7 General discussion and future perspectives 123 Nephropathic cystinosis In chapter 2, an experimental zebrafish embryo model for the autosomal recessive disease of nephropathic cystinosis is introduced. Nephropathic cystinosis is a lysosomal storage disease where the CTNS gene is mutated, which leads to the accumulation of cystine in lysosomes. If left untreated, the disease is fatal.(106, 107) Currently, specific treatment is limited to cysteamine, which prevents further cystine accumulation but does not reverse the damage. Moreover, drug compliance is relatively low due to adverse effects of bad breath, skin odour, gastro-intestinal complaints such as nausea, vomiting, diarrhoea, and abdominal pain.(108-110) A Ctns knockout mouse has been developed, but this model lacks the glomerular changes also seen in nephropathic cystinosis.(111-113) The ctns -/- zebrafish mutant introduced in this study is presented as a promising model for the investigation of new therapeutic options and the pathophysiology of nephropathic cystinosis. The model displays a phenotype similar to that of the human disease, including cystine accumulation, increased glomerular permeability, and decreased proximal tubular reabsorption. They have higher mortality than wild-type animals. These last symptoms were preventable by treating mutant embryos with cysteamine. Renal and extrarenal manifestations of cystinosis have also been described in the adult model of this mutant.(114) The zebrafish cystinosis model has already been used to test novel treatment strategies for nephropathic cystinosis, such as luteolin, disulfiram, and bicalutamide-cysteamine.(115-118) Heparan sulphate glycosaminoglycans Chapters 3 and 4 discuss the previously held paradigm that heparan sulphate glycosaminoglycans are essential to glomerular filtration barrier function. This hypothesis was formulated by Kanwar and Farquhar several decades ago and was based on the finding that enzymatic removal of HS-GAG resulted in the loss of GFB integrity.(7, 8) Also, HS-GAG expression has been found to be reduced in various proteinuric renal diseases.(22) However, based on the results presented in chapters 3 and 4, homozygous germline mutations in zebrafish and, respectively, heterozygous mutations in humans of HS backbone elongating enzymes are shown not to result in proteinuria, nor a renal phenotype. The role of HS-GAG has long been thought to provide the GFB its charge selectivity due to the negatively charged sulphate groups of heparan sulphate. In chapter 3, we show that a significant reduction in negatively charged sites in the glomerular basement membrane does not result in proteinuria. Results from other experimental animal models with HS-GAG deficiencies are in line with this notion.(23-25, 27, 119) In chapter 4, the effect of heterozygous germline mutations on the backbone elongating
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