published_or_final_version / abstract / Surgery / Doctoral / Doctor of Philosophy

published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

Vaccination is one of the most effective public health interventions in the world, saving millions of lives and preventing the onset of debilitating diseases. With widespread emergence of multi-drug resistant pathogens, the importance of preventative medicine has become even more apparent. However, one of the limiting factors in developing novel vaccines that are both safe and highly immunogenic is the availability of adjuvant delivery systems licensed for human use. The purpose of this study was to investigate the role gold nanoparticles could play as an effective vaccine delivery system. A variety of coupling chemistries were explored for their ability to conjugate protein and polysaccharide antigens onto the surface of gold nanoparticles for the development of vaccines against a number of biologically important human pathogens including Y. pestis, B. mallei and S. pneumoniae. Retention of antigenicity and coupling efficiency of conjugated molecules was measured using characterisation techniques such as localised surface plasmon resonance and immunoblotting. Gold nanoparticle coupled antigens were then used to immunise mice and to measure the protective efficacy and the immunological response induced. The findings indicate antigen-specific immune responses are elevated when an antigen is coupled onto gold nanoparticles. Moreover, immunological data from nanoparticle coupled glycoconjugate vaccines against B. mallei and S. pneumoniae indicate the likely presence of a strong T cell immune response which is essential for providing immunological memory. Finally, an intracellular trafficking assay was carried out to identify some of the mechanisms that might be involved in uptake of gold nanoparticles into professional phagocytes. Confocal imaging of receptors associated with endosomal compartments revealed that gold nanoparticles may enter cells through multiple pathways. The findings reported in this study suggest that gold nanoparticles may be an excellent candidate for further investigation as a novel vaccine delivery system.

Cancer is one of the most notorious diseases affecting the human population today with very few effective treatments. Due to the disparate nature of cancers, it is difficult to obtain a treatment that can cure cancer. Thus, there is a large influx of research towards cancer therapies, leading to one of the discovery that cancer cells (tumors) have a low thermotolerance in comparison to normal cells. If the temperature of the cancer cells is increased into the hyperthermia range (~45°C) thermal damage occurs, causing cell death by protein denaturation and membrane disruption. A recent development in this field has been in the photothermal treatment of tumors, which is starting to utilize plasmonic particles to enhance the specificity of the treatment. The plasmonic nanoparticles, specifically gold, can reach the tumor site using passive targeting and when irradiated with a tuned laser will emit heat localized to a small region around the nanoparticle killing the surrounding cancer cells. This process has been shown to reduce tumor size in vivo with gold nanoshells and gold nanorods. However, it has not been shown which particle is better at delivering the heat to the tumor site. Therefore in this study, it will be shown which particle generates the most heat. Solutions of tissue simulating phantom and different concentrations of nanoparticles were irradiated with a laser to measure the increase in temperature. Additionally, simulations were performed using Mie Theory for nanoshells and the Discrete Dipole Approximation for nanorods. Based on the physical parameters of the nanoshells and nanorods used in this experiment, the adjusted absorption cross-section was determined. It was found that nanoshells generated the most amount of heat on a per particle basis, and that it was necessary to have a nanorod concentration of 5.5 times the concentration of nanoshells to generate the same amount of heat as nanoshells. These results were confirmed using Monte Carlo and Finite Difference Modeling of the nanoparticle heating experiments. However, the choice of nanoparticle still depends on the application and the targeting efficiency in vivo. / text

Glycerol is a very promising alternative fuel to hydrogen in fuel cells. However, the utilisation of glycerol as a fuel requires a good catalyst, due to the slow kinetics of glycerol electrooxidation. Gold has been identified as a promising catalyst due to its high activity and stability for glycerol electrooxidation – although the overpotentials are higher than on platinum and palladium. Modification of a nano-Au/C catalyst by the addition of MnO2, in an attempt to further improve the activity and lower the overpotential for glycerol oxidation, was therefore first explored. This was followed by investigations into the effects of gold particle size and loading. Finally, the effect of gold particle size on oxidation of gold-catalysed glycerol oxidation intermediates was also briefly explored. Studies into MnO2 addition showed that the pre-deposition of MnO2 yielded catalysts with smaller, more uniform gold particles, and catalysts with MnO2 contents of 5 and 9 wt % had higher mass activities and lower onset- and peak- potentials than Au/C. All the Au/xMnO2/C catalysts were more active than the palladium- and platinum-based catalysts reported in literature, which effectively demonstrated the advantage of using a gold-based catalyst for glycerol oxidation – especially when supported by MnO2 which lowered the overpotential for glycerol oxidation over gold. For the study into gold particle size, small gold particles of average diameter ≤ 4.7 nm had higher gold mass-based activities than medium-sized (14.7 nm) particles and were at least twice as active as catalysts containing large (≥ 43 nm) gold particles. The small gold particles also gave lower glycerol oxidation onset potentials, which was attributed to the predominance of Au(110) planes on those particles. Glycerol oxidation also appeared to proceed further along the oxidation pathway over small gold particles, which was confirmed in preliminary studies into the oxidation of glycerol oxidation intermediates. However, specific activity increased with increasing gold particle size, due mainly to the higher intrinsic activity of the Au(111) plane, which increased relative to Au(110) with increasing gold particle size. The important requirements for fuel cell applications are factors such as high mass activity, low overpotentials and high stability – all of which were met by the catalysts containing small gold particles defined by predominantly Au(110) facets. Investigations into the gold loading effect showed similar mass- and specific- activities for catalysts with 5-20 % gold loading. However, only the catalysts with higher gold loadings (15-20 %) did not deactivate early during CV, indicating that a larger gold surface area is necessary to resist poisoning at high potentials. On the basis of low onset potentials, high mass activity, and stability at low overpotentials, a minimum gold loading of 12.5 % appears to be necessary for a supported gold catalyst with small gold nanoparticles; although even higher loadings may be preferable for a higher power output in a fuel cell. Importantly, the insights gleaned from this study on the fundamental properties required for early activation, activity and stability of the gold catalysts could lead to a more intelligent design of gold-based catalysts in future.

The integrity of groundwater sources is constantly threatened by contaminant plumes generated by accidental gasoline leakages and leachates escaping landfills. These plumes are of concern due to their proven toxicity to living organisms. Aromatic and chlorinated hydrocarbons, volatile fatty acids, phenols, and ammonia have been found in these leachates. In addition, benzene, toluene, and xylenes (BTX) are major components of gasoline. The lack of oxygen in groundwater makes anaerobic bioremediation desired for the treatment of groundwater contaminated with BTX and chlorinated solvents. With the objective of finding microorganisms capable of BTX and cis-dichloroethylene (cis-DCE) degradation under anaerobic conditions for their use in permeable reactive barriers, different inocula were tested in batch experiments. Toluene was rapidly degraded by several inocula in the presence of alternative electron acceptors. Benzene and m-xylene were eliminated by few of the inocula tested after incubation periods ranging from 244 to 716 days. cis-DCE was highly recalcitrant as no degradation was observed over 440 days. Biological processes have been successfully applied for the treatment of landfill leachates as well. In an effort to provide an effective and economical alternative, an anaerobic-aerobic system was evaluated using a synthetic media simulating the organic and ammonia content of real leachates. The removal of the organic content reached 98% in an upflow anaerobic sludge blanket reactor, and resulted in the formation of methane. During the aerobic process, in an innovative down-flow sponge reactor, ammonia was highly transformed to nitrite and nitrate. Complete nitrification was eventually achieved.The capacity of current wastewater treatment plants for removing nanoparticles has been questioned during the last years. Nanoparticles have been incorporated into numerous applications and their presence in wastewater seems to be inevitable. A laboratory-scale secondary treatment system was set-in to study the behavior of cerium and aluminum oxide nanoparticles during wastewater treatment. The nanoparticles were highly removed, suggesting that secondary treatment is suitable for their elimination. The removal of these nanoparticles was influenced by the pH and organic content of the wastewater. Aluminum nanoparticles proved to be toxic; however the performance of the system for eliminating the organic content was recovered over time.

Intracellular hyperthermic therapy may prove to be a unique and novel approach to the management of pancreatic cancer. Utilizing the principle of photothermal destruction, selective killing of cancer cells with minimal injury to normal tissues may be possible. This dissertation investigated the role of antibody targeted metal nanoparticles and the cytotoxic effects of nonionizing radiofrequency fields in pancreatic cancer. Cancer cell death was induced by heat release from intracellular metal nanoparticles after radiofrequency field exposure. Fluorescent and gold nanoparticles were delivered with two antibodies, cetuximab and PAM-4, to pancreatic cancer cells in vitro and mouse xenografts in vivo. Selective delivery of these nanoparticles induced cell death in vitro and decreased tumor burden in vivo after whole animal RF field exposure. This occurred through both apoptosis and necrosis. In addition, activated caspase-3 was increased after antibody treatment and RF field exposure. Furthermore, although there was non-specific uptake by the liver and spleen in vivo, there was no evidence of acute or chronic toxicity in the animals. These results are in agreement with the principle that malignant cells are more thermally sensitive than normal cells or tissues. Selective intracellular delivery of metal nanoparticles coupled with whole body RF field exposure may be a beneficial therapy against micrometastases and unresectable pancreatic cancer in the future. Further studies are planned with more specific antibodies, other nanoparticles, and other cancer targets.

Chiral molecules, which exist under enantiomers with non-mirror-symmetrical structures, have been the subject of intense research for their linear and nonlinear optical activities. Cysteine is such a chiral amino acid found as a building block of proteins throughout human bodies. Second harmonic generation (SHG) has been considered to investigate chiral molecules. SHG from metallic nanoparticles is promising for nanoplasmonics and photonic nanodevice applications. Therefore, it’s desirable to combine and study nonlinear properties due to both chirality and metallic nanoparticles, and help developing an alternatively optical diagnostic of chiral molecules. Our experiments are carried out with the FemtoFiber Scientific FFS laser system. SHG of silver nanoparticles (Ag NPs) modified by either L-Cysteine (L-C) or D-Cysteine (D-C) is observed, where L-Cysteine and D-Cysteine are a pair of enantiomers. Ag NPs are deposited through Vacuum Thermal Evaporation, controlled under different deposition thicknesses. UV-Vis/IR spectra and AFM are used to characterize Ag NPs under different conditions. Transmitted SHG measurements dependent on incidence are recorded with standard lock-in techniques. Deposition thickness of vacuum thermal evaporation plays an important role in forming diverse Ag NPs, which strongly imparts the intensity of SHG. Second harmonic intensity as a function of the incident angle presents similar results for Ag NPs with or without L-Cysteine or D-Cysteine modification, in the output of p- and s-polarization. However, we monitor reversed rotation difference in second harmonic intensities at linearly +45° and -45° polarization for L-C/Ag NPs and D-C/Ag NPs, while there’s no difference at linearly +45° and -45° polarization for Ag NPs alone. This optical rotation difference in SHG is termed as SHG-ORD. Also, for second harmonic light fixed at p-polarization, L-C/Ag NPs and D-C/Ag NPs exhibit a reversely net difference for SHG excited by right and left circular polarization, which is termed as SHG-CD. Experiments on SHG-ORD of chiral-modified Ag NPs by a mixture of L-Cysteine and D-Cysteine further help verifying the existence of chirality in chiral-modified Ag NPs. As a conclusion, SHG efficiently probed and distinguished L-Cysteine from D-Cysteine in chiral-modified Ag NPs. / Thesis (Master, Physics, Engineering Physics and Astronomy) -- Queen's University, 2013-09-29 21:27:23.112

Silica nanoparticles are effective fluorophore carriers with high potential in imaging, diagnostics, and therapy. The particles are resistant to drastic change of environmental conditions (pH, temperature etc.) and insulate the dyes so as to protect them from photobleaching. Silica chemistry is also versatile and affords an easy modification of the particle composition and surface to integrate targeting ligands or to integrate other nanoparticles. Regardless of their advantages, there exists a lack of dye diversity in the literature that is connected to a low affinity for potential tools for biology and medicineThis thesis describes the development of an alternative method for the synthesis of fluorescent silica nanoparticles and their modification to incorporate iron oxide and gold. cont/d.

The use of nanoscale materials is growing exponentially, but there are also concerns about the environmental hazard to aquatic biota. Metal-containing engineered nanoparticles (NPs) are an important group of these new materials, and whilst there are undoubtedly a plethora of beneficial uses for these NPs, it is essential that an appropriate risk assessment is carried out in order to protect the environment and human health, with the consumption of contaminated fish a distinct possibility. The current study aimed to assess the bioavailability, uptake and toxicological effects of two metal-NPs (TiO2 NPs and Cu-NPs) to fish from both dietary and waterborne exposure routes and where appropriate compare them to their bulk counterpart. Whole body system effects were assessed along with the influences of the life stage of exposed fish and abiotic factors on toxicity. A technique to improve the quantification of Ti from TiO2 NPs in fish tissue was also developed. Effects from exposure to dietary TiO2 NPs manifested similarly to traditional dietary metal exposure, with no reduction in growth, but some sublethal affects. Exposure to waterborne Cu-NPs showed that rainbow trout were more acutely sensitive to CuSO4 than the NPs, but that despite limited uptake several body systems were affected (most notably ionoregulation). Larvae were more sensitive to CuSO4 than Cu-NPs, but no differences were seen with embryos, whilst larvae were more sensitive than embryos. Abiotic factors did have an effect on acute Cu-NP toxicity, though not always in a predictable manner, with some effects more pronounced than with CuSO4. Overall, it appears that metal-NPs are not as acutely toxic as their bulk counterparts, but sublethal effects, were routinely observed. As TiO2 NPs appear more toxic than its bulk counterpart, current legislation governing safe environmental limits may have to be adjusted, though the situation with Cu-NPs isn’t as clear and further investigation is required. However, the risk of human exposure via the consumption of NP contaminated fish fillets is extremely low.