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Studies on glycosaminoglycans (GAG) in mammalian embryonic tissue

I. A micromethod for the fractionation and characterisation of GAG according to their anionic behaviour

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Summary

  1. 1.

    A micromethod is presented which allows a separation of GAG according to their anionic behaviour on ECTEOLA-cellulose columns in routine studies. Three fractions with increasing anionic behaviour (F I, FII, FIII) have been chosen to characterise GAG. After labelling GAG in vivo the uronic acid content (carbazol reaction) and35S incorporation have been measured in each fraction.

  2. 2.

    The uronic acid content and35S incorporation have been studied at different stages of embryonic development in the rat. The total amount of GAG per dry weight is found to increase with embryonic age. This is accompanied by an increase in the percentage of high-anionic GAG (F II and F III).

  3. 3.

    Several experimental conditions have been found to effect the kinetics of35S within the maternal organism, thus leading to a difference in the availability of35S in embryonic GAG. So total35S incorporation rates into GAG often do not represent the rates of biosynthesis of GAG.

  4. 4.

    After labelling GAG with35S in vivo, the35S-pattern of the three fractions obtained from Ecteola-cellulose columns is characteristic of a given tissue and furthermore may be used as a measure for changes occuring within the GAG under different experimental conditions.

  5. 5.

    Since in pharmacological studies the rate of incorporation of35S into GAG may not directly reflect GAG metabolism, the distribution of uronic acids as well as35S radioactivity within the three GAG fractions is suggested as a measure for changes occuring in GAG metabolism. This takes into account that under pharmacological or pathological conditions a change of the anionic behaviour of GAG may take place. The ratio of35S incorporation into two fractions\(\left( {R{}^{35}Si\frac{{III}}{{II}}or\frac{{III}}{I}} \right)\) is taken as an indication of a change in the GAG pattern. A number of parameters influencing these ratios are discussed.

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References

  • Anseth, A., Laurent, T. C.: Studies on corneal polysaccharides. I. Separation. Exp. Eye Res.1, 25 (1961).

    Google Scholar 

  • Antonopoulos, C. A., Fransson, L.-A., Heinegard, D., Gardell, S.: Chromatography of glyoosaminoglycans on Eoteola-cellulose columns. Biochim. biophys. Acta (Amst.)148, 158 (1967).

    Google Scholar 

  • —, Gardell, S., Szirmai, J. A., De Tyssonsk, E. R.: Determination of glycosaminoglyoans (mucopolysaccharides) from tissues on the microgram scale. Biochim. biophys. Acta (Amst.)83, 1 (1964).

    Google Scholar 

  • Bathgate, G. N.: An automated gel filtration system for determining the molecular weight of polysacoharides. J. Chromatogr.47, 92 (1970).

    Google Scholar 

  • Bitter, T., Muir, H. M.: A modified uronic acid carbazole reaction. Analyt. Biochem.4, 330 (1962).

    Google Scholar 

  • Boström, H., Bernstein, K., Whitehouse, M. W.: Biochemical properties of antiinflammatory drugs. II. Some effects on sulphate-35S metabolism in vivo. Biochem. Pharmacol.13, 413 (1964).

    Google Scholar 

  • Burton, K.: A study of the conditions and mechanisms of the diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem. J.62, 315 (1956).

    Google Scholar 

  • Cleland, R. L., Cleland, M. C., Lipsky, J. J., Lyn, V. E.: Ionic polysaccharides. I. Adsorption and fractionation of polyelectrolytes on (diethylamino) ethyl cellulose. J. Amer. chem. Soc.90, 3141 (1968).

    Google Scholar 

  • Constantopoulos, G., Dekaban, A. S., Carroll, W. R.: Determination of molecular weight distribution of acid mucopolysaccharides by sephadex gel filtration. Analyt. Biochem.31, 59 (1969).

    Google Scholar 

  • Davies, M., Mchol, L. W., Ogstron, A. G.: Frictional effects in the migration of mixtures of hyaluronic acid and serum albumin. Biochim. biophys. Acta (Amst.)75, 436 (1963).

    Google Scholar 

  • Gibian, H.: Mucopolysaccharide und Mucopolysaccharidasen. Wien: Franz Deuticke 1959.

    Google Scholar 

  • Greiling, H., Herbertz, Th., Stuhlsatz, H. W.: Chromatographische Untersuchungen über die Mucopolysaccharidverteilung im hyalinen Knorpel. Hoppe-Seylers Z. physiol. Chem.336, 149 (1964).

    Google Scholar 

  • —: Struktur und Stoffwechsel von Glykosaminoglykan-Proteinen. Die Keratansulfat-Peptide der Rinder-Cornea. Hoppe-Seylers Z. physiol. Chem.345, 236 (1966).

    Google Scholar 

  • — —: Glykosaminoglykan-Peptide aus dem humanen Kniegelenkknorpel. Hoppe-Seylers physiol. Chem.350, 449 (1969).

    Google Scholar 

  • Kleine, O. T., Hilz, H.: Metabolically different proteoglycans in calf rip cartilage. In: Chemistry and molecular biology of the intercellular matrix, vol. 2, pp. 907 to 919. Ed. E. A. Balazs. London-New York: Academic Press 1970.

    Google Scholar 

  • Köhler, E., Merker, H.-J., Ehmke, W., Wojnorowiez, F.: Growth kinetics os mammalian embryos during the phase of organogenesis. Naunyn-Schmiedeberg's Arch. Pharmac. (in press) 1972.

  • Kresse, H., Buddecke, E.: Stoffwechselheterogenität von Chondroitin-4-suIfat-Proteinen und Kollagenstoffwechsel in Rindernasenknorpel. Hoppe-Seylers Z. physiol. Chem.349, 1497 (1968).

    Google Scholar 

  • Kühn, K., Grassmann, W., Hofmann, U.: Über die Bildung der Kollagenflbrillen aus gelöstem Kollagen und die Funktion der kohlenhydrathaltigen Begleitkomponenten. Z. Naturforsch.14b, 436 (1959).

    Google Scholar 

  • Laurent, T. C.: Studies on fractionated heparin. Arch. Biochem.92, 224 (1961).

    Google Scholar 

  • —, Scott, J. E.: Molecular weight fractionation of polyanions by cytylpyridinium Chloride in salt solutions. Nature (Lond.)202, 661 (1964).

    Google Scholar 

  • Lindahl, U.: Comment on the use of cetylpyridinium chloride in the isolation of connective-tissue proteoglycan. J. Biochem.113, 569 (1969).

    Google Scholar 

  • Lowther, D. A., Toole, B. P., Herrington, A. C.: Interaction of proteoglycans with tropocollagen. Chemistry and molecular biology of the intercellular matrix, p. 1135. London-New York: Academic Press 1970.

    Google Scholar 

  • Martin, C. J., Bhatnagar, G. M.: Unfolding reactions of proteins. II. Spectral and optical rotatory dispersion studies in urea, guanidinium chloride and 2-chloroethanol. Biochemistry6, 1638 (1967).

    Google Scholar 

  • Mathews, M. B.: The macromolecular organization of connective tissue. In: Chemical physiology of mucopolysaccharides. Boston: Little, Brown & Co. 1968.

    Google Scholar 

  • —: The interactions of proteoglycans and collagen-model systems. Chemistry and molecular biology of the intercellular matrix, p. 1155. London-New York: Academic Press 1970.

    Google Scholar 

  • —, Decker, L.: The distribution of sulfate groups in chondroitin sulfate chains. Detection of “oversulfated” and “undersulfated” repeating periods. Biochim. biophys. Acta (Amst.)156, 419 (1968).

    Google Scholar 

  • Öbrink, B.: Studies on the electrostatic interaction between collagen and chondroitin 4-sulfate. Chemistry and molecular biology of the intercellular matrix, p. 1171. London-New York: Academic Press 1970.

    Google Scholar 

  • Partridge, S. M.: The chondroitin sulfate-protein complex from bovine cartilage. In: Chemical physiology of mucopolysaccharides. Boston: Little, Brown & Co. 1968.

    Google Scholar 

  • Pearce, R. H., Mathieson, J. M., Grimmer, B. J.: Fractionation of anionic glycosaminoglycans by ion-exchange chromatography. Analyt. Biochem.24, 141 (1968).

    Google Scholar 

  • Ringertz, N. R., Reichard, P.: Chromatography on Ecteola of sulphate containing mucopolysaccharides and nucleotides. Acta chem. scand.13, 1467 (1959).

    Google Scholar 

  • — —: Chromatography on Ecteola of sulfate containing mucopolysaccharides. Acta chem. scand.14, 303 (1960).

    Google Scholar 

  • Robertson, D. W.: The effect of sodium tolbutamide on the uptake of sodium-35S-sulfate and35S-methionine in inbred strains of mice. Teratology1, 387 (1968).

    Google Scholar 

  • Schimmelpfennig, K.: Problems connected with the in vivo labelling of embryonic glycosaminoglycans with Naa 35SO4 in teratological studies. Naunyn-Schmiedebergs Arch. Pharmak.271, 320 (1971).

    Google Scholar 

  • - Baumann, I., Kaufmann, C.: Studies on glycosaminoglycans (GAG) in mammalian embryonic tissue. II. Influence of vitamin A and Na-salicylate on embryonic GAG. Naunyn-Schmiedeberg's Arch. Pharmac. (in press) 1972.

  • - Schweichel, J.-U.: A hypothesis on the teratogenic mode of action of A-hypervitaminosis. Teratology (in press).

  • Scott, J. E.: Aliphatic ammonium salts in the assay of acidic polysaccharides from tissues. Meth. biochem. Anal.8, 145 (1960).

    Google Scholar 

  • Stefanovich, V., Gore, I.: A micromethod for the determination of acid mucopolysaccharides in vascular tissue. J. Chromatogr.31, 473 (1967).

    Google Scholar 

  • Svejcar, J.: The content of glycosaminoglycans (mucopolysaccharides) in mouse fetus and placenta. Biochim. biophys. Acta (Amst.)165, 84 (1968).

    Google Scholar 

  • —, Robertson, W. B. van: Micro separation and determination of mammalian acidic glycosaminoglycans (mucopolysaccharides). Analyt. Biochem.18, 333 (1967).

    Google Scholar 

  • Tomlinson, R. V., Tener, G. M.: The use of urea to eliminate the secondary binding forces in ion exchange chromatography of polynucleotides. J. Amer. chem. Soc.84, 2644 (1962).

    Google Scholar 

  • Trundle, J. S., Mann, G. U.: Characterization and measurement of acid mucopolysaccharides in tissues. Biochim. biophys. Acta (Amst.)101, 127 (1965).

    Google Scholar 

  • Wasteson, A.: A method for the determination of molecular weight dispersion in chondroitin sulphate on a microgram level. Biochim. biophys. Acta (Amst.)177, 152 (1969).

    Google Scholar 

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Authors and Affiliations

  1. Abteilung “Embryonal-Pharmakologie”, Pharmakologisches Institut der Freien Universität Berlin, Germany

    K. Schimmelpfennig, I. Baumann & C. Kaufmann

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  1. K. Schimmelpfennig
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  2. I. Baumann
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  3. C. Kaufmann
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Additional information

This work was generously supported by grants from the Deutsche Forschungsgemeinschaft given to the Sonderforschungsbereich 29, “Embryonale Entwicklung und Differenzierung (Embryonal-Pharmakologie)”.

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Schimmelpfennig, K., Baumann, I. & Kaufmann, C. Studies on glycosaminoglycans (GAG) in mammalian embryonic tissue. Naunyn-Schmiedebergs Arch. Pharmak. 271, 430–456 (1971). https://doi.org/10.1007/BF00997129

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