidation Debris from Multi-walled Carbon Nanotubes

Conventional liquid phase oxidation of multiwall carbon nanotubes (MWCNTs) using concentrated acids generates contaminating debris that should be removed using aqueous base before further reaction.

Particle-stabilized Surfactant-free Medium Internal Phase Emulsions As Templates for Porous Nanocomposite Materials: Poly-Pickering-Foams

We report on the successful use of particle-stabilized Medium Internal Phase Emulsion (MIPE) templates for the synthesis of porous polymer foams. In this case, carbon nanotubes (CNTs) were used to stabilize the minority phase as the continuous phase, through adsorption at the interface. The addition of the CNTs not only provides processing advantages (no need for traditional non-ionic molecular surfactants) but also enhances the mechanical and electrical properties of the final polyFoams. This approach allows the manufacture of both closed- and open-celled porous polymer foams in a one-pot process with porosities up to 66%.

Atmospheric Plasma Treatment of Porous Polymer Constructs for Tissue Engineering Applications

Porous 3D polymer scaffolds prepared by TIPS from PLGA (53:47) and PS are intrinsically hydrophobic which prohibits the wetting of such porous media by water. This limits the application of these materials for the fabrication of scaffolds as supports for cell adhesion/spreading. Here we demonstrate that the interior surfaces of polymer scaffolds can be effectively modified using atmospheric air plasma (AP). Polymer films (2D) were also modified as control. The surface properties of wet 2D and 3D scaffolds were characterised using zeta-potential and wettability measurements. These techniques were used as the primary screening methods to assess surface chemistry and the wettability of wet polymer constructs prior and after the surface treatment. The surfaces of the original polymers are rather hydrophobic as highlighted but contain acidic functional groups. Increased exposure to AP improved the water wetting of the treated surfaces because of the formation of a variety of oxygen and nitrogen containing functions. The morphology and pore structure was assessed using SEM and a liquid displacement test. The PLGA and PS foam samples have central regions which are open porous interconnected networks with maximum pore diameters of 49 microm for PLGA and 73 microm for PS foams.

Inducing PH Responsiveness Via Ultralow Thiol Content in Polyacrylamide (micro)gels with Labile Crosslinks

Here we present the synthesis and characterization of pH responsive polyacrylamide microgels, synthesized via free radical polymerization of acrylamide and bis (acryloylcystamine) (BAC). The gels were made with ultralow amounts of thiol functional groups incorporated into the polymer. The resulting gel monoliths were mechanically chopped into microgel particles with size distributions ranging from 80 to 200 mum. The gels exhibit an interesting reversible pH-dependent rheological behavior which led to gelling of the colloidal suspension when the pH was increased, and a low-viscosity suspension was obtained when the pH was taken back to the original value. The viscosity of the colloidal system containing MBA crosslinked microgels remained insensitive to pH. This observation motivated further analysis; viscosity measurements of the highly viscous (gel-like) state of the BAC crosslinked microgel colloidal suspension were carried out to further understand the rheological behavior of the colloidal system. Electrophoretic mobility measurements as function of pH of the BAC and MBA crosslinked colloidal polyacrylamide microgel suspensions were performed. The swelling behavior of the microgels for both colloidal systems was also determined as function of pH using static light scattering. This swelling behavior was used to rationalize the observed rheological behavior. The work presented here demonstrates that free thiol groups present within a polymer gel matrix confer pH responsive behavior to the gel in solution. The viscosity of a BAC crosslinked microgel suspension was also measured under reducing conditions. The viscosity of the microgel suspension reduced with time, due to the breakage of the disulfide bonds in the crosslinkers.

Interfacial Behavior Between Atmospheric-plasma-fluorinated Carbon Fibers and Poly(vinylidene Fluoride)

Atmospheric-plasma fluorination was used to introduce fluorine functionalities onto the surface of carbon fibers without affecting their bulk properties. The interfacial adhesion between atmospheric-plasma-fluorinated carbon fibers and poly(vinylidene fluoride) (PVDF) was studied by means of direct wetting measurements and single fiber pullout tests. Measured contact angles of PVDF melt droplets on modified carbon fibers show that short exposure times of carbon fibers to atmospheric-plasma fluorination (corresponding to a degree of surface fluorination of F/C = 0.01 (1.1%)) leads to improved wettability of the fibers by PVDF melts. The apparent interfacial shear strength as a measure of practical adhesion, determined by the single-fiber pullout test, increases by 65% under optimal treatment conditions. The improved practical adhesion is not due to the formation of transcrystalline regions around the fibers or a change of the bulk matrix crystallinity or to an increased surface roughness; it seems to be due to the compatibilization of the interface caused of the atmospheric-plasma fluorination of the carbon fibers.

High Internal Phase Emulsion Templates Solely Stabilised by Functionalised Titania Nanoparticles

Porous polymer foams (poly-Pickering-HIPEs) have been synthesised from stable high internal phase emulsion templates solely stabilised by low concentrations of functionalised titania nanoparticles.

Characterisation of a Soft Elastomer Poly(glycerol Sebacate) Designed to Match the Mechanical Properties of Myocardial Tissue

The myocardial tissue lacks significant intrinsic regenerative capability to replace the lost cells. Therefore, the heart is a major target of research within the field of tissue engineering, which aims to replace infarcted myocardium and enhance cardiac function. The primary objective of this work was to develop a biocompatible, degradable and superelastic heart patch from poly(glycerol sebacate) (PGS). PGS was synthesised at 110, 120 and 130 degrees C by polycondensation of glycerol and sebacic acid with a mole ratio of 1:1. The investigation was focused on the mechanical and biodegrading behaviours of the developed PGS. PGS materials synthesised at 110, 120 and 130 degrees C have Young's moduli of 0.056, 0.22 and 1.2 MPa, respectively, which satisfy the mechanical requirements on the materials applied for the heart patch and 3D myocardial tissue engineering construction. Degradation assessment in phosphate buffered saline and Knockout DMEM culture medium has demonstrated that the PGS has a wide range of degradability, from being degradable in a couple of weeks to being nearly inert. The matching of physical characteristics to those of the heart, the ability to fine tune degradation rates in biologically relevant media and initial data showing biocompatibility indicate that this material has promise for cardiac tissue engineering applications.

Through-thickness Plasma Modification of Biodegradable and Nonbiodegradable Porous Polymer Constructs

Pure poly(lactide-co-glycolide) and polystyrene surfaces are not very suitable to support cell adhesion/spreading owing to their hydrophobic nature and low surface energy. The interior surfaces of large porous 3D scaffolds were modified and activated using radio-frequency, low-pressure air plasma. An increase in the wettability of the surface was observed after exposure to air plasma, as indicated by the decrease in the contact angles of the wet porous system. The surface composition of the plasma-treated polymers was studied using X-ray photoelectron spectroscopy. pH-dependent zeta-potential measurements confirm the presence of an increased number of functional groups. However, the plasma-treated surfaces have a less acidic character than the original polymer surfaces as seen by a shift in their isoelectric point. Zeta-potential, as well as contact angle measurements, on 3D scaffolds confirm that plasma treatment is a useful tool to modify the surface properties throughout the interior of large scaffolds.

Surface Modification of Natural Fibers Using Bacteria: Depositing Bacterial Cellulose Onto Natural Fibers to Create Hierarchical Fiber Reinforced Nanocomposites

Triggered biodegradable composites made entirely from renewable resources are urgently sought after to improve material recyclability or be able to divert materials from waste streams. Many biobased polymers and natural fibers usually display poor interfacial adhesion when combined in a composite material. Here we propose a way to modify the surfaces of natural fibers by utilizing bacteria ( Acetobacter xylinum) to deposit nanosized bacterial cellulose around natural fibers, which enhances their adhesion to renewable polymers. This paper describes the process of modifying large quantities of natural fibers with bacterial cellulose through their use as substrates for bacteria during fermentation. The modified fibers were characterized by scanning electron microscopy, single fiber tensile tests, X-ray photoelectron spectroscopy, and inverse gas chromatography to determine their surface and mechanical properties. The practical adhesion between the modified fibers and the renewable polymers cellulose acetate butyrate and poly(L-lactic acid) was quantified using the single fiber pullout test.

Direct Measurement of the Wetting Behavior of Individual Carbon Nanotubes by Polymer Melts: the Key to Carbon Nanotube-polymer Composites

Carbon nanotube wetting is the critical parameter for the development of nanocomposites yet, due to the lack of suitable methods of qualifying and, more importantly, quantifying nanoscale wetting and adhesion phenomena, it is often overlooked. Here, we discuss a qualitative approach that provides wetting/nonwetting information and present a microfluidics method to produce "nanodroplets" of polymers on individual carbon nanotubes which enable the direct quantification of contact angles.

High Internal Phase Emulsions Stabilized Solely by Functionalized Silica Particles

Polyaniline Hollow Fibres for Organic Solvent Nanofiltration

Intrinsically-skinned asymmetric PANi hollow fibres, fabricated using a process of directly adding large organic acids to highly concentrated PANi solutions, show stability in a wide variety of organic solvents and have shown promising nanofiltration properties, giving high rejections of nanosolutes in acetone.

Inverse Gas Chromatography of As-received and Modified Carbon Nanotubes

The surface properties of chemical vapor deposition (CVD)-grown, multiwalled carbon nanotubes (CNTs) have been studied using inverse gas chromatography (IGC). By adapting known IGC methodologies to these challenging materials, the surface character of a broad range of CNT materials can be reliably compared and quantified in terms of dispersive and specific surface energies, electron acceptor and donor numbers, and adsorption capacities. The effect of CNT surface modification by high temperature annealing, thermal oxidation, and grafting of methyl methacrylate was explored. The IGC surface characterization of these materials was consistent with results from other surface-sensitive analytical techniques, including X-ray photoelectron spectroscopy (XPS), titration, and electron microscopy, confirming the validity and sensitivity of our approaches. The same IGC methodologies were successfully applied to characterize three as-received CNT materials which differed significantly in their specific surface areas and functional surface group concentrations.

Reactive Polyurethane Carbon Nanotube Foams and Their Interactions with Osteoblasts

The remarkable intrinsic properties of carbon nanotubes, including their high mechanical strength, electrical conductivity, and nanoscale 3D architecture, create promising opportunities for the use of nanotube composites in a number of fields, particularly for composites in which conventional fillers cannot be accommodated. In the current study, 3D polyurethane (PU) nanocomposite foams were developed, and their potential biomedical applications were investigated. Multiwalled carbon nanotubes (CNTs) were synthesized by chemical vapor deposition and, following suitable chemical modification, uniformly distributed within the walls of PU foams produced by direct reaction. Although the loading fraction was too low to observe significant mechanical effects, CNT incorporation improved the wettability of the nanocomposite surfaces in a concentration-dependent manner, supporting the claim that the nanotubes are active at the pore surface. Studies of bone cell interactions with the nanocomposite foams revealed that increasing CNT loading fraction did not cause osteoblast cytotoxicity nor have any detrimental effects on osteoblast differentiation or mineralization. The application of "fixed" or embedded CNTs in nondegradable scaffolds is likely advantageous over "loose" or unattached CNTs from a toxicological point of view.

Premature Degradation of Poly(alpha-hydroxyesters) During Thermal Processing of Bioglass-containing Composites

Bioactive, biodegradable composites are increasingly being explored as bone replacement materials and as scaffolds for tissue engineering. Their properties are not only dependent on the properties of the filler and matrix, but are also determined by their interaction. This study investigated the effect on poly(D,L-lactide) (PDLLA) matrix when processed at high-temperatures in the presence of Bioglass particulate filler. Composites with different filler contents were compounded at elevated temperatures by co-extrusion followed by compression moulding and compared with composites of similar composition prepared by thermally induced phase separation (TIPS), a low-temperature processing route. It was found that the inclusion of Bioglass in PDLLA under elevated temperatures resulted in the degradation of the matrix, leading to a reduction in the mechanical properties of the composites and in the molecular weight of the matrix. Attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy showed the presence of a peak at 1600 cm(-1) in the composite material, particularly when processed at elevated temperatures, whereas no peak at this wavelength was discernible for the pure PDLLA. Furthermore, time-based ATR-FTIR spectra taken at elevated temperatures on the TIPS-processed composites showed an increase in the intensity of the peak at 1600 cm(-1) and a concomitant reduction of the CO stretch peak at 1745 cm(-1) with time. This suggested the formation of a carboxylate salt by-products as a consequence of a reaction at the interface between the Bioglass filler and the PDLLA matrix. Therefore, the results confirmed that this degradation was not solely due to shear effects during the extrusion process. This work thereby supports the assertion that, in the presence of Bioglass filler particles, poly(alpha-hydroxyester)-based composites should not be processed at elevated temperatures.

Tissue Engineering of Lung: the Effect of Extracellular Matrix on the Differentiation of Embryonic Stem Cells to Pneumocytes

We have previously differentiated lung epithelium from human and murine embryonic stem cells (mESCs) and are now exploring the potential applications of these cells, including in the engineering of lung tissue constructs. In this study, we hypothesized that the differentiation and maintenance of lung epithelium derived from mESCs can be enhanced by extracellular matrix (ECM) proteins. Our established differentiation protocol was applied to mESCs grown on a range of ECMs: collagen I, laminin 332, fibronectin, Matrigel, and, as an experimental control, gelatin. The ECMs were coated onto tissue culture plastic (TCP) and poly-DL-lactic acid (PDLLA), a biodegradable polymer we have previously shown to support the growth of mature pneumocytes. Matrigel or Laminin-332 coating of either TCP or PDLLA film resulted in enhanced surfactant protein C gene expression in differentiating mESCs, a direct indication of the upregulation of lung epithelial differentiation. For each combination, changes in the contact angle and zeta potential of protein-coated TCP and PDLLA film confirmed protein adsorption. We conclude that the choice of the coating protein can greatly affect the differentiation of ESCs, and laminin-332-coated PDLLA provided an ECM-degradable scaffold combination that is suitable for engineering of lung tissue constructs.

The Development of a Three-dimensional Scaffold for Ex Vivo Biomimicry of Human Acute Myeloid Leukaemia

Acute myeloid leukaemia (AML) is a cancer of haematopoietic cells that develops in three-dimensional (3-D) bone marrow niches in vivo. The study of AML has been hampered by lack of appropriate ex vivo models that mimic this microenvironment. We hypothesised that fabrication and optimisation of suitable biomimetic scaffolds for culturing leukaemic cells ex vivo might facilitate the study of AML in its native 3-D niche. We evaluated the growth of three leukaemia subtype-specific cell lines, K-562, HL60 and Kasumi-6, on highly porous scaffolds fabricated from biodegradable and non-biodegradable polymeric materials, such as poly (L-lactic-co-glycolic acid) (PLGA), polyurethane (PU), poly (methyl-methacrylate), poly (D, L-lactade), poly (caprolactone), and polystyrene. Our results show that PLGA and PU supported the best seeding efficiency and leukaemic growth. Furthermore, the PLGA and PU scaffolds were coated with extracellular matrix (ECM) proteins, collagen type I (62.5 or 125 microg/ml) and fibronectin (25 or 50 microg/ml) to provide biorecognition signals. The 3 leukaemia subtype-specific lines grew best on PU scaffolds coated with 62.5 microg/ml collagen type I over 6 weeks in the absence of exogenous growth factors. In conclusion, PU-collagen scaffolds may provide a practical model to study the biology and treatment of primary AML in an ex vivo mimicry.

High-porosity Macroporous Polymers Sythesized from Titania-particle-stabilized Medium and High Internal Phase Emulsions

Particle-stabilized high internal phase emulsions have been used to synthesize tough and very high porosity macroporus polymers with a closed-cell pore structure. In this study, we show that Pickering water-in-oil emulsion templates with up to an 85 vol % internal phase can be stabilized by only 1 wt % of titania particles with their surfaces suitably modified by the adsorption of 3.5 +/- 0.5 wt % oleic acid. The pore structure and mechanical properties of the resulting macroporous polymers were tailored by altering the internal phase volume ratio of the emulsion template and the titania particle concentration used to stabilize the emulsion templates. The pore size and pore size distributions increase with increasing internal phase volume of the emulsion template as well as decreasing titania particle concentration used to stabilize the emulsion template. The mechanical properties, namely, Young's modulus and the crush strength of the macroporous polymers, increased with decreasing porosity and increasing foam density. The toughest macroporous polymer had the lowest porosity but also the smallest pore size and narrowest pore size distribution.

Effects of Fibroblast Growth Factors on the Differentiation of the Pulmonary Progenitors from Murine Embryonic Stem Cells

The fibroblast growth factors (FGFs) play an important role in the development of embryonic lung. In this study, we investigated the effects of mainly FGF 1, 2, and 10 at concentrations selected on the basis of data obtained from previous in vitro culture on the derivation of the pulmonary progenitors from murine embryonic stem cells cultured on gelatin or Matrigel-coated plates. For cells cultured on a gelatin-coated plate, high concentrations of FGF1 were found to enhance the expression of mRNAs for SPC and CC10, markers of distal airway epithelium, while high levels of FGF2 decreased the expression of RNAs for not only SPC, CC10 but also for the additional markers SPD and aquaporin 5. FGF10 at all tested concentrations was found to have no effect on the differentiation of pneumocytes when ESCs were grown on gelatin-coated plates. However, when differentiation was performed on Matrigel-coated plates, the addition of 60 ng/ml FGF10 enhanced the expression of pneumocyte markers, suggesting a synergic effect of FGF10 and extracellular matrix. In conclusion, growth factors were proven to be effective in the differentiation of pulmonary progenitors from mESCs. The need of signals from extracellular matrix proteins depends on the growth factors supplemented.

Highly Permeable Macroporous Polymers Synthesized from Pickering Medium and High Internal Phase Emulsion Templates

Structure, Morphology and Thermal Characteristics of Banana Nano Fibers Obtained by Steam Explosion

In this work, cellulose nanofibers were extracted from banana fibers via a steam explosion technique. The chemical composition, morphology and thermal properties of the nanofibers were characterized to investigate their suitability for use in bio-based composite material applications. Chemical characterization of the banana fibers confirmed that the cellulose content was increased from 64% to 95% due to the application of alkali and acid treatments. Assessment of fiber chemical composition before and after chemical treatment showed evidence for the removal of non-cellulosic constituents such as hemicelluloses and lignin that occurred during steam explosion, bleaching and acid treatments. Surface morphological studies using SEM and AFM revealed that there was a reduction in fiber diameter during steam explosion followed by acid treatments. Percentage yield and aspect ratio of the nanofiber obtained by this technique is found to be very high in comparison with other conventional methods. TGA and DSC results showed that the developed nanofibers exhibit enhanced thermal properties over the untreated fibers.

Cross-linked Bacterial Cellulose Networks Using Glyoxalization

In this study, we demonstrate that bacterial cellulose (BC) networks can be cross-linked via glyoxalization. The fracture surfaces of samples show that, in the dry state, less delamination occurs for glyoxalized BC networks compared to unmodified BC networks, suggesting that covalent bond coupling between BC layers occurs during the glyoxalization process. Young's moduli of dry unmodified BC networks do not change significantly after glyoxalization. The stress and strain at failure are, however, reduced after glyoxalization. However, the wet mechanical properties of the BC networks are improved by glyoxalization. Raman spectroscopy is used to demonstrate that the stress-transfer efficiency of deformed dry and wet glyoxalized BC networks is significantly increased compared to unmodified material. This enhanced stress-transfer within the networks is shown to be a consequence of the covalent coupling induced during glyoxalization and offers a facile route for enhancing the mechanical properties of BC networks for a variety of applications.

Macroporous Polymers with Hierarchical Pore Structure from Emulsion Templates Stabilised by Both Particles and Surfactants

Inspired by natural porous materials, such as wood, bamboo and spongy bone consisting of individual structural units that are hierarchically arranged to optimise mechanical properties such as strength and toughness, synthetic macroporous polymers with enhanced physical properties were created by emulsion templating. Hierarchical poly(merised) high internal phase emulsions (HIPE) were created from HIPEs stabilised simultaneously by particles and a surfactant. In these HIPEs, surfactant stabilised and particle stabilised water droplets coexist, which upon polymerisation of the minority oil phase gives rise to macroporous polymers with a hierarchical pore structure. An improvement of the mechanical properties of our hierarchically structured macroporous polymers at equal porosity was observed, due to a more efficient packing of pores in a configuration that improves mechanical strength despite the presence of interconnecting pore throats. Moreover, the permeability of the hierarchically structured polyHIPEs are exceeding those measured for conventional polyHIPEs made from surfactant only stabilised HIPEs.

Macroporous Polymers Obtained in Highly Concentrated Emulsions Stabilized Solely with Magnetic Nanoparticles

Magnetic macroporous polymers have been successfully prepared using Pickering high internal phase ratio emulsions (HIPEs) as templates. To stabilize the HIPEs, two types of oleic acid-modified iron oxide nanoparticles (NPs) were used as emulsifiers. The results revealed that partially hydrophobic NPs could stabilize W/O HIPEs with an internal phase above 90%. Depending upon the oleic acid content, the nanoparticles showed either an arrangement at the oil-water interface or a partial dispersion into the oil phase. Such different abilities to migrate to the interface had significant effects on the maximum internal phase fraction achievable and the droplet size distribution of the emulsions. Highly macroporous composite polymers were obtained by polymerization in the external phase of these emulsions. The density, porosity, pore morphology and magnetic properties were characterized as a function of the oleic acid content, concentration of NPs, and internal phase volume of the initial HIPEs. SEM imaging indicated that a close-cell structure was obtained. Furthermore, the composite materials showed superparamagnetic behavior and a relatively high magnetic moment.

Long-term Cytokine-free Expansion of Cord Blood Mononuclear Cells in Three-dimensional Scaffolds

Cord blood expansion ex vivo can be achieved in liquid suspension through the addition of cytokines at the expense of often undesirable cell differentiation. In order to derive a cytokine-free dynamic culture system, we hypothesised that a three-dimensional (3D) environment in the form of highly porous scaffolds made of poly (D,L-lactide-co-glycolide) (PLGA) or polyurethane (PU) for the biomimetic growth of cord blood mononuclear cells (CBMNCs), would facilitate expansion of hematopoietic cells without exogenous cytokines. Both scaffolds supported cellular expansion ex vivo. Cytokine-free, long-term culture was best in PU coated with collagen type I (54-fold expansion). In contrast, traditional 2D well-plate cultures collapsed within 4 days in the absence of cytokines. CBMNCs cultured in the scaffolds were visualised by scanning electron microscopy and immunophenotypic/immunostaining analysis and the studies validated the presence of a dynamic culture containing erythroid precursors (CD45(-)/CD71(+)/CD235a(+)), hematopoietic stem/progenitor cells (CD38(-)CD34(+), CD117(+)), maturing myeloid cells (CD38(+), MPO(+)), CD4(+) and CD8(+) T-lymphocytes and megakaryocytes (FVIII(+)). Colony forming unit (CFU) assays indicated that BFU-E and CFU-GM increased (p < 0.05) whereas CFU-GEMM were maintained at week 4. In conclusion, this 3D culture system is capable of long-term, cytokine-free expansion of CBMNCs, enabling the study of hematopoiesis and providing a potential platform for drug discovery and therapeutic applications ex vivo.

Mapping Local Microstructure and Mechanical Performance Around Carbon Nanotube Grafted Silica Fibres: Methodologies for Hierarchical Composites

The introduction of carbon nanotubes (CNTs) modifies bulk polymer properties, depending on intrinsic quality, dispersion, alignment, interfacial chemistry and mechanical properties of the nanofiller. These effects can be exploited to enhance the matrices of conventional microscale fibre-reinforced polymer composites, by using primary reinforcing fibres grafted with CNTs. This paper presents a methodology that combines atomic force microscopy, polarised Raman spectroscopy, and nanoindentation techniques, to study the distribution, alignment and orientation of CNTs in the vicinity of epoxy-embedded micrometre-scale silica fibres, as well as, the resulting local mechanical properties of the matrix. Raman maps of key features in the CNT spectra clearly show the CNT distribution and orientation, including a 'parted' morphology associated with long grafted CNTs. The hardness and indentation modulus of the epoxy matrix were improved locally by 28% and 24%, respectively, due to the reinforcing effects of CNTs. Moreover, a slower stress relaxation was observed in the epoxy region containing CNTs, which may be due to restricted molecular mobility of the matrix. The proposed methodology is likely to be relevant to further studies of nanocomposites and hierarchical composites.

Ionic Liquids As Internal Phase for Non-aqueous PolyHIPEs

Stable high internal phase emulsions (HIPEs) with the ionic liquid 1-ethyl-3-methylimidazolium bis(trifluoromethyl-sulfonyl)imide as dispersed phase were prepared and polymerised thermally into polyHIPEs. All polyHIPEs exhibited pore morphologies similar to that of polyHIPEs obtained with an aqueous dispersed phase. PolyHIPEs containing the dispersed phase possess a low T(g) and are thermally stable in excess of 200 °C, offering the potential for new porous materials where water as dispersed phase is chemically or physically undesirable.

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