Advertisement

Cellular and Molecular Bioengineering

, Volume 9, Issue 1, pp 107–115 | Cite as

Biomimetic Microstructure Morphology in Electrospun Fiber Mats is Critical for Maintaining Healthy Cardiomyocyte Phenotype

  • Rutwik Rath
  • Jung Bok Lee
  • Truc-Linh Tran
  • Sean F. Lenihan
  • Cristi L. Galindo
  • Yan Ru Su
  • Tarek Absi
  • Leon M. Bellan
  • Douglas B. SawyerEmail author
  • Hak-Joon SungEmail author
Article

Abstract

Despite recent advances in biomimetic substrates, there is still only limited understanding of how the extracellular matrix (ECM) functions in the maintenance of cardiomyocyte (CM) phenotype. In this study, we designed electrospun substrates inspired by morphologic features of non-failing and failing human heart ECM, and examined how these substrates regulate phenotypes of adult and neonatal rat ventricular CMs (ARVM and NRVM, respectively). We found that poly(ε-caprolactone) fiber substrates designed to mimic the organized ECM of a non-failing human heart maintained healthy CM phenotype (evidenced by cell morphology, organized actin/myomesin bands and expression of β-MYH7 and SCN5A.1 and SCN5A.2) compared to both failing heart ECM-mimetic substrates and tissue culture plates. Moreover, culture of ARVMs and NRVMs on aligned substrates showed differences in m- and z-line alignment; with ARVMs aligning parallel to the ECM fibers and the NRVMs aligning perpendicular to the fibers. The results provide new insight into cardiac tissue engineering by illustrating the importance models that mimic the cardiac ECM microenvironment in vitro.

Keywords

Electrospinning Poly(ε-caprolactone) Cell–matrix interaction Cardiomyocyte Cell phenotype 

Notes

Acknowledgments

This study was supported by NIH HL091465, NSF DMR 1006558, U0100398, and AHA 13GRNT16690019. The authors would also like to acknowledge the use of resources at the Vanderbilt Institute of Nanoscale Science and Engineering (VINSE), a facility renovated under NSF ARI-R2 DMR-0963361.

Conflict of interest

Rutwik Rath, Jung Bok Lee, Truc-Linh Tran, Sean F. Lenihan, Cristi L. Galindo, Yan Ru Su, Tarek Absi, Leon M. Bellan, Douglas B. Sawyer, and Hak-Joon Sung declare that they have no conflicts of interest.

Human and Animal Rights and Informed Consent

All human subject research was carried out in accordance with the guidelines of Vanderbilt IRB and human research protection program and approved by approved by the Vanderbilt Institutional Review Board. All animal studies were carried out in accordance with the guidelines of the Vanderbilt University Animal Care and Use Committee and approved by the Vanderbilt Institutional Review Board.

Supplementary material

12195_2015_412_MOESM1_ESM.docx (3.8 mb)
Supplementary material 1 (DOCX 3888 kb)

References

  1. 1.
    Akhyari, P., H. Kamiya, A. Haverich, M. Karck, and A. Lichtenberg. Myocardial tissue engineering: the extracellular matrix. Eur. J. Cardio-Thorac. Surg. 34:229–241, 2008.CrossRefGoogle Scholar
  2. 2.
    Ausma, J., and M. Borgers. Dedifferentiation of atrial cardiomyocytes: from in vivo to in vitro. Cardiovasc. Res. 55:9–12, 2002.CrossRefGoogle Scholar
  3. 3.
    Bhana, B., R. K. Iyer, W. L. Chen, R. Zhao, K. L. Sider, M. Likhitpanichkul, C. A. Simmons, and M. Radisic. Influence of substrate stiffness on the phenotype of heart cells. Biotechnol. Bioeng. 105:1148–1160, 2010.Google Scholar
  4. 4.
    Bird, S. D., P. A. Doevendans, M. A. van Rooijen, A. de Brutel la Riviere, R. J. Hassink, R. Passier, and C. L. Mummery. The human adult cardiomyocyte phenotype. Cardiovasc. Res. 58:423–434, 2003.CrossRefGoogle Scholar
  5. 5.
    Bugaisky, L. B., and R. Zak. Differentiation of adult rat cardiac myocytes in cell culture. Circ. Res. 64:493–500, 1989.CrossRefGoogle Scholar
  6. 6.
    Bursac, N., M. Papadaki, R. J. Cohen, F. J. Schoen, S. R. Eisenberg, R. Carrier, G. Vunjak-Novakovic, and L. E. Freed. Cardiac muscle tissue engineering: toward an in vitro model for electrophysiological studies. Am. J. Physiol. 277:H433–H444, 1999.Google Scholar
  7. 7.
    Chlopcikova, S., J. Psotova, and P. Miketova. Neonatal rat cardiomyocytes: a model for the study of morphological, biochemical and electrophysiological characteristics of the heart. Biomed. Pap. Med. Fac. Univ. Palacký Olomouc Czechoslovakia 145:49–55, 2001.CrossRefGoogle Scholar
  8. 8.
    Crapo, P. M., T. W. Gilbert, and S. F. Badylak. An overview of tissue and whole organ decellularization processes. Biomaterials 32:3233–3243, 2011.CrossRefGoogle Scholar
  9. 9.
    Duling, R. R., R. B. Dupaix, N. Katsube, and J. Lannutti. Mechanical characterization of electrospun polycaprolactone (pcl): a potential scaffold for tissue engineering. J. Biomech. Eng. 130:011006–011006, 2008.CrossRefGoogle Scholar
  10. 10.
    Ellingsen, O., A. J. Davidoff, S. K. Prasad, H. J. Berger, J. P. Springhorn, J. D. Marsh, R. A. Kelly, and T. W. Smith. Adult rat ventricular myocytes cultured in defined medium: phenotype and electromechanical function. Am. J. Physiol. 265:H747–H754, 1993.Google Scholar
  11. 11.
    Engelmayr, Jr., G. C., M. Cheng, C. J. Bettinger, J. T. Borenstein, R. Langer, and L. E. Freed. Accordion-like honeycombs for tissue engineering of cardiac anisotropy. Nat. Mater. 7:1003–1010, 2008.CrossRefGoogle Scholar
  12. 12.
    Farouz, Y., Y. Chen, A. Terzic, and P. Menasche. Concise review: growing hearts in the right place: on the design of biomimetic materials for cardiac stem cell differentiation. Stem Cells 33:1021–1035, 2015.CrossRefGoogle Scholar
  13. 13.
    Galie, P. A., N. Khalid, K. E. Carnahan, M. V. Westfall, and J. P. Stegemann. Substrate stiffness affects sarcomere and costamere structure and electrophysiological function of isolated adult cardiomyocytes. Cardiovasc. Pathol. 22:219–227, 2013.CrossRefGoogle Scholar
  14. 14.
    Galindo, C. L., E. Kasasbeh, A. Murphy, S. Ryzhov, S. Lenihan, F. A. Ahmad, P. Williams, A. Nunnally, J. Adcock, Y. Song, F. E. Harrell, T. L. Tran, T. J. Parry, J. Iaci, A. Ganguly, I. Feoktistov, M. K. Stephenson, A. O. Caggiano, D. B. Sawyer, and J. H. Cleator. Anti-remodeling and anti-fibrotic effects of the neuregulin-1beta glial growth factor 2 in a large animal model of heart failure. J. Am. Heart Assoc. 3:e000773, 2014.CrossRefGoogle Scholar
  15. 15.
    Golden, H. B., D. Gollapudi, F. Gerilechaogetu, J. Li, R. J. Cristales, X. Peng, and D. E. Dostal. Isolation of cardiac myocytes and fibroblasts from neonatal rat pups. Methods Mol. Biol. 843:205–214, 2012.CrossRefGoogle Scholar
  16. 16.
    Gupta, M. K., J. M. Walthall, R. Venkataraman, S. W. Crowder, D. K. Jung, S. S. Yu, T. K. Feaster, X. Wang, T. D. Giorgio, C. C. Hong, F. J. Baudenbacher, A. K. Hatzopoulos, and H.-J. Sung. Combinatorial polymer electrospun matrices promote physiologically-relevant cardiomyogenic stem cell differentiation. PLoS One 6:e28935, 2011.CrossRefGoogle Scholar
  17. 17.
    Hang, C. T., J. Yang, P. Han, H. L. Cheng, C. Shang, E. Ashley, B. Zhou, and C. P. Chang. Chromatin regulation by Brg1 underlies heart muscle development and disease. Nature 466:62–67, 2010.CrossRefGoogle Scholar
  18. 18.
    Herron, T. J., F. S. Korte, and K. S. McDonald. Loaded shortening and power output in cardiac myocytes are dependent on myosin heavy chain isoform expression. Am. J. Physiol. Heart Circ. Physiol. 281(3):H1217–H1222, 2001.Google Scholar
  19. 19.
    Huang, Y., L. Zheng, X. Gong, X. Jia, W. Song, M. Liu, and Y. Fan. Effect of cyclic strain on cardiomyogenic differentiation of rat bone marrow derived mesenchymal stem cells. PLoS One 7:e34960, 2012.CrossRefGoogle Scholar
  20. 20.
    Huyghe, J. M., D. H. van Campen, T. Arts, and R. M. Heethaar. The constitutive behaviour of passive heart muscle tissue: a quasi-linear viscoelastic formulation. J. Biomech. 24:841–849, 1991.CrossRefGoogle Scholar
  21. 21.
    Inserte, J., V. Hernando, M. Ruiz-Meana, M. Poncelas-Nozal, C. Fernandez, L. Agullo, C. Sartorio, U. Vilardosa, and D. Garcia-Dorado. Delayed phospholamban phosphorylation in post-conditioned heart favours ca2+ normalization and contributes to protection. Cardiovasc. Res. 103:542–553, 2014.CrossRefGoogle Scholar
  22. 22.
    Kabaeva, Z., M. Zhao, and D. E. Michele. Blebbistatin extends culture life of adult mouse cardiac myocytes and allows efficient and stable transgene expression. Am. J. Physiol. Heart Circ. Physiol. 294:H1667–H1674, 2008.CrossRefGoogle Scholar
  23. 23.
    Kim, D. H., E. A. Lipke, P. Kim, R. Cheong, S. Thompson, M. Delannoy, K. Y. Suh, L. Tung, and A. Levchenko. Nanoscale cues regulate the structure and function of macroscopic cardiac tissue constructs. Proc. Natl. Acad. Sci. USA 107:565–570, 2010.CrossRefGoogle Scholar
  24. 24.
    Knöll, R. A role for membrane shape and information processing in cardiac physiology. Pflug. Arch. 467:167–173, 2015.CrossRefGoogle Scholar
  25. 25.
    Kovács, M., J. Tóth, C. Hetényi, A. Málnási-Csizmadia, and J. R. Sellers. Mechanism of blebbistatin inhibition of myosin II. J. Biol. Chem. 279:35557–35563, 2004.CrossRefGoogle Scholar
  26. 26.
    Kubin, T., J. Pöling, S. Kostin, P. Gajawada, S. Hein, W. Rees, A. Wietelmann, M. Tanaka, H. Lörchner, S. Schimanski, M. Szibor, H. Warnecke, and T. Braun. Oncostatin m is a major mediator of cardiomyocyte dedifferentiation and remodeling. Cell Stem Cell 9:420–432, 2011.CrossRefGoogle Scholar
  27. 27.
    Kuo, P.-L., H. Lee, M.-A. Bray, N. A. Geisse, Y.-T. Huang, W. J. Adams, S. P. Sheehy, and K. K. Parker. Myocyte shape regulates lateral registry of sarcomeres and contractility. Am. J. Pathol. 181:2030–2037, 2012.CrossRefGoogle Scholar
  28. 28.
    Lakshmanan, R., U. M. Krishnan, and S. Sethuraman. Living cardiac patch: the elixir for cardiac regeneration. Expert Opin. Biol. Ther. 12:1623–1640, 2012.CrossRefGoogle Scholar
  29. 29.
    Louch, W. E., K. A. Sheehan, and B. M. Wolska. Methods in cardiomyocyte isolation, culture, and gene transfer. J. Mol. Cell. Cardiol. 51:288–298, 2011.CrossRefGoogle Scholar
  30. 30.
    Lutolf, M. P., and J. A. Hubbell. Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering. Nat. Biotechnol. 23:47–55, 2005.CrossRefGoogle Scholar
  31. 31.
    McCain, M. L., and K. K. Parker. Mechanotransduction: the role of mechanical stress, myocyte shape, and cytoskeletal architecture on cardiac function. Pflug. Arch. 462:89–104, 2011.CrossRefGoogle Scholar
  32. 32.
    Mitcheson, J. S., J. C. Hancox, and A. J. Levi. Action potentials, ion channel currents and transverse tubule density in adult rabbit ventricular myocytes maintained for 6 days in cell culture. Pflug. Arch. 431:814–827, 1996.CrossRefGoogle Scholar
  33. 33.
    Modis, L. Organization of the extracellular matrix. Taylor & Francis: CRC Press, 1990.Google Scholar
  34. 34.
    Morrisey, E. E. Rewind to recover: dedifferentiation after cardiac injury. Cell Stem Cell 9:387–388, 2011.CrossRefGoogle Scholar
  35. 35.
    Norris, R. A., T. K. Borg, J. T. Butcher, T. A. Baudino, I. Banerjee, and R. R. Markwald. Neonatal and adult cardiovascular pathophysiological remodeling and repair. Ann. N. Y. Acad. Sci. 1123:30–40, 2008.CrossRefGoogle Scholar
  36. 36.
    Parameswaran, S., S. Kumar, R. S. Verma, and R. K. Sharma. Cardiomyocyte culture: an update on the in vitro cardiovascular model and future challenges. Can. J. Physiol. Pharmacol. 91:985–998, 2013.CrossRefGoogle Scholar
  37. 37.
    Patel, A., B. Fine, M. Sandig, and K. Mequanint. Elastin biosynthesis: the missing link in tissue-engineered blood vessels. Cardiovasc. Res. 71:40–49, 2006.CrossRefGoogle Scholar
  38. 38.
    Sander, V., G. Suñe, C. Jopling, C. Morera, and J. C. I. Belmonte. Isolation and in vitro culture of primary cardiomyocytes from adult zebrafish hearts. Nat. Protoc. 8:800–809, 2013.CrossRefGoogle Scholar
  39. 39.
    Shutova, M., C. Yang, J. M. Vasiliev, and T. Svitkina. Functions of nonmuscle myosin II in assembly of the cellular contractile system. PLoS One 7:e40814, 2012.CrossRefGoogle Scholar
  40. 40.
    Simpson, D. G., L. Terracio, M. Terracio, R. L. Price, D. C. Turner, and T. K. Borg. Modulation of cardiac myocyte phenotype in vitro by the composition and orientation of the extracellular matrix. J. Cell. Physiol. 161:89–105, 1994.CrossRefGoogle Scholar
  41. 41.
    Sreejit, P., and R. S. Verma. Natural ECM as biomaterial for scaffold based cardiac regeneration using adult bone marrow derived stem cells. Stem Cell Rev. 9:158–171, 2013.CrossRefGoogle Scholar
  42. 42.
    Stout, D. A., J. Yoo, A. N. Santiago-Miranda, and T. J. Webster. Mechanisms of greater cardiomyocyte functions on conductive nanoengineered composites for cardiovascular application. Int. J. Nanomed. 7:5653–5669, 2012.Google Scholar
  43. 43.
    Valencik, M. L., D. Zhang, B. Punske, P. Hu, J. A. McDonald, and S. E. Litwin. Integrin activation in the heart: a link between electrical and contractile dysfunction? Circ. Res. 99:1403–1410, 2006.CrossRefGoogle Scholar
  44. 44.
    Wang, N., K. Burugapalli, W. Song, J. Halls, F. Moussy, Y. Zheng, Y. Ma, Z. Wu, and K. Li. Tailored fibro-porous structure of electrospun polyurethane membranes, their size-dependent properties and trans-membrane glucose diffusion. J. Membr. Sci. 427:207–217, 2013.CrossRefGoogle Scholar
  45. 45.
    Wendel, J. S., L. Ye, P. Zhang, R. T. Tranquillo, and J. J. Zhang. Functional consequences of a tissue-engineered myocardial patch for cardiac repair in a rat infarct model. Tissue Eng. Part A. 20:1325–1335, 2014.CrossRefGoogle Scholar

Copyright information

© Biomedical Engineering Society 2015

Authors and Affiliations

  • Rutwik Rath
    • 1
  • Jung Bok Lee
    • 1
  • Truc-Linh Tran
    • 2
  • Sean F. Lenihan
    • 2
  • Cristi L. Galindo
    • 2
  • Yan Ru Su
    • 2
  • Tarek Absi
    • 3
  • Leon M. Bellan
    • 1
    • 4
  • Douglas B. Sawyer
    • 2
    Email author
  • Hak-Joon Sung
    • 1
    • 2
    Email author
  1. 1.Department of Biomedical EngineeringVanderbilt UniversityNashvilleUSA
  2. 2.Cardiovascular Division, Department of MedicineVanderbilt UniversityNashvilleUSA
  3. 3.Department of Cardiothoracic SurgeryVanderbilt UniversityNashvilleUSA
  4. 4.Department of Mechanical EngineeringVanderbilt UniversityNashvilleUSA

Personalised recommendations