Histochemistry and Cell Biology

, Volume 124, Issue 2, pp 123–130 | Cite as

Osteocalcin and matrix GLA protein in developing teleost teeth: identification of sites of mRNA and protein accumulation at single cell resolution

  • J. B. Ortiz-Delgado
  • D. C. Simes
  • P. Gavaia
  • C. Sarasquete
  • M. L. CancelaEmail author
Original paper


In this study, the tissue distribution and accumulation of osteocalcin or bone Gla protein (BGP) and matrix Gla protein (MGP) were determined during tooth development in a teleost fish, Argyrosomus regius. In this species, the presence of BGP and MGP mRNA in teeth was revealed by in situ hybridization. mRNA for BGP was detected in the odontoblasts as well as in its cytoplasmic processes emerging through dentinal tubules, while mRNA for MGP was expressed in the enamel portion within the apical portion of the elongated cell bodies of enameloblasts, adjacent to the root of the teeth as well as in cells within the pulpal space. Immunolocalization of BGP and MGP demonstrated that these proteins accumulate mainly in the mineralized dentin or in enameloblastic processes, confirming in situ hybridization results. In this study, we examined for the first time the localization of both BGP and MGP gene expression and protein accumulation within the different regions of the vertebrate tooth. We clearly demonstrated that although the overall pattern of BGP and MGP gene expression and protein accumulation in A. regius teeth was in general agreement to what is known for other vertebrates such as rats or rodents, our study provided novel information and highlighted some species-differences between fish and higher vertebrates.


Osteocalcin BGP Matrix Gla protein MGP Teleost fish Immunohistochemistry Teeth In situ hybridization 



This work was supported in part by CCMAR (Portugal) funding and by a research grant from the Spanish Foundation for Science and Technology (MCYT; AGL2003-03558). J.B. Ortiz Delgado and P. Gavaia were recipients, respectively, of postdoctoral (SFRH/BPD/7151/2001) and PhD (PRAXIS XXI BD/99/19665) fellowships, awarded by the Portuguese Science and Technology Foundation (FCT). Part of this work was performed while D. Simes was the recipient of a PRODEP fellowship from the Portuguese Ministry of Education.


  1. Bawden JW, Deaton TG, Koch GG, Crawford BP (1989) Effect of an acute maternal fluoride dose on fetal plasma fluoride levels and enamel fluoride uptake in guinea pigs. J Dent Res 68(7):1169–1172PubMedGoogle Scholar
  2. Bronckers AL, Gay S, DiMuzio MT, Butler WT (1985) Immunolocalization of gamma-carboxyglutamic acid containing proteins in developing molar tooth germs of the rat. Collagen 5:17–22PubMedGoogle Scholar
  3. Bronckers AL, D’Souza RN, Butler WT, Lyaruu DM, van Dijk S, Gay S, Woltgens JH (1993) Dentin sialoprotein: biosynthesis and developmental appearance in rat tooth germs in comparison with amelogenins, osteocalcin and collagen type-I. Cell Tissue Res 272(2):237–247CrossRefPubMedGoogle Scholar
  4. Bronckers AL, Farach-Carson MC, Van Waveren E, Butler WT (1994) Immunolocalization of osteopontin, osteocalcin, and dentin sialoprotein during dental root formation and early cementogenesis in the rat. J Bone Miner Res 9(6):833–841PubMedCrossRefGoogle Scholar
  5. Cole DE, Hanley DA (1991) Osteocalcin. In: Hall BK (ed) Bone matrix and bone specific products. CRC Press, London, pp 239–294Google Scholar
  6. Ducy P, Desbois C, Boyce B, Pinero G, Story B, Dunstan C, Smith E, Bonadio J, Goldstein S, Gundberg C, Bradley A, Karsenty G (1996) Increased bone formation in osteocalcin-deficient mice. Nature 382(6590):448–452CrossRefPubMedGoogle Scholar
  7. Fraser JD, Price PA (1988) Lung heart and kidney express high levels of mRNA for the vitamin K-dependent MGP. J Biol Chem 263:11033–10036PubMedGoogle Scholar
  8. Gorter V, Coomans D, Wisse E (1988) Immunocytochemical localization of OC in human teeth. Calcif Tissue Int 43:128–130PubMedCrossRefGoogle Scholar
  9. Hale JE, Fraser JD, Price PA (1988) The identification of matrix Gla protein in cartilage. J Biol Chem 263(12):5820–5824PubMedGoogle Scholar
  10. Hashimoto F, Kobayashi Y, Kobayashi ET, Sakai E, Kobayashi K, Shibata M, Kato Y, Sakai H (2001) Expression and localization of MGP in rat tooth cementum. Arch Oral Biol 46(7):585–592CrossRefPubMedGoogle Scholar
  11. Hauschka PV, Reddi AH (1980) Correlation of the appearance of gamma-carboxyglutamic acid with the onset of mineralization in developing endochondral bone. Biochem Biophys Res Commun 92(3):1037–1041CrossRefPubMedGoogle Scholar
  12. Hauschka PV, Lian JB, Cole DE, Gundberg CM (1989) Osteocalcin and matrix Gla protein: vitamin K-dependent proteins in bone. Physiol Rev 69(3):990–1047PubMedGoogle Scholar
  13. Heersche JN, Reimers SM, Wrana JL, Waye MM, Gupta AK (1992) Changes in expression of alpha 1 type 1 collagen and osteocalcin mRNA in osteoblasts and odontoblasts at different stages of maturity as shown by in situ hybridization. Proc Finn Dent Soc 88(1):173–182PubMedGoogle Scholar
  14. Luo G, Ducy P, McKee MD, Pinero GJ, Loyer E, Behringer RR, Karsenty G (1997) Spontaneous calcification of arteries and cartilage in mice lacking MGP. Nature 386:78–81CrossRefPubMedGoogle Scholar
  15. Nishimoto SK, Waite JH, Nishimoto M, Kriwacki RW (2003) Structure, activity, and distribution of fish osteocalcin. J Biol Chem 278(14):11843–11848CrossRefPubMedGoogle Scholar
  16. Otawara Y, Price PA (1986) Developmental appearance of matrix GLA protein during calcification in the rat. J Biol Chem 261(23):10828–10832PubMedGoogle Scholar
  17. Papagerakis P, Berdal A, Mesbah M, Peuchmaur M, Malaval L, Nydegger J, Simmer J, Macdougall M (2002) Investigation of osteocalcin, osteonectin, and dentin sialophosphoprotein in developing human teeth. Bone 30(2):377–385CrossRefPubMedGoogle Scholar
  18. Pinto JP, Ohresser MC, Cancela ML (2001) Cloning of the bone Gla protein gene from the teleost fish Sparus aurata. Evidence for overall conservation in gene organization and bone-specific expression from fish to man. Gene 270(1–2):77–91CrossRefPubMedGoogle Scholar
  19. Pinto JP, Conceicao N, Gavaia PJ, Cancela ML (2003) Matrix Gla protein gene expression and protein accumulation colocalize with cartilage distribution during development of the teleost fish Sparus aurata. Bone 32(3):201–210CrossRefPubMedGoogle Scholar
  20. Price PA (1989) Gla-containing proteins of bone. Connect Tissue Res 21:51–60PubMedCrossRefGoogle Scholar
  21. Price PA, Urist MR, Otawara Y (1983) Matrix Gla protein, a new γ-carboxyglutamic acid-containing protein which is associated with the organic matrix of bone. Biochem Biophys Res Commun 117:756–771CrossRefGoogle Scholar
  22. Sarasquete C, Gutiérrez M (2005) New tetrachromic VOF stain (Type III-G.S) for normal and pathological fish tissues. Eur J Histochem (in press)Google Scholar
  23. Simes DC, Williamson MK, Schaff BJ, Gavaia PJ, Ingleton PM, Price PA, Cancela ML (2003) Characterization of osteocalcin (BGP) and matrix Gla protein (MGP) fish specific antibodies: validation for immunodetection studies in lower vertebrates. Calcif Tissue Int 74(2):170–180CrossRefPubMedGoogle Scholar
  24. Sire JY, Huysseune A (2003) Formation of dermal skeletal and dental tissues in fish: a comparative and evolutionary approach. Biol Rev 78:219–249PubMedCrossRefGoogle Scholar
  25. Sommer B, Bickel M, Hofstetter W, Wetterwald A (1996) Expression of matrix proteins during the development of mineralized tissues. Bone 19(4):371–380CrossRefPubMedGoogle Scholar
  26. Yamamoto TT, Takemura T, Kitamura Y, Nomura S (1994) Site-specific expression of mRNA for osteonectin, osteocalcin and osteopontin revealed by in situ hybridization in rat periodontal ligament during physiological tooth movement. J Histochem Cytochem 42(7):885–896PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • J. B. Ortiz-Delgado
    • 1
    • 2
  • D. C. Simes
    • 1
  • P. Gavaia
    • 1
  • C. Sarasquete
    • 2
  • M. L. Cancela
    • 1
    Email author
  1. 1.CCMarUniversidade do AlgarveFaroPortugal
  2. 2.Superior Council for Scientific InvestigationInstitute of Marine Sciences of Andalucia - CSICPuerto Real (Cádiz)Spain

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