The work described in this dissertation contributes to a better mechanistic understanding of nanoparticles in vivo. To achieve that goal, we used the zebrafish as a highly predictive pre-screening... Show moreThe work described in this dissertation contributes to a better mechanistic understanding of nanoparticles in vivo. To achieve that goal, we used the zebrafish as a highly predictive pre-screening model of nanoparticles. This approach enables the investigation of the fundamental behavior of nanoparticles, correlation of the physicochemical properties of the formulated nanoparticles with their biodistribution and identification of important nano-bio interactions. Zebrafish established transgenic lines were used to study specific interactions. In addition, genetically modified zebrafish applying CRISPR/Cas9 were generated. These strategies not only show key mechanistic features of nanoparticles in circulation, but also promote the rational design of more efficient nanoparticles systems.After understanding the fundamental behavior of nanoparticles, this thesis describes the identification of a key interaction between stabilins receptors (expressed in liver sinusoidal endothelial cells) and nanoparticles. Next, the scope is changed to design nano-systems that target specific cell types showing liposomes capable of switching the surface charge in situ and in vivo using light as an external trigger and a rationally designed lipid nanoparticle formulation containing mRNA able to preferentially target the hepatic reticuloendothelial system. In addition, a phase-separated liposomes hijacking a lipase mediated transport to selectively target endothelial lipase in vivo was studied. Show less
This thesis is focused on the innate immune defence mechanisms responsible for controlling mycobacterial growth after infection. To provide a detailed description of the host__s innate immune... Show moreThis thesis is focused on the innate immune defence mechanisms responsible for controlling mycobacterial growth after infection. To provide a detailed description of the host__s innate immune response to M. marinum infection, zebrafish gene expression levels were analysed by RNA sequencing at various time points during infection and correlated with imaging data of the process of pathogenesis. We demonstrate that the scavenger receptor Marco (macrophage receptor with collagenous structure) is a key player in the rapid phagocytosis of M. marinum and we use gene expression analysis in combination with gene knockdown studies to show that it is also essential in the establishment of an initial transient pro-inflammatory response to M. marinum infection. Once phagocytosed, M. marinum is capable of avoiding killing mechanisms of the host cell and can continue to grow within macrophages. This is the period when Membrane Attack Complex/Perforin proteins are involved in killing intracellular bacteria by their pore-forming activities. We reveal the regulatory mechanisms and function of two macrophage specific genes, mpeg1 and mpeg1.2 (macrophage expressed gene 1.2). The results from this thesis complement knowledge obtained from other model organisms by providing new insights into both counteracting and supporting mechanisms underlying the innate immune response. Show less
