Enhanced efficacy in drug-resistant cancer cells through synergistic nanoparticle mediated delivery of cisplatin and decitabine.

There are several limitations with monodrug cancer therapy, including poor bioavailability, rapid clearance and drug resistance. Combination therapy addresses these by exploiting synergism between different drugs against cancer cells. In particular, the combination of epigenetic therapies with conventional chemotherapeutic agents can improve the initial tumour response and overcome acquired drug resistance. Co-encapsulation of multiple therapeutic agents into a single polymeric nanoparticle is one of the many approaches taken to enhance therapeutic effect and improve the pharmacokinetic profile. In this study, different types of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs), matrix and core-shell (CS), were investigated for simultaneous encapsulation of a demethylating drug, decitabine, and a potent anticancer agent, cisplatin. It was shown that by altering the configuration of the CS structure, the release profile could be tuned. In order to investigate whether this could enhance the anticancer effect compared to cisplatin, human ovarian carcinoma cell line (A2780) and its cisplatin resistant variant (A2780cis) were exposed to free cisplatin and the CS-NPs. A better response was obtained in both cell lines (11% and 51% viability of A2780 and A2780cis, respectively) using CS-NPs than cisplatin alone (27%, 82% viability of A2780 and A2780cis, respectively) or in combination with decitabine (22%, 96% viability of A2780 and A2780cis, respectively) at equivalent doses (10 µM).

The influence of drug solubility and sampling frequency on metformin and glibenclamide release from double-layered particles: experimental analysis and mathematical modelling.

Co-axial electrohydrodynamic atomization was used to prepare core/shell polymethylsilsesquioxane particles for co-delivery of metformin and glibenclamide in a sustained release manner. The drug-loaded microparticles were mostly spherical and uniformly distributed in size, with average diameters between 3 and 5 µm across various batches. FTIR was used to confirm the presence of drugs within the particles while X-ray diffraction studies revealed drugs encapsulated existed predominantly in the amorphous state. Intended as systems that potentially can act as depot formulations for long-term release of antidiabetics, a detailed analysis of drug release from these particles was necessary. Drugs of different solubilities were selected in order to study the effects of drug solubility from a core/shell particle system. Further analyses to determine how conditions such as release into a limited volume of media, sampling rate and partitioning of drug between the core and shell layers influenced drug release were conducted by comparing experimental and mathematically modelled outcomes. It was found that while the solubility of drug may affect release from such systems, rate of removal of drug (sampling frequency) which upsets local equilibrium at the particle/solution interface prompting a rapid release to redress the equilibrium influenced release more.

Electrosprayed microparticles for intestinal delivery of prednisolone.

Single and coaxial electrospraying was used to prepare Eudragit L100-55 polymer microparticles containing prednisolone as the active pharmaceutical ingredient. Different compositions of prednisolone and Eudragit L100-55 were used to develop five different formulations with different polymer : drug ratios. The resultant microparticles had a toroidal shape with a narrow size distribution. Prednisolone was present in an amorphous physical state, as confirmed by X-ray diffraction analysis. Dissolution studies were carried out in order to investigate the feasibility of the proposed system for site-specific release of prednisolone. The release rates were interpreted in terms of diffusion-controlled release. It was shown that utilization of pH-responsive Eudragit L100-55 could minimize the release of prednisolone in the acidic conditions of the stomach, which was followed by rapid release as the pH of the release medium was adjusted to 6.8 after the first 2 h. This is especially desirable for the treatment of conditions including inflammatory bowel disease and colon cancer.

Gyrospun antimicrobial nanoparticle loaded fibrous polymeric filters.

A one step approach to prepare hybrid nanoparticle embedded polymer fibres using pressurised gyration is presented. Two types of novel antimicrobial nanoparticles and poly(methylmethacrylate) polymer were used in this work. X-ray diffraction analysis of the nanoparticles revealed Ag, Cu and W are the main elements present in them. The concentration of the polymer solution and the nanoparticle concentration had a significant influence on the fibre diameter, pore size and morphology. Fibres with a diameter in the range of 6-20µm were spun using 20wt% polymer solutions containing 0.1, 0.25 and 0.5 wt% nanoparticles under 0.3MPa working pressure and a rotational speed of 36,000rpm. Continuous, bead-free fibre morphologies were obtained for each case. The pore size in the fibres varied between 36 and 300nm. Successful incorporation of the nanoparticles in polymer fibres was confirmed by energy dispersive x-ray analysis. The fibres were also gyrospun on to metallic discs to prepare filters which were tested for their antibacterial activity on a suspension of Pseudomonas aeruginosa. Nanoparticle loaded fibres showed higher antibacterial efficacy than pure poly(methylmethacrylate) fibres.

Porous Polymeric Films from Microbubbles Generated Using a T-Junction Microfluidic Device.

In this work, a simple microfluidic junction with a T geometry and coarse (200 µm diameter) capillaries was used to generate monodisperse microbubbles with an alginate polymer shell. Subsequently, these bubbles were used to prepare porous alginate films with good control over the pore structure. The lack of pore size, shape, and surface control in scalable forming of polymeric films is a major application-limiting drawback at present. Controlling the thinning process of the shell of the bubbles to tune the surface of the resulting structures was also explored. Films were prepared with nanopatterned surfaces by controlling the thinning of the bubble shell, with the aid of surfactants, to induce efficient bursting (fragmentation) of bubbles to generate nanodroplets, which become embedded within the film surface. This novel feature greatly expands and enhances the use of hydrophilic polymers in a wide range of biomedical applications, particularly in drug delivery and tissue engineering, such as studying cellular responses to different morphological surfaces.

Investigating the particle to fibre transition threshold during electrohydrodynamic atomization of a polymer solution.

Electrohydrodynamic atomization (EHDA) is a key research area for producing micro and nano-sized structures. This process can be categorized into two main operating regimes: electrospraying for particle generation and electrospinning for fibre production. Producing particles/fibres of the desired size or morphology depends on two main factors; properties of the polymeric solution used and the processing conditions including flow rate, applied voltage and collection distance. In this work the particle-fibre transition region was analyzed by changing the polymer concentration of PLGA poly (lactic-co-glycolic acid) in acetone between 2 and 25wt%. Subsequently the processing conditions were adjusted to study the optimum transition parameters. Additionally the EHDA configuration was also modified by adding a metallic plate to observe the deposition area. The diameter and the distance of the plate from the capillary tip were adjusted to investigate variations in particle and fibre morphologies as well. It was found that complete transition from particles to fibres occurs at 20wt% indicating concentration to be the dominant criterion. Low flow rates yielded fibres without beads. However the applied voltage and distance between the tip of the nozzle jetting the polymer solution and collector (working distance) did not yield definitive results. Reducing the collector distance and increasing applied voltages produces smooth as well as beaded fibres. Addition of a metal plate reduces particle size by ~1µm; the fibre size increases especially with increasing plate diameter while bead density and size reduces when the disc is fixed closer to the capillary tip. Additionally, the deposition area is reduced by 70% and 57% with the addition of metal plates of 30mm and 60mm, respectively. The results indicate that a metal plate can be utilized further to tune the particle/fibre size and morphology and this also significantly increases the yield of EHDA process which is currently a limitation in adopting it as a mass production technique.

Physio-chemical and antibacterial characteristics of pressure spun nylon nanofibres embedded with functional silver nanoparticles.

A novel and facile approach to prepare hybrid nanoparticle embedded polymer nanofibers using pressurised gyration is presented. Silver nanoparticles and nylon polymer were used in this work. The polymer solution's physical properties, rotating speed and the working pressure had a significant influence on the fibre diameter and the morphology. Fibres in the range of 60-500nm were spun using 10wt.%, 15wt.% and 20wt.% nylon solutions and these bead-free fibres were processed under 0.2MPa and 0.3MPa working pressure and a rotational speed of 36,000rpm. 1-4wt.% of Ag was added to these nylon solutions and in the case of wt.% fibres in the range 50-150nm were prepared using the same conditions of pressurised gyration. Successful incorporation of the Ag nanoparticles in nylon nanofibres was confirmed by using a combination of advanced microscopical techniques and Raman spectrometry was used to study the bonding characteristics of nylon and the Ag nanoparticles. Inductively coupled plasma mass spectroscopy showed a substantial concentration of Ag ions in the nylon fibre matrix which is essential for producing effective antibacterial properties. Antibacterial activity of the Ag-loaded nanofibres shows higher efficacy than nylon nanofibres for Gram-negative Escherichia coli and Pseudomonas aeruginosa microorganisms, and both Ag nanoparticles and the Ag ions were found to be the reason for enhanced cell death in the bacterial solutions.

Rheology and pressurised gyration of starch and starch-loaded poly(ethylene oxide).

This work investigates the rheology and spinning of starch and starch-loaded poly(ethylene oxide) (PEO) by pressurised gyration in order to prepare nanofibres. The spinning dope's rheological properties played a crucial role in fibre formation. Newtonian behaviour is observed in 1-20 wt% starch suspensions and non-Newtonian behaviour is found in all the PEO-starch mixtures. Pressurised gyration of the starch suspensions produced beads only but PEO-starch mixtures generated fibres. The fibre diameter of the PEO-starch samples is shown to be a function of polymer concentration and rotating speed of the gyration system. Fibre formation can only be facilitated below a certain working pressure. The concentration of starch in the PEO-starch mixtures is crucial in defining whether beaded or continuous fibres were generated and this is related to the composition of the spinning dope. FT-IR, XRD and microscopy studies indicated very good miscibility of starch and PEO in the nanofibres. The storage modulus of the PEO-starch were also studied as a function of temperature (30-150°C) and showed interesting results but it was not possible to deduce general trends valid for the entire temperature range.

Facile synthesis of both needle-like and spherical hydroxyapatite nanoparticles: effect of synthetic temperature and calcination on morphology, crystallite size and crystallinity.

Synthetic hydroxyapatite (HA) nanoparticles, that mimic natural HA, are widely used as biocompatible coatings on prostheses to repair and substitute human bones. In this study, HA nanoparticles are prepared by precipitating them from a precursor solution containing calcium sucrate and ammonium dihydrogen orthophosphate, at a Ca/P mole ratio of 1.67:1, at temperatures, ranging from 10°C to 95°C. A set of products, prepared at different temperatures, is analyzed for their crystallinity, crystallite size, morphology, thermal stability and composition, by X-ray diffraction (XRD), scanning electron microscopy (SEM), thermogravimetric analysis (TGA) and Fourier transform infrared (FT-IR) spectroscopic techniques, while the other set is analyzed after calcining the respective products, soon after their synthesis, for 3h, at 700°C. The as-prepared products, after 2h of drying, without any calcination, are not crystalline, but they grow very slowly into needle-like morphologies, as they are ripened with time. The percentage crystallinity of the final products increases from 15% to 52%, with increasing the preparative temperature. The calcined samples always produce spherical nanoparticles of essentially the same diameter, between 90 nm and 100 nm, which does not change due to aging and preparative temperatures. Therefore, the same method can be utilized to synthesize both spherical and needle-like nanoparticles of hydroxyapatite, with well-defined sizes and shapes. The ability to use readily available cheap raw materials, for the synthesis of such well-defined crystallites of hydroxyapatite, is an added advantage of this method, which may be explored further for the scaling up of the procedures to suit to industrial scale synthesis of such hydroxyapatite nanoparticles.

Formation, stability, and mechanical properties of bovine serum albumin stabilized air bubbles produced using coaxial electrohydrodynamic atomization.

Bovine serum albumin (BSA) microbubbles were generated using coaxial electrohydrodynamic atomization (CEDHA) using various concentrations of BSA solutions. The bubble characteristics and the long-term stability of the microbubbles were studied through adjustment of processing parameters and the collection media. Bubbles in the range of 40-800 µm were obtained in a controlled fashion, and increasing the flow rate of the BSA solution reduced the polydispersity of the microbubbles. Use of distilled water-glutaraldehyde, glycerol, and glycerol-Tween 80 collection media allowed a remarkable improvement in bubble stability compared to BSA solution collection medium. Possible physical mechanisms were developed to explain the stability of the microbubbles. The collection distance showed a marked influence on stability of the microbubbles. Near-monodisperse particle-reinforced microbubbles were formed with various concentrations of 2,2'-azobis(isobutyramidine) dihydrochloride (AIBA)-polystyrene particle in BSA solution. The bubble size and the size distribution showed negligible change over a period of time irrespective of the concentration of particles at the bubble surface. The compression stiffness of the microbubbles was determined using nanoindentation at ambient temperature and showed that the stiffness of the microbubbles increased from 8 N/m to 20 N/m upon changing the concentration of BSA solution from 5 wt % to 15 wt %.

Creating "hotels" for cells by electrospinning honeycomb-like polymeric structures.

It is well established that three-dimensional honeycomb-like nanofibrous structures enhance cell activity. In this work, we report that electrospun polymer nanofibres self-assemble into three-dimensional honeycomb-like structures. The underlying mechanism is studied by varying the polymer solution concentration, collecting substrates and working distance. The polymer solution concentration has a significant effect on the size of the electrospun nanofibres. The collection substrate and working distance affect the electric field strength, the evaporation of solvent and the discharging of nanofibres and consequently these two had a significant influence on the self-assembly of nanofibres.

Print head design and control for electrohydrodynamic printing of silk fibroin.

This study investigates the effect of print head design on the electrohydrodynamic printed resolution of silk fibroin. Needles with large orifices measuring at 800 µm were used to build five different print heads. The print heads were manufactured, tested, and optimized using four different silk fibroin solution concentrations of 10 wt.%, 15 wt.%, 20 wt.%, and 22 wt.% at applied voltages that ranged from 10 to 20 kV with two different flow rates of 1.5 µl/min and 2.0 µl/min. Each print head design behaved in a unique manner in terms of printed line characteristics as the flow rate, voltage and concentration were varied. The highest printed resolution of the order of 1 µm was achieved using the pinhole reservoir print head. Possible explanations for each of the observed behaviors and design criteria for future print heads are discussed.

Electrospinning versus fibre production methods: from specifics to technological convergence.

Academic and industrial research on nanofibres is an area of increasing global interest, as seen in the continuously multiplying number of research papers and patents and the broadening range of chemical, medical, electrical and environmental applications. This in turn expands the size of the market opportunity and is reflected in the significant rise of entrepreneurial activities and investments in the field. Electrospinning is probably the most researched top-down method to form nanofibres from a remarkable range of organic and inorganic materials. It is well known and discussed in many comprehensive studies, so why this review? As we read about yet another "novel" method producing multifunctional nanomaterials in grams or milligrams in the laboratory, there is hardly any research addressing how these methods can be safely, consistently and cost-effectively up-scaled. Despite two decades of governmental and private investment, the productivity of nanofibre forming methods is still struggling to meet the increasing demand. This hinders the further integration of nanofibres into practical large-scale applications and limits current uses to niche-markets. Looking into history, this large gap between supply and demand of synthetic fibres was seen and addressed in conventional textile production a century ago. The remarkable achievement was accomplished via extensive collaborative research between academia and industry, applying ingenious solutions and technological convergence from polymer chemistry, physical chemistry, materials science and engineering disciplines. Looking into the present, current advances in electrospinning and nanofibre production are showing similar interdisciplinary technological convergence, and knowledge of industrial textile processing is being combined with new developments in nanofibre forming methods. Moreover, many important parameters in electrospinning and nanofibre spinning methods overlap parameters extensively studied in industrial fibre processing. Thus, this review combines interdisciplinary knowledge from the academia and industry to facilitate technological convergence and offers insight for upscaling electrospinning and nanofibre production. It will examine advances in electrospinning within a framework of large-scale fibre production as well as alternative nanofibre forming methods, providing a comprehensive comparison of conventional and contemporary fibre forming technologies. This study intends to stimulate interest in addressing the issue of scale-up alongside novel developments and applications in nanofibre research.

Effect of deposition parameters and post-deposition annealing on the morphology and cellular response of electrosprayed TiO2 films.

Titania films (TiO(2)) were deposited on Ti alloy substrates by electrohydrodynamic atomization (EHDA), also called electrospraying, and the morphology and phase composition of the coatings were evaluated. A range of TiO(2) sols (2-8 wt%) were prepared via the hydrolysis of polymeric precursors using isopropanol (prOH) as a solvent carrier. Stable cone-jet formation during the EHDA process was greatly influenced by varying the liquid physical properties. Deposition parameters such as sol concentration, needle-to-substrate distance and spray time were found to affect film morphology. Dense and continuous films were obtained under optimized conditions whereby a 2 wt% TiO(2) sol was atomized at a flow rate of 5 µl min(-1) by a needle of 300 µm inner diameter, kept at a distance of 20 mm from the grounded substrate and operating at an applied voltage of 3.5-4.2 kV. The films were then crystallized by heating to 300-600 °C. Annealing increased the hydrophilicity of the films but did not significantly affect the surface roughness of the films. In vitro cellular response was determined by studying the interaction between MG63 cells and the films annealed at the various temperatures. MG63 cells were able to grow and proliferate on all the TiO(2) films, while the highest proliferation rate was found on the TiO(2) films annealed at 600 °C. Our results indicate that electrosprayed TiO(2) films possess great potential for biomedical applications.

The pathway to intelligent implants: osteoblast response to nano silicon-doped hydroxyapatite patterning.

Bioactive hydroxyapatite (HA) with addition of silicon (Si) in the crystal structure (silicon-doped hydroxyapatite (SiHA)) has become a highly attractive alternative to conventional HA in bone replacement owing to the significant improvement in the in vivo bioactivity and osteoconductivity. Nanometre-scaled SiHA (nanoSiHA), which closely resembles the size of bone mineral, has been synthesized in this study. Thus, the silicon addition provides an extra chemical cue to stimulate and enhance bone formation for new generation coatings, and the next stage in metallic implantation design is to further improve cellular adhesion and proliferation by control of cell alignment. Topography has been found to provide a powerful set of signals for cells and form contact guidance. Using the recently developed novel technique of template-assisted electrohydrodynamic atomization (TAEA), patterns of pillars and tracks of various dimensions of nanoSiHA were achieved. Modifying the parameters of TAEA, the resolution of pattern structures was controlled, enabling the topography of a substrate to be modified accordingly. Spray time, flow rate and distance between the needle and substrate were varied to improve the pattern formation of pillars and tracks. The 15 min deposition time provided the most consistent patterned topography with a distance of 50 mm and flow rate of 4 µl min(-1). A titanium substrate was patterned with pillars and tracks of varying widths, line lengths and distances under the optimized TAEA processing condition. A fast bone-like apatite formation rate was found on nanoSiHA after immersion in simulated body fluid, thus demonstrating its high in vitro bioactivity. Primary human osteoblast (HOB) cells responded to SiHA patterns by stretching of the filopodia between track and pillar, attaching to the apex of the pillar pattern and stretching between two. HOB cells responded to the track pattern by elongating along and between the track, and the length of HOB cells was proportional to the gaps between track patterns, but this relationship was not observed on the pillar patterns. The study has therefore provided an insight for future design of next generation implant surfaces to control and guide cellular responses, while TAEA patterning provides a controllable technique to provide topography to medical implants.

Ceramic encapsulation with polymer via co-axial electrohydrodynamic jetting.

Co-flowing media of a polymeric solution (30 wt% polymethylsilsesquioxane in ethanol) and a ceramic suspension (10 wt% alumina in glycerol) were subjected to an electric field. The flow rates of the media (10-30 microL min(-1)) and the applied voltage (0-11 kV) were varied systematically during the experimentation by making gradual increments to each variable, which enabled the construction of a mode selection map. Under co-flowing conditions, with the flow rate of polymer solution (outer needle) twice that of the ceramic suspension (inner needle), encapsulated droplets of polymer-coated alumina were produced within stable cone-jet mode. These were collected in a thin film of water and the resultant particle size varied between 1 and 38 microm. Encapsulation was confirmed with scanning electron microscopy and element analysis.

Size mapping of electric field-assisted production of polycaprolactone particles.

In this investigation, biodegradable polycaprolactone polymeric particles (300-4500 nm in diameter) were prepared by jetting a solution in an electric field. An extensive study has been carried out to determine how the size and size distribution of the particles generated can be controlled by systematically varying the polymer concentration in solution (and thereby its viscosity and electrical conductivity), and also the selected flow rate (2-50 microl min(-1)) and applied voltage (0-15 kV) during particle generation. Change in these parameters affects the mode of jetting, and within the stable cone-jet mode window, an increase in the applied voltage (approx. 15 kV) resulted in a reduction in particle size and this was more pronounced at high flow rates (such as; 30, 40 and 50 microl min(-1)) in the same region. The carrier particles were more polydisperse at the peripheral regions of the stable cone-jet mode, as defined in the applied voltage-flow rate parametric map. The effect of loading a drug on the particle size, size distribution and encapsulation efficiency was also studied. Release from drug-loaded particles was investigated using UV spectrophotometry over 45 days. This work demonstrates a powerful method of generating drug-loaded polymeric particles, with the ability to control size and polydispersivity, which has great potential in several categories of biotechnology requiring carrier particles, such as drug delivery and gene therapy.

The role of surface wettability and surface charge of electrosprayed nanoapatites on the behaviour of osteoblasts.

A new deposition method is presented, based on electrospraying, that can build bioceramic structures with desirable surface properties. This technology allows nanoapatite crystals, including hydroxyapatite (nHA), carbonate-substituted HA (nCHA) and silicon-substituted HA (nSiHA), to be electrosprayed on glass substrates. Human osteoblast cells cultured on nSiHA showed enhanced cell attachment, proliferation and protein expression, namely alkaline phosphatase, type 1 collagen and osteocalcin, as compared to nHA and nCHA. The modification of nanoapatite by the addition of silicon into the HA lattice structure renders the electrosprayed surface more hydrophilic and electronegatively charged.

Novel preparation of transdermal drug-delivery patches and functional wound healing materials.

The application of an electric field to a flowing medium can result in the formation of microscale and nanoscale structures suitable for drug delivery applications. We show that the design of the drug carrier can be varied and the release mechanism can be controlled by changing the physical state of the component containing the active agent. The structures formed include loaded micrometer-scale tubes and microcapsules and nanocapsules, which can also be utilized together to fabricate patches and wound healing materials. The aim of this study was to demonstrate novel processing of such patches and wound dressings. The processing used to generate these structures is carried out at the ambient temperature and is a versatile one-step operation suitable for a range of materials with low running costs and set-up costs without the degradation of the active drug component. The process can be multiplexed and requires no solvent extraction. It also offers pharmaceutical applications outside the remit of the potential uses presented.