A high-performance liquid chromatography approach for isolation and sequencing of chondroitin sulfate oligosaccharides

Delaney, Stephen R.; Conrad, H. Edward; Glaser, J H

doi:10.1016/0003-2697(80)90689-2

Cited by 55 publications

(17 citation statements)

References 17 publications

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“…The percentage that is ACII-resistant reflects iduronic acid content. Radiolabeled disaccharides were analyzed by HPLC using a SAX column exactly as described (20).…”

Section: Age-related Changes In Proteoglycan Structurementioning

confidence: 99%

Age-related Changes in Human Bone Proteoglycan Structure

Grzesik

Frazier

Shapiro

et al. 2002

Journal of Biological Chemistry

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Proteoglycans (PGs) are a family of molecules that undergo extensive post-translational modifications that include addition of glycosaminoglycan (GAG) chains as well as N-and O-linked oligosaccharides to the protein core. PG composition and structure have been reported to alter with age. To test whether the post-translational modifications to PGs can serve as in vitro surrogate end point markers for chronological age, the extent of GAG modifications was determined for PGs derived from normal human bone cells of 14 donors (age range, fetal to 60 years). Isolated cells were steady state radiolabeled with 35 SO 4 2؊ and [ 3 H]GlcN. For biglycan and decorin, iduronate content was linearly correlated with age (increased 1.5؋ between fetal and age 60 years). For the syndecan-like heparan sulfate PG, the N-sulfation of post-natal cells increased over 3.5-fold until reaching a plateau during the 4th decade of life. The amount of O-linked oligosaccharides was also found to decrease as a function of increasing normal donor age, whereas the specific activity of the metabolic precursor pool remained constant regardless of donor age. These age-related changes in post-translational modifications were then used to demonstrate that osteoblasts derived from patients with osteogenesis imperfecta did not exhibit facets of a pre-mature aging, but rather were arrested in a fetallike phenotypic state. A growth matrix rich in thrombospondin altered PG metabolism in osteoblastic cells, resulting in the production and secretion of the fetal-like (rich in O-linked oligosaccharides) forms of decorin and biglycan. This effect was qualitatively different from the effect of transforming growth factor-␤, which predominantly altered GAGs rather than O-linked oligosaccharides. No other Arg-Gly-Asp protein (fibronectin, vitronectin, type I collagen, osteopontin, and bone sialoprotein) showed any detectable effect on PG metabolism in bone cells. These results indicate that a proper matrix stoichiometry is critical for metabolism of PGs.

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“…The percentage that is ACII-resistant reflects iduronic acid content. Radiolabeled disaccharides were analyzed by HPLC using a SAX column exactly as described (20).…”

Section: Age-related Changes In Proteoglycan Structurementioning

confidence: 99%

Age-related Changes in Human Bone Proteoglycan Structure

Grzesik

Frazier

Shapiro

et al. 2002

Journal of Biological Chemistry

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“…After 2 hr at 37"C, both portions were heated at 100°C for 10 min. Unsaturated disaccharide products were then separated and quantitated by a modified HPLC method of Delaney et a1 [25]. A Beckman Model llOA liquid chromatograph equipped with a model 2 10 injector ( 2 0 4 loop) and variable wavelength detector was used.…”

Section: Analysis Of Glycosaminoglycans By High-pressure Liquid Chrommentioning

confidence: 99%

Stimulation of glycosaminoglycan production in murine tumors

Knudson

Biswas

Toole

1984

J of Cellular Biochemistry

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Three types of murine tumors, B-16 melanoma, A-10 carcinoma, and S-180 sarcoma, were shown to contain elevated glycosaminoglycan (GAG) concentrations in vivo as compared to normal muscle or subcutaneous tissue. Hyaluronate was especially concentrated in the A-10 carcinoma, which contained approximately six times more hyaluronate than subcutaneous tissue and 18 times more than muscle. In all three tumors, chondroitin sulfates, especially chondroitin-4-sulfate, were present in higher concentrations than in the normal tissues. In culture, however, all three tumor cell lines produced less than 5% as much GAG as mouse fibroblasts, when measured by incorporation of [3H] acetate or by chemical analysis. Varying the culture passage number or the medium composition, ie, glucose, serum, and insulin concentrations, had little effect on GAG synthesis by the tumor cells. The low GAG levels in the tumor cell cultures were not due to hyaluronidase activity in their media. In an attempt to mimic possible host-tumor cell interactions that could account for the elevated GAG levels in vivo, tumor cells were cocultured with fibroblasts, but no stimulation above the amount made by the tumor cells alone plus that by the fibroblasts alone was observed. Conditioned media from the tumor cells, either dialyzed or not against fresh complete medium, had no effect on fibroblast GAG synthesis. Tumor extracts, however, were found to stimulate synthesis of hyaluronate by fibroblasts. Stimulation by extracts of A-10 carcinoma was greater than and additive to that of serum. The above results strongly suggest that GAG production in these tumors is in part regulated by host-tumor interactions.

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“…1 paper in glacial acetic acid-butanol-1 M NH40H (3:2:1, vol/vol) (15,20). The relative amounts of different disaccharides were quantitated by high-pressure liquid chromatography (2). Breakdown of proteoglycan by intact cells.…”

Section: Methodsmentioning

confidence: 99%

Digestion of proteoglycan by Bacteroides thetaiotaomicron

Kuritza¹,

Salyers²

1983

J Bacteriol

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It has been shown previously that Bacteroides thetaiotaomicron, a human colonic anaerobe, can utilize the tissue mucopolysaccharide chondroitin sulfate as a source of carbon and energy and that the enzymes involved in this utilization are all cell associated (A. A. Salyers and M. B. O'Brien, J. Bacteriol. 143:772-780, 1980). Since chondroitin sulfate does not generally occur in isolated form in tissue, but rather is bound covalently in proteoglycan, we investigated the extent to which chondroitin sulfate which is bound in such a sterically hindered complex can be utilized by intact bacteria. Intact cells of B. thetaiotaomicron were able to digest chondroitin sulfate in proteoglycan, although at a slightly slower rate than free chondroitin sulfate. Prior digestion of proteoglycan with trypsin to produce small fragments of protein with several chondroitin sulfate chains attached did not increase the rate at which the bound chondroitin sulfate was digested. Accordingly, the slower rate of digestion was probably due to attachment of chondroitin sulfate chains to the protein backbone rather than to steric hindrance by other components of the proteoglycan. When proteoglycan which had been incubated with intact bacteria was treated with sodium borohydride to release the undigested fragments of chondroitin sulfate from the protein backbone, the size and composition of the fragments indicated that intact bacteria were able to digest all but three monosaccharides of the chondroitin sulfate chains. Thus, despite steric hindrance due to attachment of the chondroitin sulfate chains to the protein backbone, digestion of bound chondroitin sulfate by intact bacteria was nearly complete.

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A high-performance liquid chromatography approach for isolation and sequencing of chondroitin sulfate oligosaccharides

Cited by 55 publications

References 17 publications

Age-related Changes in Human Bone Proteoglycan Structure

Age-related Changes in Human Bone Proteoglycan Structure

Stimulation of glycosaminoglycan production in murine tumors

Digestion of proteoglycan by Bacteroides thetaiotaomicron

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