1-Deoxymannojirimycin inhibits capillary tube formation in vitro. Analysis of N-linked oligosaccharides in bovine capillary endothelial cells.
Capillary endothelial cells can be induced to form capillary-like structures in vitro by plating on fibronectin-coated dishes (Ingber, D. E., and Folkman, J. (1989) J. Cell Biol. 109, 317-330), thereby mimicking angiogenesis. To assess the role of glycoproteins bearing asparagine-linked oligosaccharides in this process, we tested the effect of oligosaccharide processing inhibitors on the formation of capillary tubes. Deoxymannojirimycin, a compound that prevents synthesis of hybrid and complex-type oligosaccharides, inhibited the formation of capillary tubes. In contrast, swainsonine, an inhibitor that blocks synthesis of complex- but not hybrid-type oligosaccharides, did not inhibit tube formation. Lectin affinity chromatography of 2-[3H] mannose-labeled glycopeptides from endothelial cells induced to form tubes did not reveal a striking difference in the spectrum of oligosaccharides compared to uninduced cells. Since endothelial cells formed tubes normally in the presence of swainsonine, we analyzed glycopeptides from swainsonine-treated induced and uninduced cells. Cells induced to form tubes were enriched in monosialylated hybrid-type oligosaccharides sensitive to alpha-fucosidase, beta-galactosidase, and beta-N-acetylhexosaminidase, suggestive of sialyl Lewis-X determinants. We used an enzyme-linked immunoassay to measure sialyl Lewis-X epitopes on capillary endothelial cells and found that both induced and uninduced cells expressed sialyl Lewis-X epitopes. Deoxymannojirimycin and, to a lesser extent, swainsonine reduced the level of sialyl Lewis-X epitopes in cells induced to form capillary tubes, but neither compound affected the level of epitopes in cell monolayers. We conclude that synthesis of at least hybrid-type oligosaccharides is required for capillary tube formation in vitro and that an increase in monosialylated, fucosylated glycans on asparagine-linked oligosaccharides occurs during this process.
Nguyen
M
M
Surgical Research Laboratory, Children's Hospital, Boston, Massachusetts.
Folkman
J
J
Bischoff
J
J
D017485:1-Deoxynojirimycin / Q000494:pharmacology*; D000302:Adrenal Cortex / Q000098:blood supply; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D002196:Capillaries / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002999:Clone Cells; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006020:Glycopeptides / Q000096:biosynthesis* / Q000302:isolation & purification; D008358:Mannose / Q000378:metabolism*; D008361:Mannosidases / Q000037:antagonists & inhibitors; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000096:biosynthesis* / Q000302:isolation & purification; D017026:Swainsonine / Q000494:pharmacology
D000470:Alkaloids; D006020:Glycopeptides; D009844:Oligosaccharides; C065629:kifunensine; D017485:1-Deoxynojirimycin; D008361:Mannosidases; D008358:Mannose; D017026:Swainsonine