Biomimetic perfusion and electrical stimulation applied in concert improved the assembly of engineered cardiac tissue

Maintenance of normal myocardial function depends intimately on synchronous tissue contraction, driven by electrical activation and on adequate nutrient perfusion in support thereof. Bioreactors have been used to mimic aspects of these factors in vitro to engineer cardiac tissue but, due to design limitations, previous bioreactor systems have yet to simultaneously support nutrient perfusion, electrical stimulation and unconstrained (i.e. not isometric) tissue contraction. To the best of our knowledge, the bioreactor system described herein is the first to integrate these three key factors in concert. We present the design of our bioreactor and characterize its capability in integrated experimental and mathematical modelling studies. We then cultured cardiac cells obtained from neonatal rats in porous, channelled elastomer scaffolds with the simultaneous application of perfusion and electrical stimulation, with controls excluding either one or both of these two conditions. After 8 days of culture, constructs grown with simultaneous perfusion and electrical stimulation exhibited substantially improved functional properties, as evidenced by a significant increase in contraction amplitude (0.23 ± 0.10% vs 0.14 ± 0.05%, 0.13 ± 0.08% or 0.09 ± 0.02% in control constructs grown without stimulation, without perfusion, or either stimulation or perfusion, respectively). Consistently, these constructs had significantly improved DNA contents, cell distribution throughout the scaffold thickness, cardiac protein expression, cell morphology and overall tissue organization compared to control groups. Thus, the simultaneous application of medium perfusion and electrical conditioning enabled by the use of the novel bioreactor system may accelerate the generation of fully functional, clinically sized cardiac tissue constructs. Copyright © 2011 John Wiley & Sons, Ltd.

[1]  Jin Gao,et al.  Macroporous elastomeric scaffolds with extensive micropores for soft tissue engineering. , 2006, Tissue engineering.

[2]  Fen Chen,et al.  Biomimetic approach to cardiac tissue engineering: oxygen carriers and channeled scaffolds. , 2006, Tissue engineering.

[3]  J. Nichol,et al.  Co-culture induces alignment in engineered cardiac constructs via MMP-2 expression. , 2008, Biochemical and biophysical research communications.

[4]  Aldo R Boccaccini,et al.  Characterisation of a soft elastomer poly(glycerol sebacate) designed to match the mechanical properties of myocardial tissue. , 2008, Biomaterials.

[5]  Milica Radisic,et al.  Functional assembly of engineered myocardium by electrical stimulation of cardiac myocytes cultured on scaffolds , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[6]  J. Voldman.  Electrical forces for microscale cell manipulation. , 2006, Annual review of biomedical engineering.

[7]  Samuel Bernard,et al.  Evidence for Cardiomyocyte Renewal in Humans , 2008, Science.

[8]  Andreas Hess,et al.  Engineered heart tissue grafts improve systolic and diastolic function in infarcted rat hearts , 2006, Nature Medicine.

[9]  Nenad Bursac,et al.  Cardiac tissue engineering using stem cells. , 2009, IEEE engineering in medicine and biology magazine : the quarterly magazine of the Engineering in Medicine & Biology Society.

[10]  Doris A Taylor,et al.  Perfusion-decellularized matrix: using nature's platform to engineer a bioartificial heart , 2008, Nature Medicine.

[11]  M. Radisic,et al.  Pulsatile perfusion bioreactor for cardiac tissue engineering , 2008, Biotechnology progress.

[12]  Milica Radisic,et al.  Medium perfusion enables engineering of compact and contractile cardiac tissue. , 2004, American journal of physiology. Heart and circulatory physiology.

[13]  Richard A. Williams,et al.  Communication systems analysis and design , 1987 .

[14]  Eric D. Adler,et al.  Human cardiovascular progenitor cells develop from a KDR+ embryonic-stem-cell-derived population , 2008, Nature.

[15]  M. Moretti,et al.  Insulin-like growth factor-I and slow, bi-directional perfusion enhance the formation of tissue-engineered cardiac grafts. , 2009, Tissue engineering. Part A.

[16]  Milica Radisic,et al.  High-density seeding of myocyte cells for cardiac tissue engineering. , 2003, Biotechnology and bioengineering.

[17]  Lisa E. Freed,et al.  Accordion-Like Honeycombs for Tissue Engineering of Cardiac Anisotropy , 2008, Nature materials.

[18]  Milica Radisic,et al.  Mathematical model of oxygen distribution in engineered cardiac tissue with parallel channel array perfused with culture medium containing oxygen carriers. , 2005, American journal of physiology. Heart and circulatory physiology.

[19]  Masayuki Yamato,et al.  Endothelial Cell Coculture Within Tissue-Engineered Cardiomyocyte Sheets Enhances Neovascularization and Improves Cardiac Function of Ischemic Hearts , 2008, Circulation.

[20]  Tal Dvir,et al.  Electric field stimulation integrated into perfusion bioreactor for cardiac tissue engineering. , 2010, Tissue engineering. Part C, Methods.

[21]  M. Radisic,et al.  Biomimetic approach to tissue engineering. , 2009, Seminars in cell & developmental biology.

[22]  L. Samuelson,et al.  Troponin I isoform expression is developmentally regulated in differentiating embryonic stem cell‐derived cardiac myocytes , 1996, Developmental dynamics : an official publication of the American Association of Anatomists.

[23]  Milica Radisic,et al.  Challenges in cardiac tissue engineering. , 2010, Tissue engineering. Part B, Reviews.

[24]  Milica Radisic,et al.  Cardiac tissue engineering using perfusion bioreactor systems , 2008, Nature Protocols.

[25]  Gordana Vunjak-Novakovic,et al.  Perfusion improves tissue architecture of engineered cardiac muscle. , 2002, Tissue engineering.

[26]  J. Vacanti,et al.  Contractile cardiac grafts using a novel nanofibrous mesh. , 2004, Biomaterials.

[27]  Joseph D. Bronzino,et al.  The Biomedical Engineering Handbook , 1995 .

[28]  Hyoungshin Park,et al.  Pre-treatment of synthetic elastomeric scaffolds by cardiac fibroblasts improves engineered heart tissue. , 2008, Journal of biomedical materials research. Part A.

[29]  W. Zimmermann,et al.  Tissue Engineering of a Differentiated Cardiac Muscle Construct , 2002, Circulation research.

[30]  R. Langer,et al.  A tough biodegradable elastomer , 2002, Nature Biotechnology.

[31]  Gordana Vunjak-Novakovic,et al.  Alignment and elongation of human adipose-derived stem cells in response to direct-current electrical stimulation , 2009, 2009 Annual International Conference of the IEEE Engineering in Medicine and Biology Society.

[32]  C. Yerebakan,et al.  Cell Sources for Cardiovascular Tissue Regeneration and Engineering , 2009, The Thoracic and cardiovascular surgeon.

[33]  G. Vunjak-Novakovic,et al.  Design of electrical stimulation bioreactors for cardiac tissue engineering , 2008, 2008 30th Annual International Conference of the IEEE Engineering in Medicine and Biology Society.

[34]  N. Bursac,et al.  Cardiac tissue engineering using stem cells [Cellular/Tissue Engineering] , 2009, IEEE Engineering in Medicine and Biology Magazine.

[35]  G. Vunjak‐Novakovic,et al.  A Biocompatible Endothelial Cell Delivery System for in Vitro Tissue Engineering , 2009, Cell transplantation.

[36]  G. Koh,et al.  Genetically selected cardiomyocytes from differentiating embronic stem cells form stable intracardiac grafts. , 1996, The Journal of clinical investigation.

[37]  C. Cannizzaro,et al.  Practical aspects of cardiac tissue engineering with electrical stimulation. , 2007, Methods in molecular medicine.

[38]  Thomas Eschenhagen,et al.  Engineered heart tissue for regeneration of diseased hearts. , 2004, Biomaterials.

[39]  Austin G Smith,et al.  Mouse and human induced pluripotent stem cells as a source for multipotent Isl1+ cardiovascular progenitors , 2010, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[40]  Kumaraswamy Nanthakumar,et al.  Interrogating functional integration between injected pluripotent stem cell-derived cells and surrogate cardiac tissue , 2009, Proceedings of the National Academy of Sciences.

[41]  Nicola Elvassore,et al.  Electrical stimulation of human embryonic stem cells: cardiac differentiation and the generation of reactive oxygen species. , 2009, Experimental cell research.

[42]  D. Mozaffarian,et al.  Heart disease and stroke statistics--2009 update: a report from the American Heart Association Statistics Committee and Stroke Statistics Subcommittee. , 2009, Circulation.

[43]  Gordana Vunjak-Novakovic,et al.  Perfusion seeding of channeled elastomeric scaffolds with myocytes and endothelial cells for cardiac tissue engineering , 2010, Biotechnology progress.

[44]  S. Schiaffino,et al.  Troponin T switching in the developing rat heart. , 1988, The Journal of biological chemistry.

[45]  Milica Radisic,et al.  Oxygen gradients correlate with cell density and cell viability in engineered cardiac tissue , 2006, Biotechnology and bioengineering.

[46]  Milica Radisic,et al.  Electrical stimulation systems for cardiac tissue engineering , 2009, Nature Protocols.

[47]  Jianwen Luo,et al.  A fast normalized cross-correlation calculation method for motion estimation , 2010, IEEE Transactions on Ultrasonics, Ferroelectrics and Frequency Control.