Skip to main content
SpringerLink
Log in

Synergistic Effects of Conditioned Media and Hydrostatic Pressure on the Differentiation of Mesenchymal Stem Cells

  • Published:
Cellular and Molecular Bioengineering Aims and scope Submit manuscript

Abstract

The objective of this study was to investigate the effect of mechanical stimuli in combination with conditioned media (CM) on the osteogenic and chondrogenic differentiation of mesenchymal stem cells (MSCs). Two “opposing” groups were defined for the study. The first, designated the bone group, was formed by seeding MSCs onto porous poly(l-lactide-co-ε-caprolactone)/hydroxyapatite scaffolds. The second group, designated the cartilage group, was formed by encapsulating MSCs in 2% agarose. The samples were either subjected to hydrostatic pressure (HP), administered CM from the opposing group at a concentration of 50%, or given a combination of the two stimuli. The sulfated glycosaminoglycan (sGAG) content and SOX9, aggrecan, and collagen type II expressions suggest that both osteoblast CM and intermittent HP individually enhanced the chondrogenic differentiation of MSCs. In addition, there is evidence of a synergistic interaction between CM and HP on chondrogenic differentiation. The alkaline phosphatase (ALP) activity and osteocalcin and bone sialoprotein expression suggest that the combination of chondrocyte CM and HP significantly enhanced the osteogenic differentiation of MSCs. However, the individual stimuli did not have a significant effect on osteogenic differentiation. Enzyme-linked immunosorbent assay (ELISA) results revealed that both the chondrocyte CM and osteoblast CM contained considerable amounts of TGF-β1 and BMP-2.

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
Figure 4
Figure 5

Similar content being viewed by others

References

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

    Article  Google Scholar 

  2. Barnes, G. L., P. J. Kostenuik, L. C. Gerstenfeld, and T. A. Einhorn. Growth factor regulation of fracture repair. J. Bone Miner. Res. 14:1805–1815, 1999.

    Article  Google Scholar 

  3. Beris, A. E., M. G. Lykissas, C. D. Papageorgiou, and A. D. Georgoulis. Advances in articular cartilage repair. Injury 36:S14–S23, 2005.

    Article  Google Scholar 

  4. Bismar, H., T. Kloppinger, E. M. Schuster, S. Balbach, I. Diel, R. Ziegler, and J. Pfeilschifter. Transforming growth factor beta (TGF-beta) levels in the conditioned media of human bone cells: relationship to donor age, bone volume, and concentration of TGF-beta in human bone matrix in vivo. Bone 24:565–569, 1999.

    Article  Google Scholar 

  5. Bostrom, M. P. G. Expression of bone morphogenetic proteins in fracture healing. Clin. Orthop. Relat. Res. 355:S116–S123, 1998.

    Article  Google Scholar 

  6. Elder, B. D., and K. A. Athanasiou. Hydrostatic pressure in articular cartilage tissue engineering: from chondrocytes to tissue regeneration. Tissue Eng. B 15:43–53, 2009.

    Article  Google Scholar 

  7. Elder, S. H., S. W. Sanders, W. R. McCulley, M. L. Marr, J. W. Shim, and K. A. Hasty. Chondrocyte response to cyclic hydrostatic pressure in alginate versus pellet culture. J. Orthop. Res. 24:740–747, 2006.

    Article  Google Scholar 

  8. Festuccia, C., M. Bologna, G. L. Gravina, F. Guerra, A. Angelucci, I. Villanova, D. Millimaggi, and A. Teti. Osteoblast conditioned media contain TGF-beta 1 and modulate the migration of prostate tumor cells and their interactions with extracellular matrix components. Int. J. Cancer 81:395–403, 1999.

    Article  Google Scholar 

  9. Gerstenfeld, L. C., G. L. Barnes, C. M. Shea, and T. A. Einhorn. Osteogenic differentiation is selectively promoted by morphogenetic signals from chondrocytes and synergized by a nutrient rich growth environment. Connect. Tissue Res. 44:85–91, 2003.

    Google Scholar 

  10. Gimble, J. M., and M. E. Nuttall. Bone and fat—old questions, new insights. Endocrine 23:183–188, 2004.

    Article  Google Scholar 

  11. Goldring, M. B., K. Tsuchimochi, and K. Ijiri. The control of chondrogenesis. J. Cell. Biochem. 97:33–44, 2006.

    Article  Google Scholar 

  12. Gunja, N. J., R. K. Uthamanthil, and K. A. Athanasiou. Effects of TGF-beta 1 and hydrostatic pressure on meniscus cell-seeded scaffolds. Biomaterials 30:565–573, 2009.

    Article  Google Scholar 

  13. Gunther, M., H. D. Haubeck, E. Vandeleur, J. Blaser, S. Bender, I. Gutgemann, D. C. Fischer, H. Tschesche, H. Greiling, P. C. Heinrich, and L. Graeve. Transforming growth-factor-beta-1 regulates tissue inhibitor of metalloproteinases-1 expression in differentiated human articular chondrocytes. Arthritis Rheum. 37:395–405, 1994.

    Article  Google Scholar 

  14. Hall, A. C. Differential effects of hydrostatic pressure on cation transport pathways of isolated articular chondrocytes. J. Cell. Physiol. 178:197–204, 1999.

    Article  Google Scholar 

  15. Heng, B. C., T. Cao, and E. H. Lee. Directing stem cell differentiation into the chondrogenic lineage in vitro. Stem Cells 22:1152–1167, 2004.

    Article  Google Scholar 

  16. Hunziker, E. B. Articular cartilage repair: basic science and clinical progress. A review of the current status and prospects. Osteoarthritis Cartilage 10:432–463, 2002.

    Article  Google Scholar 

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

    Article  Google Scholar 

  18. Jortikka, M. O., J. J. Parkkinen, R. I. Inkinen, J. Karner, H. T. Jarvelainen, L. O. Nelimarkka, M. I. Tammi, and M. J. Lammi. The role of microtubules in the regulation of proteoglycan synthesis in chondrocytes under hydrostatic pressure. Arch. Biochem. Biophys. 374:172–180, 2000.

    Article  Google Scholar 

  19. Kim, D. H., S. J. Heo, J. W. Shin, S. H. Park, and J. W. Shin. Effects of mechanical stimulation on chondrogenesis of mesenchymal stem cells via 3D co-culturing system. Tissue Eng. Regen. Med. 6:739–745, 2009.

    Google Scholar 

  20. Krampera, M., G. Pizzolo, G. Aprili, and M. Franchini. Mesenchymal stem cells for bone, cartilage, tendon and skeletal muscle repair. Bone 39:678–683, 2006.

    Article  Google Scholar 

  21. Langer, F., A. E. Gross, M. West, and E. P. Urovitz. Immunogenicity of allograft knee–joint transplants. Clin. Orthop. Relat. Res. 132:155–162, 1978.

    Google Scholar 

  22. Langer, R., and J. P. Vacanti. Tissue engineering. Science 260:920–926, 1993.

    Article  Google Scholar 

  23. Lefebvre, V., R. R. Behringer, and B. de Crombrugghe. L-Sox5, Sox6 and Sox9 control essential steps of the chondrocyte differentiation pathway. Osteoarthritis Cartilage 9:S69–S75, 2001.

    Article  Google Scholar 

  24. Liu, J., Z. H. Zhao, J. Li, L. Zou, C. Shuler, Y. W. Zou, X. J. Huang, M. L. Li, and J. Wang. Hydrostatic pressures promote initial osteodifferentiation with ERK1/2 not p38 MAPK signaling involved. J. Cell. Biochem. 107:224–232, 2009.

    Article  Google Scholar 

  25. Majumdar, M. K., E. Wang, and E. A. Morris. BMP-2 and BMP-9 promote chondrogenic differentiation of human multipotential mesenchymal cells and overcome the inhibitory effect of IL-1. J. Cell. Physiol. 189:275–284, 2001.

    Article  Google Scholar 

  26. Maxson, S., and K. J. L. Burg. Conditioned media cause increases in select osteogenic and adipogenic differentiation markers in mesenchymal stem cell cultures. J. Tissue Eng. Regen. Med. 2:147–154, 2008.

    Article  Google Scholar 

  27. Maxson, S., and K. J. L. Burg. Conditioned media enhance osteogenic differentiation on poly(l-lactide-co-epsilon-caprolactone)/hydroxyapatite scaffolds and chondrogenic differentiation in alginate. J. Biomater. Sci. Polym. Ed. 21:1441–1458, 2010.

    Article  Google Scholar 

  28. Miyanishi, K., M. C. D. 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-beta 3 on adult human mesenchymal stem cell chondrogenesis in vitro. Tissue Eng. 12:1419–1428, 2006.

    Article  Google Scholar 

  29. Mizuno, S., T. Tateishi, T. Ushida, and J. Glowacki. Hydrostatic fluid pressure enhances matrix synthesis and accumulation by bovine chondrocytes in three-dimensional culture. J. Cell. Physiol. 193:319–327, 2002.

    Article  Google Scholar 

  30. Orr, D. E., and K. J. L. Burg. Design of a modular bioreactor to incorporate both perfusion flow and hydrostatic compression for tissue engineering applications. Ann. Biomed. Eng. 36:1228–1241, 2008.

    Article  Google Scholar 

  31. Parkkinen, J. J., J. Ikonen, M. J. Lammi, J. Laakkonen, M. Tammi, and H. J. Helminen. Effects of cyclic hydrostatic-pressure on proteoglycan synthesis in cultured chondrocytes and articular-cartilage explants. Arch. Biochem. Biophys. 300:458–465, 1993.

    Article  Google Scholar 

  32. Raghunath, J., H. J. Salacinski, K. M. Sales, P. E. Butler, and A. M. Seifalian. Advancing cartilage tissue engineering: the application of stem cell technology. Curr. Opin. Biotechnol. 16:503–509, 2005.

    Article  Google Scholar 

  33. Rahaman, M. N., and J. J. Mao. Stem cell-based composite tissue constructs for regenerative medicine. Biotechnol. Bioeng. 91:261–284, 2005.

    Article  Google Scholar 

  34. Ripamonti, U. Soluble osteogenic molecular signals and the induction of bone formation. Biomaterials 27:807–822, 2006.

    Article  Google Scholar 

  35. Robling, A. G., A. B. Castillo, and C. H. Turner. Biomechanical and molecular regulation of bone remodeling. Annu. Rev. Biomed. Eng. 8:455–498, 2006.

    Article  Google Scholar 

  36. Roos, E. M. Joint injury causes knee osteoarthritis in young adults. Curr. Opin. Rheumatol. 17:195–200, 2005.

    Article  Google Scholar 

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

    Google Scholar 

  38. Sharma, G., R. K. Saxena, and P. Mishra. Differential effects of cyclic and static pressure on biochemical and morphological properties of chondrocytes from articular cartilage. Clin. Biomech. 22:248–255, 2007.

    Article  Google Scholar 

  39. Spalazzi, J. P., K. L. Dionisio, J. Jiang, and H. H. Lu. Osteoblast and chondrocyte interactions during coculture on scaffolds. IEEE Eng. Med. Biol. Mag. 22:27–34, 2003.

    Article  Google Scholar 

  40. Thompson, A. D., M. W. Betz, D. M. Yoon, and J. P. Fisher. Osteogenic differentiation of bone marrow stromal cells induced by coculture with chondrocytes encapsulated in three-dimensional matrices. Tissue Eng. A 15:1181–1190, 2009.

    Article  Google Scholar 

  41. Toyoda, T., B. B. Seedhom, J. Kirkham, and W. A. Bonass. Upregulation of aggrecan and type II collagen mRNA expression in bovine chondrocytes by the application of hydrostatic pressure. Biorheology 40:79–85, 2003.

    Google Scholar 

  42. Tsuchiya, K., G. P. Chen, T. Ushida, T. Matsuno, and T. Tateishi. The effect of coculture of chondrocytes with mesenchymal stem cells on their cartilaginous phenotype in vitro. Mater. Sci. Eng. C 24:391–396, 2004.

    Article  Google Scholar 

  43. Wagner, D. R., D. P. Lindsey, K. W. Li, P. Tummala, S. E. Chandran, R. L. Smith, M. T. Longaker, D. R. Carter, and G. S. Beaupre. Hydrostatic pressure enhances chondrogenic differentiation of human bone marrow stromal cells in osteochondrogenic medium. Ann. Biomed. Eng. 36:813–820, 2008.

    Article  Google Scholar 

  44. Wicks, S. J., S. Lui, N. Abdel-Wahab, R. M. Mason, and A. Chantry. Inactivation of SMAD-transforming growth factor beta signaling by Ca2+-calmodulin-dependent protein kinase II. Mol. Cell. Biol. 20:8103–8111, 2000.

    Article  Google Scholar 

  45. Zeiter, S., P. Lezuo, and K. Ito. Effect of TGF beta(1), BMP-2 and hydraulic pressure on chondrogenic differentiation of bovine bone marrow mesenchymal stromal cells. Biorheology 46:45–55, 2009.

    Google Scholar 

Download references

Acknowledgments

The authors are grateful for financial support of this work which was provided by the Hunter Endowment and the AO Foundation.

Sponsors/Grant Number

Hunter Endowment; AO Foundation.

Author information

Authors and Affiliations

  1. Department of Bioengineering and Institute for Biological Interfaces of Engineering, Clemson University, 401 Rhodes Engineering Research Center, Clemson, SC, 29634, USA

    Scott Maxson & Karen J. L. Burg

Authors
  1. Scott Maxson
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Karen J. L. Burg
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Karen J. L. Burg.

Additional information

Associate Editor Helen Lu oversaw the review of this article.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Maxson, S., Burg, K.J.L. Synergistic Effects of Conditioned Media and Hydrostatic Pressure on the Differentiation of Mesenchymal Stem Cells. Cel. Mol. Bioeng. 5, 414–426 (2012). https://doi.org/10.1007/s12195-012-0248-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12195-012-0248-5

Keywords

Search

Navigation