Chapter 7 126 processes, including renal physiology. They are even part of the aquatic habitat on the International Space Station and are one of the few vertebrates to have lived a full life cycle in space.(125) In chapters 2, 3, 5, and 6, an experimental zebrafish (Danio rerio) embryo model is used to assess whether knocking down mRNA translation of a single gene results in the development of proteinuria and whether tubular reabsorption mechanisms remain intact. Using this model presents several advantages compared to other experimental animal models. First, zebrafish embryos develop rapidly. Most major organs are formed within 40 hours post-fertilization. Due to their mostly transparent appearance, this development can be visualized relatively easily. Secondly, a single pair of adult zebrafish can lay over 200 eggs. Thus, in controlled conditions, it is possible to create high throughput models. The zebrafish embryo kidney consists of a pronephros with two nephrons that share a fused glomerulus in the midline of the body. Despite its simple structure compared to the more complex human metanephros, the zebrafish kidney shares many similar features with the kidneys of higher vertebrates and as such, is increasingly used as an experimental model for the study of cellular and molecular mechanisms of renal pathophysiology.(105, 126) Because of these characteristics, these animals are highly suited for investigating individual components of the pathways leading to proteinuria. (39, 40, 43, 82, 105, 126, 127) In chapters 2 and 3, genetically mutated zebrafish were used as experimental models. In chapters 5 and 6, gene knockdown was effectuated by injecting zebrafish embryos with morpholino constructs. These constructs bind to mRNA and thus inhibit translation, leading to a functional knockdown of the targeted gene and its mRNA. In all these models, functional assays of glomerular filtration barrier integrity and tubular reabsorption were assessed by injecting a mixture of TRITC-labelled 3 kDa and FITC-labelled 70-kDa dextrans. As 3 kDa dextrans can freely pass the glomerular filtration barrier, they are reabsorbed in endosomes in the proximal tubule under physiological conditions. On the other hand, 70 kDa dextrans do not readily pass the GFB and as such, are only reabsorbed when GFB integrity is compromised. Thus, the presence of 3 kDa droplets was used to assess whether tubular reabsorption mechanism functions properly. The presence of 70 kDa was used to assess the loss of GFB integrity. This model was developed in these studies after adapting it from Hentschel et al. (40) Other methods to assess GFB integrity in zebrafish embryos have also been developed by others. For example, a transgenic zebrafish expresses its main serum protein, vitamin D binding protein, bound with green fluorescent protein (EGFP). This model removes
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