Oligodendrocyte precursor cells (OPCs) are a major transplant population to promote myelin repair and central nervous system (CNS) regeneration in conditions such as Multiple Sclerosis and spinal cord injury. Magnetic particles (MPs) can offer a multifunctional platform for cell therapies, facilitating labelling for cell tracking (e.g. by MRI and histopathology); biomolecule delivery (including nonviral gene delivery, enhanceable by novel ‘magnetofection’ strategies); and magnetic cell targeting of transplant populations. However, MP-based applications for neural tissue engineering have received limited attention to date. This thesis demonstrates that ~60% of OPCs (derived from a primary source) can be safely labelled using two well-characterised MP formulations, including a novel multimodal MP with transfection plus cell labelling capabilities. A rapid, technically simple, high-throughput ultrastructural imaging technique, OTOTO SEM, has been developed to study the surface interactions of MPs with precursor cells. Safe MP-mediated transfection of OPCs was demonstrated, including with multiple and therapeutic genes. Transfection efficiency was enhanced by static/oscillating ‘magnetofection’ techniques (~21%; competitive with nonviral alternatives). Organotypic cerebellar slice cultures were developed as a model of ‘host’ neural tissue, and ‘magnetofected’ OPCs exhibited normal migration, proliferation and differentiation profiles following transplantation onto such slices. Safe labelling (~45%) and transfection (enhanced by static/oscillating magnetofection strategies: ~6%) of oligodendrocytes was achieved utilising identical protocols to those developed for OPCs. A comparative intralineage analysis demonstrated that MP-uptake and amenability to transfection were significantly lower in oligodendrocytes compared to OPCs. Inter-cellular comparisons of MP-handling by the four major CNS glial subtypes (viz. OPCs, oligodendrocytes, astrocytes, microglia; derived from the same primary source) revealed major differences in the rate/extent of MP uptake, amenability to transfection, optimal magnetofection frequency, and MP-associated toxicity. Finally, a stoichiometrically-defined glial co-culture model was developed and utilised to test the hypothesis that microglia represent an ‘extracellular barrier’ to MP uptake by other glia.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:702500 |
| Date | January 2013 |
| Creators | Jenkins, Stuart Iain |
| Publisher | Keele University |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://eprints.keele.ac.uk/3821/ |
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