Transient expression of type 2 chain in A-active hexaglycosylceramide of rat small intestine at weaning time. Demonstration by affinity chromatography and ceramide glycanase hydrolysis of A-active glycosphingolipids followed by gas chromatography and mass spectrometry of permethylated hexasaccharides.
The small intestine of 15- to 23-day-old rats was cut into four segments from the duodenum to the ileum. Neutral glycosphingolipids were purified from each segment and submitted to thin-layer chromatography and immunostaining with the A005 monoclonal anti-A antibody. This antibody detected an hexaglycosylceramide located mainly in the duodenum during the postnatal development. In order to characterize hexaglycosylceramides, blood group A-active glycolipids were purified by affinity chromatography on immobilized Helix pomatia lectin in organic solvent. Hexaglycosylceramides (A-6) were subsequently isolated by preparative thin-layer chromatography and hydrolyzed with ceramide glycanase. The free hexasaccharides were permethylated and analyzed by gas chromatography. Two peaks were detected in varying ratios during development, corresponding to type 1 and type 2 chain A hexasaccharides. Gas chromatography clearly demonstrated that type 2 A-6 occurred in the duodenum of developing rats, and that a shift from type 2 to type 1 A-6 occurred with growing age. The change from type 2 to type 1 chain was also assessed by methylation analysis, and by the variation of the characteristic fragmentations of type 1 and type 2 chain hexasaccharides upon mass spectometry of the permethylated A-6 oligosaccharides from the duodenum of 19-day-old and adult rats.
Bouhours
D
D
Institut National de la Santé et de la Recherche Médicale, Unité 76, Paris, France.
Bouhours
J F
JF
Larson
G
G
Karlsson
H
H
Pimlott
W
W
Hansson
G C
GC
D000375:Aging; D000818:Animals; D000911:Antibodies, Monoclonal; D002518:Ceramides / Q000096:biosynthesis* / Q000302:isolation & purification; D002846:Chromatography, Affinity; D002849:Chromatography, Gas; D002855:Chromatography, Thin Layer; D004386:Duodenum / Q000254:growth & development; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry; D006028:Glycosphingolipids / Q000302:isolation & purification; D007082:Ileum / Q000254:growth & development; D007421:Intestine, Small / Q000254:growth & development* / Q000378:metabolism; D007583:Jejunum / Q000254:growth & development; D013058:Mass Spectrometry; D008745:Methylation; D024510:Muscle Development; D009130:Muscle, Smooth / Q000254:growth & development / Q000378:metabolism; D009844:Oligosaccharides / Q000096:biosynthesis* / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains; D014886:Weaning
D000911:Antibodies, Monoclonal; D002518:Ceramides; D006028:Glycosphingolipids; D009844:Oligosaccharides; C065996:blood group A-active hexaglycosylceramide type 1 chain; C065997:blood group A active hexaglycosylceramide type 2 chain