BACKGROUND AND AIMS\nMETHODS\nRESULTS\nCONCLUSIONS\nScavenger receptors form a superfamily of membrane-bound receptors that bind and internalize different types of ligands, including pro... Show moreBACKGROUND AND AIMS\nMETHODS\nRESULTS\nCONCLUSIONS\nScavenger receptors form a superfamily of membrane-bound receptors that bind and internalize different types of ligands, including pro-atherogenic oxidized low-density lipoproteins (oxLDLs). In vitro studies have indicated a role for the liver sinusoidal endothelial cell receptors stabilin 1 (stab1) and 2 (stab2) in oxLDL clearance. In this study, we evaluated the potential role of stab1 and stab2 in lipoprotein uptake in zebrafish, an upcoming model for studying cholesterol metabolism and atherosclerosis.\nLipoproteins were injected in the duct of Cuvier of wild-type (ABTL) or stab1 and stab2 mutant (stab1-/-stab2-/-) zebrafish larvae at 3 days post-fertilization. To examine the effect of stabilin deficiency on lipoprotein and cholesterol metabolism, zebrafish larvae were challenged with a high cholesterol diet (HCD; 4% w/w) for 10 days.\nLipoprotein injections showed impaired uptake of both LDL and oxLDL into the vessel wall of caudal veins of stab1-/-stab2-/- zebrafish, which was paralleled by redistribution to tissue macrophages. Total body cholesterol levels did not differ between HCD-fed stab1-/-stab2-/- and ABTL zebrafish. However, stab1-/-stab2-/- larvae exhibited 1.4-fold higher mRNA expression levels of ldlra involved in (modified) LDL uptake, whereas the expression levels of scavenger receptors scarb1 and cd36 were significantly decreased.\nWe have shown that stabilins 1 and 2 have an important scavenging function for apolipoprotein B-containing lipoproteins in zebrafish and that combined deficiency of these two proteins strongly upregulates the clearance of lipoproteins by macrophages within the caudal vein. Our current study highlights the use of zebrafish as model to study lipoprotein metabolism and liver sinusoidal endothelial cell function. Show less
Background: Recently, an emerging flavivirus, Usutu virus (USUV), has caused an epidemic among birds in Europe, resulting in a massive die-off in Eurasian blackbirds. Currently found only in Europe... Show moreBackground: Recently, an emerging flavivirus, Usutu virus (USUV), has caused an epidemic among birds in Europe, resulting in a massive die-off in Eurasian blackbirds. Currently found only in Europe and Africa, it can be envisioned that Usutu virus will follow the path of other flaviviruses, like West Nile virus and Zika virus, and will spread via its mosquito vectors and bird hosts to other parts of the world. Several cases of human infections by Usutu virus have already been published. Anticipating this spread, development of an efficacious vaccine would be highly desirable. Method: This study describes the production in E. coli, purification, and refolding of a partial USUV envelope protein. Prior to immunization, the protein was characterized using size exclusion chromatography, transmission electron microscopy and dynamic light scattering, showing the limited presence of virus-like structures, indicating that the protein solution is probably a mixture of mono and multimeric envelope proteins. Results: Immunizations of two rabbits with the refolded E-protein fraction, mixed with a strong adjuvant, resulted in the generation of neutralizing antibodies, as evidenced in an in vitro assay. Discussion: The way forward towards a subunit vaccine against Usutu virus infection is discussed. Show less
Verwilligen, R.A.F.; Dijke, A. van; Hoekstra, M.; Bussmann, J.; Eck, M. van 2019
Atherosclerosis, the main underlying cause of cardiovascular disease and related morbidities, is a multifactorial disease in which liver endothelial cells and macrophages have shown to play an... Show moreAtherosclerosis, the main underlying cause of cardiovascular disease and related morbidities, is a multifactorial disease in which liver endothelial cells and macrophages have shown to play an important role. Recently, it has been demonstrated that oxidized phospholipids in oxidized LDL (oxLDL) are pro-inflammatory and pro-atherogenic in hypercholesterolaemic mice. Multiple scavenger receptors are known for oxLDL. They form a superfamily of membrane bound receptors that bind and internalize different types of ligands including other modified lipoproteins, endogenous proteins and pathogens. Several studies, including a microarray study of our research group, have already identified scavenger receptors stabilin-1 and -2 (STAB1 and STAB2) as a risk factor for atherosclerosis, and therefore as potential targets for treatment. An upcoming and powerful model organism to study early atherogenesis and that allows non-invasive cell tracking studies is the zebrafish. Recent studies have demonstrated the possibility to induce (early) atherosclerosis in zebrafish.We have used Crispr/Cas9 mutagenesis to generate zebrafish stab1 and stab2 mutants.Our results show that oxLDL clearance by zebrafish scavenger endothelial cells (homologous to liver sinusoidal endothelial cells) is diminished in stab2 zebrafish mutants and is absent in stab1/2 double mutants. In stabilin mutants, macrophage oxLDL uptake is increased, suggesting a protective role for stabilins in foam cell formation.To study this further, we are currently performing an atherosclerosis study to examine foam cell formation and vascular lipid accumulation in stabilin mutant zebrafish. Show less
Horeman, T.; Blikkendaal, M.D.; Feng, D.; Dijke, A. van; Jansen, F.; Dankelman, J.; Dobbelsteen, J.J. van den 2014
