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Enhancement of calcification by osteoblasts cultured on hydroxyapatite surfaces with adsorbed inorganic polyphosphate

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Abstract

Inorganic polyphosphate has been expected to accelerate bone regeneration. However, there are limited evidences to prove that polyphosphate adsorbed on the surface of a hydroxyapatite plate enhances calcification of cultured osteoblasts. In this study, we examined the effect of polyphosphate adsorbed onto the surface of a hydroxyapatite plate on the attachment, proliferation, differentiation, and calcification of osteoblasts. After hydroxyapatite plates were soaked in solutions of polyphosphate, the plate surfaces were analyzed by scanning electron microscopy and toluidine blue staining to confirm adsorption of polyphosphate. The hydroxyapatite plates were further subjected to the measurements of surface roughness, water contact angle, and the binding capacity of calcium ions. Cell culture experiments were carried out using MC3T3-E1 pre-osteoblastic cells. It was found that soaking a hydroxyapatite plate in a polyphosphate solution gave rise to an increase in surface roughness and reduction in water contact angle in a concentration-dependent manner, suggesting the adsorption of polyphosphate onto the surface of a hydroxyapatite plate. It was further observed that surface-adsorbed polyphosphate exhibited an inhibitory effect on cell adhesion and proliferation. In contrast, cell differentiation was promoted on hydroxyapatite plates with adsorbed polyphosphate, when assessed from expression of differentiation marker genes including alkaline phosphatase, osteopontin, and osteocalcin. In addition, calcification of the culture was enhanced on hydroxyapatite plates with relatively low density of adsorbed polyphosphate. Our results as a whole provided an evidence to show that there is a narrow window with regard to the surface density of adsorbed polyphosphate for the enhancement of osteoblast calcification.

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References

  • Arends J, Jongebloed WL (1977) Dislocations and dissolution in apatites: theoretical considerations. Caries Res 11(3):186–188

    Article  PubMed  CAS  Google Scholar 

  • Arima Y, Iwata H (2007) Effects of surface functional groups on protein adsorption and subsequent cell adhesion using self-assembled monolayers. J Mater Chem 17:4079–4087

    Article  CAS  Google Scholar 

  • Beck GR Jr, Zerler B, Moran E (2000) Phosphate is a specific signal for induction of osteopontin gene expression. Proc Natl Acad Sci U S A 97: 8352–8357

  • Caswell AM, Russell RG (1988) Evidence that ecto-nucleosidetriphosphate pyrophosphatase serves in the generation of extracellular inorganic pyrophosphate in human bone and articular cartilage. Biochim Biophys Acta 966:310–317

    Article  PubMed  CAS  Google Scholar 

  • Doi K, Kubo T, Takeshita R, Kajihara S, Kato S, Kawazoe Y, Shiba T, Akagawa Y (2014) Inorganic polyphosphate adsorbed onto hydroxyapatite for guided bone regeneration: an animal study. Dent Mater J 33:179–186

    Article  PubMed  CAS  Google Scholar 

  • Dunn T, Gable K, Beeler T (1994) Regulation of cellular Ca2+ by yeast vacuoles. J Biol Chem 269:7273–7278

    PubMed  CAS  Google Scholar 

  • Hoac B, Kiffer-Moreira T, Millán JL, McKee MD (2013) Polyphosphates inhibit extracellular matrix mineralization in MC3T3-E1 osteoblast cultures. Bone 53:478–486

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Inaba M (2002) Reguration of adhesion molecules by calcium. Clin Calcium 12:583–586

    PubMed  CAS  Google Scholar 

  • Kawazoe Y, Shiba T, Nakamura R, Mizuno A, Tsutsumi K, Uematsu T, Yamaoka M, Shindoh M, Kohgo T (2004) Induction of calcification in MC3T3-E1 cells by inorganic polyphosphate. J Dent Res 83:613–618

    Article  PubMed  CAS  Google Scholar 

  • Leyhausen G, Lorenz B, Zhu H, Geurtsen W, Bohnensack R, Müller WE, Schröder HC (1998) Inorganic polyphosphate in human osteoblast-like cells. J Bone Miner Res 13:803–812

    Article  PubMed  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2–ΔΔCt method. Methods 25:402–408

    Article  PubMed  CAS  Google Scholar 

  • Morita K, Doi K, Kubo T, Takeshita R, Kato S, Shiba T, Akagawa Y (2010) Enhanced initial bone regeneration with inorganic polyphosphate-adsorbed hydroxyapatite. Acta Biomater 6:2808–2815

    Article  PubMed  CAS  Google Scholar 

  • Nakamura M, Sekijima Y, Nakamura S, Kobayashi T, Niwa K, Yamashita K (2006) Role of blood coagulation components as intermediators of high osteoconductivity of electrically polarized hydroxyapatite. J Biomed Mater Res Part A 79A(3):627–634

    Article  CAS  Google Scholar 

  • Nilen RWN, Richter PW (2008) The thermal stability of hydroxyapatite in biphasic calcium phosphate ceramics. J Mater Sci Mater Med 19(4):1693–1702

    Article  PubMed  CAS  Google Scholar 

  • Nørgaard R, Kassem M, Rattan SI (2006) Heat shock-induced enhancement of osteoblastic differentiation of hTERT-immortalized mesenchymal stem cells. Ann N Y Acad Sci 48:207–214

    Google Scholar 

  • Puleo DA, Nanci A (1999) Understanding and controlling the bone–implant interface. Biomaterials 20:2311–2321

    Article  PubMed  CAS  Google Scholar 

  • Schliephake H, Dard M, Planck H, Hierlemann H, Stern U (2000) Alveolar ridge repair using resorbable membranes and autogenous bone particles with simultaneous placement of implant: an experimental pilot study in dogs. Int J Oral Maxillofac Implants 15:364–373

    PubMed  CAS  Google Scholar 

  • Schröder HC, Lorenz B, Kurz L, Müller WE (1999) Inorganic polyphosphate in eukaryotes: enzymes, metabolism and function. Prog Mol Subcell Biol 23:45–81

    Article  PubMed  Google Scholar 

  • Shiba T, Nishimura D, Kawazoe Y, Onodera Y, Tsutsumi K, Nakamura R, Ohshiro M (2003) Modulation of mitogenic activity of fibroblast growth factors by inorganic polyphosphate. J Biol Chem 278:26788–26792

    Article  PubMed  CAS  Google Scholar 

  • Tsutsumi K, Saito N, Kawazoe Y, Ooi HK, Shiba T (2014) Morphogenetic study on the maturation of osteoblastic cell as induced by inorganic polyphosphate. PLoS One 9:e86834

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Wang X, Schröder HC, Feng Q, Diehl-Seifert B, Grebenjuk VA, Müller WE (2014) Isoquercitrin and polyphosphate co-enhance mineralization of human osteoblast-like SaOS-2 cells via separate activation of two RUNX2 cofactors AFT6 and Ets1. Biochem Pharmacol 89:413–421

    Article  PubMed  CAS  Google Scholar 

  • Wu AT, Aoki T, Sakoda M, Ohta S, Ichimura S, Ito T, Ushida T, Furukawa KS (2015) Enhancing osteogenic differentiation of MC3T3-E1 cells by immobilizing inorganic polyphosphateonto hyaluronic acid hydrogel. Biomacromolecules 16:166–173

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

This study was supported by a Grant-in-Aid for Scientific Research from the National Institute of Biomedical Innovation in Japan (Project ID 25861845).

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Authors and Affiliations

  1. Department of Advanced Prosthodontics, Graduate School of Biomedical & Health Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima, 734-8553, Japan

    Kan Kato, Koji Morita, Kazuya Doi, Takayasu Kubo & Kazuhiro Tsuga

  2. Department of Biomaterials, Graduate School of Biomedical & Health Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima, 734-8553, Japan

    Isao Hirata & Koichi Kato

Authors
  1. Kan Kato
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  2. Koji Morita
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  3. Isao Hirata
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  4. Kazuya Doi
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  5. Takayasu Kubo
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  6. Koichi Kato
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  7. Kazuhiro Tsuga
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Correspondence to Kan Kato.

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Editor: Tetsuji Okamoto

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Kato, K., Morita, K., Hirata, I. et al. Enhancement of calcification by osteoblasts cultured on hydroxyapatite surfaces with adsorbed inorganic polyphosphate. In Vitro Cell.Dev.Biol.-Animal 54, 449–457 (2018). https://doi.org/10.1007/s11626-018-0257-3

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  • DOI: https://doi.org/10.1007/s11626-018-0257-3

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