title: An in vitro model of Fabry disease.
abstract: Fabry disease is an X-linked inherited loss of alpha-galactosidase A (alpha-Gal A). Affected patients experience complications that include neuropathy, renal failure, and cardiovascular disease. Although the genetic and biochemical basis of this sphingolipidosis is well studied, the basis for the vascular disease remains poorly understood. In an attempt to create a suitable in vitro model of this disease, conditions for the growth of primary cultures of aortic endothelial cells from wild-type and alpha-Gal A -/0 mice were established. The cultured cells demonstrated CD-31 expression by flow cytometry and LDL binding by immunofluorescence. The glycolipid expression patterns were compared between wild-type and alpha-Gal A null cells. Importantly, cells from alpha-Gal A -/0 mice but not alpha-Gal A +/0 mice expressed high levels of the globo-series glycosphingolipid globotriaosylceramide (Gb3). The age-dependent elevation in Gb3 was measured. By 4 mo of age, alpha-Gal A -/0 mouse aortic endothelial cells achieved their peak Gb3 levels. The ability to lower Gb3 levels pharmacologically was assessed next. The glucosylceramide synthase inhibitor ethylenedioxyphenyl-P4 significantly lowered but did not eliminate Gb3 levels by 96 h of treatment. Gb3 synthesis was completely blocked as measured by [14C]galactose labeling. Recombinant alpha-Gal A more significantly lowered Gb3 levels by 48 h but had a more limited effect on de novo synthesis. Together, both agents eliminated detectable Gb3. In summary, primary cultures of aortic endothelial cells from Fabry mice retain the phenotype of elevated globo-series glycosphingolipids. These cells provide a useful model for comparing pharmacologic agents used for glycolipid reduction.
authors: Nephrology Division, Department of Internal Medicine, University of Michigan, Ann Arbor, MI 48109-0676, USA.
authors: Liming
authors: L
authors: Shu
authors: Hedwig S
authors: HS
authors: Murphy
authors: Laura
authors: L
authors: Cooling
authors: James A
authors: JA
authors: Shayman
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004195:Disease Models, Animal; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D000795:Fabry Disease / Q000188:drug therapy / Q000209:etiology* / Q000235:genetics / Q000378:metabolism; D005964:Glucosyltransferases / Q000037:antagonists & inhibitors; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D011412:Propanolamines / Q000494:pharmacology; D011759:Pyrrolidines / Q000494:pharmacology; D011994:Recombinant Proteins / Q000494:pharmacology; D014281:Trihexosylceramides / Q000378:metabolism; D000519:alpha-Galactosidase / Q000235:genetics / Q000494:pharmacology
chemical_list: C404730:3',4'-ethylenedioxyphenyl-2-palmitoylamino-3-pyrrolidino-1-propanol; D004791:Enzyme Inhibitors; D011412:Propanolamines; D011759:Pyrrolidines; D011994:Recombinant Proteins; D014281:Trihexosylceramides; C018549:globotriaosylceramide; D005964:Glucosyltransferases; C023246:ceramide glucosyltransferase; D000519:alpha-Galactosidase