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Vet Comp Orthop Traumatol 1996; 09(03): 95-100
DOI: 10.1055/s-0038-1632511
Original Research
Schattauer GmbH

Cell Proliferation as Assessed by Expression of Proliferating Cell Nuclear Antigen or the Uptake of Bromodeoxyuridine in a Rabbit Model of Leg-lengthening

G. Li
1   From the Nuffield Department of Orthopaedic Surgery
,
A. H. R. W. Simpson
1   From the Nuffield Department of Orthopaedic Surgery
,
J. Kenwright
1   From the Nuffield Department of Orthopaedic Surgery
,
J. T. Triffitt
2   MRC Bone Research Laboratory, Nuffield Orthopaedic Centre, University of Oxford, Headington, Oxford, UK
› Author Affiliations
Further Information

Publication History

Received for publication 15 August 1995

Publication Date:
23 February 2018 (online)

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Summary

An experimental model of leg lengthening has been used to study the cellular responses of the regenerating bone to different rates of distraction. Cell proliferation were assessed by detection of proliferating cell nuclear antigen using a monoclonal antibody, PC10. The technique was verified by comparison with bromodeoxyuridine uptake and subsequent detection with specific antibody (Bu20A). The positive staining index (PSI) was calculated for a variety of tissues and the PC10 PSI was greater than that of Bu20A, as described by the expression: PC10 PSI = 1.6 Bu20A PSI + 12.9, with a correlation coefficient 0.79. The results suggest that PC10 may be used as an alternative marker to assess cell proliferation in rabbit regenerating bone tissue. In addition, the rate of cell proliferation during leg-lengthening was found to reach a maximum at a distraction rate of 0.7 mm/day without further change at higher rates.

Cell proliferation was assessed in an experimental system of leg-lengthening by two separate methods. The presence of proliferating cell nuclear antigen or the uptake of bromodeoxyuridine were determined immuno-histochemically. Both methods indicated cell proliferation during leg-lengthening reaches a maximum at a distraction rate of 0.7 mm/day.

Keywords

Cell proliferation - immunohistochemistry - PC10 - bromodeoxyuridine - rabbit model - leg-lengthening
 

  • REFERENCES

  • 1 Alberty A, Pelton J. Proliferation of the hypertrophic chondrocytes of the growth plate after physeal distraction. Clin Orthop 1993; 297: 7-11.
    PubMedGoogle Scholar
  • 2 Apte SS. Validation of bromodeoxyuridine immunohistochemistry for localization of S-phase cells in decalcified tissues: A comparative study with tritiated thymidine autoradiography. J Histochem 1990; 22: 401
    CrossrefPubMedGoogle Scholar
  • 3 Apte SS. Physeal distraction and cell proliferation in the growth plate. J Bone Joint Surg (Br) 1994; 76 B 837-43.
    PubMedGoogle Scholar
  • 4 Bravo R, Macoonald-Bravo H. Changes in the nuclear distribution of cyclin (PCNA) but not its synyhesis depend on DNA replication. EMBO J 1985; 4: 655-61.
    PubMedGoogle Scholar
  • 5 Bravo R, Macoonald-Bravo H. Existence of two populations of cyclin/proliferating cell nuclear antigen during the cell cycle association with DNA replication sites. J Cell Biol 1987; 105: 1549-54.
    CrossrefPubMedGoogle Scholar
  • 6 Codivilla A. On the means of lengthening in the lower limbs, the muscles and tissues which are shortened through deformity. Am J Orthop Surg 1905; 2: 353-69.
    PubMedGoogle Scholar
  • 7 Galand P, Degraef C. Cyclin / PCNA immunostaining as an alternative to tritiated thymidine pulse labelling for marking S phase cells in paraffin sections from animals and human tissues. Cell Tissue Kinet 1989; 22: 383-92.
    PubMedGoogle Scholar
  • 8 Gratzner HG. Monoclonal antibody to 5-bromo and 5-iododeoxyuridine: a new reagent for detection of DNA replication. Science 1982; 218: 474-5.
    CrossrefPubMedGoogle Scholar
  • 9 Hall PA, Levison DA, Woods AL. et al Proliferating cell nuclear antigen (PCNA) immuno-localisation in paraffin sections: an index of cell proliferation with evidence of deregulated expression in some neoplasms. J Pathol 1990; 162: 285-94.
    CrossrefPubMedGoogle Scholar
  • 10 Ilizarov GA. The tension-stress effect on the genesis and growth of tissues. Part I. The influence of stability of fixation and soft tissue preservation. Clin Orthop 1989; 238: 249-81.
    CrossrefPubMedGoogle Scholar
  • 11 Ilizarov GA. The tension-stress effect on the genesis and growth of tissues. Part II. The influence of the rate and frequency of distraction. Clin Orthop 1989; 239: 263
    PubMedGoogle Scholar
  • 12 Jaskulski D, Gatti C, Travali S. et al Regulation of the proliferating cell nuclear antigen/ cyclin and thymidine kinase mRNA levels by growth factors. J Biol Chem 1988; 263 (10) 175-9.
    PubMedGoogle Scholar
  • 13 Kawamura B, Hosono S, Takahashi T. et al Limb lengthening by means of subcutaneous osteotomy. J Bone Joint Surg (Am) 1968; 50 A (05) 851-78.
    CrossrefPubMedGoogle Scholar
  • 14 Matsunaga S, Ijiri K, Sakou T, Morimoto N. Immunohistochemical study of activating osteoblasts from undifferentiated mesenchymal cells by electrical stimulation. Bone 1993; 14: 327-32.
    CrossrefPubMedGoogle Scholar
  • 15 Miyachi K, Fritzler MJ, Tan EM. Autoantibody to a nuclear antigen in proliferating cells. J Immunol 1978; 121: 2228-34.
    PubMedGoogle Scholar
  • 16 Morris GF, Mathews MB. Regulation of proliferating cell nuclear antigen during the cell cycle. J Cell Biol 1989; 264 (13) 856-64.
    PubMedGoogle Scholar
  • 17 Prelich G, Tan CK, Kostura M. et al Functional identity of proliferating cell nuclear antigen and DNA polymerase delta auxiliary protein. Nature 1987; 326: 517-20.
    CrossrefPubMedGoogle Scholar
  • 18 Scott RJ, Hall PA, Haldane JS. et al A comparison of immunohistological markers of cell proliferation with experimentally determined growth fraction. J Pathol 1991; 165: 173-8.
    CrossrefPubMedGoogle Scholar
  • 19 Shivji KK, Kenny MK, Wood RD. Proliferating cell nuclear antigen is required for DNA excision repair. Cell 1992; 69: 367-74.
    CrossrefPubMedGoogle Scholar
  • 20 Toschi L, Bravo R. Changes in cyclin/proliferating cell nuclear antigen distribution during DNA repair synthesis. J Cell Biol 1988; 107: 1623-8.
    CrossrefPubMedGoogle Scholar
  • 21 Wagner H. Operative lengthening of the femur. Clin Orthop 1978; 136: 125-42.
    PubMedGoogle Scholar
  • 22 Weibel ER. Practical methods for biological morphometry. In: Stereological Methods. Vol. 1. London: Academic Press Inc; 1979: 95-115.
    Google Scholar
  • 23 Wijsman JH, Van Dierendonck JH, Keizzer R. et al Immunoreactivity of proliferatng cell nuclear antigen compared with bromodeoxyurdine incorporation in normal and neoplastic rat tissue. J Pathol 1992; 168: 75-83.
    CrossrefPubMedGoogle Scholar
  • 24 Woods AL, Hall PA, Shepherd NA. et al The assessment of proliferating cell nuclear antigen (PCNA) immunostaining in primary gastro-intestinal lymphomas and its relationship to histological grade, S + G2M phase fraction (flow cytometric analysis) and prognosis. Histopathology 1991; a 19: 21-7.
    CrossrefPubMedGoogle Scholar
  • 25 Yu CC-W, Woods AL, Levison DA. The assessment of cellular proliferation by immunohistochemistry: a review of currently available methods and their applications. Histochem J 1992; (b) 24: 121-31.
    CrossrefPubMedGoogle Scholar
  • 26 Yu CC-W, Filipe MI. Update on proliferation- associated antibodies applicable to formalin-fixed paraffin-embedded tissue and their clinical applications. Histochem J 1993; 25: 843-53.
    CrossrefPubMedGoogle Scholar