Trans-differentiation of human adipose-derived mesenchymal stem cells into cardiomyocyte-like cells on decellularized bovine myocardial extracellular matrix-based films

  • Yavuz Emre ArslanEmail author
  • Yusuf Furkan Galata
  • Tugba Sezgin Arslan
  • Burak Derkus
Tissue Engineering Constructs and Cell Substrates Original Research
Part of the following topical collections:
  1. Tissue Engineering Constructs and Cell Substrates


In this study, we aimed at fabricating decellularized bovine myocardial extracellular matrix-based films (dMEbF) for cardiac tissue engineering (CTE). The decellularization process was carried out utilizing four consecutive stages including hypotonic treatment, detergent treatment, enzymatic digestion and decontamination, respectively. In order to fabricate the dMEbF, dBM were digested with pepsin and gelation process was conducted. dMEbF were then crosslinked with N-hydroxysuccinimide/1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide (NHS/EDC) to increase their durability. Nuclear contents of native BM and decellularized BM (dBM) tissues were determined with DNA content analysis and agarose-gel electrophoresis. Cell viability on dMEbF for 3rd, 7th, and 14th days was assessed by MTT assay. Cell attachment on dMEbF was also studied by scanning electron microscopy. Trans-differentiation capacity of human adipose-derived mesenchymal stem cells (hAMSCs) into cardiomyocyte-like cells on dMEbF were also evaluated by histochemical and immunohistochemical analyses. DNA contents for native and dBM were, respectively, found as 886.11 ± 164.85 and 47.66 ± 0.09 ng/mg dry weight, indicating a successful decellularization process. The results of glycosaminoglycan and hydroxyproline assay, and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), performed in order to characterize the extracellular matrix (ECM) composition of native and dBM tissue, showed that the BM matrix was not damaged during the proposed method. Lastly, regarding the histological study, dMEbF not only mimics native ECM, but also induces the stem cells into cardiomyocyte-like cells phenotype which brings it the potential of use in CTE.



Work on this paper was financially supported by Ministry of Science, Industry and Technology, Republic of Turkey (Project ID. 0089.TGSD.2013) and the Scientific and Technological Research Council of Turkey (Project ID. 114S851). We wish to thank Canakkale Onsekiz Mart University, Science and Technology Application & Research Center and MER-TER Medical for collaborating with analyses.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Supplementary material

10856_2018_6135_MOESM1_ESM.docx (1.2 mb)
Supplementary Materials.


  1. 1.
    Wang B, Borazjani A, Tahai M, De Jongh Curry AL, Simionescu DT, Guan J, et al. Fabrication of cardiac patch with decellularized porcine myocardial scaffold and bone marrow mononuclear cells. J Biomed Mater Res Part A. 2010;94:1100–10.Google Scholar
  2. 2.
    Wang RM, Christman KL. Decellularized myocardial matrix hydrogels: In basic research and preclinical studies. Adv Drug Deliv Rev. 2016;96:77–82.CrossRefGoogle Scholar
  3. 3.
    Wu KH, Liu YL, Zhou B, Han ZC. Cellular therapy and myocardial tissue engineering: the role of adult stem and progenitor cells. Eur J Cardiothorac Surg. 2006;30:770–81.CrossRefGoogle Scholar
  4. 4.
    Yeong WY, Sudarmadji N, Yu HY, Chua CK, Leong KF, Venkatraman SS, et al. Porous polycaprolactone scaffold for cardiac tissue engineering fabricated by selective laser sintering. Acta Biomater. 2010;6:2028–34.CrossRefGoogle Scholar
  5. 5.
    Venugopal JR, Prabhakaran MP, Mukherjee S, Ravichandran R, Dan K, Ramakrishna S. Biomaterial strategies for alleviation of myocardial infarction. J R Soc Interface. 2012;9:1–19.CrossRefGoogle Scholar
  6. 6.
    Weymann A, Loganathan S, Takahashi H, Schies C, Claus B, Hirschberg K, et al. Development and evaluation of a perfusion decellularization porcine heart model. Circ J. 2011;75:852–60.CrossRefGoogle Scholar
  7. 7.
    Nugent HM, Edelman ER. Tissue engineering therapy for cardiovascular disease. Circ Res. 2003;92:1068–78.CrossRefGoogle Scholar
  8. 8.
    Beltrami AP, Barlucchi L, Torella D, Baker M, Limana F, Chimenti S, et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell. 2003;114:763–76.CrossRefGoogle Scholar
  9. 9.
    Shimizu K, Ito A, Arinobe M, Murase Y, Iwata Y, Narita Y, et al. Effective cell-seeding technique using magnetite nanoparticles and magnetic force onto decellularized blood vessels for vascular tissue engineering. J Biosci Bioeng. 2007;103:472–8.CrossRefGoogle Scholar
  10. 10.
    Langer R, Vacanti JP. Tissue Engineering. Science. 1993;260:920–6.CrossRefGoogle Scholar
  11. 11.
    Demirbag B, Huri PY, Kose GT, Buyuksungur A, Hasirci V. Advanced cell therapies with and without scaffolds. Biotechnol J. 2011;6:1437–53.CrossRefGoogle Scholar
  12. 12.
    Arslan YE, Hiz MM, Sezgin Arslan T. The use of decellularized animal tissues in regenerative therapies. Kafkas Univ Vet Fak Derg. 2015;21:139–45.Google Scholar
  13. 13.
    Shin H, Jo S, Mikos AG. Biomimetic materials for tissue engineering. Biomaterials. 2003;24:4353–64.CrossRefGoogle Scholar
  14. 14.
    Ma Z, Kotaki M, Inai R, Ramakrishna S. Potential of nanofiber matrix as tissue-engineering scaffolds. Tissue Eng. 2005;11:101–9.CrossRefGoogle Scholar
  15. 15.
    Arslan YE, Sezgin Arslan T, Derkus B, Emregul E, Emregul KC. Fabrication of human hair keratin/jellyfish collagen/eggshell-derived hydroxyapatite osteoinductive biocomposite scaffolds for bone tissue engineering: From waste to regenerative medicine products. Colloids Surf B Biointerfaces. 2017;154:160–70.CrossRefGoogle Scholar
  16. 16.
    Chen Q, Liang S, Thouas GA. Elastomeric biomaterials for tissue engineering. Prog Polym Sci. 2013;38:584–671.CrossRefGoogle Scholar
  17. 17.
    Shadrin IY, Allen BW, Qian Y, Jackman CP, Carlson AL, Juhas ME, et al. Cardiopatch platform enables maturation and scale-up of human pluripotent stem cell-derived engineered heart tissues. Nat Commun. 2017;8:1825.CrossRefGoogle Scholar
  18. 18.
    Song JJ, Ott HC. Organ engineering based on decellularized matrix scaffolds. Trends Mol Med. 2011;17:424–32.CrossRefGoogle Scholar
  19. 19.
    Pagoulatou E, Triantaphyllidou IE, Vynios DH, Papachristou DJ, Koletsis E, Deligianni D, et al. Biomechanical and structural changes following the decellularization of bovine pericardial tissues for use as a tissue engineering scaffold. J Mater Sci Mater Med. 2012;23:1387–96.CrossRefGoogle Scholar
  20. 20.
    Erten E, Sezgin Arslan T, Derkus B, Arslan YE. Detergent-free decellularization of bovine costal cartilage for chondrogenic differentiation of human adipose mesenchymal stem cells in vitro. RSC Adv. 2016;6:94236–46.CrossRefGoogle Scholar
  21. 21.
    Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials. 2006;27:3675–83.Google Scholar
  22. 22.
    Mendoza-Novelo B, Avila EE, Cauich-Rodríguez JV, Jorge-Herrero E, Rojo FJ, Guinea GV, Mata-Mata HL. Decellularization of pericardial tissue and its impact on tensile viscoelasticity and glycosaminoglycan content. Acta Biomater. 2011;7:1241–8.CrossRefGoogle Scholar
  23. 23.
    Pourfarhangia KE, Mashayekhana S, Asla SG, Hajebrahimic Z. Construction of scaffolds composed of acellular cardiac extracellular matrix for myocardial tissue engineering. Biologicals. 2018;53:10–8.CrossRefGoogle Scholar
  24. 24.
    Ye X, Wang H, Gong W, Li S, Li H, Wang Z, Zhao Q. Impact of decellularization on porcine myocardium as scaffold for tissue engineered heart tissue. J Mater Sci Mater Med. 2016;27:70CrossRefGoogle Scholar
  25. 25.
    Poornejad N, Nielsen JJ, Morris RJ, Gassman JR, Reynolds PR, Roeder BL, et al. Comparison of four decontamination treatments on porcine renal decellularized extracellular matrix structure, composition, and support of human renal cortical tubular epithelium cells. J Biomater Appl. 2016;30:1154–67.CrossRefGoogle Scholar
  26. 26.
    Wilshaw S-P, Kearney JN, Fisher J, Ingham E. Production of an acellular amniotic membrane matrix for use in tissue engineering. Tissue Eng. 2006;12:2117–29.CrossRefGoogle Scholar
  27. 27.
    Sawkins MJ, Bowen W, Dhadda P, Markides H, Sidney LE, Taylor AJ, et al. Hydrogels derived from demineralized and decellularized bone extracellular matrix. Acta Biomater. 2013;9:7865–73.CrossRefGoogle Scholar
  28. 28.
    Buttafoco L, Kolkman NG, Engbers-Buijtenhuijs P, Poot AA, Dijkstra PJ, Vermes I, et al. Electrospinning of collagen and elastin for tissue engineering applications. Biomaterials. 2006;27:724–34.CrossRefGoogle Scholar
  29. 29.
    Singelyn JM, DeQuach JA, Seif-Naraghi SB, Littlefield RB, Schup-Magoffin PJ, Christman KL. Naturally derived myocardial matrix as an injectable scaffold for cardiac tissue engineering. Biomaterials. 2009;30:5409–16.CrossRefGoogle Scholar
  30. 30.
    Derkus B, Arslan YE, Bayrac AT, Kantarcioglu I, Emregul KC, Emregul E. Development of a novel aptasensor using jellyfish collagen as matrix and thrombin detection in blood samples obtained from patients with various neurodisease. Sens Actuators B Chem. 2016;228:725–36.CrossRefGoogle Scholar
  31. 31.
    Seif-Naraghi SB, Horn D, Schup-Magoffin PJ, Christman KL. Injectable extracellular matrix derived hydrogel provides a platform for enhanced retention and delivery of a heparin-binding growth factor. Acta Biomater. 2012;8:3695–703.CrossRefGoogle Scholar
  32. 32.
    Pati F, Jang J, Ha DH, Won Kim S, Rhie JW, Shim JH, et al. Printing three-dimensional tissue analogues with decellularized extracellular matrix bioink. Nat Commun. 2014;5:1–11.CrossRefGoogle Scholar
  33. 33.
    Porzionato A, Sfriso MM, Pontini A, Macchi V, Petrelli L, Pavan PG, et al. Decellularized human skeletal muscle as biologic scaffold for reconstructive surgery. Int J Mol Sci. 2015;16:14808–31.CrossRefGoogle Scholar
  34. 34.
    Bonvillain RW, Scarritt ME, Pashos NC, Mayeux JP, Meshberger CL, Betancourt AM, et al. Non human primate lung decellularization and recellularization using a specialized large-organ bioreactor. J Vis Exp. 2012;18:2437–52.Google Scholar
  35. 35.
    Yang B, Zhang Y, Zhou L, Sun Z, Zheng J, Chen Y, et al. Development of a porcine bladder acellular matrix with well-preserved extracellular bioactive factors for tissue engineering. Tissue Eng Part C Methods. 2010;16:1201–11.CrossRefGoogle Scholar
  36. 36.
    Kaiser NJ, Coulombe KLK. Physiologically inspired cardiac scaffolds for tailored in vivo function and heart regeneration. Biomed Mater. 2015;10:1–26.CrossRefGoogle Scholar
  37. 37.
    Abd-Elgaliel WR, Tung CH. A cardiac tissue-specific binding agent of troponin I. Mol Biosyst. 2012;8:2629–32.CrossRefGoogle Scholar
  38. 38.
    Wiles K, Fishman JM, De Coppi P, Birchall MA. The host immune response to tissue-engineered organs: Current problems and future directions. Tissue Eng Part B Rev. 2016;22:208–19.CrossRefGoogle Scholar
  39. 39.
    Wang Y, Bao J, Wu X, Wu Q, Li Y, Zhou Y, et al. Genipin crosslinking reduced the immunogenicity of xenogeneic decellularized porcine whole-liver matrices through regulation of immune cell proliferation and polarization. Sci Rep. 2016;6:1–16.CrossRefGoogle Scholar
  40. 40.
    Badylak SF, Gilbert TW. Immune response to biologic scaffold materials. Semin Immunol. 2009;20:109–16.CrossRefGoogle Scholar

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© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Regenerative Biomaterials Laboratory, Department of Bioengineering, Engineering FacultyCanakkale Onsekiz Mart UniversityCanakkaleTurkey
  2. 2.Department of Biomedical Engineering, Engineering FacultyEskisehir Osmangazi UniversityEskisehirTurkey

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