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Tissue Engineering and Regenerative Medicine

, Volume 16, Issue 1, pp 69–80 | Cite as

Induction of Chondrogenic Differentiation in Human Mesenchymal Stem Cells Cultured on Human Demineralized Bone Matrix Scaffold under Hydrostatic Pressure

  • Saeid Reza Shahmoradi
  • Maryam Kabir SalmaniEmail author
  • Hamid Reza Soleimanpour
  • Amir Hossein Tavakoli
  • Kazem Hosaini
  • Nooshin Haghighipour
  • Shahin BonakdarEmail author
Original Article
  • 56 Downloads

Abstract

Background:

Articular cartilage damage is still a troublesome problem. Hence, several researches have been performed for cartilage repair. The aim of this study was to evaluate the chondrogenicity of demineralized bone matrix (DBM) scaffolds under cyclic hydrostatic pressure (CHP) in vitro.

Methods:

In this study, CHP was applied to human bone marrow mesenchymal stem cells (hBMSCs) seeded on DBM scaffolds at a pressure of 5 MPa with a frequency of 0.5 Hz and 4 h per day for 1 week. Changes in chondrogenic and osteogenic gene expressions were analyzed by quantifying mRNA signal level of Sox9, collagen type I, collagen type II, aggrecan (ACAN), Osteocalcin, and Runx2. Histological analysis was carried out by hematoxylin and eosin, and Alcian blue staining. Moreover, DMMB and immunofluorescence staining were used for glycosaminoglycan (GAG) and collagen type II detection, respectively.

Results:

Real-time PCR demonstrated that applying CHP to hBMSCs in DBM scaffolds increased mRNA levels by 1.3-fold, 1.2-fold, and 1.7-fold (p < 0.005) for Sox9, Col2, and ACAN, respectively by day 21, whereas it decreased mRNA levels by 0.7-fold and 0.8-fold (p < 0.05) for Runx2 and osteocalcin, respectively. Additionally, in the presence of TGF-β1 growth factor (10 ng/ml), CHP further increased mRNA levels for the mentioned genes (Sox9, Col2, and ACAN) by 1.4-fold, 1.3-fold and 2.5-fold (p < 0.005), respectively. Furthermore, in histological assessment, it was observed that the extracellular matrix contained GAG and type II collagen in scaffolds under CHP and CHP with TGF-β1, respectively.

Conclusion:

The osteo-inductive DBM scaffolds showed chondrogenic characteristics under hydrostatic pressure. Our study can be a fundamental study for the use of DBM in articular cartilage defects in vivo and lead to production of novel scaffolds with two different characteristics to regenerate both bone and cartilage simultaneously.

Graphical abstract

Keywords

Bone marrow mesenchymal stem cells Chondrogenic differentiation Hydrostatic pressure Demineralized bone matrix scaffold 

Notes

Acknowledgements

The authors would like to express their appreciation to Alborz Balk Pharmaceutical Company, Nano Zist Arrayeh, and Dr. Kayvan pathology lab for their generous support. This study was supported by Iran Pasteur Institute research (Grant No. 861).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

There are no animal experiments carried out in this article. However, tissue removal from donors was carried out according to Iran Brain death law and Iran Tissue Bank Standards. Ethical and technical proses had been approved by Iran Food and Drug Administration (664/188547).

References

  1. 1.
    Mobasheri A, Csaki C, Clutterbuck AL, Rahmanzadeh M, Shakibaei M. Mesenchymal stem cells in connective tissue engineering and regenerative medicine: applications in cartilage repair and osteoarthritis therapy. Histol Histopathol. 2009;24:347–66.Google Scholar
  2. 2.
    Redman SN, Oldfield SF, Archer CW. Current strategies for articular cartilage repair. Eur Cell Mater. 2005;9:23–32.CrossRefGoogle Scholar
  3. 3.
    Werkmeister JA, Adhikari R, White JF, Tebb TA, Le TP, Taing HC, et al. Biodegradable and injectable cure-on-demand polyurethane scaffolds for regeneration of articular cartilage. Acta Biomater. 2010;6:3471–81.CrossRefGoogle Scholar
  4. 4.
    Hwang NS, Varghese S, Elisseeff J. Cartilage tissue engineering: directed differentiation of embryonic stem cells in three-dimensional hydrogel culture. Methods Mol Biol. 2007;407:351–73.CrossRefGoogle Scholar
  5. 5.
    Getgood A, Brooks R, Fortier L, Rushton N. Articular cartilage tissue engineering: today's research, tomorrow's practice? J Bone Joint Surg Br. 2009;91:565-76.CrossRefGoogle Scholar
  6. 6.
    Shao XX, Hutmacher DW, Ho ST, Goh JC, Lee EH. Evaluation of a hybrid scaffold/cell construct in repair of high-load-bearing osteochondral defects in rabbits. Biomaterials. 2006;27:1071–80.CrossRefGoogle Scholar
  7. 7.
    Lutolf MP, Hubbell JA. Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat Biotechnol. 2005;23:47–55.CrossRefGoogle Scholar
  8. 8.
    Zheng L, Fan HS, Sun J, Chen XN, Wang G, Zhang L, et al. Chondrogenic differentiation of mesenchymal stem cells induced by collagen-based hydrogel: an in vivo study. J Biomed Mater Res A. 2010;93:783–92.Google Scholar
  9. 9.
    Zheng L, Yang J, Fan H, Zhang X. Material-induced chondrogenic differentiation of mesenchymal stem cells is material-dependent. Exp Ther Med. 2014;7:1147–50.CrossRefGoogle Scholar
  10. 10.
    Bonakdar S, Emami SH, Shokrgozar MA, Farhadi A, Ahmadi SAH, Amanzadeh A. Preparation and characterization of polyvinyl alcohol hydrogels crosslinked by biodegradable polyurethane for tissue engineering of cartilage. Mater Sci Eng C Mater Biol Appl. 2010;30:636–43.CrossRefGoogle Scholar
  11. 11.
    Derakhshan ZH, Shaghaghi B, Asl MP, Majidi M, Ghazizadeh L, Chegini A, et al. In situ forming hydrogel based on chondroitin sulfate–hydroxyapatite for bone tissue engineering. Int J Polym Mater. 2015;64:919–26.CrossRefGoogle Scholar
  12. 12.
    Karkhaneh A, Naghizadeh Z, Shokrgozar MA, Bonakdar S. Evaluation of the chondrogenic differentiation of mesenchymal stem cells on hybrid biomimetic scaffolds. J Appl Polym Sci. 2014;131:40635.CrossRefGoogle Scholar
  13. 13.
    Shokrgozar MA, Bonakdar S, Dehghan MM, Emami SH, Montazeri L, Azari S, et al. Biological evaluation of polyvinyl alcohol hydrogel crosslinked by polyurethane chain for cartilage tissue engineering in rabbit model. J Mater Sci Mater Med. 2013;24:2449–60.CrossRefGoogle Scholar
  14. 14.
    Downes S, Archer RS, Kayser MV, Patel MP, Braden M. The regeneration of articular cartilage using a new polymer system. J Mater Sci Mater Med. 1994;5:88–95.CrossRefGoogle Scholar
  15. 15.
    Bauer TW, Muschler GF. Bone graft materials: an overview of the basic science. Clin Orthop Relat Res. 2000;371:10–27.CrossRefGoogle Scholar
  16. 16.
    Harakas NK. Demineralized bone-matrix-induced osteogenesis. Clin Orthop Relat Res. 1984;188:239–51.Google Scholar
  17. 17.
    Chakkalakal DA, Strates BS, Garvin KL, Novak JR, Fritz ED, Mollner TJ, et al. Demineralized bone matrix as a biological scaffold for bone repair. Tissue Eng. 2001;7:161–77.CrossRefGoogle Scholar
  18. 18.
    Gruskin E, Doll BA, Futrell FW, Schmitz JP, Hollinger JO. Demineralized bone matrix in bone repair: history and use. Adv Drug Deliv Rev. 2012;64:1063–77.CrossRefGoogle Scholar
  19. 19.
    Kelly DJ, Jacobs CR. The role of mechanical signals in regulating chondrogenesis and osteogenesis of mesenchymal stem cells. Birth Defects Res C Embryo Today. 2010;90:75–85.CrossRefGoogle Scholar
  20. 20.
    Choi JR, Yong KW, Choi JY. Effects of mechanical loading on human mesenchymal stem cells for cartilage tissue engineering. J Cell Physiol. 2018;233:1913–28.CrossRefGoogle Scholar
  21. 21.
    Fahy N, Alini M, Stoddart MJ. Mechanical stimulation of mesenchymal stem cells: implications for cartilage tissue engineering. J Orthop Res. 2018;36:52–63.Google Scholar
  22. 22.
    Thorpe S, Buckley C, Vinardell T, O’Brien FJ, Campbell V, Kelly DJ. Dynamic compression can inhibit chondrogenesis of mesenchymal stem cells. Biochem Biophys Res Commun. 2008;377:458–62.CrossRefGoogle Scholar
  23. 23.
    Thorpe S, Buckley C, Vinardell T, O’Brien FJ, Campbell V, Kelly DJ. The response of bone marrow-derived mesenchymal stem cells to dynamic compression following TGF-β3 induced chondrogenic differentiation. Ann Biomed Eng. 2010;38:2896–909.CrossRefGoogle Scholar
  24. 24.
    Huang AH, Farrell MJ, Kim M, Mauck RL. Long-term dynamic loading improves the mechanical properties of chondrogenic mesenchymal stem cell-laden hydrogels. Eur Cells Mater. 2010;19:72–85.CrossRefGoogle Scholar
  25. 25.
    Angele P, Yoo JU, Smith C, Mansour J, Jepsen KJ, Nerlich M, et al. Cyclic hydrostatic pressure enhances the chondrogenic phenotype of human mesenchymal progenitor cells differentiated in vitro. J Orthop Res. 2003;21:451–7.CrossRefGoogle Scholar
  26. 26.
    Miyanishi K, Trindade MC, Lindsey DP, Beaupré GS, Carter DR, Goodman SB, et al. Effects of hydrostatic pressure and transforming growth factor-β3 on adult human mesenchymal stem cell chondrogenesis in vitro. Tissue Eng. 2006;12:1419–28.CrossRefGoogle Scholar
  27. 27.
    Wagner DR, Lindsey DP, Li KW, Tummala P, Chandran SE, Smith RL, et al. Hydrostatic pressure enhances chondrogenic differentiation of human bone marrow stromal cells in osteochondrogenic medium. Ann Biomed Eng. 2008;36:813–20.CrossRefGoogle Scholar
  28. 28.
    Ogawa R, Mizuno S, Murphy GF, Orgill DP. The effect of hydrostatic pressure on three-dimensional chondroinduction of human adipose-derived stem cells. Tissue Eng Part A. 2009;15:2937–45.CrossRefGoogle Scholar
  29. 29.
    Meyer EG, Buckley CT, Steward AJ, Kelly DJ. The effect of cyclic hydrostatic pressure on the functional development of cartilaginous tissues engineered using bone marrow derived mesenchymal stem cells. J Mech Behav Biomed Mater. 2011;4:1257–65.CrossRefGoogle Scholar
  30. 30.
    Karkhaneh A, Naghizadeh Z, Shokrgozar MA, Bonakdar S, Solouk A, Haghighipour N. Effects of hydrostatic pressure on biosynthetic activity during chondrogenic differentiation of MSCs in hybrid scaffolds. Int J Artif Organs. 2014;37:142–8.CrossRefGoogle Scholar
  31. 31.
    Puetzer J, Williams J, Gillies A, Bernacki S, Loboa EG. The effects of cyclic hydrostatic pressure on chondrogenesis and viability of human adipose-and bone marrow-derived mesenchymal stem cells in three-dimensional agarose constructs. Tissue Eng Part A. 2012;19:299–306.CrossRefGoogle Scholar
  32. 32.
    Wagers AJ, Sherwood RI, Christensen JL, Weissman IL. Little evidence for developmental plasticity of adult hematopoietic stem cells. Science. 2002;297:2256–9.CrossRefGoogle Scholar
  33. 33.
    Prockop DJ. Marrow stromal cells as stem cells for nonhematopoietic tissues. Science. 1997;276:71–4.CrossRefGoogle Scholar
  34. 34.
    Bruder SP, Fink DJ, Caplan AI. Mesenchymal stem cells in bone development, bone repair, and skeletal regenaration therapy. J Cell Biochem. 1994;56:283–94.CrossRefGoogle Scholar
  35. 35.
    Lien SM, Ko LY, Huang TJ. Effect of pore size on ECM secretion and cell growth in gelatin scaffold for articular cartilage tissue engineering. Acta Biomater. 2009;5:670–9.CrossRefGoogle Scholar
  36. 36.
    Hall AC, Urban JP, Gehl KA. The effects of hydrostatic pressure on matrix synthesis in articular cartilage. J Orthop Res. 1991;9:1–10.CrossRefGoogle Scholar
  37. 37.
    Zimmermann G, Moghaddam A. Allograft bone matrix versus synthetic bone graft substitutes. Injury. 2011;42:S16–21.CrossRefGoogle Scholar
  38. 38.
    Cao L, Yang F, Liu G, Yu D, Li H, Fan Q et al. The promotion of cartilage defect repair using adenovirus mediated Sox9 gene transfer of rabbit bone marrow mesenchymal stem cells. Biomaterials. 2011;32:3910–20.CrossRefGoogle Scholar

Copyright information

© The Korean Tissue Engineering and Regenerative Medicine Society and Springer Science+Business Media B.V., part of Springer Nature 2018

Authors and Affiliations

  1. 1.Biomaterials and Tissue Engineering Department, Stem Cell DivisionNational Institute of Genetic Engineering and BiotechnologyTehranIran
  2. 2.Iranian Tissue Bank, Imam khomani HospitalUniversity of Medical SciencesTehranIran
  3. 3.National Cell Bank DepartmentPasteur Institute of Iran (IPI)TehranIran

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