Nanoparticle therapy in a zebrafish model of tuberculosis
In the past ten years, our group at IBV has established the tuberculosis (TB) model of infection in zebrafish embryos by using the fish TB pathogen Mycobacterium marinum. For this, the mycobacteria are microinjected via the blood, or directly in the tissue of interest, causing a chronic infection lasting for about 10 days and characterized by the presence of infected macrophage aggregates called granulomas, the main focus of the infection in the zebrafish, as it is in humans. In this system, we are investigating drug nanocarriers as possible alternatives to conventional free drug administration. In human TB the standard treatment involves the use of four different antibiotics at high doses every day for at least 6 months, and up to 2 years in multi-drug resistant TB; these treatments are associated with severe toxicity issues. Extensive studies in mouse and guinea pig models infected with Mycobacterium tuberculosis show that when the same drugs are encapsulated inside nanoparticles made from lipids or polymers the frequency of dosing can be greatly reduced leading to better therapy with less toxicity. However, it has proven difficult to follow the fate of the nanoparticles in these animals, that are opaque. In contrast, zebrafish larvae are especially suited for this kind of study, thanks to their optical transparency and for the availability of transgenic lines that have fluorescently labelled specific cell types, such as macrophages, neutrophils or endothelial cells. In this vertebrate model it is therefore possible and simple to study the fate of nanoparticles, bacteria and immune cells in real time, simultaneously and non-invasively. As in humans the focus of the infection involves the granulomas. In TB, the diseased tissue stimulates new blood vessels formation (angiogenesis) that are induced to sprout in response to signals originating from the granuloma. Nanoparticles are known to passively accumulate in the newly-forming vessels during cancer development and we have recently established this to be the case also for angiogenic vasculature of granulomas in tuberculosis. There are many parallels between tumors and granulomas.
Focus of this Masters project
- In depth study of the mechanism of how nanoparticles get transported from the blood vessels across the endothelium (extravasation) to access the TB granulomas using state of art microscopy techniques (confocal, spinning disc, electron microscopy). A collaboration with a group of UiO mathematicians will be instrumental to further rationalize how NP flow characteristics influence the ability of the NP to concentrate in TB granulomas.
- Analysis of the accumulation of NP in the diseased site using vasoconstrictor drug co-treatment; this is hypothesized to enhance NP permeability through the blood vessels and to increase access of the NP to the granulomas.
- Once we have found the best conditions for therapy, drug-loaded NP will be tested. We currently collaborate with different international groups (In India, New Zealand, Belgium and Germany) who provide new tuberculosis drugs and innovative nanoparticle formulations.
Students in our group will find a friendly and stimulating environment and will learn:
- Microscopy techniques: 1)Light microscopy: stereomicroscope/confocal/spinning disc, Software: Imagej and IMARIS 2)Electron microscopy: TEM and SEM 3)Histology/anatomy of zebrafish and antibody labeling of sections.
- Handling of zebrafish embryos and Mycobacterium marinum, including microinjections
- Simple nanoparticle preparation including encapsulation of a fluorescent dye and a drug