Aqueous outflow pathway complex carbohydrate synthesis in vitro.

A technique is described for examining the in vitro synthesis of glycosaminoglycans and glycoproteins in the aqueous outflow pathway (AOP). New Zealand red rabbit eyes were maintained at near-physiologic conditions and were infused by anterior chamber exchange with 30 muCi [3H] glucosamine and 100 muCi [35S] sulfate. At 0.5, 1.0, and 2.0 hours, the anterior segment tissues were dissected, isolated, and fractionated by gel filtration chromatography on Sephadex G-50 columns. The elution profiles demonstrated an increase of labeled material with time in the glycosaminoglycan (GAG) fractions and in the glycopeptide fractions. The relative rate of synthesis of labeled material was: central cornea greater than peripheral cornea greater than iris-ciliary body greater than AOP greater than anterior sclera. In order to characterize the profiles of each tissue, the isolated material was analyzed for hexosamine, hexuronic acid, and sulfate. The AOP cells synthesized a heterogenous mixture of GAGs and glycoproteins which biochemically appeared to be distinct from other anterior segment tissues. In addition, the percent distribution of GAG polymers in each tissue was determined by selective GAG degradative procedures and by gel filtration chromatography. The AOP cells synthesized four types of GAGs, and the percent distribution of the labeled GAGs was different from the other tissues. The present results suggest that this technique may provide a well-controlled method to probe the metabolic activity of complex carbohydrates in the AOP and in the anterior segment.

Age-related changes in the cerebrospinal fluid outflow glycosaminoglycans.

The glycosaminoglycan distribution patterns of the cerebrospinal fluid (CSF) outflow pathway, dura mater and cerebral cortex of young New Zealand red rabbits and 1-, 3- and 12-week-old C-57 mice were identified by analyses of the glycosaminoglycan moieties and by the use of zone electrophoresis. The glycosaminoglycans were identified by specific degradation procedures, i.e., hyaluronate lyase, chondroitin ABC lyase, endo-beta-D-galactosidase and nitrous acid treatment. The CSF outflow pathway and dura mater glycosaminoglycan components were primarily hyaluronic acid and chondroitin sulfate-dermatan sulfate, whereas the cerebral cortex glycosaminoglycan components were hyaluronic acid, chondroitin sulfate-dermatan sulfate, keratan sulfate and heparan sulfate. The glycosaminoglycan components of the dura mater and cerebral cortex decreased and those of the CSF outflow pathway increased as a function of age. These results demonstrate the feasibility of analyses of the CSF outflow pathway glycosaminoglycan components and suggest that topographical changes in the glycosaminoglycan distribution profiles may contribute to the pattern of cerebrospinal fluid outflow.

Keratan sulfate-like glycosaminoglycan in the cerebral cortex of the brain and its variation with age.

A keratan sulfate-like substance was found kn the cerebral cortex proteoglycans of rats. This substance tripled during the rapid growth phase from 1 to 3 months of age and then decreased steadily to a negligible quantity in the senescent rat, aged 25 months. In contrast, the uronic acid-containing glycosaminoglycans remained constant during the life span studied. The marked decrease observed in the proportion of keratan sulfate-like moiety to the other glycosaminoglycans in the cerebral cortex of the senescent rat (aged 25 months) compared with the young adult rat (aged 3 months) may explain the reduced extracellular volume observed by others in the senescent brain.

An anti-A1 lectin in the seeds of Amphicarpaea bracteata.

An anti-A1 lectin has been isolated from the extract of Amphicarpaea bracteata seeds by affinity chromatography on Epoxy-activated Sepharose 6B coupled to N-acetyl-D-galactosamine. The yield of the purified lectin was 86 microgram/g of seeds. The purified lectin shows one main band on electrophoresis in sodium dodecyl sulfate-polyacrylamide. The amino acid and neutral sugar composition indicate that this lectin is an acidic glycoprotein with a neutral sugar content of approx. 2%. The composition of the lectin is different from that of the Dolichos biflorus lectin but the two lectins have some common characteristics. The most powerful inhibitors of the agglutination of A1 red blood cells by the A. bracteata lectin is N-acetyl-D-galactosamine. Much weaker inhibitors of the agglutination are alpha-lactose, D-fucose, and five other sugars.