Qualitative and quantitative analyses in sulfated glycosaminoglycans, chondroitin sulfate/dermatan sulfate, during 3T3-L1 adipocytes differentiation

Abstract

Abstract Chondroitin sulfate/dermatan sulfate (CS/DS) is a member of glycosaminoglycans (GAGs) found in animal tissues. Major CS/DS subclasses, O, A, C, D, and E units, exist based on the sulfation pattern in d-glucuronic acid (GlcA) and N-acetyl-d-galactosamine (GalNAc) repeating units. Dermatan sulfate (DS) chains are formed when GlcA is epimerized into l-iduronic acid (IdoA). Our study aimed to analyze the CS/DS profile in 3T3-L1 cells before and after adipogenic induction. Their CS/DS contents, molecular weight (Mw), and sulfation pattern were analyzed by using a high-performance liquid chromatography system. CS/DS synthesis/degradation- and sulfotransferase-related gene expression was also analyzed by reverse transcription real-time PCR. The CS/DS amount was significantly decreased in the differentiated (DI) group compared to the non-differentiated (ND) group, along with a lower expression of CS biosynthesis-related genes, chondroitin sulfate N-acetylgalactosaminyltransferase 1, chondroitin sulfate N-acetylgalactosaminyltransferase 2, and chondroitin polymerizing factor. The GAGs in the DI group also showed lower Mw than those of ND. Furthermore, the A unit was the major CS/DS disaccharide in both groups, with a proportionally higher CS-A ratio in the DI group. This was consistent with the expression of carbohydrate sulfotransferase 12 that encodes chondroitin 4-O-sulfotransferase, for CS-A formation. Unlike the ND group, both GlcA and IdoA residues in the O unit of CS/DS from the DI group were absent. These qualitative and quantitative changes in CS/DS and CS/DS-synthases/hydrolases before and after adipocyte differentiation reveal valuable insights into adipocyte development.