Skip to main content

Mineralization process during acellular cementogenesis in rat molars: a histochemical and immunohistochemical study using fresh-frozen sections

Histochemistry and Cell Biology volume 127, pages 303–311 (2007)Cite this article

Abstract

This study was designed to detect tissue non-specific alkaline phosphatase (TNSALP) by Azo-dye staining, calcium by glyoxal bis (2-hydroxyanil) (GBHA) staining, bone sialoprotein (BSP) and osteopontin (OPN) by immunoperoxidase staining in developing rat molars, and also to discuss the mineralization process during acellular cementogenesis. To restrain a reduction in histochemical and immunohistochemical reactions, fresh-frozen undemineralized sections were prepared. Where the epithelial sheath was intact, TNSALP reaction was observed in the dental follicle, but not in the epithelial sheath. With the onset of dentin mineralization, the BSP- and OPN-immunoreactive, initial cementum layer appeared. At this point, cementoblasts had shown intense TNSALP reaction and GBHA reactive particles (=calcium-GBHA complex) appeared on the root surface. With further development, the reaction of TNSALP and GBHA became weak on the root surface. Previous studies have shown that the initial cementum is fibril-poor and that matrix vesicles and calciferous spherules appear on the root surface only during the initial cementogenesis. The findings mentioned above suggest that: during the initial cementogenesis, cementoblasts release matrix vesicles which result in calciferous spherules, corresponding to the GBHA reactive particles. The calciferous spherules trigger the mineralization of the initial cementum. After principal fiber attachment, mineralization advances along collagen fibrils without matrix vesicles.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  • Anderson HC (2003) Matrix vesicles and calcification. Curr Rheumatol Rep 5:222–226

    PubMed  Google Scholar 

  • Arambawatta AKS, Yamamoto T, Wakita M (2006) An immunohistochemical study of the attachment mechanisms in different kinds of adhesive interfaces in teeth and alveolar bone of the rat. J Periodont Res 41:259–265

    PubMed  Article  CAS  Google Scholar 

  • Balcerzak M, Hamade E, Zhang L, Pikula S, Azzar G, Radisson J, Bandorowicz-Pikula J, Buchet R (2003) The roles of annexins and alkaline phosphatase in mineralization process. Acta Biochim Polon 50:1019–1038

    PubMed  CAS  Google Scholar 

  • Bosshardt DD (2005) Are cementoblasts a subpopulation of osteoblasts or a unique phenotype? J Dent Res 84:390–406

    PubMed  CAS  Google Scholar 

  • Bosshardt DD, Nanci A (1997) Immunodetection of enamel- and cementum-related (bone) proteins at the enamel-free area and cervical portion of tooth in rat molars. J Bone Miner Res 12:367–379

    PubMed  Article  CAS  Google Scholar 

  • Bosshardt DD, Zalzal S, McKee MD, Nanci A (1998) Developmental appearance and distribution of bone sialoprotein and osteopontin in human and rat cementum. Anat Rec 250:13–33

    PubMed  Article  CAS  Google Scholar 

  • Burstone MS (1962) Alkaline phosphatase, naphthol AS phosphate method. In: Burstone MS (ed) Enzyme histochemistry and its application on the study of neoplants. Academic, New York, pp 275–276

    Google Scholar 

  • Cho MI, Garant PR (1988) Ultrastructural evidence of directed cell migration during initial cementoblast differentiation in root formation. J Periodont Res 23:268–276

    PubMed  Article  CAS  Google Scholar 

  • Cho MI, Garant PR (1989) Radioautographic study of (3H) mannose utilization during cementoblast differentiation, formation of cementum, and development of periodontal ligament principal fibers. Anat Rec 223:209–222

    PubMed  Article  CAS  Google Scholar 

  • Cho MI, Garant PR (2000) Development and general structure of the periodontium. Periodontol 2000 24:9–27

    PubMed  Article  CAS  Google Scholar 

  • Diekwisch TSH (2001) The developmental biology of cementum. Int J Dev Biol 45:695–706

    PubMed  CAS  Google Scholar 

  • Embery G, Waddington RJ, Hall RC, Last KS (2000) Connective tissue elements as diagnostic aids in periodontology. Periodontol 2000 24:193–214

    PubMed  Article  CAS  Google Scholar 

  • Embery G, Hall R, Waddington R, Septier D, Goldberg M (2001) Proteoglycans in dentinogenesis. Crit Rev Oral Biol Med 12:331–349

    PubMed  CAS  Google Scholar 

  • Ganss B, Kim RH, Sodek J (1999) Bone sialoprotein. Crit Rev Oral Biol Med 10:79–98

    PubMed  CAS  Google Scholar 

  • Goldberg M, Septier D, Lécole S, Chardin H, Quintana MA, Acevedo AC, Gafni G, Dillouya D, Vermelin L, Thonemann B, Schmaltz G, Bissila-Mapahou P, Carreau JP (1995) Dental mineralization. Int J Dev Biol 39:93–110

    PubMed  CAS  Google Scholar 

  • Hammarström L (1997) Enamel matrix, cementum development, and regeneration. J Clin Periodontol 24:658–668

    PubMed  Article  Google Scholar 

  • Hammarström L, Alatli I, Fong CD (1996) Origins of cementum. Oral Dis 2:63–69

    PubMed  Article  Google Scholar 

  • Hirata A, Nakamura H (2006) Localization of perlkan and heparanase in Hertwig’s epithelial root sheath during root formation in mouse molars. J Histochem Cytochem DOI 10.1369/jhc.5A6883.2006

  • Hosoya A, Yoshiba K, Yoshiba N, Hoshi K, Iwaku M, Ozawa H (2003) An immunohistochemical study on hard tissue formation in a subcutaneously transplanted rat molar. Histochem Cell Biol 119:27–35

    PubMed  CAS  Google Scholar 

  • Hosoya A, Hoshi K, Sahara N, Ninomiya T, Akahane S, Kawamoto T, Ozawa H (2005) Effects of fixation and decalcification on the immunohistochemical localization of bone matrix proteins in fresh-frozen bone sections. Histochem Cell Biol 123:639–646

    PubMed  Article  CAS  Google Scholar 

  • Janones DS, Massa LF, Arana-Chavez VE (2005) Immunohistochemical examination of the presence of amelogenin during the root development of rat molars. Arch Oral Biol 50:527–532

    PubMed  Article  CAS  Google Scholar 

  • Kaneko H, Hashimoto S, Enokiya Y, Ogiuchi H, Shimono M (1999) Cell proliferation and death of Hertwig’s epithelial root sheath in the rat. Cell Tissue Res 298:95–103

    PubMed  Article  CAS  Google Scholar 

  • Kashiwa HK (1966) Calcium in cells of fresh bone stained with glyoxal bis (2-hydroxyanil). Stain Technol 41:49–55

    PubMed  CAS  Google Scholar 

  • Kawamoto T (2003) Use of a new adhesive film for the preparation of multi-purpose fresh-frozen sections from hard tissues, whole-animals, insects and plants. Arch Histol Cytol 66:123–143

    PubMed  Article  Google Scholar 

  • Kawamoto T, Shimizu M (2000) A method for preparing 2- to 50-μm-thick fresh-frozen sections of large samples and undecalcified hard tissues. Histochem Cell Biol 113:331–339

    PubMed  CAS  Google Scholar 

  • Limeback H (1991) Molecular mechanisms in dental hard tissue mineralization. Curr Opin Dent 1:826–835

    PubMed  CAS  Google Scholar 

  • Linde A (1995) Dentin mineralization and the role of odontoblasts in calcium transport. Connect Tissue Res 33:163–170

    PubMed  CAS  Google Scholar 

  • McKee MD, Zalzal S, Nanci A (1996) Extracellular matrix in tooth cementum and mantle dentin: localization of osteopontin and other noncollagenous proteins, plasma proteins, and glycoconjugates by electron microscopy. Anat Rec 245:293–312

    PubMed  Article  CAS  Google Scholar 

  • Nanci A (1999) Content and distribution of noncollagenous matrix proteins in bone and cementum: relationship to speed of formation and collagen packing density. J Struct Biol 126:256–269

    PubMed  Article  CAS  Google Scholar 

  • Owens PDA (1980) A light and electron microscopic study of the early stages of root surface formation in molar teeth in the rat. Arch Oral Biol 24:901–907

    Article  Google Scholar 

  • Paynter KJ, Pudy G (1958) A study of the structure, chemical nature, and development of cementum in the rat. Anat Rec 131:233–251

    PubMed  Article  CAS  Google Scholar 

  • Sasano Y, Maruya Y, Sato H, Zhu J-X, Takahashi I, Mizoguchi I, Kagayama M (2001) Distinctive expression of extracellular matrix molecules at mRNA and protein levels during formation of cellular and acellular cementum in the rat. Histochem J 33:91–99

    PubMed  Article  CAS  Google Scholar 

  • Schroeder HE (1992) Biological problems of regenerative cementogenesis: synthesis and attachment of collagenous matrices on growing and established root surfaces. Int Rev Cytol 142:1–59

    PubMed  CAS  Article  Google Scholar 

  • Schroeder HE (1993) Human cellular mixed stratified cementum: a tissue with alternating layers of acellular extrinsic- and cellular intrinsic fiber cementum. Schweiz Monatsschr Zahnmed 103:550–560

    PubMed  CAS  Google Scholar 

  • Slavkin HC, Bessem C, Fincham AG, Bringas P, Santos V, Snead M, Zeichner-David M (1989) Human and mouse cementum proteins immunologically related to enamel proteins. Biochim Biophys Acta 991:12–18

    PubMed  CAS  Google Scholar 

  • Sodek J, Ganss B, McKee MD (2000) Osteopontin. Crit Rev Oral Biol Med 11:279–303

    PubMed  CAS  Article  Google Scholar 

  • Tadatomo Y, Komatsu M, Kagayama M (2002) Immunohistochemical studies on physiological root resorption of rat molars (in Japanese). Jpn J Oral Biol 44:282–292

    Google Scholar 

  • Takano Y, Matsuo S, Wakisaka S, Ichikawa H, Nishikawa S (1988) A histochemical demonstration of calcium in the maturation stage enamel organ of rat incisors. Arch Histol Cytol 51:241–248

    PubMed  CAS  Google Scholar 

  • Ten Cate AR (1994) Hard tissue formation and destruction. In: Ten Cate AR (ed) Oral histology, 4th edn. Mosby, St. Louis, pp 111–119

    Google Scholar 

  • Thomas HF (1995) Root formation. Int J Dev Biol 39:231–237

    PubMed  CAS  Google Scholar 

  • Waddington RJ, Embery G (2001) Proteoglycans and orthodontic movement. J Orthod 28:281–290

    PubMed  Article  CAS  Google Scholar 

  • Whyte MP (1994) Hypophosphatasia and the role of alkaline phosphatase in skeletal mineralization. Endocr Rev 15:439–461

    PubMed  Article  CAS  Google Scholar 

  • Wiesmann HP, Meyer U, Plate U, Höhling HJ (2005) Aspects of collagen mineralization in hard tissue formation. Int Rev Cytol 242:121–156

    PubMed  Article  CAS  Google Scholar 

  • Yamamoto T (1986) The innermost layer of cementum: its ultrastructure, development, and calcification. Arch Histol Jap 49:459–481

    PubMed  CAS  Google Scholar 

  • Yamamoto T, Wakita M (1990) Initial attachment of principal fibers to the root dentin surface in rat molars. J Periodont Res 25:113–119

    PubMed  Article  CAS  Google Scholar 

  • Yamamoto T, Domon T, Takahashi S, Islam MN, Suzuki R, Wakita M (2000a) The structure of the cemento–dentinal junction in rat molars. Ann Anat 182:185–190

    CAS  Google Scholar 

  • Yamamoto T, Domon T, Takahashi S, Islam MN, Suzuki R (2000b) The fibrillar structure of cementum and dentin at the cemento–dentinal junction in rat molars. Ann Anat 182:499–503

    CAS  Article  Google Scholar 

  • Yamamoto T, Domon T, Takahashi S, Islam MN, Suzuki R (2001) The initial attachment of cemental fibrils to the root dentin surface in acellular and cellular cementogenesis in rat molars. Ann Anat 183:123–128

    PubMed  CAS  Google Scholar 

  • Yamamoto T, Domon T, Takahashi S, Arambawatta AKS, Wakita M (2004) Immunolocalization of proteoglycans and bone-related noncollagenous glycoproteins in developing acellular cementum of rat molars. Cell Tissue Res 317:299–312

    PubMed  Article  CAS  Google Scholar 

  • Yamamoto T, Domon T, Takahashi S, Arambawatta AKS, Anjuman KAY, Fukushima C Wakita M (2005) Determination of two different types of cellular cementogenesis in rat molars: a histological and immunohistochemical study. Mat Biol 24:295–305

    Article  CAS  Google Scholar 

  • Yoshiki S, Umeda T, Kurihashi Y (1972) An effective reactivation of alkaline phosphatase in hard tissues completely decalcified for light and electron microscopy. Histochemie 29:296–304

    PubMed  CAS  Google Scholar 

  • Zeichner-David M, Oishi K, Su Z, Zakartchenco V, Chen L-S, Arzate H, Bringas P. Jr (2003) Role of Hertwig’s epithelial root sheath cells in tooth root development. Dev Dyn 228:651–663

    PubMed  Article  CAS  Google Scholar 

Download references

Acknowledgments

This study was supported by a Grant for Scientific Research from the Japanese Ministry of Education, Culture, Sports, Science and Technology (No.17591900).

Author information

Affiliations

  1. Division of Oral Health Science, Hokkaido University Graduate School of Dental Medicine, Kita 13 Nishi 7 Kita-ku, Sapporo, 060-8586, Japan

    Tsuneyuki Yamamoto, Shigeru Takahashi, Khan Ara Yasmin Anjuman, Chifumi Fukushima & Minoru Wakita

  2. Division of Oral Functional Science, Hokkaido University Graduate School of Dental Medicine, Kita 13 Nishi 7 Kita-ku, Sapporo, 060-8586, Japan

    Takanori Domon

Authors
  1. Tsuneyuki Yamamoto
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Takanori Domon
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Shigeru Takahashi
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Khan Ara Yasmin Anjuman
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Chifumi Fukushima
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Minoru Wakita
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tsuneyuki Yamamoto.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Yamamoto, T., Domon, T., Takahashi, S. et al. Mineralization process during acellular cementogenesis in rat molars: a histochemical and immunohistochemical study using fresh-frozen sections. Histochem Cell Biol 127, 303–311 (2007). https://doi.org/10.1007/s00418-006-0242-x

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00418-006-0242-x

Keywords

  • Acellular cementum
  • Fresh-frozen sections
  • Matrix vesicles
  • Mineralization
  • Tissue non-specific alkaline phosphatase