In vivo accuracy of a macrophage-based drug delivery system demonstrated in zebrafish

Journal Title
Journal ISSN
Volume Title
Stellenbosch : Stellenbosch University
ENGLISH ABSTRACT: Targeted drug delivery systems are widely regarded as being the ‘magic bullet’ in the drug delivery research niche. Coined by Paul Ehrlich in 1900, the 'magic bullet’ is an immunological concept for a treatment intervention that targets only damaged or diseased cells in the body, leaving healthy cells intact. Targeted drug delivery systems achieve this by combining a drug carrier capable of precisely and accurately targeting specific tissues, cells, or mechanisms with a therapeutic agent. Many drug carriers have been developed and tested within the last decade with some being more effective than others. Based on prior in vitro work completed by our group, showing that human macrophages could be modified to ingest matter, appropriately translocate and expel the matter, all without degrading said cargo, we selected these cells as potential drug carriers. Based on the high degree of physiological conservation between humans and zebrafish, high fecundity, cheap maintenance, optical transparency - allowing tracking of injected cells - and the fact that primary human macrophages have been found to survive for extended periods in zebrafish larvae, we set out to experimentally determine whether these would be suitable model organisms in which to study and develop a human macrophage-based drug delivery system. First, cell microinjection and fluorescent staining parameters were optimised for long-term tracking of cells in zebrafish larval circulation. The latter parameter was optimised using immortalised, undifferentiated human monocytic leukemia cells (THP-1) which were shown to exhibit endothelial adherence to the caudal hematopoietic region of the larval blood vessels. This behaviour is similar to that of endogenous zebrafish macrophages/monocytes and illustrates a significant degree of conservation between human and zebrafish immune cells. The undifferentiated human THP-1 monocytes also underwent rapid proliferation in response to zebrafish inflammatory stimuli as a result of tail fin transections, suggesting some degree of cross-species reactivity to inflammatory cues. Next, the THP-1 cells were differentiated and polarised to M1 macrophage-like cells to determine if these cells were suitable for drug delivery in zebrafish larvae. These cells were also shown to exhibit adhesion to the zebrafish caudal hematopoietic tissue (CHT) and associated blood vessels, but all became unviable and lost fluorescent signal within 24 hours post injection, without undergoing migration to the transected inflammatory site. The experiment was repeated with unpolarised THP-1 macrophages and yielded similar results, suggesting that THP-1-derived macrophages may be unsuitable for drug delivery research in zebrafish larvae. Finally, the experiments were repeated employing primary human M1 polarised macrophages. These cells proved to exhibit a more suitable survival capacity, maintaining cellular viability to the experimental endpoint, however migrational capacity ultimately remained insufficient for drug delivery. This series of experiments lays a solid foundation for future studies which could definitively answer whether zebrafish larvae are suitable models in which to investigate a macrophage-based drug delivery system.
AFRIKAANSE OPSOMMING: Geen opsomming beskikbaar.
Thesis (MSc)--Stellenbosch University, 2022.
Zebrafish, Drug delivery systems, Macrophages -- Effect of drugs on, UCTD