L’ingénierie tissulaire est un domaine pluridisciplinaire visant l’élaboration de prothèses biologiques autologues. Dans cette stratégie, la fabrication de structures, appelées scaffolds, utilisées pour la culture cellulaire est nécessaire. Nous avons développé plusieurs méthodes de fabrication de ces structures tridimensionnelles. La première méthode (solvant casting/particulate leaching out) utilisant une solution de polymère et des particules sphérolisées de glucose comme porogène, permet l’obtention de structures possédant des pores sphériques et bien interconnectés. Nous avons montré que ces scaffolds sont biocompatibles et que leurs propriétés mécaniques en compression peuvent être ajustées. La seconde méthode, l’electrospinning, permet la fabrication de membranes fibreuses biocompatibles, dont le diamètre des fibres peut être contrôlé (de 800 nm à plusieurs micromètres). La troisième méthode de fabrication consiste à tricoter des fils de suture, élaborant ainsi des matrices hautement poreuses, dont le comportement en traction est similaire, dans sa forme, à celui d’un tendon ou d’un ligament. En couplant le procédé de tricotage et celui d’electrospinning, il est possible de construire des scaffolds, où des microfibres alignées sont déposées sur la surface des structures tricotées. Ce procédé innovant autorise un ensemencement cellulaire facile et efficace des scaffolds et nous avons montré que les cellules s’orientent spontanément selon la direction des fibres, imitant ainsi la morphologie des tendons et des ligaments. Dans une future utilisation, dans un bioréacteur appliquant de la traction-torsion cyclique, les microfibres vont pouvoir transmettre les déformations aux cellules et stimuler la synthèse de la matrice extracellulaire / Tissue engineering is a pluridisciplinary domain aiming at elaborating biological autologous prosthesis. In this strategy, the fabrication of structures, called scaffolds, used for cell culture is necessary. We developed several fabrication techniques of these three-dimensional structures. The first technique (solvent casting/particulate leaching out), involving a polymer solution and spherolized glucose particles, allows the elaboration of scaffolds, owing spherical and well interconnected pores. We showed that the scaffolds are biocompatible and that their mechanical properties in compression can be adjusted. The second technique, electrospinning, leads to the elaboration of biocompatible fibrous membranes whose fiber diameter can be controlled from 800 nm to several micrometers. The third technique of scaffold fabrication proceeds by the elaboration of knitted scaffolds from suture threads. The knitted scaffolds are highly porous and their tensile behavior is similar, in its shape, to the ligaments and tendons stress-strain curves. Using knitting and electrospinning, it has been possible to fabricate knitted scaffolds where aligned microfibers are deposited on their surface. This innovative process allows an easy and efficient cell seeding and we showed that cells are orientated along the fibers, mimicking thus tendons and ligaments morphology. In the future, theses scaffolds will be used in a bioreactor where cyclic traction and torsion will be applied. The aligned microfibers will be able to fully transmit the deformation to the cells, stimulating by this mean the extracellular matrix synthesis

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Thesis (MSc)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Trace amounts of soluble Pt(II/IV) ions are not recovered using current refining processes. There are both economic and environmental incentives to recover these Pt(II/IV) ions from effluent. The work presented in this dissertation was aimed at producing functionalised electrospun nanofibre webs for the extraction of trace amounts of Pt(II/IV) ions in the form of [PtCl6]2- from acidic solutions. An insoluble, low molecular weight oligomeric compound, poly(N-terephthaloylthiourea)-N’,N’-piperazine, was synthesised from relatively inexpensive starting reagents using a “one-pot” two step synthesis procedure. Interest in this compound lies in its ability to extract Pt(II/IV) ions from acidic, chloride-rich solutions, as may be encountered in real process solutions in platinum group metal refineries. The product was isolated and characterised with an array of techniques, including GPC, elemental analysis, 1H and 13C NMR, as well as FTIR, and it was found to be a mixture of various molecular weight fractions with a degree of chemical variance between oligomer chains. The poly(N-terephthaloylthiourea)-N’,N’-piperazine was blended with polyacrylonitrile (PAN) and electrospun using both the classical single needle approach as well as a high throughput free-surface electrospinning process, called ball electrospinning. The nanofibres consisted of the oligomer which provided the affinity for [PtCl6]2- while PAN provided sufficient polymer chain entanglement which allowed the formation of fibrous structures. Two different solutions were found to produce nanofibres with the desired dimensions, namely: 6 wt% and 8 wt% PAN solution, both having a PAN to oligomer ratio of 7:3. The fibres produced by needle electrospinning and ball electrospinning had average fibre diameters of 172 ± 35 nm and 210 ± 49 nm, respectively. The ball electrospinning process had 86 times greater fibre production rates compared to needle electrospinning. The effects of three experimental conditions on the recovery of Pt(II/IV) ions by the poly(N-terephthaloylthiourea)-N’,N’-piperazine-containing nanofibres were determined. The conditions were: (i) the effects of specific surface area and available coordination sites over time, (ii) the effect of extraction temperature, and (iii) the effect of hydrochloric acid (HCl) concentration on [PtCl6]2- extraction. Increased availability of coordination sites caused an increase in Pt ion extraction. The Pt ion extraction also increased from 0.007 g to 0.023 g for each gram of nanofibres used as the temperature was increased from 20 °C to 60 °C when using a 114 mg/L Pt stock solution. The HCl concentration had no effect on Pt ion extraction when varied between 1.0 x 10-3 M to 1 M, while increased extraction as well as fibre damage was caused at HCl concentrations greater than 1 M. Nanofibres containing an oligomeric compound with affinity for [PtCl6]2- in acidic solutions were successfully synthesised and used to extract trace amounts of Pt(II/IV) ions from solutions under various conditions. / AFRIKAANSE OPSOMMING: In huidige verfynings prosesse word spoorelemente van oplosbare Pt(II/IV) nie herwin nie. Daar is beide ekonomiese en omgewings insentiewe om hierdie Pt(II/IV) ione te verhaal uit die afval oplossings. Hierdie tesis was gemik daarop om funksionele elektrospinde nanovesel webbe te produseer vir die herwinning van Pt(II/IV) ioon spoorelemente in die vorm van [PtCl6]2- uit aangesuurde oplossings. ‘n Onoplosbare oligomeriese verbinding met ‘n lae molukulêre gewig, poly(N-terephthaloylthiourea)-N’,N’-piperazine, was uit relatief goedkoop begin reagense gesintetiseer deur gebruik te maak van ‘n “een-pot” twee stap prosedure. Die belangrikheid van die verbinding lê in sy vermoë om Pt(II/IV) ione uit aangesuurde, chloried-ryke oplossing te onttrek, soos wat in alledaagse afval oplossings van platinum-groep metalurgiese raffinaderye ondervind kan word. Die sintese produk was geisoleer en gekarakariseer deur gebruik te maak van ‘n verskeidenheid tegnieke, waaronder GPC, elementêre analise, 1H en 13C NMR sowel as FTIR, en daar was bepaal dat die produk bestaan uit ‘n mengsel van verskeie molukulêre gewig kettings met ‘n mate van chemiese variansie tussen hulle. Die gesintetiseerde oligometriese verbinding was gemeng met poliakrielonitriel (PAN) en elektrospin deur gebruik te maak van beide die klasieke naald spin proses, sowel as ‘n hoë-produksie vrye oppervlak spin proses, genaamd die bal elektrospin proses. Die nanovesels bestaan uit die oligomeer wat die affiniteit vir die [PtCl6]2- voorsien terwyl die PAN genoegsame polimeer ketting verstrengeling veroorsaak het om die veselagtige struktuur te vorm. Nanovesels met die gewensde dimensies was gevorm deur die elektrospin proses toe te pas op twee verskillende oplossings, naamlik: ‘n 6 massa persent PAN en ‘n 8 massa persent PAN oplossing, beide met ‘n PAN tot oligomeer verhouding van 7:3. Die vesels geproduseer deur die naald en bal elektrospin prosesse het ‘n gemiddelde vesel deursneë gehad van 172 ± 35 nm en 210 ± 49 nm, onderskeidelik. Die bal spin proses het egter ‘n 86 keer groter produksie kapasiteit van vesels gehad in vergelyking met die naald spin proses. Die effek van drie verskillende toestande op die effektiwiteit van die nanovesels, wat poly(N-terephthaloylthiourea)-N’,N’-piperazine bevat, om Pt(II/IV) ione te onttrek uit die oplossings was ondersoek. Die toestande was: (i) die effekte van spesifieke oppervlak area asook beskikbare ontginnings setels oor tyd, (ii) die effek van die ontginnings temperatuur, en (iii) die effek van die soutsuur (HCl) konsentrasie op die Pt ioon ontginning. ‘n Toename in die beskikbaarheid van die ontginnings setels het gelei tot ‘n toename in die Pt ioon ontginning. Die Pt ioon ontginning het toegeneem van 0.007 g tot 0.023 g vir elke gram van nanovesels gebruik soos die temperatuur verhoog was van 20 °C tot 60 °C wanneer ‘n 114 ppm (m/v) Pt ioon oplossing gebruik was. Daar was geen effek op die Pt ioon ontginning toe die HCl konsentrasie tussen 1.0 x 10-3 M en 1 M HCl varieer was nie, alhoewel daar by konsentrasies hoër as 1M ‘n verhoogde ontginning sowel as vesel skade was. Nanovesels wat ‘n oligemetriese verbinding bevat met ‘n affiniteit vir [PtCl6]2- in ‘n aangesuurde oplossing, was suksesvol gesintetiseerd en gebruik om spoorelemente van Pt(II/IV) te onttrek onder verskillende omstandighede.

Electrospinning process is a simple and widely used method for producing polymeric nanofibres. However, despite its popularity, significant challenges remain in controlling the fibre deposition due to the complex nature of electrospinning process. The process is renowned for its chaotic motion of fibre deposition, also known as the whipping instability. This instability is caused by electrostatic and fluid dynamics interactions of the charged jet and it is partly responsible for the thinning of the fibres into nanoscale diameters. Due to the instability, an electrospinning process typically deposits random orientated fibres in a circular deposition area. Furthermore, there is no control over the location where the fibres land on the collector electrode except that the fibres always travel through the shortest trajectory between the source and the collector electrodes. In this study, an alternative controlled deposition technique was proposed based on electric field manipulation (EFM). The main hypothesis of this study is that a consistent and repeatable method of controlled deposition can be achieved by using EFM. EFM was achieved by introducing a pair of charged auxiliary electrodes positioned adjacent and perpendicular to the fibre deposition direction. The applied voltage of either direct current (dc) or time-varying (ac) voltage at the auxiliary electrodes act as control to influence the spatial location and size of the deposition area. Samples were produced on black paper substrates and scanned into greyscale images. An image analysis technique was developed to measure the shift and size of the deposition area. A computer simulation was used to calculate the electric field strength and to simulate the behaviour of fibre response based on the trajectory of a charged particle. An image analysis based on greyscale intensity measurement was also developed to examine the uniformity of the deposition area. Finally, fibre characterisation was carried out to examine the fibre morphology, diameter, and orientation based on scanning electron micrographs. The results from this study showed that EFM can provide a consistent and repeatable control of the deposition area. When the auxiliary electrodes were independently charged with two dc voltages, it was observed that the deposition area moved away from the most positive electrode. The magnitude of shift of the deposition area was found to increase linearly with voltage difference between the auxiliary electrodes. Furthermore, the aspect ratio of the deposition area (ratio of width over height) decreased linearly with base voltage i.e. lower of the two auxiliary electrode voltages. These two controls were found to act independently from each other and can be described as two separate controls i.e. voltage difference for spatial location and base voltage for aspect ratio of the deposition area. A similar response was observed in simulation i.e. the particle moved away from the most positive electrode. Simulation results also showed that the x-axis component of the electric field (Ex) was responsible for the shift in location and the reduction of aspect ratio of the deposition area. When the auxiliary electrodes were charged with two antiphase time-varying voltages, continuous scanning of the electrospinning jet was observed producing a wide electrospun fibre mat. It was first thought the smooth oscillation of a sine wave would produce a more uniform deposition pattern compared to a triangle wave, but the results showed otherwise. The inferior uniformity of the sine wave sample was found due to the variability of the jet scanning speed when compared to the constant speed achieved when using a triangle wave. It was also observed that the deposition pattern can be further improved by using two clipped triangle wave voltages. The results open up the possibility for further exploiting the control voltage to achieve the desired deposition pattern. Two case studies were presented to demonstrate the applicability of the technique in real electrospinning applications. In the first case study, it was demonstrated that the continuous scanning of electrospinning jet was capable of eliminating the stripe deposition pattern which is commonly associated to a multi-spinneret electrospinning system. In the second case study, it was found that the alignment and distribution of aligned fibres in a gap electrospinning system can be improved by using the EFM technique. A new technique was also introduced to produce a multi-layer orientated fibre construct. These application examples showed that the EFM technique is ready for the production of engineered electrospun fibre constructs. This would extend the use of electrospun fibres to applications which is currently limited by geometrical constraints of the fibre constructs.

Chemical sensors based on arrays of polymer-coated surface acoustic wave (SAW) devices are required for defense applications that provide a combination of sensitivity, selectivity, portability and response time. The primary challenge in the development of these polymer-based microsensors is the need to reproducibly deposit high quality, defect-free polymer coatings onto microelectrodes. Coating methods such as air brushing and solvent casting have proven unreliable and I have investigated the possibility of depositing polymer films on microelectrodes using electrostatic processing methods. In this work AC electrospraying was used to deposit nanoscale polymer films onto the surface of microelectrodes. The alternating polarity of the electric field eliminates surface charge accumulation and the polymers were deposited uniformly across both electrically insulating and conducting surface regions. In a different work, DC electrospraying was used to deposit patterned organic coatings onto the surface of microelectrodes. The surface of the microelectrode array consisted of an alternating pattern of insulating, grounded-metallic and ungrounded metallic regions, each with a width of 15µm. The charged particles were deposited only onto the grounded-metallic surface regions where there is an electrical path for charge dissipation. No polymer deposition was observed on the insulating or ungrounded-metallic regions due to the effects of surface charge accumulation. Also, I, DC electrodeposited organic molecules within the pores of ceramic film. Due to electrospraying, this film has a strong built-in electric field that induces Stark effect in the organic molecules, providing a unique new technology for bio and chemical sensing. Electrospinning has been used to produce polymer nanofibers with diameters ranging from a few microns to less than 100 nanometers. Due to mechanical oscillations of the electrically charged fibers during electrospinning, they are usually collected in the form of a non-woven mat without any significant fiber orientation. I have developed a new method for making highly aligned arrays of polymer nanofibers by using an AC coupled DC field to drive the electrospinning process. This new "biased AC electrospinning" method can be used to deposit aligned arrays of polymer nanofibers onto virtually any substrate. Potential applications of well-ordered nanofiber materials include tissue engineering, filtration, drug delivery and microelectronics.

This study focused on the comparison of the electrospun silk scaffolds to the electrospun silk fibroin gel scaffolds. Moreover, this study examined the differences in cross-linking effects of genipin and methanol as well as solvents on the mechanical properties and cell compatibility of the scaffolds. Silk scaffolds were electrospun from an aqueous solution or 1,1,1,3,3-hexafluoro-2-propanol (HFIP) without genipin, immediately after 8 % (wt) genipin was added to the solution, and 18 hours after genipin blended with the solution. Uniaxial tensile testing determined that the silk scaffolds electrospun from water exhibit a higher modulus and peak stress than that of the silk scaffolds electrospun from HFIP. In vitro cell culture was conducted to determine the cell compatibility of the various silk fibroin-based scaffolds. 4'-6-Diamidino-2-phenylindole (DAPI) staining and histology suggest that genipin may enhance cell compatibility, and that neither ethanol nor methanol inhibit cell interactions.

Tissue engineering is an interdisciplinary field that aims to restore, maintain, or improve diseased or damaged tissues. Electrospinning has become one of the most popular means to fabricate a scaffold for various tissue engineering applications as the process is extremely versatile and inexpensive. The ability for electrospinning to consistently create nanofibrous structures capable of mimicking the native extracellular matrix (ECM) is the basis behind why this technique is so successful in tissue engineering. Cardiovascular disease has been the leading cause of death in the United States for over 100 years, and because of this, the need for coronary artery replacements is in serious demand. More specifically, small diameter vessels (<6 mm I.D.) are most needed, due to the fact that they are most often affected and the current clinical replacements provide less than optimal long-term patency and regenerative ability. Tissue engineering of vascular grafts has been investigated for over 50 years, however, synthetic replacements made of Dacron® and expanded-poly(tetrafluoroethylene) (e-PTFE) still remain the clinical standard. This study examines a variety of different ways to alter different characteristics of electrospun constructs, to create scaffolds that would be favorable for use as a blood vessel replacement; the end goal being the creation of an acellular bioresorbable vascular graft that would provide sufficient mechanical support to withstand physiological forces, as well as ample biocompatibility to allow host cells to infiltrate and regenerate the graft as the structure degrades. As a way of tailoring the mechanical and thermal properties of a scaffold to be more conducive to that of a native artery, a novel co-polymer was created from the random copolymerization of two monomers; 1,4-Dioxan-2-one (DX) and DL-3-methyl-1,4-dioxan-2-one (DL-3-MeDX) were mixed at different ratios and electrospun, forming nanofibrous scaffolds that exhibited different mechanical and thermal properties. Next, scaffolds were electrospun from natural and synthetic polymers, and the potential for these materials to elicit the formation of an acute thrombotic occlusion was investigated by quantifying tissue factor expression from monocytes using a novel technique. Tissue factor expression by monocytes on the electrospun natural and synthetic polymer scaffolds was compared to that of e-PTFE to determine their potential for use as vascular graft materials. Platelet-rich plasma (PRP), a naturally occurring blood component which is comprised of supraphysiologic concentrations of autologous growth factors, was activated and lyophilized to form a preparation rich in growth factors (PRGF). PRGF was electrospun for the first time, to create a scaffold that would mimic the role of the native ECM in the wound healing cascade. Characterization of these scaffolds proved their bioactivity was enhanced, with cell infiltration occurring throughout the structures in as little as 3 days. Lastly, PRP/PRGF and/or heparin were incorporated into electrospun PCL scaffolds as a means of enhancing the regenerative potential and reducing the thrombogenic potential of the scaffolds, while supplying the constructs with mechanical stability. The release of several pro-regenerative growth factors and chemokines from the PRP incorporated scaffolds was analyzed and the effect of PRP and heparin on scaffold degradation characteristics was determined. Additionally, cell proliferation, migration, sprout formation, and chemokine release were evaluated, and results from these experiments proved the addition of PRP could enhance the regenerative potential of the electrospun scaffolds. The results from this study reveal the variety of ways in which a number of characteristics of an electrospun scaffold can be altered to create a more ideal bioresorbable vascular graft that has the potential to be regenerated within the body, while providing enough mechanical support for this to occur over time.

Tissue engineering scaffolds come in many shapes and sizes, however, due to difficulty manufacturing the microstructure architecture required in tissue engineering, most scaffolds are architecturally non-dynamic in nature. Because the microstructural architecture of all biological tissues is inherently complicated, non-dynamic tissue engineering scaffolds tend to be a poor platform for tissue regeneration. The current method for manufacturing dynamic tissue engineering scaffolds involves electrospinning successive layers of different fibers, an approach that exhibits no fiber transition between layers and subsequent delamination problems. In this study we aim to address the design challenges of tissue engineering scaffolds through our novel integrated fiber electrospinning technique. Developed in our lab, this electrospinning technique makes it possible to manufacture complex electrospun scaffolds tailorable to specific tissue engineering needs while minimizing delamination tendencies. Our goal is to enhance the capabilities of the tissue engineering field by increasing the manufacturable scaffold complexity and overall structural integrity of electrospun scaffolds.

Electrospinning is a technique used to generate scaffolds composed of nano- to micron-sized fibers for use in tissue engineering. This technology possesses several key weaknesses that prevent it from adoption into the clinical treatment regime. One major weakness is the lack of porosity exhibited in most electrospun scaffolds, preventing cellular infiltration and thus hosts tissue integration. Another weakness seen in the field is the inability to physically cut electrospun scaffolds in the frontal plane for subsequent microscopic analysis (current electrospun scaffold analysis is limited to sectioning in the cross-sectional plane). Given this it becomes extremely difficult to associate spatial scaffold dynamics with a specific cellular response. In an effort to address these issues the research presented here will discuss modifications to electrospinning technology, cryosectioning technology, and our understanding of cellular infiltration mechanisms into electrospun scaffolds. Of note, the hypothesis of a potentially significant passive phase of cellular infiltration will be discussed as well as modifications to cell culture protocols aimed at establishing multiple passive infiltration phases during prolonged culture to encourage deep cellular infiltration.

Electrospinning is a process by which polymer fibers can be produced using an electrostatically driven fluid jet. Electrospun fibers can be produced at the micro- or nano-scale and are, therefore, very promising for air filtration applications. However, because electrospun fibers are electrically charged, it is difficult to control the morphology of filtration media. Fiber size, alignment and uniformity are very important factors that affect filter performance. The focus of this project is to understand the relationship between filter morphology and performance and to develop new methods to create filtration media with optimum morphology. This study is divided into three focus areas: unimodal and bimodal microscale fibrous media with aligned, orthogonal and random fiber orientations; unimodal and bimodal nanoscale fibers in random orientations; bimodal micrometer and nanometer fiber media with orthogonally aligned orientations. The results indicate that the most efficient filters, which are those with the highest ratio of particle collection efficiency divided by pressure drop, can be obtained through fabricating filters in orthogonal layers of aligned fibers with two different fiber diameters. Moreover, our results show that increasing the number of layers increases the performance of orthogonally layered fibers. Also, controlling fiber spacing in orthogonally layered micrometer fiber media can be an alternative way to study the filtration performance. Finally, such coatings presented throughout this research study can be designed and placed up-stream, down-stream, and/or in between conventional filters.