Electrospinning has attracted great attention in the fields of tissue engineering and controlled release of drugs/biomolecules. The aim of this project was to investigate electrospinning of nanofibers with core-shell structures using emulsion electrospinning, the formation of monolithic and core-shell structured nanofibrous drug/biomolecule delivery vehicles using polymers such as poly(D,L-lactic acid) (PDLLA) and poly(lactic-co-glycolic acid) (PLGA), and the formation of multicomponent bone tissue engineering scaffolds with angiogenic property, osteoinductivity and osteoconductivity. The foundation of this project was laid by investigating monocomponent scaffolds. First, effects of properties of polymer solutions and water-in-oil (w/o) emulsions and electrospinning parameters on the morphology, diameter and structure of fibers were systematically investigated. Second, drugs (vancomycin and rifamycin) and a model protein (bovine serum albumin) were incorporated in monolithic or core-shell nanofibers via blend electrospinning or emulsion electrospinning to form single or dual delivery systems, providing fundamental understandings. Growth factors such as recombinant human bone morphogenetic protein-2 (rhBMP-2) and basic-fibroblast growth factor (b-FGF) were then incorporated in PLGA or PDLLA nanofibrous delivery vehicles. The in vitro release behaviour of drugs and biomolecules was studied. Third, calcium phosphate (Ca-P) nanoparticles were synthesized and used for fabricating Ca-P/PLGA and Ca-P/PDLLA nanocomposite scaffolds. Homogeneous distribution of Ca-P in fibrous scaffolds could be achieved. With the assistance of emulsion electrospinning and nanocomposite electrospinning, bicomponent scaffolds containing rhBMP-2 and Ca-P nanoparticles were fabricated using dual-source dual-power electrospinning. The fibrous component ratio could be varied by using multiple syringes for electrospinning fibers. The structure and properties, including in vitro release behaviour, of mono- and bicomponent scaffolds were studied in detail. Tricomponent scaffolds incorporated with recombinant human vein endothelial growth factor (rhVEGF), rhBMP-2 and Ca-P nanoparticles were subsequently fabricated using multi-source dual-power electrospinning. To achieve a sequential release of firstly rhVEGF and then rhBMP-2, PLGA/polyethylene glycol (PEG) blends and PLGA were used for incorporating rhVEGF and rhBMP-2, respectively. For tricomponent scaffolds with different component ratios, different release amounts but similar release profiles could be achieved for the growth factors. In vitro biological investigations were conducted for mono-, bi- and tricomponent scaffolds. Pre-osteoblast cells (MC3T3-E1) were found to attach, spread, proliferate and express alkaline phosphatase (ALP) activity on rhBMP-2 and Ca-P nanoparticle incorporated bicomponent scaffolds. Calcium deposition was also observed in cells cultured with bicomponent scaffolds. Human umbilical vein endothelial cells (HUVECs) were found to attach, spread, proliferate on tricomponent scaffolds and rhVEGF released from mono-, bi- and tricomponent scaffolds could facilitate cell proliferation and migration, indicating released rhVEGF could promote angiogenesis. C3H10T1/2 cell line and human bone marrow derived mesenchymal stem cells (hBMSCs) were found to attach, spread and proliferate on bi- and tricomponent scaffolds. As compared with cells seeded on monocomponent scaffolds, C3H10T1/2 cells and hBMSCs on bi- and tricomponent scaffolds expressed higher ALP activity. Enhanced mineralization was observed for C3H10T1/2 cells and hBMSCs seeded bicomponent scaffolds comprising rhBMP-2/PLGA and Ca-P/PLGA fibers and also tricomponent scaffolds. hBMSCs seeded on rhBMP-2/PLGA and Ca-P/PLGA monocomponent scaffolds expressed abundant F-actin and vinculin, while bicomponent and tricomponent scaffolds induced much more F-actin and vinculin expression. / published_or_final_version / Mechanical Engineering / Doctoral / Doctor of Philosophy

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published_or_final_version / abstract / toc / Orthopaedics and Traumatology / Master / Master of Philosophy

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The aims of this study were to prepare poly (ethylcyanoacrylate) (PECA) nanoparticles on the basis of different types of microemulsions, to investigate the entrapment within and release of a bioactive from these particles and to establish a set of delivery systems with varying entrapment and release characteristics, thereby giving the formulator the opportunity of a more tailor-made approach in the development of a delivery system. Furthermore the scale up of particle preparation and the possible enhancement of the immunogenic properties of PECA particles by incorporation of the adjuvant Quil A was investigated. Methods: Four phase triangles were established and microemulsion samples, used as a template to prepare nanoparticles, were characterised by viscosity and conductivity measurements, polarising light microscopy, freeze fracture transmission electron microscopy (TEM), cryo field emission electron microscopy (cryo FESEM) and self-diffusion NMR to determine their microemulsion type (droplet, bicontinuous, solution type). PECA nanoparticles were prepared from different types of microemulsions by interfacial polymerisation. Particle size, polydispersity index (PI) and [zeta]-potential were measured by photon correlation spectroscopy and electrophoretic mobility respectively. Normal scanning electron microscopy (SEM) and cryo FESEM were used to visualise particles. Fluorescently labelled ovalbumin (FITC-OVA) was used as a model protein/antigen and entrapment within and release from nanoparticles was investigated. To scale up nanoparticle preparation an instrumental set-up with reactor, peristaltic pump and stirrer was used. A 2⁷ fractional factorial study was designed to observe possible factors or their interactions that could influence particle formation under scale up conditions. For an immunological study freeze dried formulations of PECA nanoparticles, having FITC-OVA and Quil A entrapped, were prepared, and activation and uptake of formulations by murine bone marrow derived dendritic cells (DCs) and T cells in vitro were monitored. Results: Results obtained from the measurements described above, for formulations from the four different phase triangles, indicated that microemulsions of w/o droplet, bicontinuous or solution type could be formed. It was possible to prepare PECA nanoparticles from all of the different types of microemulsions. Particles had an average size of 265 nm � 24, with an average PI of 0.18 � 0.05 and an average negative [zeta]-potential of -17 mV � -5. Particle size, PI and [zeta]-potential were not influenced by the type of microemulsion that was used as a polymerisation template. Entrapment and release were however influenced by the type of microemulsion and although entrapment of FITC-OVA was generally high for PECA particles, it was highest for particles prepared from a droplet type microemulsion. Entrapment could also be increased by increasing amounts of monomer. The rate of release was dependent on the amount of monomer used for polymerisation and the type of microemulsion used for particle preparation, with nanoparticles prepared from a w/o droplet type microemulsion showing the slowest release. Furthermore it was shown that particle preparation could be scaled-up with the instrumental set-up used in this study, but conditions need to be refined as the average particle size and polydispersity index were considerably larger (441 nm � 101, 0.68 � 0.14) when compared to particles prepared by the beaker-pipette method (see above). The adjuvant Quil A could efficiently be entrapped into PECA nanoparticles together with FITC-OVA. Incubation of DCs and T cells with the various formulations did, however, not result in increased uptake or activation. Conclusions: PECA nanoparticles with high entrapment efficiency of antigen and adjuvant can be prepared from different types of microemulsions. Particles show different rates of entrapment and release depending on the type of microemulsion used as a polymerisation template, possibly because two different types of nanoparticles form. Nanocapsules are believed to form on the basis of droplet type microemulsions and nanospheres form on the basis of bicontinuous and solution type microemulsions. Freeze dried formulations of PECA nanoparticles, containing Quil A and FITC-OVA, were not able to induce an immune response, which might be due to charge repulsion effects between the negatively charged PECA nanoparticles and the negatively charged surface of dendritic cells. Moreover, no adjuvant effect of Quil A was apparent, perhaps caused by total encapsulation of the compound into the particle matrix with no active groups extending out displaying adjuvanticity.

Different studies and reports have shown that conventional wet chemical cleaning and sterilization methods of used biomaterials and medical devices are inadequate. There is significant risk of infections and associated diseases that arise due to the use of reprocessed surgical equipment during medical procedures. For example, transmissible spongiform encephalopathy infectivity, which is highly resistant to conventional methods of deactivation, can be transmitted iatrogenically. However, a complete ban on the reuse of medical devices will have major implications for health care costs and will also be environmentally unsound. / Thesis (MApSc)--University of South Australia, 2006.

Thesis (PhD)--University of South Australia, 2002.