Lysosomal storage disorders (LSDs) are a group of orphan diseases characterized by lysosomal dysfunction or impaired lysosomal catabolism and affect collectively about 1 in 5000 live births. A... Show moreLysosomal storage disorders (LSDs) are a group of orphan diseases characterized by lysosomal dysfunction or impaired lysosomal catabolism and affect collectively about 1 in 5000 live births. A common LSD is Gaucher disease, which is characterized by a defect in glucocerebrosidase (GCase) degrading glucosylceramide (GlcCer) in lysosomes. In this thesis, the zebrafish is evaluated as vertebrate animal model for the investigation of lysosomal storage disorders, in particular Gaucher disease. Zebrafish are an appealing model organism to study genetic disorders with a high evolutionary conservation of genes and proteins compared to humans, easy maintenance and simple genetic and pharmacological manipulation. Zebrafish larvae are of particular use as zebrafish can generate hundreds of off-spring which have a rapid embryonal development, are transparent and fit in a 96-wells plate. In this thesis several biochemical and genetic techniques have been developed in order to 1) compare the catalytic features of zebrafish GCase with human GCase, 2) investigate the consequences of its defect in zebrafish larvae and adults as well as a concomitant defect in non-lysosomal GBA2 and 3) study the potential toxicity of excessive glucosylsphingosine during GCase deficiency as consequence of a defect in lysosomal acid ceramidase. GCase-deficient zebrafish showed similar symptoms and affected molecular mechanisms as patients and mouse models. Therefore the zebrafish offers exciting new possibilities to study molecular mechanisms underlying pathological processes during lysosomal hydrolase deficiencies. Show less
Aerts, J.M.F.G.; Kuo, C.L.; Lelieveld, L.T.; Boer, D.E.C.; Lienden, M.J.C. van der; Overkleeft, H.S.; Artola, M. 2019
Glycosphingolipids are important building blocks of the outer leaflet of the cell membrane. They are continuously recycled, involving fragmentation inside lysosomes by glycosidases. Inherited... Show moreGlycosphingolipids are important building blocks of the outer leaflet of the cell membrane. They are continuously recycled, involving fragmentation inside lysosomes by glycosidases. Inherited defects in degradation cause lysosomal glycosphingolipid storage disorders. The relatively common glycosphingolipidosis Gaucher disease is highlighted here to discuss new insights in the molecular basis and pathophysiology of glycosphingolipidoses reached by fundamental research increasingly using chemical biology tools. We discuss improvements in the detection of glycosphingolipid metabolites by mass spectrometry and review new developments in laboratory diagnosis and disease monitoring as well as therapeutic interventions. Show less
Advanced mass spectrometry of glycosphingolipids takes the central stage in this thesis. Investigations focus on characterization of glycosphingolipid metabolism in health and disease with... Show moreAdvanced mass spectrometry of glycosphingolipids takes the central stage in this thesis. Investigations focus on characterization of glycosphingolipid metabolism in health and disease with emphasis to the detection and accurate quantitation of known and so far unknown glycosphingolipids and closely regulated metabolites. Inherited defects in lysosomal degradation of glycosphingolipids, in particular the glycosphingolipidoses Gaucher disease (GD) and Fabry disease (FD), relatively common lysosomal storage disorders, are key topics of examination. The thesis provides an introductory background on the field of research and contains three different sections describing conducted experimental work. Section one consists of studies reporting on the discovery of excessive occurrence of glycosphingoid bases in lysosomal storage diseases, the development of methods for their accurate quantitation in biological samples with UPLC-ESI-MS/MS and the use of these methods in diagnosis and disease monitoring. The great value of identical 13C-encoded (glyco)sphingolipids and their bases as internal standards in mass spectrometric quantitation of these lipids in biological materials is described. Section two introduces clinical aspects and challenges of GD and FD and provides examples of the practical value of lipid analyses in the GD and FD clinic. Section three concerns the pathophysiology of lysosomal disorders in glycosphingolipid metabolism and related fundamental investigations. Show less
