Clinical Rheumatology

, Volume 15, Issue 6, pp 563–572 | Cite as

Intermittent compressive load stimulates osteogenesis and improves osteocyte viability in bones cultured “in vitro”

  • E. Lozupone
  • C. Palumbo
  • A. Favia
  • M. Ferretti
  • S. Palazzini
  • F. P. Cantatore


The effect of mechanical stresses on osteogenesis, the viability of osteocytes and their metabolic activity in organ culture of bones intermittently loaded “in vitro” are reported.

Metatarsal bones, isolated from 12-day-old rats, were cultured in BGJb medium (with 10% foetal calf serum, 75µg/ml of ascorbic acid, 100 U/ml of penicillin and 100µg/ml of streptomycin), in humidified air enriched by 5% CO2 and 30% O2, and loaded in our original device for 1/2 an hour at 1 Hz. homotypic isolated and unloaded bones, cultured in the same medium, were taken as controls.

The ALP (alkaline phophatase activity) increases in the media of loaded bones in comparison with the control bones. The percentage of viable osteocytes is significantly greater in loaded than in control bones. TEM observations demonstrate that in both loaded and control unloaded bones, osteocytes show well developed organelle machinery and several gap junctions with adjacent cellular processes. In the cells of loaded bones, however, a higher number of cytoplasmic organelles and gap junctions were found. In particular, RER increases twice, gap junctions three times. The induced osteogenesis and the TEM observations demonstrate the suitability of this experimental model and support the recent advanced hypothesis according to which the mechanical loading may exert a trophic function on osteocytes, stimulating both the proteic synthesis in the above-mentioned cells and the cell-to-cell communication. Furthermore, the loading is likely to exert a biological stimulus on osteoblasts via signalling molecules produced by osteocytes.

Key words

Organ Culture Intermittent Compressive Force Osteogenesis Ultrastructure Osteocyte 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    Smith, E.L. Gilligan, C. Mechanical forces and bone. In: Bone and Mineral Research. Eds.: Peck W.A., Amsterdam, NY, Oxford, Elsevier 1989, 139–173.Google Scholar
  2. 2.
    Skerry, T.M., Bitenski, L., Chayen, J., Lanyon, L.E. Early strain-related changes in enzyme activity in osteocytes following bone loading in vivo. J Bone Min Res 1989, 4, 783–788.Google Scholar
  3. 3.
    Dodds, R.A., Ali, N., Pead, M.J., Lanyon, L.E. Early loading-related changes in the activity of G6PD and ALP in osteocytes and periosteal osteoblasts in rat fibulae in vivo. J Bone Min Res 1993, 8, 261–267.Google Scholar
  4. 4.
    El Hai, A.J., Minter, S.L., Rawlinson, S.C.F., Suswillo, R., Lanyon, L.E. Cellular responses to mechanical loading in vitro. J Bone Min Res 1990, 5, 923–932.Google Scholar
  5. 5.
    Rawlinson, S.C.F., El Haj, A.J., Minter, S.L., Tavares, I.A., Bennett, A., Lanyon, L.E. Load-related increases of prostaglandin production in cores of adult canine cancellous bone in vitro — a role for prostacyclin in adaptive bone remodelling? J Bone Min Res 1991, 6, 1345–1351.Google Scholar
  6. 6.
    Lozupone, E., Favia, A., Grimaldi, A. Effects of intermittent mechanical force on bone tissue in vitro: preliminary results. J Bone Min Res 1992, 7 (suppl. 2), S407-S409.Google Scholar
  7. 7.
    Lozupone, E., Favia, A., Grimaldi, A., Coluccia, R. Sopravvivenza del tessuto osseo in cultura organotipica, sotto carico meccanico intermittente. Risultati preliminari. Boll Soc Ital Biol Sper 1990, 66, 1043–1050.PubMedGoogle Scholar
  8. 8.
    Rodan, G.A., Mensi, T., Harvey, A. A quantitative method for the application of compressive forces to bone in tissue culture. Calcif Tiss Res 1975, 18, 125–131.Google Scholar
  9. 9.
    Weinreb, M., Shinar, O., Rodan, G.A. Different pattern of alkaline phosphatase, osteopontin and osteocalcin expression in developing rat bone visualized by in situ hybridization. J Bone Min Res 1990, 5, 831–842.Google Scholar
  10. 10.
    Collin, P. Nefussi, J.R., Wetterwald, A., Nicolas, V., Boy-Lefevre, M.L., Fleisch, H., Forest, N. Expression of collagen, osteocalcin and bone alkaline phosphatase in a mineralizing rat osteoblastic cell culture. Calcif Tissue Int 1992, 50, 175–183.CrossRefPubMedGoogle Scholar
  11. 11.
    Schmidt, R., Kulbe, K.D. Long-term cultivation of human osteoblasts. Bone and Mineral 1993, 20, 211–221.PubMedGoogle Scholar
  12. 12.
    Piekarski, K., Munro, M. Transport mechanism operating between blood supply and osteocytes in long bones. Nature 1977, 269, 80–82.CrossRefPubMedGoogle Scholar
  13. 13.
    Kufahl, R.H., Saha, S. A theoretical model for stress-generated fluid flow in the canaliculi-lacunae network in bone tissue. J. Biomechan 1990, 23, 171–180.CrossRefGoogle Scholar
  14. 14.
    Baldwin, K.M. The fine structure and electrophysiology of heart muscle cell injury. J Cell Biol 1970, 46, 455–476.CrossRefPubMedGoogle Scholar
  15. 15.
    Marotti, G., Canè, V., Palazzini, S., Palumbo, C. Structure-function relationships in the osteocyte. Ital J Miner Electrol Metab 1990, 4, 93–106.Google Scholar
  16. 16.
    Farris, E.J. The rat as an experimental animal. In: The care and breeding of laboratory animals. New York: Wiley, 1950.Google Scholar
  17. 17.
    Pead, M.D., Suswillo, R., Skerry, T.M., Vedi, S., Lanyon, L.E. Increased 3H-uridine levels in osteocytes following a single short period of dynamic bone loading in vivo. Calcif Tissue Int 1988, 43, 92–96.PubMedGoogle Scholar
  18. 18.
    Zaman, G., Dallas, S.L., Lanyon, L.E. Cultured embryonic bone shafts show osteogenic responses to mechanical loading. Calcif Tissue Int 1992, 51, 132–136.CrossRefPubMedGoogle Scholar
  19. 19.
    Klein-Nulend, J., van der Plas, A., Semeins, C.M., Ajubi, N.E., Frangos, J.A., Nijweide, P.J., Burger, E.H. Sensitivity of osteocytes to mechanical stress in vitro. FASEB J 1995, 9, 441–445.PubMedGoogle Scholar
  20. 20.
    Palumbo, C. A three-dimensional ultrastructural study of osteoidosteocytes in the tibia of chick embryos. Cell Tissue Res 1986, 246, 125–131.CrossRefPubMedGoogle Scholar
  21. 21.
    Stanka, P. Occurrence of cell junctions and microfilaments in osteoblasts. Cell Tissue Res 1975, 159, 413–422.CrossRefPubMedGoogle Scholar
  22. 22.
    Jeansonne, B.G., Feafin, F.F., McMinn, R.W., Shoemaker, R.L., Rehm, V.S. Cell-to-cell communication of osteoblasts. J Dent Res 1979, 58, 1415–1423.PubMedGoogle Scholar
  23. 23.
    Knese, H.K. Stutzgewebe und Skelettsystem. In: Handbuch der mikroskopischen Anatomie des Menschen. Bd 2/5 Ed.: H.K. Knese, Berlin, Springer 1979, 513–594.Google Scholar
  24. 24.
    Matthews, J.L. Bone structure and ultrastructure. In: Fundamental and Clinical Bone Physiology. Eds.: Marshall, R., Urist, M.D., Philadelphia, Lippincott JB Company, 1980, 4–44.Google Scholar
  25. 25.
    Miller, S.C., Bowman, B.M., Smith, J.M., Jee, W.S.S. Characterization of endosteal bone-lining cells from fatty marrow bone sites in adult beagles. Anat Rec 1980, 198, 163–173.CrossRefPubMedGoogle Scholar
  26. 26.
    Doty, S.B. Morphological evidence of gap junctions between bone cells. Calcif Tissue Int 1981, 33, 509–512.PubMedGoogle Scholar
  27. 27.
    Doty, S.B. Cell-to-cell communication in bone tissue. In: The Biological Mechanism of Tooth Eruption and Tooth Resorption. Ed.: V. Davidovitch, Birmingham, EBSCO Media, 1988. 61–69Google Scholar
  28. 28.
    Bhargawa, U., Bar-Lev, M., Bellows, C.G., Audin, J.E. Ultrastructural analysis of bone nodules formed in vitro by isolated fetal rat calvaria cells. Bone 1988, 9, 155–163.CrossRefPubMedGoogle Scholar
  29. 29.
    Palumbo, C., Palazzini, S., Marotti, G. Morphological study of intercellular junctions during osteocyte differentiation. Bone 1990, 11, 401–406.CrossRefPubMedGoogle Scholar
  30. 30.
    Palumbo, C., Palazzini, S., Zaffe, D., Marotti, G. Osteocyte differentiation in the tibia of newborn rabbit: an ultrastructural study of the formation of cytoplasmic processes. Acta Anat 1990, 137, 350–358.PubMedGoogle Scholar
  31. 31.
    Lozupone, E., Favia, A., Cantatore, F.P. Overloading inhibits osteogenesis and induces osteocyte death in bones cultured “in vitro”. Calcif Tissue Int 1995, 56, 436 (abstract).Google Scholar

Copyright information

© Clinical Rheumatology 1996

Authors and Affiliations

  • E. Lozupone
    • 3
  • C. Palumbo
    • 2
  • A. Favia
    • 3
  • M. Ferretti
    • 2
  • S. Palazzini
    • 2
  • F. P. Cantatore
    • 1
  1. 1.Institute of RheumatologyUniversity of BariItaly
  2. 2.Dept of Morphological and Forensic Sciences, Section of Human AnatomyUniversity of ModenaItaly
  3. 3.Institute of Human AnatomyUniversity of Bari, PoliclinicoBariItaly

Personalised recommendations