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Inhibition of calcium phosphate mineral growth by proteoglycan aggregate fractions in a synthetic lymph

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Abstract

The effect of proteoglycans on growth of seeding minerals in synthetic lymph was studied with special reference to regulation of endochondral calcification. Proteoglycans were isolated from bovine nasal cartilage by three published methods. By each method two fractions were separated which differed in respect to presence or absence of fast-sedimenting components on analytical ultracentrifugation. Each fraction was tested for its capacity to inhibit mineral growth in a buffered synthetic lymphin vitro. At concentrations of proteoglycans estimated to occur in the interstitial fluid of endochondral plates from 6- to 7-week-old rats, the fractions containing fast-sedimenting components were inhibitory to mineral growth; whereas fractions containing the slow-sedimenting components and a glycoprotein (link protein) had no inhibitory activity demonstrable in these systems. Comparison of calcium-binding capacity of certain proteoglycan fractions as well as their computed effect upon reduction of calcium activity under conditions of equilibrium dialysis revealed no differences in the behavior of a proteoglycan fraction rich in fast—as opposed to fractions composed entirely of slow-sedimenting components. An increased degree of shielding of mineral embryos provided by adjacent protein cores of aggregated proteoglycans is hypothesized to explain the inhibitory action of fast-sedimenting proteoglycans on mineral growthin vitro.

Résumé

L'effet des protéoglycanes sur la croissance de minéraux d'ensemencement dans un milieu synthétique est étudié sous l'angle de la régulation de l'ossification enchondrale. Les protéoglycanes sont isolés à partir ducartilage nasal bovin à l'aide de trois méthodes publiées. A l'aide de chacune de ces méthodes, deux fractions sont isolées qui se distinguent par la présence ou l'absence de composés qui se sédimentent rapidement par ultracentrifugation analytique. Chaque fraction est étudiée en fonction de sa possibilité d'inhiber la croissance minérale dans un milieu tamponné synthétiquein vitro. A des concentrations de protéoglycanes qui se retrouvent dans le liquide interstitielle de la métaphyse de rats de 6 à 7 semaines, les fractions contenant des composés qui se sédimentent rapidement, inhibent la croissance minérale; alors que les fractions contenant des composés, qui sédimentent lentement, ainsi qu'une glycoprotéine (protéine de liaison) n'ont pas d'activité d'inhibition dans ces systèmes.

La comparaison de la capacité de fixation du calcium de certaines fonctions de protéoglycanes ainsi que leur effet sur la diminution de l'activité calcique dans des conditions de dialyse équilibrées ne montrent aucune différence sur le comportement des fractions de protéoglycanes comportant des produits sédimentant rapidement ou lentement. Un degré plus élevé de protection des minéraux naissants, fournie par les portions protéiques adjacentes de protéoglycanes agrégés, pourrait être responsable de l'action d'inhibition de croissance minéralein vitro de protéoglycanes sédimentant rapidement.

Zusammenfassung

Die Wirkung von Proteoglykanen auf das Wachstum von Impfkristallen in synthetischer Lymphe wurde, mit besonderer Berücksichtigung der Regulation von endochondraler Verkalkung, studiert. Die Proteoglykane wurden nach drei publizierten Methoden aus dem Nasenknorpel des Rindes isoliert. Bei jeder Methode wurden zwei Fraktionen abgetrennt, welche sich bei der analytischen Ultrazentrifugation in bezug auf An- oder Abwesenheit von schnellsedimentierenden Komponenten unterschieden. Jede Fraktion wurde darauf geprüft, ob siein vitro das Mineralwachstum in einer gepufferten synthetischen Lymphe zu hemmen vermochte. Bei Proteoglykan-Konzentrationen, wie sie in der interstitiellen Flüssigkeit endochondraler Platten von 6 bis 7 Wochen alten Ratten vermutet werden, hatten diejenigen Fraktionen, welche schnell-sedimentierende Komponenten enthielten, eine Hemmwirkung auf das Mineralwachstum; Fraktionen mit langsam-sedimentierenden Komponenten und mit einem Glycoprotein („link protein”) hingegen zeigten in diesen Systemen keine Hemmwirkung.

Der Vergleich der Calcium-bindenden Fähigkeit bestimmter Proteoglykan-Fraktionen sowie deren vereinte Wirkung auf die Herabsetzung der Calcium-Aktivität unter Bedingungen der Gleichgewichtsdialyse zeigte keine Unterschiede im Verhalten von Proteoglykan-Fraktionen, die reich an schnell-sedimentierenden Komponenten waren im Gegensatz zu Fraktionen, welche ausschließlich langsam-sedimentierende Komponenten enthielten. Die Hemmwirkung von schnell-sedimentierenden Proteoglykanen auf das Mineralwachstumin vitro wird mit folgender Hypothese erklärt: Die Mineralkeime werden in zunehmendem Maße durch angrenzende Proteinkerne angehäufter Proteoglykane geschützt.

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Authors and Affiliations

  1. Arthritis Division, Department of Medicine, Veterans Administration Hospital, Miami, Florida

    L. A. Cuervo, J. C. Pita & D. S. Howell M. D.

  2. Arthritis Division, Department of Medicine, University of Miami School of Medicine, Biscayne Annex, P.O. Box 875, 33152, Miami, Florida, USA

    L. A. Cuervo, J. C. Pita & D. S. Howell M. D.

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  3. D. S. Howell M. D.
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Cuervo, L.A., Pita, J.C. & Howell, D.S. Inhibition of calcium phosphate mineral growth by proteoglycan aggregate fractions in a synthetic lymph. Calc. Tis Res. 13, 1–10 (1973). https://doi.org/10.1007/BF02015390

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