Identification of decorin proteoglycan in bovine tracheal serous cells in culture and localization of decorin mRNA in situ.

Bovine tracheal submucosal gland serous cells in culture synthesize and secrete proteoglycans and not mucin glycoconjugates. We are interested in the characterization and role of these proteoglycans in airway secretions. The major [35S]methionine-labeled proteoglycan present is identified as the small chondroitin/dermatan sulfate proteoglycan decorin (PG II. PG40). Consistent with its identity as decorin this proteoglycan showed average apparent molecular weights of 75,000 to 130,000 with a core protein of an average, M(r) of about 40,000 and with glycosaminoglycan chains sensitive to chondroitinase ABC lyase of an average M(r) of about 25,000. These data were obtained from gel chromatographic and SDS-PAGE analyses. Northern blot analysis and partial amino acid sequencing of the purified protein further confirmed its identity as decorin. In situ hybridization studies using a decorin riboprobe revealed no expression of decorin in the surface epithelium and only low levels of expression in submucosal gland epithelial cells of bovine tracheal tissue. However, high levels of expression were localized to cells which are peripheral to tracheal submucosal gland epithelial cells and which contact with the extracellular matrix.

Interrelationship between the Na+/glucose cotransporter and CFTR in Caco-2 cells: relevance to cystic fibrosis.

Both the Na+-dependent glucose cotransporter (SGLT1) and the cystic fibrosis transmembrane conductance regulator (CFTR) modulate Na+ and fluid movement, although in opposite directions. Yet few studies have investigated a possible interrelationship between these two transporters. By using the Caco-2 human colon carcinoma cell line, we confirmed that the activities of these transporters increased with spontaneous differentiation to the enterocytic phenotype. We showed that SGLT1 was positively regulated by Cl- and that optimal activity of CFTR was dependent on the presence of glucose. We also demonstrated that inhibition of CFTR by glibenclamide or diphenylamine-2-carboxylate did not modify the activity of SGLT1 and inhibition of SGLT1 by phlorizin did not modify the activity of CFTR, although it resulted in inhibition of glycoconjugate synthesis. These results point to positive substrate-cross regulation of SGLT1 and CFTR and suggest that NaCl and glucose are important for not only Na+ absorption and fluid movement, but also for cAMP-dependent Cl- efflux, and glycoconjugate synthesis, functions that are known to be anomalous in cystic fibrosis.

Glycoconjugate metabolism in a cystic fibrosis knockout mouse model.

Cystic fibrosis knockout mice (cftr(-/-)) die prematurely of obstruction of the intestine which may result from accumulation of dehydrated glycoconjugate-containing mucus. We noted an increase in the specific activity of [(14)C]glucosamine-labeled high-molecular weight glycoconjugates, probably mucin, in the lumen of the intestine of cftr(-/-) (homozygous) mice compared to cftr(+/+) (wild-type) and cftr(+/-) (heterozygous) mice and a decrease in the turnover of glycoconjugates of several organs of the cftr(-/-) mice. No difference in the anionic composition of secreted intestinal glycoconjugates was detected and no difference in the amount of mucin 1 (Muc1) was found in the small intestine, colon, pancreas, and lungs of the different genotypes. In addition, the spleen of the cftr(-/-) mice was significantly smaller than that of control mice and the small intestine and colon were, respectively, longer and shorter compared to control mice. These results indicate modified glycoconjugate metabolism in cystic fibrosis knockout mice and morphologic changes to the spleen and intestine where the latter modifications are possibly related to the intestinal malabsorption associated with cystic fibrosis.