Glycosaminoglycans from the rooster comb and wattle.

Glycosaminoglycans (GAG) were isolated from the rooster comb and wattle by digestion with papain and analyzed by cellulose acetate electrophoresis and enzymatic digestion. The results indicated that the concentration of GAG uronic acid is approximately two-fold greater (P less than .01) in the comb than in the wattle. In both tissues, hyaluronic acid was the major GAG, with a small proportion of dermatan sulfate.

Feedback control of selected biosynthetic activities of chondrocytes in culture.

The syntheses of proteoglycans and proteins by chondrocytes from the Swarm rat chondrosarcoma in primary cultures were modulated on the addition of matrical molecules. In the presence of hyaluronan, collagen or proteoglycan aggregates the synthesis of proteoglycans was depressed. The synthesis of collagen was also depressed in the presence of hyaluronan or collagen. In the presence of proteoglycan monomers, the incorporation of 35S-sulfate was enhanced in proportion to the concentration of the additive in the medium; the synthesis of protein was unaffected. The proteoglycan monomers synthesized in the presence of proteoglycan monomers were larger than those synthesized in their absence. In combinations, the exogenous macromolecules did not affect the selected biosynthetic activities to an extent greater than that which they exerted separately. The data suggest, however, that the proteoglycan monomers can counteract the inhibitory effects of the macromolecules which are inhibitory. The data, moreover, suggest that the chondrocytes of the Swarm rat chondrosarcoma have the potential to discriminate between proteoglycan monomers produced by self and those produced by chondrocytes of hyaline cartilages.

A new chromatographic method for the fractionation of hyaluronic acid.

Sodium hyaluronate from rooster comb, umbilical cord, bovine vitreous and a special commercial hyaluronic acid preparation, "HealonR" have been fractionated at 4 degrees C on DEAE-Sephacel columns using salt gradients from 0 leads to 0.4M NaCl. All samples examined displayed a high degree of polydispersity. The number of well separated fractions observed in these chromatograms ranges from 10-30 depending on the source of the material. A molecular weight distribution effect does not seem to be the main factor controlling this fractionation as no differences could be observed when the limiting viscosity numbers (eta) for numerous fractions isolated from the same sample were compared with each other. As no galactosamine, sulfate or phosphate is found directly linked to these fractions, it is suggested that disaggregation of an hyaluronate molecular aggregate could be the basis for this fractionation.

Depolymerization of glycosaminoglycuronans into di- and higher molecular-weight oligo-saccharides: improved preparation of N-acetyldermosine and oligomeric N-acetylchondrosines.

Nonsulfated di- to octadeca-saccharides having 2-acetamido-2-deoxy-D-galactose at the reducing end were prepared, in 81% yield, by treatment of chondroitin 6-sulfate (pyridinium salt) with dimethyl sulfoxide containing 10% of water for 14 h at 90 degrees. N-Acetylchondrosine and N-acetyldermosine were obtained from dermatan sulfate of rooster comb, in 30% and 38% yields, respectively, by solvolysis with dimethyl sulfoxide, containing 10% of water, for 30 h at 105 degrees. Hyaluronic acid was also depolymerized by the same solvent in the presence of an equimolar amount of pyridinium sulfate or chloride per disaccharide unit to give reducing di- and higher molecular weight oligo-saccharides. The results of solvolytic desulfation and depolymerization are compared with those of the conventional methods by acid hydrolysis.

Sequential hydrolysis of hyaluronate by beta-glucuronidase and beta-N-acetylhexosaminidase.

1. Hyaluronate extracted from rooster comb was digested by a mixture of beta-N-acetylhexosaminidase and beta-glucuronidase with simultaneous dialysis for 96 h. 2. The produjct, yielding 99.6% of a mixture of mono- and oligo-saccharides, was purified by gel chromatography and analysed for glucuronic acid, N-acetylglucosamine and other sugars. 3. The oligosaccharide portion was chromatographed on DEAE-cellulose, and the effluent fractions were analysed for glucuronic acid and N-acetylglucosamine, reduced with NaBH4, digested by beta-N-acetylhexosaminidase and subjected to acid hydrolysis and glucosamine determination. 4. GlcNAc-GlcA-GlcNAc, GlcA-GlcA-GlcNAc and GlcA-GlcA-GlcA-GlcNAc were the oligosaccharides obtained, which resulted from the transferase activity of the enzymes and represented 57% of the digestion products. The results demonstrate that this hyaluronate is an unbranched polymer of approximatey equal amounts of glucuronic acid and N-acetylglucosamine. The data also indicate that if this glycosaminoglycan contains any of the neutral sugars for which it was analysed, their concentration must be less than 0.020% of the sum of the known components.

Effect of diflubenzuron (Dimilin; TH--6040) on the hyaluronic acid concentration in chicken combs.

The addition of N-[[(4-chlorphenyl)amino]carbonyl]-2,6-difluorobenzamide(diflubenzuron; Dimilin; TH-6040) up to levels of 250 ppm of the total diet of both male broilers and layers for 98 days did not affect the hyaluronic acid (HA) concentration (microgram/g tissue) in the combs. The concentration was measured at 21, 28, 42, 56, and 98 days on feed. The concentration in the combs of the layers were not significantly different at any sampling period regardless of diet. At the end of 56 days on feed, the combs of the broiler controls had a significantly (P less than .025) higher concentration than did any of the groups fed diflubenzuron. There were no differences observed at 21, 28, 42, or 98 days in the broilers. The HA concentration increased as the chickens matured and became larger; however, large variations were observed within the various groups at a given sample period.

Further evidence for the presence of androgen receptors in the lungs and in the head appendages of the cock.

The presence of cytoplasmic dihydrotestosterone receptors in the lungs, the comb, the wattle, and the ear lobes of the cock was demonstrated by sucrose density-gradient centrifugation. All these tissues exhibited saturable 'in vitro' binding of dihydrotestosterone in the 8-11S region of the gradient. When 0.5 M KCl was added to the lung, wattle, and comb cytosol and to the gradients, the radioactive dihydrotestosterone migrated in the 4-5S region. These studies suggest that the mechanism of action of androgens in the head appendages of the cock and in other target tissues is similar.