John S. Forsythe

Review Paper: A Review of the Cellular Response on Electrospun Nanofibers for Tissue Engineering

Electrospinning has been employed extensively in tissue engineering to generate nanofibrous scaffolds from either natural or synthetic biodegradable polymers to simulate the cellular microenvironment. Electrospinning rapidly produces fibers of the nanolength scale and the process offers many opportunities to tailor the physical, chemical, and biological properties of a material for specific applications and cellular environments. There is growing evidence that nanofibers amplify certain biological responses such as contact guidance and differentiation, however this has not been fully exploited in tissue engineering. This review addresses the cellular interactions with electrospun scaffolds, with particular focus on neural, bone, cartilage, and vascular tissue regeneration. Some aspects of scaffold design, including architectural properties, surface functionalization and materials selection are also addressed.

The radiation chemistry of fluoropolymers

This review covers some of the pioneering work of the radiolysis of fluoropolymers, which started in the mid 1950s and has continued up to the present day. In recent years, there has been renewed interest in the radiolysis of fluoropolymers-extending from the standpoint of radiation degradation in, for example, space environments and nuclear facilities, to the deliberate use of radiation for the modification of the fluoropolymers. In compiling this literature review, a fact that stood out was the dearth of information regarding the mechanisms for chain scission and especially crosslinking reactions. The lack of current, fundamental knowledge of the radiolysis of fluoropolymers, in the authors opinion, has limited the use of radiation in many applications, which are only now being realised, e.g. exciting new applications of crosslinked poly(tetrafluoroethylene)

Three-Dimensional Nanofibrous Scaffolds Incorporating Immobilized BDNF Promote Proliferation and Differentiation of Cortical Neural Stem Cells

Attempts to repair the central nervous system damaged as a result of trauma or disease will depend on the ability to restore the appropriate neuronal connectivity. This will rely on establishing appropriate chemical and physical environments for supporting neural cells and their processes and in this regard, engineering of biomaterials is of increasing interest. It will be important to understand how cells behave on these biomaterials in vitro, prior to future in vivo application. We reveal that modification of 3-dimensional (3D) electrospun poly-epsilon-caprolactone (PCL) nanofiber scaffolds by fiber alignment and aminolysation is superior to classical 2-dimensional (2D) culture-ware in promoting in vitro proliferation and differentiation of cortical cells. Many studies have examined the importance of exogenous soluble factors to promote cell fate specification. Here, we demonstrate that tethering the neurotrophin, brain-derived neurotrophic factor (BDNF), onto modified nanofibers is superior to culturing in the presence of soluble BDNF. Functional immobilization of BDNF to polymer nanofibers enhances neural stem cell (NSC) proliferation and directs cell fate toward neuronal and oligodendrocyte specification, essential for neural tissue repair. These findings indicate that modified PCL nanofibrous 3D scaffolds are capable of supporting NSCs and their derivatives and may present a new avenue for encouraging neural repair in the future.

Characterization of neural stem cells on electrospun poly(epsilon-caprolactone) submicron scaffolds: evaluating their potential in neural tissue engineering.

Development of biomaterials with specific characteristics to influence cell behaviour has played an important role in exploiting strategies to promote nerve regeneration. The effect of three-dimensional (3D) non-woven electrospun poly(ε-caprolactone) (PCL) scaffolds on the behaviour of rat brain-derived neural stem cells (NSCs) is reported. The interaction of NSCs on the randomly orientated submicron (PCL) fibrous scaffolds, with an average fibre diameter of 750 ± 100 nm, was investigated. The PCL scaffolds were modified with ethylenediamine (ED) to determine if amino functionalisation and changes in surface tension of the fibrous scaffolds affected the proliferation and differentiation characteristics of NSCs. Surface tension of the fibrous scaffold increased upon treatment with ED which was attributed to amine moieties present on the surface of the fibres. Although surface treatment did not change the differentiation of the NSCs, the modified scaffolds were more hydrophilic, resulting in a significant increase in the number of adhered cells, and increased spreading throughout the entirety of the scaffold. When the NSCs were seeded on the PCL scaffolds in the presence of 10% FBS, the stem cells differentiated primarily into oligodendrocytes, indicating that electrospun PCL has the capacity to direct the differentiation of NSCs towards a specific lineage. The data presented here is useful for the development of electrospun biomaterial scaffolds for neural tissue engineering, to regulate the proliferation and differentiation of NSCs.

Interaction of embryonic cortical neurons on nanofibrous scaffolds for neural tissue engineering

The interaction of murine embryonic cortical neurons on randomly orientated electrospun scaffolds of poly(L-lactide) (P(L)LA) and poly(lactide-co-glycolide) (PLGA) is investigated in this study. The scaffolds were surface treated with different concentrations of KOH to partially hydrolyze the surface and therefore change the surface tension. Hydrophilicity did not significantly influence the number of primary and secondary branches; however, it had a considerable effect on neurite extension. For scaffolds with surface tensions of 40-47 dyn cm(-1) there was a significantly greater overall neurite length for both the primary and secondary branches compared with more hydrophilic scaffolds. Another major finding of this work was that the interfibre distance influenced how the neurites extended. When the interfibre distance was greater than approximately 15 microm the neurites followed the fibres and avoided regions of very high fibre density. At interfibre distances less than approximately 15 microm, the neurites traversed between the fibres. Therefore, this study provided little evidence that contact guidance was the dominating cue in directing neurite extension, instead inferring that chemical cues, possibly from adjacent neurons had induced directional change.

Kinetics and network structure of thermally cured vinyl ester resins

Abstract Bisphenol-A diglycidyl ether dimethacrylate was blended with styrene at varying concentrations and this model vinyl ester resin (VER) was compared with two commercial VERs. The VERs were characterized using gravimetry, FTIR spectroscopy, NMR spectroscopy, differential scanning calorimetry (DSC) and DMTA. NMR spectroscopy differentiated between a novolac epoxy-based multimethacrylate oligomer and the two bisphenol-A epoxy-based dimethacrylate oligomers. Reaction kinetics were studied using scanning and isothermal DSC and isothermal FTIR spectroscopy using benzoyl peroxide as the thermal initiator. The presence of oxygen was found to inhibit significantly the polymerization. Increased initiator concentration raised the rate of isothermal polymerization, but did not affect the final conversion while increased styrene concentration reduced the polymerization rate constant and increased the total conversion. This was interpreted in terms of the variations in the termination rate and the stability of the styryl radical on the cure rate and the effect of vitrification on the extent of cure. From measurements of the dynamic mechanical properties as a function of temperature, the breadth of the glass transition tan δ curve and the magnitude of the rubbery modulus was found to increase while the tan δ maximum decreased with increased crosslink density. The T g , as measured by DSC, and the temperature of the tan δ maximum, as measured by DMTA, were not significantly affected by the styrene content in the resin per se , but were dependent on the combined effects of composition and crosslink density of the network.

Biosurface engineering through ink jet printing

The feasibility of thermal ink jet printing as a robust process for biosurface engineering was demonstrated. The strategy investigated was to reconstruct a commercial printer and take advantage of its colour management interface. High printing resolution was achieved by formulating bio-inks of viscosity and surface tension similar to those of commercial inks. Protein and enzyme denaturation during thermal ink jet printing was shown to be insignificant. This is because the time spent by the biomolecules in the heating zone of the printer is negligible; in addition, the air and substrate of high heat capacity absorb any residual heat from the droplet. Gradients of trophic/tropic factors can serve as driving force for cell growth or migration for tissue regeneration. Concentration gradients of proteins were printed on scaffolds to show the capability of ink jet printing. The printed proteins did not desorb upon prolonged immersion in aqueous solutions, thus allowing printed scaffold to be used under in vitro and in vivo conditions. Our group portrait was ink jet printed with a protein on paper, illustrating that complex biopatterns can be printed on large area. Finally, patterns of enzymes were ink jet printed within the detection and reaction zones of a paper diagnostic.

Biomaterials for Brain Tissue Engineering

Neurological disorders such as traumatic brain injuries or stroke result in neuronal loss and disruption of the brain parenchyma. Current treatment strategies are limited in that they can only mitigate the degeneration process or alleviate the symptoms but do not reverse the condition. In contrast, regenerative cell-based therapies offer long-term hope for many patients. Bioactive scaffolds are likely to reinforce the success of cell replacement therapies by providing a microenvironment that facilitates the survival, proliferation, differentiation, and connectivity of transplanted and/or endogenous cells. This Review outlines various biomaterials (including hydrogels, self-assembling peptides, and electrospun nanofibres) that have been investigated for the repair of brain tissue, and discusses strategies for the immobilization of biomolecules. An overview of the potential clinical applications of such scaffolds in neurodegenerative diseases is also provided.

Self-assembly of ciprofloxacin and a tripeptide into an antimicrobial nanostructured hydrogel

This work reports the self-assembly of a sparingly soluble antibiotic (ciprofloxacin) and a hydrophobic tripeptide ((D)Leu-Phe-Phe) into supramolecular nanostructures that yield a macroscopic hydrogel at physiological pH. Drug incorporation results in modified morphology and rheological properties of the self-assembled hydrogel. These changes can be correlated with intermolecular interactions between the drug and the peptide, as confirmed by spectroscopic analysis (fluorescence, circular dichroism, IR). The drug appears bound within the hydrogel by non-covalent interactions, and retains its activity over a prolonged release timescale. Antimicrobial activity of the ciprofloxacin-peptide self-assembled hydrogel was evaluated against Staphylococcus aureus, Escherichia coli, and a clinical strain of Klebsiella pneumoniae. Interestingly, the peptide hydrogel alone exhibited a mild anti-bacterial activity against Gram-negative bacteria. While toxic to bacteria, no major cytotoxicity was seen in haemolysis assays of human red blood cells or in mouse fibroblast cell cultures. This new approach of drug incorporation into the nanostructure of a simple tripeptide hydrogel by self-assembly may have important applications for cost-effective wound dressings and novel antimicrobial formulations.

Unzipping the role of chirality in nanoscale self-assembly of tripeptide hydrogels

Change of chirality is a useful tool to manipulate the aqueous self-assembly behaviour of uncapped, hydrophobic tripeptides. In contrast with other short peptides, these tripeptides form hydrogels at a physiological pH without the aid of organic solvents or end-capping groups (e.g. Fmoc). The novel hydrogel forming peptide (D)Leu-Phe-Phe ((D)LFF) and its epimer Leu-Phe-Phe (LFF) exemplify dramatic supramolecular effects induced by subtle changes to stereochemistry. Only the d-amino acid-containing peptide instantly forms a hydrogel in aqueous solution following a pH switch, generating long fibres (>100 μm) that entangle into a 3D network. However, unexpected nanostructures are observed for both peptides and they are particularly heterogeneous for LFF. Structural analyses using CD, FT-IR and fluorescent amyloid staining reveal anti-parallel beta-sheets for both peptides. XRD analysis also identifies key distances consistent with beta-sheet formation in both peptides, but suggests additional high molecular order and extended molecular length for (D)LFF only. Molecular modelling of the two peptides highlights the key interactions responsible for self-assembly; in particular, rapid self-assembly of (D)LFF is promoted by a phenylalanine zipper, which is not possible because of steric factors for LFF. In conclusion, this study elucidates for the first time the molecular basis for how chirality can dramatically influence supramolecular organisation in very short peptide sequences.