Skip to main content
Log in

Properties of osteoblast-like cells isolated from the cortical endosteal bone surface of adult rabbits

  • Laboratory Investigations
  • Published:
Calcified Tissue International Aims and scope Submit manuscript

Summary

The purpose of the present study was to isolate cells from the cortical endosteal bone surface of adult rabbits and characterize some properties of these cells in vitro. Following removal of the marrow from the medullary cavity of long bones, the cells lining the cortical endosteal bone surface were obtained by enzymatic digestion using 0.1% crude collagenase. These cells were examined immediately after isolation or following their growth for 8–10 days in primary culture. Freshly isolated cells contained 7–10 times more alkaline phosphatase (AP) than acid phosphatase activity. This AP activity was extremely heat labile (incubation at 58°C caused >90% loss of enzyme activity within 15 min) and significantly more sensitive to inhibition by levamisole and L-homoarginine than L-phenylalanine. Synthetic bPTH-(1–34) [1–100 ng/ml], native bPTH-(1–84) [1 U/ml] and prostaglandin E2 [1–10 µg/ml] all caused a significant accumulation of cAMP in cultured endosteal bone cells. In contrast, neither salmon calcitonin (1 U/ml) nor porcine insulin (10–1000 ng/ml) stimulated an increase in cAMP. These results suggest that cells isolated from the cortical endosteal bone surface of adult rabbits are osteoblasts or osteoblast-like. These cells retain their ability to respond in a specific manner to bone-seeking hormones following their growth in culture. Cortical endosteal bone cells isolated from rabbits of different ages, physiologic or pathologic conditions can be used to assess the changes in bone cell function and responsiveness which may accompany senescence or development of adult metabolic bone disease.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Similar content being viewed by others

References

  1. Avioli LV (1978) An appraisal of bone cellular function in senescence. In: Horton JE, Tarpley TM, Davis WF (eds) Proceedings, mechanisms of localized bone loss. Information Retrieval Inc, Washington DC, p 21

    Google Scholar 

  2. Cohn DV, Wong GL (1979) Isolated bone cells. In: Simmons DJ, Kunin AS (eds) Skeletal research: an experimental approach. Academic Press, New York, p 3

    Google Scholar 

  3. Peck WA, Dirksen TR (1966) The metabolism of bone tissuein vitro. Clin Orthop 48:243–265

    CAS  PubMed  Google Scholar 

  4. Stern PH, Raisz LG (1979) Organ culture of bone. In: Simmons DJ, Kunin AS (eds) Skeletal research: an experimental approach. Academic Press, New York, p 21

    Google Scholar 

  5. Peck WA, Birge SJ, Fedak SA (1964) Bone cells: biochemical and biological studies after enzymatic isolation. Science 146:1476–1477

    CAS  PubMed  Google Scholar 

  6. Wong GL, Cohn DV (1974) Separation of parathyroid hormone and calcitonin-sensitive cells from non-responsive bone cells. Nature 252:713–715

    Article  CAS  PubMed  Google Scholar 

  7. Rodan SB, Rodan GA (1974) The effect of parathyroid hormone and thyrocalcitonin on the accumulation of cyclic adenosine 3′:5′-monophosphate in freshly isolated bone cells. J Biol Chem 249:3068–3074

    CAS  PubMed  Google Scholar 

  8. Luben RA, Wong GL, Cohn DV (1976) Biochemical characterization with parathormone and calcitonin of isolated bone cells: provisional identification of osteoclasts and osteoblasts. Endocrinology 99:526–534

    CAS  PubMed  Google Scholar 

  9. Yagiela JA, Woodbury DM (1977) Enzymatic isolation of osteoblasts from fetal rat calvaria. Anat Rec 188:287–306

    Article  CAS  PubMed  Google Scholar 

  10. Nijweide PJ, Van der Plas A, Scherft JP (1981) Biochemical and histological studies on various bone cell preparations. Calcif Tissue Int 33:529–540

    CAS  PubMed  Google Scholar 

  11. Smith DM, Johnston CC, Severson AR (1973) Studies of the metabolism of separated bone cells. I. Techniques of separation and identification. Calcif Tissue Res 11:56–69

    CAS  PubMed  Google Scholar 

  12. Smith DM, Johnston CC (1974) Hormonal responsiveness of adenylate cyclase activity from separated bone cells. Endocrinology 95:130–139

    CAS  PubMed  Google Scholar 

  13. Williams DC, Boder GB, Toomey RE, Paul DC, Hillman CC, King KL, Van Frank RM, Johnston CC (1980) Mineralization and metabolic response in serially passaged adult rat bone cells. Calcif Tissue Int 30:233–246

    CAS  PubMed  Google Scholar 

  14. Bard DR, Dickens MJ, Smith AU, Zarek JM (1972) Isolation of living cells from mature mammalian bone. Nature 236:314–315

    Article  CAS  PubMed  Google Scholar 

  15. Puzas JE, Goodman DBP (1978) A rapid assay for cellular deoxyribonucleic acid. Anal Biochem 86:50–55

    Article  CAS  PubMed  Google Scholar 

  16. Kaplow LS (1963) Cytochemistry of leukocyte alkaline phosphatase. Am J Clin Pathol 39:439–449

    CAS  Google Scholar 

  17. Lowry OH, Roberts NR, Wu M, Hixon WS, Crawford EJ (1954) The quantitative histochemistry of brain. II. Enzyme measurements. J Biol Chem 207:19–37

    CAS  PubMed  Google Scholar 

  18. Horder M (1972) Colorimetric determination of orthophosphate in the assay of inorganic pyrophosphatase activity. Anal Biochem 49:37–47

    Article  CAS  PubMed  Google Scholar 

  19. McComb RB, Bowers GN, Posen S (1979) Alkaline phosphatase. Plenum Press, New York

    Google Scholar 

  20. Small CW (1969) A study of the heat inactivation of serum alkaline phosphatase by an automated technique. Clin Chim Acta 23:347–352

    Article  CAS  PubMed  Google Scholar 

  21. Van Belle H (1972) Kinetics and inhibition of alkaline phosphatases from canine tissue. Biochim Biophys Acta 289:158–168

    PubMed  Google Scholar 

  22. Fishman WH, Sie HG (1970) L-Homoarginine: an inhibitor of “bone and liver” alkaline phosphatase. Clin Chim Acta 29:339–341

    Article  CAS  PubMed  Google Scholar 

  23. Steiner AL, Parker CW, Kipnis DM (1972) Radioimmunoassay for cyclic nucleotides. II. Adenosine 3′,5′-monophosphate and guanosine 3′,5′-monophosphate in mammalian tissues and body fluids. J Biol Chem 247:1114–1120

    CAS  PubMed  Google Scholar 

  24. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    CAS  PubMed  Google Scholar 

  25. Woolfe CM (1968) Principles of biometry. D van Nostrand, Princeton

    Google Scholar 

  26. Davis WL, Matthews JL, Talmage RV, Martin JH, Kennedy JW III (1975) The endosteum as a functional membrane. In: Talmage RV, Owen M, Parsons JA (eds) Calcium regulating hormones. Excerpta Medica, Amsterdam, pp 275–283

    Google Scholar 

  27. Jones SJ, Boyde A (1976) Experimental study of changes in osteoblastic shape induced by calcitonin and parathyroid extract in an organ culture system. Cell Tissue Res 169:449–465

    Google Scholar 

  28. Jones JL, Davis WL, Jones RG, Miller GW, Matthews JL (1977) The effect of cytochalasin B on the endosteal lining cells of mammalian bone. Calcif Tissue Res 24:1–10

    Article  CAS  PubMed  Google Scholar 

  29. Bourne GH (1972) Phosphatase and calcification. In: Bourne GH (ed) The biochemistry and physiology of bone, vol 2. Academic Press, New York, p 79

    Google Scholar 

  30. Felix R, Fleisch H (1974) The pyrophosphatase and (Ca2+-Mg2+)-ATPase activity of purified calf bone alkaline phosphatase. Biochim Biophys Acta 350:84–94

    CAS  PubMed  Google Scholar 

  31. Wergedal JE, Baylink DJ (1969) Distribution of acid and alkaline phosphatase activity in undemineralized sections of the rat diaphysis. J Histochem Cytochem 17:799–806

    CAS  PubMed  Google Scholar 

  32. Porter RP, Gengozian N (1973) Marmoset bone marrow. In: Kruse PF, Patternson MK (eds) Tissue culture, methods and applications. Academic Press, New York, p 93

    Google Scholar 

  33. Wong GL, Cohn DV (1975) Target cells in bone for parathormone and calcitonin are different: enrichment for each cell type by sequential digestion of mouse calvaria and selective adhesion to polymeric surfaces. Proc Natl Acad Sci USA 72:3167–3171

    CAS  PubMed  Google Scholar 

  34. Peck WA, Burk JK, Wilkins J, Rodan SB, Rodan GA (1977) Evidence for preferential effects of parathyroid hormone, calcitonin and adenosine on bone and periosteum. Endocrinology 100:1357–1364

    Article  CAS  PubMed  Google Scholar 

  35. Peck WA, Carpenter J, Messinger K, DeBra D (1973) Cyclic 3′5′ adenosine monophosphate in isolated bone cells: response to low concentrations of parathyroid hormone. Endocrinology 92:692–697

    CAS  PubMed  Google Scholar 

  36. Yu J-H, Wells H, Ryan JR, Lloyd WS (1976) Effects of prostaglandins and other drugs on the cyclic AMP content of cultured bone cells. Prostaglandins 12:501–512

    Article  CAS  PubMed  Google Scholar 

  37. Dziak R, Hausmann E, Chang YW (1979) Effects of lipopolysaccharides and prostaglandins on rat bone cell calcium and cyclic AMP. Arch Oral Biol 24:347–353

    Article  CAS  PubMed  Google Scholar 

  38. Martin TJ, Ingleton PM, Coulton LA, Melick RA (1979) Metabolic properties of hormonally responsive osteogenic sarcoma cells. Clin Orthop 140:247–254

    CAS  PubMed  Google Scholar 

  39. Klein DC, Raisz LG (1970) Prostaglandins: stimulation of bone resorption in tissue culture. Endocrinology 86:1436–1440

    CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yee, J.A. Properties of osteoblast-like cells isolated from the cortical endosteal bone surface of adult rabbits. Calcif Tissue Int 35, 571–577 (1983). https://doi.org/10.1007/BF02405096

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF02405096

Key words

Navigation