Skip to main content

Advertisement

SpringerLink
Log in

Hydrostatic Pressure Enhances Chondrogenic Differentiation of Human Bone Marrow Stromal Cells in Osteochondrogenic Medium

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

Abstract

This study demonstrated the chondrogenic effect of hydrostatic pressure on human bone marrow stromal cells (MSCs) cultured in a mixed medium containing osteogenic and chondrogenic factors. MSCs seeded in type I collagen sponges were exposed to 1 MPa of intermittent hydrostatic pressure at a frequency of 1 Hz for 4 h per day for 10 days, or remained in identical culture conditions but without exposure to pressure. Afterwards, we compared the proteoglycan content of loaded and control cell/scaffold constructs with Alcian blue staining. We also used real-time PCR to evaluate the change in mRNA expression of selected genes associated with chondrogenic and osteogenic differentiation (aggrecan, type I collagen, type II collagen, Runx2 (Cbfa-1), Sox9, and TGF-β1). With the hydrostatic pressure loading regime, proteoglycan staining increased markedly. Correspondingly, the mRNA expression of chondrogenic genes such as aggrecan, type II collagen, and Sox9 increased significantly. We also saw a significant increase in the mRNA expression of type I collagen, but no change in the expression of Runx2 or TGF-β1 mRNA. This study demonstrated that hydrostatic pressure enhanced differentiation of MSCs in the presence of multipotent differentiation factors in vitro, and suggests the critical role that this loading regime may play during cartilage development and regeneration in vivo.

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

Access this article

Log in via an institution

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

Instant access to the full article PDF.

Institutional subscriptions

FIGURE 1
FIGURE 2
FIGURE 3

Similar content being viewed by others

References

  1. Angele P., D. Schumann, M. Angele, B. Kinner, C. Englert, R. Hente, B. Fuchtmeier, M. Nerlich, C. Neumann, R. Kujat 2004. Cyclic, mechanical compression enhances chondrogenesis of mesenchymal progenitor cells in tissue engineering scaffolds. Biorheology 41:335–346

    PubMed  CAS  Google Scholar 

  2. Angele P., J. U. Yoo, C. Smith, J. Mansour, K. J. Jepsen, M. Nerlich, B. Johnstone 2003. Cyclic hydrostatic pressure enhances the chondrogenic phenotype of human mesenchymal progenitor cells differentiated in vitro. J. Orthop. Res. 21:451–457

    Article  PubMed  CAS  Google Scholar 

  3. Barbero A., S. Grogan, D. Schafer, M. Heberer, P. Mainil-Varlet, I. Martin 2004). Age related changes in human articular chondrocyte yield, proliferation and post-expansion chondrogenic capacity. Osteoarthr. Cartil. 12:476–484

    Article  PubMed  Google Scholar 

  4. Barry F., R. E. Boynton, B. Liu, J. M. Murphy 2001. Chondrogenic differentiation of mesenchymal stem cells from bone marrow: differentiation-dependent gene expression of matrix components. Exp Cell Res. 268:189–200

    Article  PubMed  CAS  Google Scholar 

  5. Bell D. M., K. K. Leung, S. C. Wheatley, L. J. Ng, S. Zhou, K. W. Ling, M. H. Sham, P. Koopman, P. P. Tam, K. S. Cheah 1997. Sox9 directly regulates the type-II collagen gene. Nat. Genet. 16:174–178

    Article  PubMed  CAS  Google Scholar 

  6. Bosnakovski D., M. Mizuno, G. Kim, S. Takagi, M. Okumura, T. Fujinaga 2006. Chondrogenic differentiation of bovine bone marrow mesenchymal stem cells (MSCs) in different hydrogels: influence of collagen type II extracellular matrix on MSC chondrogenesis. Biotechnol. Bioeng. 93:1152–1163

    Article  PubMed  CAS  Google Scholar 

  7. Campbell J. J., D. A. Lee, D. L. Bader 2006. Dynamic compressive strain influences chondrogenic gene expression in human mesenchymal stem cells. Biorheology 43:455–470

    PubMed  Google Scholar 

  8. Carter, D. R., and G. S. Beaupre. Skeletal tissue regeneration. In: Skeletal Function and Form: Mechanobiology of Skeletal Development, Aging and Regeneration. Cambridge, UK: Cambridge University Press, pp. 161–200, 2001

  9. Carter, D. R., G. S. Beaupre, N. J. Giori, and J. A. Helms. Mechanobiology of skeletal regeneration. Clin. Orthop. Relat. Res. 355:S41–55, 1998

    Google Scholar 

  10. Centrella M., T. L. McCarthy, E. Canalis 1988. Tumor necrosis factor-alpha inhibits collagen synthesis and alkaline phosphatase activity independently of its effect on deoxyribonucleic acid synthesis in osteoblast-enriched bone cell cultures. Endocrinology 123:1442–1448

    Article  PubMed  CAS  Google Scholar 

  11. Chen G., D. Liu, M. Tadokoro, R. Hirochika, H. Ohgushi, J. Tanaka, T. Tateishi 2004. Chondrogenic differentiation of human mesenchymal stem cells cultured in a cobweb-like biodegradable scaffold. Biochem. Biophys. Res. Commun. 322:50–55

    Article  PubMed  CAS  Google Scholar 

  12. Ducy P., R. Zhang, V. Geoffroy, A. L. Ridall, G. Karsenty 1997. Osf2/Cbfa1: a transcriptional activator of osteoblast differentiation. Cell 89:747–754

    Article  PubMed  CAS  Google Scholar 

  13. Enomoto H., M. Enomoto-Iwamoto, M. Iwamoto, S. Nomura, M. Himeno, Y. Kitamura, T. Kishimoto, T. Komori 2000. Cbfa1 is a positive regulatory factor in chondrocyte maturation. J. Biol. Chem. 275:8695–8702

    Article  PubMed  CAS  Google Scholar 

  14. Finger A. R., C. Y. Sargent, K. O. Dulaney, S. H. Bernacki, E. G. Loboa 2007. Differential effects on messenger ribonucleic acid expression by bone marrow-derived human mesenchymal stem cells seeded in agarose constructs due to ramped and steady applications of cyclic hydrostatic pressure. Tissue Eng. 13:1151–1158

    Article  PubMed  CAS  Google Scholar 

  15. Giori N. J., G. S. Beaupre, D. R. Carter 1993. Cellular shape and pressure may mediate mechanical control of tissue composition in tendons. J. Orthop. Res. 11:581–591

    Article  PubMed  CAS  Google Scholar 

  16. Grinnell F. 2000. Fibroblast-collagen-matrix contraction: growth-factor signalling and mechanical loading. Trends Cell. Biol. 10:362–365

    Article  PubMed  CAS  Google Scholar 

  17. Hayami T., Q. Zhang, Y. Kapila, S. Kapila 2007. Dexamethasone’s enhancement of osteoblastic markers in human periodontal ligament cells is associated with inhibition of collagenase expression. Bone 40:93–104

    Article  PubMed  CAS  Google Scholar 

  18. Healy C., D. Uwanogho, P. T. Sharpe 1999. Regulation and role of Sox9 in cartilage formation. Dev. Dyn. 215:69–78

    Article  PubMed  CAS  Google Scholar 

  19. Huang C. Y., K. L. Hagar, L. E. Frost, Y. Sun, H. S. Cheung 2004. Effects of cyclic compressive loading on chondrogenesis of rabbit bone-marrow derived mesenchymal stem cells. Stem Cells 22:313–323

    Article  PubMed  CAS  Google Scholar 

  20. Ikenoue T., M. C. Trindade, M. S. Lee, E. Y. Lin, D. J. Schurman, S. B. Goodman, R. L. Smith 2003. Mechanoregulation of human articular chondrocyte aggrecan and type II collagen expression by intermittent hydrostatic pressure in vitro. J. Orthop. Res. 21:110–116

    Article  PubMed  CAS  Google Scholar 

  21. Johnstone B., T. M. Hering, A. I. Caplan, V. M. Goldberg, J. U. Yoo 1998. In vitro chondrogenesis of bone marrow-derived mesenchymal progenitor cells. Exp. Cell Res. 238:265–272

    Article  PubMed  CAS  Google Scholar 

  22. Kimura, T., N. Yasui, S. Ohsawa, and K. Ono. Chondrocytes embedded in collagen gels maintain cartilage phenotype during long-term cultures. Clin. Orthop. Relat. Res. 186:231–239, 1984

    PubMed  CAS  Google Scholar 

  23. Kolodney M. S., R. B. Wysolmerski 1992. Isometric contraction by fibroblasts and endothelial cells in tissue culture: a quantitative study. J. Cell Biol. 117:73–82

    Article  PubMed  CAS  Google Scholar 

  24. Kuznetsov S. A., M. H. Mankani, P. G. Robey 2000. Effect of serum on human bone marrow stromal cells: ex vivo expansion and in vivo bone formation. Transplantation 70:1780–1787

    Article  PubMed  CAS  Google Scholar 

  25. Lawrence R. C., C. G. Helmick, F. C. Arnett, R. A. Deyo, D. T. Felson, E. H. Giannini, S. P. Heyse, R. Hirsch, M. C. Hochberg, G. G. Hunder, M. H. Liang, S. R. Pillemer, V. D. Steen, F. Wolfe 1998. Estimates of the prevalence of arthritis and selected musculoskeletal disorders in the United States. Arthritis Rheum. 41:778–799

    Article  PubMed  CAS  Google Scholar 

  26. Lee C. R., H. A. Breinan, S. Nehrer, M. Spector 2000. Articular cartilage chondrocytes in type I and type II collagen-gag matrices exhibit contractile behavior in vitro. Tissue Eng. 6:555–565

    Article  PubMed  CAS  Google Scholar 

  27. Lennon D. P., S. E. Haynesworth, R. G. Young, J. E. Dennis, A. I. Caplan 1995. A chemically defined medium supports in vitro proliferation and maintains the osteochondral potential of rat marrow-derived mesenchymal stem cells. Exp. Cell Res. 219:211–222

    Article  PubMed  CAS  Google Scholar 

  28. Li K. W., D. P. Lindsey, D. R. Wagner, N. J. Giori, D. J. Schurman, S. B. Goodman, R. L. Smith, D. R. Carter, G. S. Beaupre 2006. Gene regulation ex vivo within a wrap-around tendon. Tissue Eng. 12:2611–2618

    Article  PubMed  CAS  Google Scholar 

  29. Lieb E., T. Vogel, S. Milz, M. Dauner, M. B. Schulz 2004. Effects of transforming growth factor beta1 on bonelike tissue formation in three-dimensional cell culture. II: osteoblastic differentiation. Tissue Eng. 10:1414–1425

    PubMed  CAS  Google Scholar 

  30. Mankin H. J. 1974. The reaction of articular cartilage to injury and osteoarthritis (first of two parts). N. Engl. J. Med. 291:1285–1292

    Article  PubMed  CAS  Google Scholar 

  31. Massague J. 1990. The transforming growth factor-beta family. Annu. Rev. Cell Biol. 6:597–641

    Article  PubMed  CAS  Google Scholar 

  32. Mauck R. L., B. A. Byers, X. Yuan, R. S. Tuan 2007. Regulation of cartilaginous ECM gene transcription by chondrocytes and MSCs in 3d culture in response to dynamic loading. Biomech. Model Mechanobiol. 6:113–125

    Article  PubMed  CAS  Google Scholar 

  33. McBeath R., D. M. Pirone, C. M. Nelson, K. Bhadriraju, C. S. Chen 2004. Cell shape, cytoskeletal tension, and RhoA regulate stem cell lineage commitment. Dev. Cell. 6:483–495

    Article  PubMed  CAS  Google Scholar 

  34. Miyanishi, K., M. C. Trindade, D. P. Lindsey, G. S. Beaupre, D. R. Carter, S. B. Goodman, D. J. Schurman, and R. L. Smith. Dose- and time-dependent effects of cyclic hydrostatic pressure on transforming growth factor-beta3-induced chondrogenesis by adult human mesenchymal stem cells in vitro. Tissue Eng. 12(8):2253–2262, 2006

    Article  PubMed  CAS  Google Scholar 

  35. Miyanishi, K., M. C. Trindade, D. P. Lindsey, G. S. Beaupre, D. R. Carter, S. B. Goodman, D. J. Schurman, and R. L. Smith. Effects of hydrostatic pressure and transforming growth factor-beta3 on adult human mesenchymal stem cell chondrogenesis in vitro. Tissue Eng. 12(6):1419–1428, 2006

    Article  PubMed  CAS  Google Scholar 

  36. Mouw J. K., J. T. Connelly, C. G. Wilson, K. E. Michael, M. E. Levenston 2007. Dynamic compression regulates the expression and synthesis of chondrocyte-specific matrix molecules in bone marrow stromal cells. Stem Cells 25:655–663

    Article  PubMed  CAS  Google Scholar 

  37. Otto F., A. P. Thornell, T. Crompton, A. Denzel, K. C. Gilmour, I. R. Rosewell, G. W. Stamp, R. S. Beddington, S. Mundlos, B. R. Olsen, P. B. Selby, M. J. Owen 1997. Cbfa1, a candidate gene for cleidocranial dysplasia syndrome, is essential for osteoblast differentiation and bone development. Cell 89:765–771

    Article  PubMed  CAS  Google Scholar 

  38. Pauwels, F. Theoretical foundation. In: Biomechanics of the Normal and Diseased Hip: Theoretical Foundation, Technique and Results of Treatment: An atlas. New York: Springer-Verlag, 1976, pp. 1–37

  39. Roche S., M. C. Ronziere, D. Herbage, A. M. Freyria 2001. Native and DPPA cross-linked collagen sponges seeded with fetal bovine epiphyseal chondrocytes used for cartilage tissue engineering. Biomaterials 22:9–18

    Article  PubMed  CAS  Google Scholar 

  40. Scherer K., M. Schunke, R. Sellckau, J. Hassenpflug, B. Kurz 2004. The influence of oxygen and hydrostatic pressure on articular chondrocytes and adherent bone marrow cells in vitro. Biorheology 41:323–333

    PubMed  CAS  Google Scholar 

  41. Shapiro F., S. Koide, M. J. Glimcher 1993. Cell origin and differentiation in the repair of full-thickness defects of articular cartilage J. Bone Joint Surg. Am. 75:532–553

    PubMed  CAS  Google Scholar 

  42. Silva W. A. Jr., D. T. Covas, R. A. Panepucci, R. Proto-Siqueira, J. L. Siufi, D. L. Zanette, A. R. Santos, M. A. Zago 2003. The profile of gene expression of human marrow mesenchymal stem cells. Stem Cells 21:661–669

    Article  PubMed  CAS  Google Scholar 

  43. Smith R. L., J. Lin, M. C. Trindade, J. Shida, G. Kajiyama, T. Vu, A. R. Hoffman, M. C. van der Meulen, S. B. Goodman, D. J. Schurman, D. R. Carter 2000. Time-dependent effects of intermittent hydrostatic pressure on articular chondrocyte type II collagen and aggrecan mRNA expression. J. Rehabil. Res. Dev. 37:153–161

    PubMed  CAS  Google Scholar 

  44. Smith R. L., S. F. Rusk, B. E. Ellison, P. Wessells, K. Tsuchiya, D. R. Carter, W. E. Caler, L. J. Sandell, D. J. Schurman 1996. In vitro stimulation of articular chondrocyte mRNA and extracellular matrix synthesis by hydrostatic pressure. J. Orthop. Res. 14:53–60

    Article  PubMed  CAS  Google Scholar 

  45. Staples A. K., A. H. Hsieh, J. C. Lotz 2006. Differential Effects of Mechanically and Biologically-Initiated Differentiation of Human Mesenchymal Stem Cell Mechanotransduction. Orthopaedic Research Society, Chicago IL

    Google Scholar 

  46. Takahashi I., G. H. Nuckolls, K. Takahashi, O. Tanaka, I. Semba, R. Dashner, L. Shum, H. C. Slavkin 1998. Compressive force promotes Sox9, type II collagen and aggrecan and inhibits IL-1beta expression resulting in chondrogenesis in mouse embryonic limb bud mesenchymal cells. J. Cell Sci. 111(Pt 14):2067–2076

    PubMed  CAS  Google Scholar 

  47. Wang Y., D. J. Blasioli, H. J. Kim, H. S. Kim, D. L. Kaplan 2006. Cartilage tissue engineering with silk scaffolds and human articular chondrocytes. Biomaterials 27:4434–4442

    Article  PubMed  CAS  Google Scholar 

  48. Wassersug R. J. 1976. A procedure for differential staining of cartilage and bone in whole formalin-fixed vertebrates. Stain Technol. 51:131–134

    PubMed  CAS  Google Scholar 

  49. Wong M., M. Siegrist, K. Goodwin 2003. Cyclic tensile strain and cyclic hydrostatic pressure differentially regulate expression of hypertrophic markers in primary chondrocytes. Bone 33:685–693

    Article  PubMed  CAS  Google Scholar 

  50. Woods A., F. Beier 2006. RhoA/Rock signaling regulates chondrogenesis in a context-dependent manner. J. Biol. Chem. 281:13134–13140

    Article  PubMed  CAS  Google Scholar 

  51. Yoo, J. U., and B. Johnstone. The role of osteochondral progenitor cells in fracture repair. Clin. Orthop. Relat. Res. 355S:S73–81, 1998

    Article  PubMed  Google Scholar 

  52. Zhao Q., H. Eberspaecher, V. Lefebvre, B. De Crombrugghe 1997. Parallel expression of Sox9 and col2a1 in cells undergoing chondrogenesis. Dev. Dyn. 209:377–386

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The work was supported in part by the Department of Veterans Affairs, Veterans Health Administration, Rehabilitation Research and Development Service; Merit Review grant A2723R (PI: G. S. Beaupre; D. R. Carter); and Research Career Scientist awards to G. S. Beaupre and D. R. Carter. This work was also supported in part by NIH DE14526 and the Oak Foundation to M. T. Longaker. We would like to thank Integra LifeSciences for the generous donation of the collagen I scaffolds and Ron Ingram and Jeff Brittan for their support in procuring the scaffolds. The authors thank Dr. Angelie Agarwal for advice on MSC culture and seeding.

Author information

Authors and Affiliations

  1. Department of Mechanical Engineering, Stanford University, Stanford, CA, USA

    Diane R. Wagner, Sheena E. Chandran, Dennis R. Carter & Gary S. Beaupre

  2. Department of Surgery, Stanford University, Stanford, CA, USA

    Diane R. Wagner & Michael T. Longaker

  3. Aerospace and Mechanical Engineering, 145 Multidisciplinary Research Building, Notre Dame, IN, 46556, USA

    Diane R. Wagner

  4. Bone and Joint Center of Excellence, VA Palo Alto Health Care System, Palo Alto, CA, USA

    Derek P. Lindsey, Kelvin W. Li, Padmaja Tummala, R. Lane Smith, Dennis R. Carter & Gary S. Beaupre

Authors
  1. Diane R. Wagner
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Derek P. Lindsey
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kelvin W. Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Padmaja Tummala
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sheena E. Chandran
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. R. Lane Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Michael T. Longaker
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Dennis R. Carter
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Gary S. Beaupre
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Diane R. Wagner.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Wagner, D.R., Lindsey, D.P., Li, K.W. et al. Hydrostatic Pressure Enhances Chondrogenic Differentiation of Human Bone Marrow Stromal Cells in Osteochondrogenic Medium. Ann Biomed Eng 36, 813–820 (2008). https://doi.org/10.1007/s10439-008-9448-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10439-008-9448-5

Keywords

Search

Navigation