Novel method for chase analysis of oligosaccharide metabolic error caused by xenobiotics.
Saccharide primers, such as dodecyl beta-lactoside (Lac-C12), are unique artificial precursors of glycolipid synthesis. In culture media supplemented with saccharide primers, they are taken up by the cells in the culture media and glycosylated by cellular glycosyltransferases in the Golgi apparatus to form pseudo-glycolipids. In this study, we examine the effects of various xenobiotics on glycolipid synthesis by implementing a novel method to analyze pseudo-glycolipids, mainly gangliosides, produced by ONS-76 medulloblastoma cells in a culture medium containing various xenobiotics. The ganglioside group of pseudo-glycolipids was effectively purified by using strong anion-exchange cartridges. The production of pseudo-gangliosides was stimulated by N-(n-butyl)deoxygalactonojirimycin (NB-DGJ), but was inhibited by castanospermine, 2-deoxy-2-fluoro-d-galactose, tunicamycin, and brefeldin A. Because the cells in the culture medium are exposed to the saccharide primers and xenobiotics at the same time, and are secreted in the culture medium in their glycosylated form, our method can be used to effectively analyze the direct effects of xenobiotics on ganglioside synthesis.
Kato
Tomohisa
T
Japan Chemical Innovation Institute, Chiyoda-ku, Tokyo 101-0051, Japan. tkato@iis.u-tokyo.ac.jp
Muraoka
Miho
M
Hatanaka
Kenichi
K
D017485:1-Deoxynojirimycin / Q000031:analogs & derivatives / Q000494:pharmacology; D020126:Brefeldin A / Q000494:pharmacology; D045744:Cell Line, Tumor; D002855:Chromatography, Thin Layer / Q000379:methods*; D005643:Fucose / Q000031:analogs & derivatives / Q000494:pharmacology; D005732:Gangliosides / Q000032:analysis / Q000096:biosynthesis / Q000302:isolation & purification; D006031:Glycosylation; D006801:Humans; D007212:Indolizines / Q000494:pharmacology; D009844:Oligosaccharides / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology; D015262:Xenobiotics / Q000494:pharmacology*; D054836:Indolizidines
D017485:1-Deoxynojirimycin; D020126:Brefeldin A; D005643:Fucose; D005732:Gangliosides; D007212:Indolizines; D009844:Oligosaccharides; D014415:Tunicamycin; D015262:Xenobiotics; C090092:migalastat; C049289:2-deoxy-2-fluorogalactose; C037806:castanospermine; D054836:Indolizidines