Isolation and characterization of monoclonal antibodies specific for chondroitin sulfate E.

Chondroitin sulfate E (CSE) is a polysaccharide containing mainly disaccharide units of D-glucuronic acid (GlcA) and 4,6-O-disulfated N-acetyl-D-galactosamine (GalNAc) residues (E-unit) in the amount of ∼ 60%. CSE is involved in many biological and pathological processes. In this study, we established new monoclonal antibodies, termed E-12C and E-18H, by using CSE that contained more than 70% of E-units as an immunogen. These antibodies recognized CSE but not other CSs isomers or dermatan sulfate (DS). We evaluated the reactivities of the antibodies to 6-O-sulfated CSA (6S-CSA) and DS (6S-DS) that possessed ∼ 60% of GalNAc (4S, 6S) moieties in their structures. Neither of the antibodies reacted with 6S-DS. The antibodies strictly distinguished the structural difference of GlcA and L-iduronic acid in the polysaccharide. Binding affinities of the antibodies were determined by a surface plasmon resonance assay using CSE and 6S-CSA. The binding affinities were strongly associated with the molecular weight of CSE and the E-unit content of 6S-CSA. Moreover, we demonstrated that the antibodies are applicable to histochemical analysis. In conclusion, the new anti-CSE monoclonal antibodies specifically recognize the E-unit of CSE. The antibodies will become useful tools for the investigation of the biological and pathological significance of CSE.

The sialidase inhibitor 2,3-dehydro-2-deoxy-N-acetylneuraminic acid is a glucose-dependent potentiator of insulin secretion.

Sialidase cleaves sialic acid residues from a sialoglycoconjugate: oligosaccharides, glycolipids and glycoproteins that contain sialic acid. Histochemical imaging of the mouse pancreas using a benzothiazolylphenol-based sialic acid derivative (BTP3-Neu5Ac), a highly sensitive histochemical imaging probe used to assess sialidase activity, showed that pancreatic islets have intense sialidase activity. The sialidase inhibitor 2,3-dehydro-2-deoxy-N-acetylneuraminic acid (DANA) remarkably enhances glutamate release from hippocampal neurons. Since there are many similar processes between synaptic vesicle exocytosis and secretory granule exocytosis, we investigated the effect of DANA on insulin release from ß-cells. Insulin release was induced in INS-1D cells by treatment with 8.3 mM glucose, and the release was enhanced by treatment with DANA. In a mouse intraperitoneal glucose tolerance test, the increase in serum insulin levels was enhanced by intravenous injection with DANA. However, under fasting conditions, insulin release was not enhanced by treatment with DANA. Calcium oscillations induced by 8.3 mM glucose treatment of INS-1D cells were not affected by DANA. Blood insulin levels in sialidase isozyme Neu3-deficient mice were significantly higher than those in WT mice under ad libitum feeding conditions, but the levels were not different under fasting conditions. These results indicate that DANA is a glucose-dependent potentiator of insulin secretion. The sialidase inhibitor may be useful for anti-diabetic treatment with a low risk of hypoglycemia.