David A. Steele

The geometric control of E14 and R1 mouse embryonic stem cell pluripotency by plasma polymer surface chemical gradients

Plasma polymer surfaces were fabricated such that the cell response to a range of carboxylic acid concentrations on a single sample could be investigated. Surface chemical gradients from hydrophobic plasma polymerised octadiene (OD) to a more hydrophilic plasma polymerised acrylic acid (AA) were formed on glass coverslips. Surface characterisation of the chemical gradients was performed using X-ray photoelectron spectroscopy to determine elemental composition. Following culture of E14 and R1 mouse embryonic stem cells (mES) in differing culture media, cell pluripotency was determined by alkaline phosphatase staining. The results demonstrate that for these cell lines the capacity for self-renewal is maintained if the cells are restricted in their spreading to <120 microm2.

Nanoscale deposition of chemically functionalised films via plasma polymerisation

Plasma polymerisation is a technologically important surface engineering process capable of depositing ultra-thin functionalised films for a variety of purposes. It has many advantages over other surface engineering processes, including that it is completely dry, can be used for complex geometries, and the physico-chemical properties of the film can be tailored through judicious choice of processing conditions. Despite this, the mechanisms of film growth are largely unknown, and current models are based on purely chemical arguments. Consideration of some basic plasma physics shows that some species can arrive at surfaces with energies greater than 1000 kJ mol−1 (>10 eV), and thus open a range of surface reactions that have not been considered previously. This review aims to close the gap between the physics and chemistry of reactive plasma systems.

Role of positive ions in determining the deposition rate and film chemistry of continuous wave hexamethyl disiloxane plasmas.

New data shed light on the mechanisms of film growth from low power, low pressure plasmas of organic compounds. These data rebalance the widely held view that plasma polymer formation is due to radical/neutral reactions only and that ions play no direct role in contributing mass at the surface. Ion reactions are shown to play an important role in both the plasma phase and at the surface. The mass deposition rate and ion flux in continuous wave hexamethyl disiloxane (HMDSO) plasmas have been studied as a function of pressure and applied RF power. Both the deposition rate and ion flux were shown to increase with applied power; however, the deposition rate increased with pressure while the ion flux decreased. Positive ion mass spectrometry of the plasma phase demonstrates that the dominant ionic species is the (HMDSO-CH(3))(+) ion at m/z 147, but significant fragmentation and subsequent oligomerization was also observed. Chemical analysis of the deposits by X-ray photoelectron spectroscopy and secondary ion mass spectrometry show that the deposits were consistent with deposits reported by previous workers grown from plasma and hyperthermal (HMDSO-CH(3))(+) ions. Increasing coordination of silicon with oxygen in the plasma deposits reveals the role of ions in the growth of plasma polymers. Comparing the calculated film thicknesses after a fixed total fluence of 1.5 × 10(19) ions/m(2) to results for hyperthermal ions shows that ions can contribute significantly to the total absorbed mass in the deposits.

The link between mechanisms of deposition and the physico-chemical properties of plasma polymer films

Film thickness and functional group retention are routinely measured parameters for plasma polymers. It is known that other parameters such as density, solubility and mechanical properties can affect the performance of the plasma polymer film, however such parameters are not often measured; nor is there any understanding of the link between the mechanisms of film growth and these properties. In this investigation we produced thin films from three classes of commonly used plasma polymers (hydrocarbons, glymes and carboxylic acids). By choosing the monomer structure and applied RF power, the dominant mechanism of film growth was varied between ionic deposition and neutral grafting. The density, solubility and modulus of the resulting films were then measured by atomic force microscopy. Films grown from saturated monomers had higher moduli, were less soluble, and surprisingly had lower density compared to their unsaturated analogues. The results demonstrate that cognizance of the mechanism of film growth allows the physical properties of the film to be tailored for specific applications.

Polyoctanediol Citrate/Sebacate Bioelastomer Films: Surface Morphology, Chemistry and Functionality

Elastomeric polyesters synthesized from non-toxic and biocompatible reactants are topical research materials for tissue-engineering applications. In such applications, the morphology, chemistry and functionality of the materials surfaces play a key role. While a number of papers have focused and reported on the fabrication and biological evaluation of elastic polyesters, only a few have attempted to characterise the surfaces of such materials. In this paper, we report on the preparation and surface characterization of films of a co-polyester bioelastomer, polyoctanediol citrate/sebacate (p(OCS)). The co-polymer was synthesized following the standard procedure of polyesterification using three non-toxic monomers (1,8-octanediol, citric acid and sebacic acid) in a catalyst-free environment. Nuclear magnetic resonance spectroscopy was used to monitor the chemical composition of the various p(OCS) elastomers. The p(OCS) films, prepared by both spin-coating and solvent casting of the p(OCS) pre-polymer solutions, were characterized by scanning electron microscopy, UV-Vis titration, photo-acoustic Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy, and tested for their cytocompatibility. The results obtained suggest that the surface morphology, chemistry and the concentration of the surface functional groups can be controlled by simply varying the initial acid concentration (citric/sebacic acids) in the pre-polymer. The films supported the attachment and proliferation of osteoblast-like cells (MG63). This unique approach provides an effective method of controlling and monitoring the fundamental p(OCS) surface properties important for their potential utilisation as a tissue-engineering material.

Gradient technology for high-throughput screening of interactions between cells and nanostructured materials

We present a novel substrate suitable for the high-throughput analysis of cell response to variations in surface chemistry and nanotopography. Electrochemical etching was used to produce silicon wafers with nanopores between 10 and 100nm in diameter. Over this substrate and flat silicon wafers, a gradient film ranging from hydrocarbon to carboxylic acid plasma polymer was deposited, with the concentration of surface carboxylic acid groups varying between 0.7 and 3% as measured by XPS. MG63 osteoblast-like cells were then cultured on these substrates and showed greatest cell spreading and adhesion onto porous silicon with a carboxylic acid group concentration between 2-3%. This method has great potential for high-throughput screening of cell-material interaction with particular relevance to tissue engineering.

Microplasma arrays: a new approach for maskless and localized patterning of materials surfaces

“Maskless” microplasma treatment of passivated surfaces has been developed for the micropatterning of materials surfaces. The micropatterned surfaces are used in the fabrication of arrays for protein and cell-based assays. The advantage of this micropatterning approach is that it can be easily integrated into current manufacturing practices and the resultant micropatterned surfaces used with existing life sciences techniques and instrumentation.

Submillimeter-Scale Surface Gradients of Immobilized Protein Ligands

We describe a method to produce antibody-captured ligand gradients over biologically relevant distances (hundreds of micrometers) whereby the ligand density and gradient shape may be tailored. Separation of the ligand from the solid-phase surface ensures that the biological activity of the ligand remains unaffected by immobilization. Our method involves the use of a plasma-masking method to generate a surface chemical gradient on a glass substrate to which the 9E10 antibody is covalently coupled. This antibody captures myc-tagged biomolecules. In our example, the antibody is then used to immobilize a gradient of the intercellular signaling molecule delta-like-1 (Dll1). To visualize the gradient of Dll1, we have used the multistep approach of binding with rabbit anti-Dll1 primary antibody and then adding colloidal-gold-conjugated secondary antibody.

Plasma Polymer Surfaces for Cell Expansion and Delivery

The purpose of this review is to explore the development, applications and outcomes of using plasmadeposited (polymeric) surfaces as culture and delivery vehicles for cells in the replacement, repair and regeneration of damaged tissues. After a brief introduction to biomaterials and cell therapy, examples of the use of plasma polymerization in surface engineering are presented. The use of these surfaces for both in vitro cell culture and ex vivo delivery is reviewed with a particular emphasis on the epithelium. The review concludes with a look at some of the emerging and potential future directions for this technology in mediating cell–surface interactions.

Osteoblast Biocompatibility on Poly(octanediol citrate)/Sebacate Elastomers with Controlled Wettability

This work examines the biocompatibility of poly(octanediol citrate)/sebacate (p(OCS)) biodegradable polyester elastomers. The growth of human MG63 osteoblast-like cells was studied on p(OCS) films. Three types of p(OCS) films were synthesised simply by varying the concentrations of 1,8-octanediol (OD), citric acid (CA), and sebacic acid (SA) monomers at initial molar ratios of 1:1:0, 1:0.75:0.25 and 1:0.5:0.5. At these ratios, the p(OCS) films exhibit decreasing hydrophilicity as shown by the measured water contact angle values of 31, 41 and 64°, respectively. For all the samples, no difference in cell growth was detected after 1 day of cell culture. However, after 4 days, the highest number of viable cells was detected on the p(OCS) film synthesised with the intermediate CA molar ratio of 0.75. This sample also contains the median concentration of surface carboxylic acid groups and hydrophilicity. Following long-term cell culture (18 days), a statistically significant higher density of viable cells had grown on the p(OCS) films with SA molar ratios of 0.25 (P < 0.0001) and 0.5 (P = 0.002) in comparison to the material containing 100% CA and no SA. The work demonstrated that the performance of possible p(OCS) bone tissue engineering scaffolds could be improved by simply adjusting the molar ratios of CA and SA in the pre-polymer without any requirements for post-synthesis modification.