Skip to main content

Advertisement

Log in

Attachment, Spreading, and Adhesion Strength of Human Bone Marrow Cells on Chitosan

  • Published:
Annals of Biomedical Engineering Aims and scope Submit manuscript

Abstract

The successful integration of an orthopedic implant into bone depends on the mechanisms at the tissue–implant interface and mostly on the osteoblast attachment phenomenon. Chitosan has emerged as an attractive biomacromolecule favoring osseointegration. In this study highly deacetylated chitosan coatings, with roughness of about 1 nm, were bonded to glass surfaces via silane–glutaraldehyde molecules. Human osteoblasts were used to study the development of attachment during the first 60 min. Chitosan favored the number of the attached cells compared to the uncoated surfaces for 30 min seeding time (t s). For t s up to 60 min the attached cell area was almost 210% significantly higher on the chitosan surfaces, indicating an enhanced spreading process. To determine the cell attachment strength, a micropipette aspiration method was used, where the value of the term I = ∫Fdt is representative of the single cell attachment–adhesion procedure and quantitatively reflects the strength evolution during attachment: F equals the detaching force applied on the cell. The results showed higher strength values on the chitosan surfaces. The findings reinforce the favorable environment of the biomacromolecule for the osteoblast and the new approach regarding the quantitatively evaluation of adhesion provides important contribution for the study of cell–material interaction, especially during the crucial first phase of cell attachment.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5
Figure 6
Figure 7
Figure 8
Figure 9

Similar content being viewed by others

References

  1. Anselme, K. Osteoblast adhesion on biomaterials. Biomaterials 21(7):667–681, 2000.

    Article  CAS  PubMed  Google Scholar 

  2. Athanassiou, G., and D. Deligianni. Adhesion strength of individual human bone marrow cells to fibronectin. Integrin beta1-mediated adhesion. J. Mater. Sci. Mater. Med. 12(10–12):965–970, 2001.

    Article  CAS  PubMed  Google Scholar 

  3. Bumgardner, J. D., R. Wiser, P. D. Gerard, P. Bergin, B. Chestnutt, M. Marin, V. Ramsey, S. H. Elder, and J. A. Gilbert. Chitosan: potential use as a bioactive coating for orthopaedic and craniofacial/dental implants. J. Biomater. Sci. Polym. Ed. 14(5):423–438, 2003.

    Article  CAS  PubMed  Google Scholar 

  4. Chellat, F., M. Tabrizian, S. Dumitriu, E. Chornet, C. H. Rivard, and L. Yahia. Study of biodegradation behavior of chitosan–xanthan microspheres in simulated physiological media. J. Biomed. Mater. Res. 53(5):592–599, 2000.

    Article  CAS  PubMed  Google Scholar 

  5. Cretel, E., A. Pierres, A. M. Benoliel, and P. Bongrand. How cells feel their environment: a focus on early dynamic events. Cell. Mol. Bioeng. 1(1):5–14, 2008.

    Article  PubMed  Google Scholar 

  6. Custódio, C. A., C. M. Alves, R. L. Reis, and J. F. Mano. Immobilization of fibronectin in chitosan substrates improves cell adhesion and proliferation. J. Tissue Eng. Regen. Med. 4(4):316–323, 2010.

    Article  PubMed  Google Scholar 

  7. Dalby, M. J., S. J. Yarwood, M. O. Riehle, J. H. Johnstone, S. Affrossman, and A. S. Curtis. Increasing fibroblast response to materials using nanotopography: morphological and genetic measurements of cell response to 13-nm-high polymer demixed islands. Exp. Cell Res. 276:1–9, 2002.

    Article  CAS  PubMed  Google Scholar 

  8. Di Martino, A., M. Sittinger, and M. V. Risbud. Chitosan: a versatile polymer for orthopaedic tissue-engineering. Biomaterials 26(30):5983–5990, 2005 (Review).

    Article  CAS  PubMed  Google Scholar 

  9. Fakhry, A., G. B. Schneider, R. Zaharias, and S. Senel. Chitosan supports the initial attachment and spreading of osteoblasts preferentially over fibroblasts. Biomaterials 25:2075–2079, 2004.

    Article  CAS  PubMed  Google Scholar 

  10. Fang, N., Z. Aiping, M. B. Chan-Park, and V. Chan. Adhesion contact dynamics of fibroblasts on biomacromolecular surfaces. Macromol. Biosci. 5:1022–1031, 2005.

    Article  CAS  PubMed  Google Scholar 

  11. Filho, F. C. S., and G. C. Menezes. Osteoblasts attachment and adhesion: how bone cells fit fibronectin-coated surfaces. Mater. Sci. Eng. C 24:637–641, 2004.

    Article  Google Scholar 

  12. Heinemman, C., S. Heinemann, A. Bernhardt, H. Worch, and T. Hanke. Novel textile chitosan scaffolds promote spreading, proliferation and differentiation of osteoblasts. Biomacromolecules 9(10):2913–2920, 2008.

    Article  Google Scholar 

  13. Jayakumar, R., V. V. Divya Rani, K. T. Shalumon, P. T. Kumar, S. V. Nair, T. Furuike, and H. Tamura. Bioactive and osteoblast cell attachment studies of novel α- and β-chitin membranes for tissue engineering applications. Int. J. Biol. Macromol. 45:260–264, 2009.

    Article  CAS  PubMed  Google Scholar 

  14. Kennedy, S. B., N. R. Washburn, C. G. Simon, Jr., and E. J. Armis. Combinatorial screen of the effect of surface energy on fibronectin-mediated osteoblast adhesion, spreading and proliferation. Biomaterials 27(20):3817–3824, 2006.

    Article  CAS  PubMed  Google Scholar 

  15. Klokkevold, P. R., L. Vandermark, E. B. Kenney, and G. W. Bernard. Osteogenesis enhanced by chitosan (poly-n-acetyl glucosaminoglycan) in vitro. J. Periodontol. 67:1170–1175, 1996.

    CAS  PubMed  Google Scholar 

  16. Klozov, M. M., M. Winterhalten, and D. Lerche. Elastic properties of strongly curved monolayers. Effect of electric surface charges. J. Phys. II 2(2):175–185, 1992.

    Google Scholar 

  17. LeBaron, R. G., and K. A. Athanasiou. Extracellular matrix cell adhesion peptides: functional applications in orthopaedic materials. Tissue Eng. 6(2):85–103, 2000.

    Article  CAS  PubMed  Google Scholar 

  18. Lim, J. Y., J. C. Hansen, C. A. Siedlecki, R. W. Hengstebeck, J. Cheng, N. Winograd, and H. J. Donahue. Osteoblast adhesion on poly(L-lactic acid)/polystyrene demixed thin film blends: effect of nanotopography, surface chemistry and wettability. Biomacromolecules 6(6):3319–3327, 2005.

    Article  CAS  PubMed  Google Scholar 

  19. Lim, J. Y., A. D. Dreiss, Z. Zhou, J. C. Hansen, C. A. Siedlecki, R. W. Hengstebeck, J. Cheng, N. Winograd, and H. J. Donahue. The regulation of integrin-mediated osteoblast focal adhesion and focal adhesion kinase expression by nanoscale topography. Biomaterials 28(10):1787–1797, 2007.

    Article  CAS  PubMed  Google Scholar 

  20. Lopez-Perez, P. M., R. M. P. da Silva, C. Serra, I. Pashkuleva, and R. L. Reis. Surface phosphorylation of chitosan significantly improves osteoblast cell viability, attachment and proliferation. J. Mater. Chem. 20:483–491, 2010.

    Article  CAS  Google Scholar 

  21. Mao, J. S., Y. L. Cui, X. H. Wang, Y. Sun, Y. J. Yin, H. M. Zhao, and K. D. De Yao. A preliminary study on chitosan and gelatin polyelectrolyte complex cytocompatibility by cell cycle and apoptosis analysis. Biomaterials 25:3973–3981, 2004.

    Article  CAS  PubMed  Google Scholar 

  22. Meyer, U., A. Büchter, H. P. Wiesmann, U. Joos, and D. B. Jones. Basic reactions of osteoblasts on structured material surfaces. Eur. Cell Mater. 9:39–49, 2005.

    CAS  PubMed  Google Scholar 

  23. Nagahama, H., V. V. Rani, K. T. Shalumon, R. Jayakumar, S. V. Nair, S. Koiwa, T. Furuike, and H. Tamura. Preparation, characterization, bioactive and cell attachment studies of α-chitin/gelatin composite membranes. Int. J. Biol. Macromol. 44:333–337, 2009.

    Article  CAS  PubMed  Google Scholar 

  24. Pierres, A., A. M. Benoliel, D. Touchard, and P. Bongrand. How cells tiptoe on adhesive surfaces before sticking. Biophys. J. 94:4114–4122, 2008.

    Article  CAS  PubMed  Google Scholar 

  25. Prasitslip, M., R. Jenwithisuk, K. Kongsuwan, N. Damrongchai, and P. Watts. Cellular responses to chitosan in vitro: the importance of deacetylation. J. Mater. Sci. Mater. Med. 11:773–778, 2000.

    Article  Google Scholar 

  26. Seol, Y. J., J. Y. Lee, Y. J. Park, Y. M. Lee, I. Young-Ku, C. Rhyu, S. J. Lee, S. B. Han, and C. P. Chung. Chitosan sponges as tissue engineering scaffolds for bone formation. Biotechnol. Lett. 26(13):1037–1041, 2004.

    Article  CAS  PubMed  Google Scholar 

  27. Suh, J. K., and H. W. Matthew. Application of chitosan-based polysaccharide biomaterials in cartilage tissue engineering: a review. Biomaterials 21(24):2589–2598, 2000.

    Article  CAS  PubMed  Google Scholar 

  28. Sung, K. L., M. K. Kwan, F. Maldonado, and W. H. Akeson. Adhesion strength of human ligament fibroblasts. J. Biomech. Eng. 116(3):237–242, 1994.

    Article  CAS  PubMed  Google Scholar 

  29. Tohimata, K., and Y. Ikada. In vitro and in vivo degradation of films of chitin and its deacetylated derivatives. Biomaterials 18(7):567–575, 1997.

    Article  Google Scholar 

  30. Verma, D., K. S. Katti, and D. R. Katti. Osteoblast adhesion, proliferation and growth on polyelectrolyte complex–hydroxyapatite nanocomposites. Philos. Transact. A Math. Phys. Eng. Sci. 368(1917):2083–2097, 2010.

    Article  CAS  PubMed  Google Scholar 

  31. Vitte, J., A. M. Benoliel, A. Pierres, and P. Bongrand. Regulation of cell adhesion. Clin. Hemorheol. Microcirc. 33:167–188, 2005.

    CAS  PubMed  Google Scholar 

  32. Wildt, B., D. Wirtz, and P. C. Searson. Programmed subcellular release for studying the dynamics of cell detachment. Nat. Methods. 6(3):211–213, 2009.

    Article  CAS  PubMed  Google Scholar 

  33. Wilson, C. J., R. E. Clegg, D. I. Leavesley, and M. J. Pearcy. Mediation of biomaterial–cell interactions by adsorbed proteins: a review. Tissue Eng. 11(1–2):1–18, 2005.

    Article  CAS  PubMed  Google Scholar 

  34. Yin, C., K. Liao, H. Q. Mao, K. W. Leong, R. X. Zhuo, and V. Chan. Adhesion contact dynamics of HepG2 cells on galactose-immobilized substrates. Biomaterials 24:837–850, 2003.

    Article  CAS  PubMed  Google Scholar 

  35. Yuan, Y., B. M. Chesnutt, L. Wright, W. O. Haggard, and J. D. Bumgardner. Mechanical property, degradation rate, and bone cell growth of chitosan coated titanium influenced by degree of deacetylation of chitosan. J. Biomed. Mater. Res. B Appl. Biomater. 86(1):245–252, 2008.

    PubMed  Google Scholar 

  36. Zheng, Z., L. Zhang, L. Kong, A. Wang, Y. Gong, and X. Zhang. The behavior of MC3T3-E1 cells on chitosan/poly-L-lysine composite films: Effect of nanotopography, surface chemistry, and wettability. J. Biomed. Mater. Res. 89A:453–465, 2009.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

The authors gratefully thank Korovessis Panagiotis, MD, Chief Orthopaedic Department, General Hospital “Agios Andreas”, Patras, for providing the human bone marrow. The authors would also like to acknowledge Professor Y. F. Missirlis and M. Katsikogianni, PhD, for their contribution regarding the AFM analysis as well as the graduate student P. Patsanis for his valuable help with the SEM images and the Image analysis. Finally, the authors thank Dr. E. Siokou, from the FORTH/ICE-HT, Patras, for the XPS analysis.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to George M. Athanassiou.

Additional information

Associate Editor Eric M. Darling oversaw the review of this article.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Moutzouri, A.G., Athanassiou, G.M. Attachment, Spreading, and Adhesion Strength of Human Bone Marrow Cells on Chitosan. Ann Biomed Eng 39, 730–741 (2011). https://doi.org/10.1007/s10439-010-0188-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10439-010-0188-y

Keywords

Navigation