Calcified Tissue Research

, Volume 11, Issue 1, pp 56–69 | Cite as

Studies of the metabolism of separated bone cells

I. Techniques of separation and identification
  • D. M. Smith
  • C. C. JohnstonJr.
  • A. R. Severson
Original Papers


The mechanical separation of rat skeletal tissue into viable populations of periosteal cells, osteoblasts, osteocytes, and marrow cells has been undertaken. Periosteal cells and osteocytes were obtained as tissue preparations, while osteoblasts and marrow cells were isolated as individual cells. The cell populations were identified during the preparative procedure by their histological and histochemical appearance. Viability of these preparations was demonstrated by their metabolic activity during short-term incubations and their histological appearance and vital staining before and after such incubations. The cell populations produced carbon dioxide and lactate, consumed oxygen, and incorporated uridine into RNA. Comparison of uridine incorporation into RNA of cells from control and parathyroid hormone treated rats was compatible with a differential effect of parathyroid hormone on the cell populations. These findings indicate that the technique described can be utilized to produce viable bone cells of different types, and may offer a new means for characterizing the role of each cell type in the metabolism of skeletal tissue.

Key words

Bone Osteoblasts Osteocytes Periosteum Marrow Cells 


La séparation mécanique du tissu squelettique du rat en cellules périostées, en ostéoblastes, en ostéocytes et en cellules de la moelle a été réalisée. Les cellules périostées et les ostéocytes ont été obtenues sous forme de préparations tissulaires, alors que les cellules ostéoblastiques et celles de la moelle ont été isolées sous forme de cellules individuelles. Les populations cellulaires sont identifiées par leurs caractères histologiques et histochimiques. La vitalité de ces préparations est věrifiée par leur activité métabolique pendant des incubations de courte durée et des examens. histologiques et des colorations vitales, réalisés avant et après incubation. Les cellules produisent du gaz carbonique et du lactate, consomment de l’oxygène et incorporent l’uridine en RNA. La comparaison de l’incorporation d’uridine en RNA de cellules de rats témoins et de rats traités par hormone parathyroïdienne concorde avec une action différentielle de la parathormone sur les divers types de cellules. Il semble que la technique décrite permette de produire des cellules osseuses vivantes de divers types et fournit ainsi le moyen d’élucider le rôle métabolique de chaque type cellulaire du squelette.


Die mechanische Trennung von Rattenskelett-Gewebe in lebensfähige Populationen von Periost-Zellen, Osteoblasten, Osteozyten und Markzellen ist unternommen worden. Die Periostzellen und Osteozyten wurden als Gewebepräparate erhalten, während die Osteoblasten und Markzellen als individuelle Zellen isolier wurden. Die Zell-Populationen wurden während des Herstellungsverfahrens histologisch und histochemisch identifiziert. Die Lebensfähigkeit dieser Präparate wurde festgestellt durch ihre metabolishe Aktivität während Kurzzeit-Inkubationen und durch ihr histologisches Aussehen sowie durch Vitalfärbung vor und nach solchen Inkubationen. Die Zellpopulationen produzierten Kohlendioxyd und Lactat, verbrauchten Sauerstoff und bauten Uridin in RNS ein. Der Vergleich zwischen Uridineinbau in RNS von Zellen aus Kontrolltieren und aus mit Parathormon behandelten Ratten war mit der Differentialwirkung von Parathormon auf die Zellpopulationen vereinbar. Diese Befunde deuten darauf hin, daß die beschriebene Technik benützt werden kann, um verschiedene Typen von lebensfähigen Knochenzellen zu produzieren und daß sie einen neuen Weg öffnen kann, um die Rolle jedes Zelltyps im Stoffwechsel des Skelettgewebes zu charakterisieren.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Barker, S. B., Summerson, W. N.: The colorimetric determination of lactic acid in biological material. J. biol. Chem.138, 535–554 (1941).Google Scholar
  2. Bingham, P. J., Brazell, I. A., Owen, M.: The effect of parathyroid extract on on cellular activity and plasma calcium levelsin vivo. J. Endocr.45, 387–400 (1969).PubMedCrossRefGoogle Scholar
  3. Borle, A. B., Nichols, N., Nichols, G., Jr.: Metabolic studies of bonein vitro. I. Normal bone. J. biol. Chem.235, 1206–1210 (1960).PubMedGoogle Scholar
  4. Burstone, M. S.: Histochemical comparison of naphthol-AS-phosphates for the demonstration of phosphatases. J. nat. Cancer Inst.20, 601–615 (1958).PubMedGoogle Scholar
  5. Cohn, D. V., Forscher, B. K.; Aerobic metabolism of gluycose by bone. J. biol. Chem.237, 615–618 (1962).PubMedGoogle Scholar
  6. Deiss, W. P., Jr., Holmes, L. B., Johnston, C. C., Jr.: Bone matrix biosynthesisin vitro. I. Labeling of hexosamine and collagen of normal bone. J. biol. Chem.237, 3555–3559 (1962).PubMedGoogle Scholar
  7. Field, J.; Respiration of tissue slices. Meth. med. Res.1, 289–307 (1948).PubMedGoogle Scholar
  8. Fitch, S. M., Harkness, M. L. R., Harkness, R. D.: Extraction of collagen from tissues. Nature (Lond.)176, 163 (1955).CrossRefGoogle Scholar
  9. Flanagan, B., Nichols, G., Jr.: The metabolism of cells isolated from bone. Fed. Proc.22, 553 (1963).Google Scholar
  10. Fullmer, H. M., Link, C. C., Jr., Baer, M. J.: A stain for bone illustrating apposition and absorption in two colors. Stain Technol.39, 71–73 (1964).PubMedGoogle Scholar
  11. Gesinski, R. M., Morrison, J. H., Toepfer, J. R.: Measurement of oxygen consumption of rat bone marrow cells by a polarographic method. J. appl. Physiol.24, 751–754 (1968).PubMedGoogle Scholar
  12. Girardi, A. J., McMichael, H., Henle, W.: The use of Hela cells in suspension for the quantitative study of virus propagation. Virology2, 532–544 (1956).PubMedCrossRefGoogle Scholar
  13. Hedeskov, C. J., Esmann, V.: Respiration and glycolysis of normal human lymphocytes. Blood28, 163–174 (1966).PubMedGoogle Scholar
  14. Hepp, D., Challoner, D. R., Williams, R. H.: Respiration in isolated fat cells and the effects of epinephrine. J. biol. Chem.243, 2321–2327 (1968).PubMedGoogle Scholar
  15. Howard, R. B., Pesch, L. A.: Respiratory activity of intact, isolated parenchymal cells from rat liver. J. biol. Chem.243, 3105–3109 (1968).PubMedGoogle Scholar
  16. Lilienthal, J. L., Zierler, K. L., Folk, B. P., Buka, R., Riley, M. J.: A reference base and system for analysis of muscle constitutents. J. biol. Chem.182, 501–508 (1950).Google Scholar
  17. Lowry, O. H., Rosebrough, N. S., Farr, A. L., Randell, R. J.: Protein measurement with the folin phenol reagent. J. biol. Chem.193, 265–275 (1951).PubMedGoogle Scholar
  18. Mateyko, G. M., Kopac, M. J.: Cytophysical studies on living normal and neoplastic cells. I. Separation into isolated populations. Ann. N. Y. Acad. Sci.105, 185–218 (1963).PubMedCrossRefGoogle Scholar
  19. Munro, H. N., Fleck, A.: The determination of nucleic acids. Meth. biochem. Anal.14, 113–176 (1966).Google Scholar
  20. Nachlas, M. M., Tsou, K. C., De Souz, E., Cheng, C. S., Seligman, A. M.: Cytochemical demonstration of succinic dehydrogenase by the use of a new p-nitrophenyl substitute ditetrazole. J. Histochem. Cytochem.5, 420–436 (1957).PubMedGoogle Scholar
  21. Nichols, G.; Collagen biosynthesis in bone. In: Structure and function of connective and skeletal tissue (ed. S. F. Jackson), p. 263–277. London: Butterworths 1965.Google Scholar
  22. Park, H. Z., Talmage, R. V.: Relation of endogenous parathyroid secretion to3H-Cytidine incorporation into bone cells. Endocrinology80, 552–560 (1967).PubMedCrossRefGoogle Scholar
  23. Park, H. Z., Talmage, R. V.: Comparison of the effects of calcium and endogenous parathyroid hormone on RNA synthesis in rat bone. In: Parathyroid hormone and thyrocalcitonin (calcitonin). Proc. 3rd Parathyroid Conference (R. V. Talmage and L. F. Belanger, eds.), New York: Exerpta Medica Foundation p. 203–215. 1968.Google Scholar
  24. Peck, W. A., Birge, S. J., Fedak, S. A.: Bone cells: biochemical and biological studies after enzymatic isolation. Science146, 1476–1477 (1964).PubMedCrossRefGoogle Scholar
  25. Raisz, L. B., Nieman, E.: Early effects of parathyroid hormone and thyrocalcitonin on bone in organ culture. Nature (Lond.)214, 486–487 (1967).CrossRefGoogle Scholar
  26. Schmidt, G., Thannhauser, S. J.: A method for the determination of desoxyribonucleic acid, ribonucleic acid and phosphoproteins in animal tissues. J. biol. Chem.161, 83–89 (1945).Google Scholar
  27. Shaw, W. N., Stadie, W. C.: Coexistence of insulin-responsive and insulin non-responsive glycolytic systems in rat diaphragm. J. biol. Chem.227, 115–134 (1957).PubMedGoogle Scholar
  28. Steinberg, J., Nichols, G., Jr.: Synthesis of ribonucleic acid in normal bonein vitro. Biochem. J.105, 843–856 (1967).PubMedGoogle Scholar
  29. Trudeau, D. L., Freier, E. F.: Determination of calcium in urine and serum by atomic absorption spectrophotometry (AAS). Clin. Chem.13, 101–114 (1967).PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1973

Authors and Affiliations

  • D. M. Smith
    • 3
  • C. C. JohnstonJr.
    • 1
  • A. R. Severson
    • 2
  1. 1.Division of Endocrinology and MetabolismIndiana University School of MedicineIndianapolis
  2. 2.Department of AnatomyIndiana University, School of MedicineIndianapolis
  3. 3.Department of MedicineIndiana University Medical CenterIndianapolisU.S.A.

Personalised recommendations