title: Enzyme replacement in Fabry endothelial cells and fibroblasts: uptake experiments and electron microscopical studies.
abstract: Endothelial cells are of particular interest for therapeutic strategies in Fabry's disease, because the accumulation of glycosphingolipids in the vascular endothelium as a result of alpha-galactosidase A (alpha-galA) deficiency is responsible for the major clinical manifestations of the disease. Electron microscopical observations of cultured endothelial cells obtained from the umbilical vein of a hemizygous Fabry fetus showed that the glycosphingolipids are deposited as lamellar material in the lysosomes, as has been found previously for cultured fibroblasts and many different tissues. Mannose 6-phosphate (man 6-P)-receptor mediated and Concanavalin A (ConA)-mediated uptake of purified alpha-galA was attempted in the endothelial cells as well as in cultured fibroblasts from the same fetus. Our results on high-uptake alpha-galA indicate that the endothelial cells do not internalize alpha-galA via the man 6-P receptor. Immunofluorescence studies after addition of the receptor antibody to the cells support the theory that they have no or very few man 6-P receptors on the surface. Morphological studies did not show lysosomal changes which could suggest that the enzyme is taken up into the endothelial cells; however, we found reproducible modifications of the lysosomes in Fabry fibroblasts after incubation with high-uptake alpha-galA. Cell-associated alpha-galA activity was found in both cell types, when the enzyme was added to cells preincubated with ConA; but the lectin treatment by itself induced considerable ultrastructural changes in the cytoplasm, which obscured a possible effect by the enzyme.
authors.affiliation: Institute of Medical Genetics, Panum Institute, Copenhagen, Denmark.
authors.forename: L
authors.initials: L
authors.lastname: Hasholt
authors.forename: A
authors.initials: A
authors.lastname: Wandall
authors.forename: S A
authors.initials: SA
authors.lastname: Sørensen
mesh_terms: D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism; D002478:Cells, Cultured; D003208:Concanavalin A / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D000795:Fabry Disease / Q000201:enzymology / Q000473:pathology*; D005333:Fetus; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D005696:Galactosidases / Q000378:metabolism*; D006801:Humans; D008247:Lysosomes / Q000378:metabolism / Q000648:ultrastructure; D008297:Male; D017527:Receptor, IGF Type 2; D000519:alpha-Galactosidase / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D017527:Receptor, IGF Type 2; D003208:Concanavalin A; D005696:Galactosidases; D000519:alpha-Galactosidase