In vivo accuracy of a macrophage-based drug delivery system demonstrated in zebrafish
Date
2022-12
Authors
Journal Title
Journal ISSN
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Publisher
Stellenbosch : Stellenbosch University
Abstract
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.
AFRIKAANSE OPSOMMING: Geen opsomming beskikbaar.
Description
Thesis (MSc)--Stellenbosch University, 2022.
Keywords
Zebrafish, Drug delivery systems, Macrophages -- Effect of drugs on, UCTD