Annals of Biomedical Engineering

, Volume 42, Issue 12, pp 2490–2500 | Cite as

Amniotic Fluid-Derived Stem Cells Demonstrated Cardiogenic Potential in Indirect Co-culture with Human Cardiac Cells

  • Yang Gao
  • Jennifer Petsche Connell
  • Lalita Wadhwa
  • Rodrigo Ruano
  • Jeffrey G. JacotEmail author


Amniotic fluid-derived stem cells (AFSC) have been shown to be broadly multipotent and non-tumorogenic. Previous studies of direct mixing of AFSC and neonatal rat ventricle myocytes indicated evidence of AFSC cardiogenesis. In this study, we examined human AFSC cardiogenic potential in indirect co-culture with human cardiac cells in conditions that eliminated the possibility of cell fusion. Human AFSC in contact with human cardiac cells showed expression of cardiac troponin T (cTnT) in immunohistochemistry, and no evidence of cell fusion was found through fluorescent in situ hybridization. When indirectly co-cultured with cardiac cells, human AFSC in contact with cardiac cells across a thin porous membrane showed a statistically significant increase in cTnT expression compared to non-contact conditions but lacked upregulation of calcium modulating proteins and did not have functional or morphological characteristics of mature cardiomyocytes. This suggests that contact is a necessary but not sufficient condition for AFSC cardiac differentiation in co-culture with cardiac cells.


Cardiomyocytes Cardiac progenitor cells Right ventricular out flow tract Pediatric tissue engineering Congenital heart defects 



Amniotic fluid derived stem cells


Congenital heart defects


Right ventricular out flow tract




Neonatal rat ventricular myocytes



This project is supported by the American Heart Association (Beginning Grant-in-Aid to JGJ.) Amniotic fluid samples were provided by the Fetal Center at Texas Children’s Hospital Pavilion for Women. Pediatric cardiac tissue samples were provided by Division of Congenital Heart Surgery at Texas Children’s Hospital through the Heart Center Biorepository.


There are no competing financial interests.


  1. 1.
    Acquistapace, A., T. Bru, P.-F. Lesault, F. Figeac, A. E. Coudert, O. le Coz, C. Christov, X. Baudin, F. Auber, R. Yiou, J.-L. Dubois-Randé, and A.-M. Rodriguez. Human mesenchymal stem cells reprogram adult cardiomyocytes toward a progenitor-like state through partial cell fusion and mitochondria transfer. Stem Cells (Dayton, Ohio) 29:812–824, 2011.Google Scholar
  2. 2.
    Benavides, O., and J. Petsche. Evaluation of endothelial cells differentiated from amniotic fluid-derived stem cells. Tissue Eng. Part A 18:1123–1131, 2012.Google Scholar
  3. 3.
    Bistola, V., M. Nikolopoulou, A. Derventzi, A. Kataki, N. Sfyras, N. Nikou, M. Toutouza, P. Toutouzas, C. Stefanadis, and M. M. Konstadoulakis. Long-term primary cultures of human adult atrial cardiac myocytes: cell viability, structural properties and BNP secretion in vitro. Int. J. Cardiol. 131:113–122, 2008.PubMedCrossRefGoogle Scholar
  4. 4.
    Chiavegato, A., S. Bollini, M. Pozzobon, A. Callegari, L. Gasparotto, J. Taiani, M. Piccoli, E. Lenzini, G. Gerosa, I. Vendramin, E. Cozzi, A. Angelini, L. Iop, G. F. Zanon, A. Atala, P. De Coppi, and S. Sartore. Human amniotic fluid-derived stem cells are rejected after transplantation in the myocardium of normal, ischemic, immuno-suppressed or immuno-deficient rat. J. Mol. Cell. Cardiol. 42:746–759, 2007.PubMedCrossRefGoogle Scholar
  5. 5.
    Connell, J. P., E. Augustini, K. J. Moise, A. Johnson, and J. G. Jacot. Formation of functional gap junctions in amniotic fluid-derived stem cells induced by transmembrane co-culture with neonatal rat cardiomyocytes. J. Cell Mol. Med. 17:774–781, 2013.PubMedCrossRefGoogle Scholar
  6. 6.
    De Coppi, P., G. Bartsch, M. M. Siddiqui, T. Xu, C. C. Santos, L. Perin, G. Mostoslavsky, A. C. Serre, E. Y. Snyder, J. J. Yoo, M. E. Furth, S. Soker, and A. Atala. Isolation of amniotic stem cell lines with potential for therapy. Nat. Biotechnol. 25:100–106, 2007.PubMedCrossRefGoogle Scholar
  7. 7.
    Du, Z. D., Z. M. Hijazi, C. S. Kleinman, N. H. Silverman, and K. Larntz. Comparison between transcatheter and surgical closure of secundum atrial septal defect in children and adults: results of a multicenter nonrandomized trial. J. Am. Coll. Cardiol. 39:1836–1844, 2002.PubMedCrossRefGoogle Scholar
  8. 8.
    French, K. M., A. V. Boopathy, J. A. DeQuach, L. Chingozha, H. Lu, K. L. Christman, and M. E. Davis. A naturally derived cardiac extracellular matrix enhances cardiac progenitor cell behavior in vitro. Acta Biomater. 8:4357–4364, 2012.Google Scholar
  9. 9.
    Guan, X., and D. Delo. In vitro cardiomyogenic potential of human amniotic fluid stem cells. J. Tissue Eng. Regen. Med. 5:220–228, 2011. doi: 10.1002/term.
  10. 10.
    Hamilton, B. E., D. L. Hoyert, J. A. Martin, D. M. Strobino, and B. Guyer. Annual summary of vital statistics: 2010–2011. Pediatrics 131:548–558, 2013.Google Scholar
  11. 11.
    Harris, L., and S. Balaji. Arrhythmias in the adult with congenital heart disease. In: Diagnosis and Management of Adult Congenital Heart Disease, edited by M. Gatzoulis, G. Webb, and P. E. Daubeney. Philadelphia: Elsevier Health Sciences, 2003, pp. 105–113.Google Scholar
  12. 12.
    Heallen, T., M. Zhang, J. Wang, M. Bonilla-Claudio, E. Klysik, R. L. Johnson, and J. F. Martin. Hippo pathway inhibits Wnt signaling to restrain cardiomyocyte proliferation and heart size. Science (New York, N.Y.) 332:458–461, 2011.CrossRefGoogle Scholar
  13. 13.
    Jacot, J., and J. Wong. Endothelial injury induces vascular smooth muscle cell proliferation in highly localized regions of a direct contact co-culture system. Cell Biochem. Biophys. 52:37–46, 2008.PubMedCentralPubMedCrossRefGoogle Scholar
  14. 14.
    Kehat, I., and D. Kenyagin-Karsenti. Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes. J. Clin. Invest. 108:363–364, 2001.Google Scholar
  15. 15.
    Mishra, R., K. Vijayan, E. J. Colletti, D. A. Harrington, T. S. Matthiesen, D. Simpson, S. K. Goh, B. L. Walker, G. Almeida-Porada, D. Wang, C. L. Backer, S. C. Dudley, L. E. Wold, and S. Kaushal. Characterization and functionality of cardiac progenitor cells in congenital heart patients. Circulation 123:364–73, 2011.Google Scholar
  16. 16.
    Moretti, A., L. Caron, A. Nakano, J. T. Lam, A. Bernshausen, Y. Chen, Y. Qyang, L. Bu, M. Sasaki, S. Martin-Puig, Y. Sun, S. M. Evans, K.-L. Laugwitz, and K. R. Chien. Multipotent embryonic isl1+ progenitor cells lead to cardiac, smooth muscle, and endothelial cell diversification. Cell 127:1151–1165, 2006.PubMedCrossRefGoogle Scholar
  17. 17.
    Murphy, S., J. Xu, and K. Kochanek. Deaths: final data for 2010. National Vital Statistics Reports 60, 2013.Google Scholar
  18. 18.
    Naqvi, N., M. Li, J. W. Calvert, T. Tejada, J. P. Lambert, J. Wu, S. H. Kesteven, S. R. Holman, T. Matsuda, J. D. Lovelock, W. W. Howard, S. E. Iismaa, A. Y. Chan, B. H. Crawford, M. B. Wagner, D. I. K. Martin, D. J. Lefer, R. M. Graham, and A. Husain. A proliferative burst during preadolescence establishes the final cardiomyocyte number. Cell 157:795–807, 2014.PubMedCrossRefGoogle Scholar
  19. 19.
    Nygren, J. M., S. Jovinge, M. Breitbach, P. Säwén, W. Röll, J. Hescheler, J. Taneera, B. K. Fleischmann, and S. E. W. Jacobsen. Bone marrow-derived hematopoietic cells generate cardiomyocytes at a low frequency through cell fusion, but not transdifferentiation. Nat. Med. 10:494–501, 2004.PubMedCrossRefGoogle Scholar
  20. 20.
    Oh, H., S. B. Bradfute, T. D. Gallardo, T. Nakamura, V. Gaussin, Y. Mishina, J. Pocius, L. H. Michael, R. R. Behringer, D. J. Garry, M. L. Entman, and M. D. Schneider. Cardiac progenitor cells from adult myocardium: homing, differentiation, and fusion after infarction. Proc. Natl Acad. Sci. U.S.A. 100:12313–12318, 2003.PubMedCentralPubMedCrossRefGoogle Scholar
  21. 21.
    Olson, E. N., and D. Srivastava. Molecular pathways controlling heart development. Science 272:671–676, 1996.PubMedCrossRefGoogle Scholar
  22. 22.
    Pedrotty, D. M., R. Y. Klinger, R. D. Kirkton, and N. Bursac. Cardiac fibroblast paracrine factors alter impulse conduction and ion channel expression of neonatal rat cardiomyocytes. Cardiovasc. Res. 83:688–697, 2009.PubMedCentralPubMedCrossRefGoogle Scholar
  23. 23.
    Reller, M., and M. Strickland. Prevalence of congenital heart defects in metropolitan Atlanta, 1998–2005. J. Pediatrics 153:807–813, 2008.Google Scholar
  24. 24.
    Schuldiner, M., O. Yanuka, J. Itskovitz-Eldor, D. A. Melton, and N. Benvenisty. Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells. Proc. Natl Acad. Sci. U.S.A. 97:11307–12, 2000.Google Scholar
  25. 25.
    Steeds, R. P., and D. Oakley. Predicting late sudden death from ventricular arrhythmia in adults following surgical repair of tetralogy of Fallot. QJM 97:7–13, 2003.CrossRefGoogle Scholar
  26. 26.
    Tsai, M.-S., J.-L. Lee, Y.-J. Chang, and S.-M. Hwang. Isolation of human multipotent mesenchymal stem cells from second-trimester amniotic fluid using a novel two-stage culture protocol. Hum. Reprod. (Oxford, England) 19:1450–1456, 2004.CrossRefGoogle Scholar
  27. 27.
    Wozniak, M. A., and C. S. Chen. Mechanotransduction in development: a growing role for contractility. Nature reviews. Mol. Cell Biol. 10:34–43, 2009.Google Scholar
  28. 28.
    Xin, M., Y. Kim, L. B. Sutherland, M. Murakami, X. Qi, J. McAnally, E. R. Porrello, A. I. Mahmoud, W. Tan, J. M. Shelton, J. A. Richardson, H. A. Sadek, R. Bassel-Duby, and E. N. Olson. Hippo pathway effector Yap promotes cardiac regeneration. Proc. Natl Acad. Sci. U.S.A. 110:13839–13844, 2013.Google Scholar
  29. 29.
    Zhang, P., J. Baxter, K. Vinod, T. N. Tulenko, and P. J. Di Muzio. Endothelial differentiation of amniotic fluid-derived stem cells: synergism of biochemical and shear force stimuli. Stem Cells Dev. 18:1299–1308, 2009.PubMedCentralPubMedCrossRefGoogle Scholar
  30. 30.
    Zhou, B., Q. Ma, S. Rajagopal, S. M. Wu, I. Domian, J. Rivera-Feliciano, D. Jiang, A. von Gise, S. Ikeda, K. R. Chien, and W. T. Pu. Epicardial progenitors contribute to the cardiomyocyte lineage in the developing heart. Nature 454:109–113, 2008.PubMedCentralPubMedCrossRefGoogle Scholar
  31. 31.
    Zimmermann, W.-H., I. Melnychenko, G. Wasmeier, M. Didié, H. Naito, U. Nixdorff, A. Hess, L. Budinsky, K. Brune, B. Michaelis, S. Dhein, A. Schwoerer, H. Ehmke, and T. Eschenhagen. Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts. Nat. Med. 12:452–458, 2006.PubMedCrossRefGoogle Scholar

Copyright information

© Biomedical Engineering Society 2014

Authors and Affiliations

  • Yang Gao
    • 1
  • Jennifer Petsche Connell
    • 1
  • Lalita Wadhwa
    • 2
  • Rodrigo Ruano
    • 3
    • 4
  • Jeffrey G. Jacot
    • 1
    • 2
    Email author
  1. 1.Department of BioengineeringRice UniversityHoustonUSA
  2. 2.Division of Congenital Heart SurgeryTexas Children’s HospitalHoustonUSA
  3. 3.Fetal Center, Pavilion for WomenTexas Children’s HospitalHoustonUSA
  4. 4.Department of Obstetrics and GynecologyBaylor College of MedicineHoustonUSA

Personalised recommendations