Physical Sciences of Imaging in the Biomedical Sciences CDT
Thesis project - "Development of novel imaging techniques to improve understanding and specificity of targeting and cellular uptake of super paramagnetic iron oxide nanoparticles(SPIONS)"
Professor Iseult Lynch, School of Geography, Earth and Environmental Sciences
Professor Michael Hannon, School of Chemistry
Dr Iain Styles, School of Computer Science
Engineered nanoparticles, such as super paramagnetic iron oxide nanoparticles (SPIONs) offer significant benefits for the development of various diagnostic and therapeutic strategies. These include the targeted delivery of drugs, hypothermia treatment and contrast enhancement in magnetic resonance imaging (MRI). However with the increased growth in production and the therapeutic benefit SPIONs offer, there is a paralleled increase in concerns associated with exposure. It is critical to improve the understanding of the interactions that occur following cellular exposure to these particles to ensure the safe and efficacious use of these particles within biomedecine in the future. The limitations of existing imaging methodologies in the study of NPs, such as the effects of fluorescent labeling and diffraction limited resolution, and the advantages that visualization of spatial localization can offer in studies, increases the demand for new and optimized imaging routines. Existing RCM methods were optimized and R-SIM imaging was introduced, offering a two fold increase in resolution - particularly advantageous for NP quantification and localization studies. Analysis routines were developed to enable the automated quantification of NP presence within cells via the different methodologies. Correlative procedures were also established for imaging the same sample with different reflectance methods and TEM, maximizing the information attainable from a single sample and allowing comparisons between the techniques for specific applications. These optimized techniques were used to determine NP uptake in four different cell lines, and, in combination with siRNA, to ascertain proteins that are involved in the uptake process. Preliminary work aimed at the translation of these studies into 3D culture models, and confirmation of the applicability of reflectance for imaging these 3D spheroid cultures was achieved. Reflectance imaging methods were applied to SPION and cerium trafficking studies, and analysis methods were extended to include fluorescent segmentation and quantitative colocalization measures, determining the eventual fate of SPIONS and cerium dioxide NPs within cancer cells. Mock dissolution studies that simulated the lysosomal compartment within cells to model the degradative process of SPIONs within this fluid, utilizing analytical techniques such as ICP-OES. This thesis thus provided several important tools for the future assessment of the efficacy and safety of NPs for clinical use, enabling quantitative analysis of uptake route, subcellular localization and NP intracellular fate.