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Thromb Haemost 2005; 94(04): 728-737
DOI: 10.1160/TH05-04-0268
Theme Issue Article
Schattauer GmbH

Molecular and phenotypic analyses of human embryonic stem cell-derived cardiomyocytes

Opportunities and challenges for clinical translation
Gareth Goh
1   Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham, UK
2   Division of Therapeutics and Molecular Medicine, University of Nottingham, Queens Medical Centre, Nottingham, UK
,
Tim Self
3   Institute of Cell Signalling, University of Nottingham, Queens Medical Centre, Nottingham, UK
,
Maria D. Barbadillo Muñoz
1   Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham, UK
4   Division of Obstetrics and Gynaecology, University of Nottingham, Queens Medical Centre, Nottingham, UK
,
Ian P. Hall
2   Division of Therapeutics and Molecular Medicine, University of Nottingham, Queens Medical Centre, Nottingham, UK
,
Lorraine Young
1   Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham, UK
4   Division of Obstetrics and Gynaecology, University of Nottingham, Queens Medical Centre, Nottingham, UK
,
Chris Denning
1   Institute of Genetics, University of Nottingham, Queens Medical Centre, Nottingham, UK
› Author AffiliationsFinancial support: This work was funded by MRC and the University of Nottingham.
Further Information

Publication History

Received19 April 2005

Accepted after revision09 July 2005

Publication Date:
07 December 2017 (online)

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Summary

Differentiation of human embryonic stem cells (hESCs) into cardiomyocytes in culture may offer unique opportunities for modeling genetic disorders, screening potentially cardiotoxic pharmaceutical agents or replacing cells of the diseased heart. However, before clinical utility can be realized, numerous hurdles must be overcome. Comprehensive molecular and phenotypic characterization is required but has so far been restricted to cardiomyocytes derived from a limited subset of hESC lines. Thus, we have initiated analysis of cardiomyocyte differentiation and function from a further two independently derived lines, BG01 and HUES-7. The challenge of improving cardiac cell induction, enrichment and maturation must also be addressed to meet the demands of high throughput pharmaceutical screening or to provide sufficient cells to repair an infarcted heart. Transplanted cells must functionally integrate without inducing arrhythmias, while survival and evasion of immune surveillance must be accomplished without tumorigenicity. This review evaluates the opportunities presented by hESC-derived cardiomyocytes and the progress towards surmounting the challenges of clinical translation.

Keywords

Human embryonic stem cells (hESC) - characterization - cardiomyocytes - differentiation - clinical translation
 

  • References

  • 1 Garcia-Martinez V, Schoenwolf GC. Primitivestreak origin of the cardiovascular system in avian embryos. Dev Biol 1993; 159: 706-19
    CrossrefPubMedGoogle Scholar
  • 2 Harvey RP. Patterning the vertebrate heart. Nat Rev Genet. 2002; 3: 544-56.
    CrossrefPubMedGoogle Scholar
  • 3 Olson EN. A decade of discoveries in cardiac biology. Nat Med 2004; 10: 467-74.
    CrossrefPubMedGoogle Scholar
  • 4 Caplice NM, and Gersh BJ. Stem cells to repair the heart: a clinical perspective. Circ Res 2003; 10: 6-8.
    CrossrefPubMedGoogle Scholar
  • 5 Splawski I, Shen J, Timothy K W. et al. Spectrum of mutations in long-QT syndrome genes. KVLQT1, HERG, SCN5A, KCNE1, and KCNE2. Circulation 2000; 102: 1178-85.
    CrossrefPubMedGoogle Scholar
  • 6 Schimmel KJ, Richel DJ, van den Brink RB. et al. Cardiotoxicity of cytotoxic drugs. Cancer Treat Rev 2004; 30: 181-91.
    CrossrefPubMedGoogle Scholar
  • 7 Davila JC, Cezar GG, Thiede M. et al. Use and application of stem cells in toxicology. Toxicol Sci. 2004; 79: 214-23.
    CrossrefPubMedGoogle Scholar
  • 8 Cubeddu LX. QT prolongation and fatal arrhythmias: a review of clinical implications and effects of drugs. J Ther 2003; 10: 452-7.
    PubMedGoogle Scholar
  • 9 Laugwitz KL, Moretti A, Lam J. et al. Postnatal isl1+ cardioblasts enter fully differentiated cardiomyocyte lineages. Nature 2005; 433: 647-53.
    CrossrefPubMedGoogle Scholar
  • 10 Matsuura K, Nagai T, Nishigaki N. et al. Adult cardiac Sca-1-positive cells differentiate into beating cardiomyocytes. J Biol Chem 2004; 279: 11384-91.
    CrossrefPubMedGoogle Scholar
  • 11 Beltrami AP, Barlucchi L, Torella D. et al. Adult cardiac stem cells are multipotent and support myocardial regeneration. Cell 2003; 114: 763-76.
    CrossrefPubMedGoogle Scholar
  • 12 Xu W, Zhang X, Qian H. et al. Mesenchymal stem cells from adult human bone marrow differentiate into a cardiomyocyte phenotypein vitro . Exp Biol Med 2004; 229: 623-31.
    CrossrefPubMedGoogle Scholar
  • 13 Zhao P, Ise H, Hongo M. et al. Human amniotic mesenchymal cells have some characteristics of cardiomyocytes. Transplantation 2005; 79: 528-35.
    CrossrefPubMedGoogle Scholar
  • 14 Murry CE, Soonpaa MH, Reinecke H. et al. Haematopoietic stem cells do not transdifferentiate into cardiac myocytes in myocardial infarcts. Nature 2004; 428: 664-8.
    CrossrefPubMedGoogle Scholar
  • 15 Balsam LB, Wagers A J. et al. Haematopoietic stem cells adopt mature haematopoietic fates in ischaemic myocardium. Nature 2004; 428: 668-73.
    CrossrefPubMedGoogle Scholar
  • 16 Thomson JA, Itskovitz-Eldor J, Shapiro S S. et al. Embryonic stem cell lines derived from human blastocysts. Science 1998; 282: 1145-7.
    CrossrefPubMedGoogle Scholar
  • 17 Reubinoff BE, Pera MF, Fong C Y. et al. Embryonic stem cell lines from human blastocysts: somatic differentiationin vitro . Nat Biotechnol 2000; 18: 399-404.
    CrossrefPubMedGoogle Scholar
  • 18 Mitalipova M, Calhoun J, Shin S. et al. Human embryonic stem cell lines derived from discarded embryos. Stem Cells 2003; 21: 521-6.
    CrossrefPubMedGoogle Scholar
  • 19 Cowan CA, Klimanskaya I, McMahon J. et al. Derivation of embryonic stem-cell lines from human blastocysts. N Engl J Med 2004; 350: 1353-6.
    CrossrefPubMedGoogle Scholar
  • 20 Hwang WS, Ryu YJ, Park J H. et al. Evidence of a pluripotent human embryonic stem cell line derived from a cloned blastocyst. Science 2004; 303: 1669-74.
    CrossrefPubMedGoogle Scholar
  • 21 Pickering SJ, Minger SL, Patel M. et al. Generation of a human embryonic stem cell line encoding the cystic fibrosis mutation deltaF508, using preimplantation genetic diagnosis. Reprod Biomed Online 2005; 10: 390-7.
    CrossrefPubMedGoogle Scholar
  • 22 Verlinsky Y, Strelchenko N, Kukharenko V. et al. Human embryonic stem cell lines with genetic disorders. Reprod Biomed Online 2005; 10: 105-10.
    CrossrefPubMedGoogle Scholar
  • 23 Kehat I, Kenyagin-Karsenti D, Snir M. et al. Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes. J Clin Invest 2001; 108: 407-14.
    CrossrefPubMedGoogle Scholar
  • 24 Xu C, Police S, Rao N. et al. Characterization and enrichment of cardiomyocytes derived from human embryonic stem cells. Circ Res 2002; 91: 501-8.
    CrossrefPubMedGoogle Scholar
  • 25 He JQ, Ma Y, Lee Y. et al. Human embryonic stem cells develop into multiple types of cardiac myocytes: action potential characterization. Circ Res 2003; 93: 32-9.
    CrossrefPubMedGoogle Scholar
  • 26 Mummery C, Ward-van Oostwaard D, Doevendans P. et al. Differentiation of human embryonic stem cells to cardiomyocytes: role of coculture with visceral endoderm-like cells. Circulation 2003; 107: 2733-40.
    CrossrefPubMedGoogle Scholar
  • 27 Reppel M, Boettinger C, Hescheler J. Beta-adrenergic and muscarinic modulation of human embryonic stem cell-derived cardiomyocytes. Cell Physiol Biochem 2004; 14: 187-96.
    CrossrefPubMedGoogle Scholar
  • 28 Xue T, Cho HC, Akar F G. et al.. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes: insights into the development of cell-based pacemakers. Circulation 2005; 111: 11-20.
    CrossrefPubMedGoogle Scholar
  • 29 Fijnvandraat AC, van Ginneken AC, Schumacher C A. et al. Cardiomyocytes purified from differentiated embryonic stem cells exhibit characteristics of early chamber myocardium. J Mol Cell Cardiol 2003; 35: 1461-72.
    CrossrefPubMedGoogle Scholar
  • 30 Snir M, Kehat I, Gepstein A. et al. Assessment of the ultrastructural and proliferative properties of human embryonic stem cell-derived cardiomyocytes. Am J Physiol Heart Circ Physiol 2003; 285: H2355-H2363.
    CrossrefPubMedGoogle Scholar
  • 31 Wobus AM, Wallukat G, Hescheler J. Pluripotent mouse embryonic stem cells are able to differentiate into cardiomyocytes expressing chronotropic responses to adrenergic and cholinergic agents and Ca2+ channel blockers. Differentiation 1991; 48: 173-82.
    CrossrefPubMedGoogle Scholar
  • 32 An RH, Davies MP, Doevendans P A. et al. Developmental changes in beta-adrenergic modulation of L-type Ca2+ channels in embryonic mouse heart. Circ Res 1996; 78: 371-8.
    CrossrefPubMedGoogle Scholar
  • 33 Kehat I, Gepstein A, Spira A. et al. High-resolution electrophysiological assessment of human embryonic stem cell-derived cardiomyocytes: a novelin vitro model for the study of conduction. Circ Res 2002; 91: 659-61.
    CrossrefPubMedGoogle Scholar
  • 34 Kehat I, Khimovich L, Caspi O. et al. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol 2004; 22: 1282-9.
    CrossrefPubMedGoogle Scholar
  • 35 Davies MP, An RH, Doevendans P. et al. Developmental changes in ionic channel activity in the embryonic murine heart. Circ Res 1996; 78: 15-25.
    CrossrefPubMedGoogle Scholar
  • 36 Shih HT. Anatomy of the action potential in the heart. Tex Heart Inst J. 1994; 21: 30-41.
    PubMedGoogle Scholar
  • 37 Snyders DJ. Structure and function of cardiac potassium channels. Cardiovasc Res 1999; 42: 377-90.
    CrossrefPubMedGoogle Scholar
  • 38 Satin J, Kehat I, Caspi O. et al. Mechanism of spontaneous excitability in human embryonic stem cell derived cardiomyocytes. J Physiol 2004; 559: 479-96.
    CrossrefPubMedGoogle Scholar
  • 39 Strehler EE, Treiman M. Calcium pumps of plasma membrane and cell interior. Curr Mol Med 2004; 4: 323-35.
    CrossrefPubMedGoogle Scholar
  • 40 Tohse N, Seki S, Kobayashi T. et al. Development of excitation-contraction coupling in cardiomyocytes. Jpn J Physiol 2004; 54: 1-6.
    CrossrefPubMedGoogle Scholar
  • 41 Anderson ME, Al-Khatib SM, Roden D M. et al. Cardiac repolarization: current knowledge, critical gaps, and new approaches to drug development and patient management. Am Heart J 2002; 144: 769-81.
    CrossrefPubMedGoogle Scholar
  • 42 Tuganowski W, Tendera M. Components of the action potential of human embryonic auricle. Am J Physiol 1973; 224: 803-8.
    CrossrefPubMedGoogle Scholar
  • 43 Hechheler J, Fleischmann BK, Lentini S. et al. Embryonic stem cells: a model to study structural and functional properties in cardiomyogenesis. Cardiovasc. Res 1997; 36: 149-62.
    PubMedGoogle Scholar
  • 44 Blake DJ, Weir A, Newey S E. et al. Function and genetics of dystrophin and dystrophin-related proteins in muscle. Physiol Rev 2002; 82: 291-329.
    CrossrefPubMedGoogle Scholar
  • 45 Tuffery-Giraud S, Saquet C, Chambert S. et al. The role of muscle biopsy in analysis of the dystrophin gene in Duchenne muscular dystrophy: experience of a national referral centre. Neuromuscul Disord 2004; 14: 650-8.
    CrossrefPubMedGoogle Scholar
  • 46 Buzin CH, Feng J, Yan J. et al. Mutation rates in the dystrophin gene: a hotspot of mutation at a CpG dinucleotide. Hum Mutat 2005; 25: 177-88.
    CrossrefPubMedGoogle Scholar
  • 47 Wilde AA, Antzelevitch C, Borggrefe M. et al. Proposed Diagnostic Criteria for the Brugada Syndrome. Circulation 2002; 106: 2514-9.
    CrossrefPubMedGoogle Scholar
  • 48 Viskin S. Long QT syndromes and torsade de pointes. Lancet 1999; 354: 1625-33.
    CrossrefPubMedGoogle Scholar
  • 49 Brand T. Heart development: molecular insights into cardiac specification and early morphogenesis. Dev Biol 2003; 258: 1-19.
    CrossrefPubMedGoogle Scholar
  • 50 van den Eijnden-van Raaij AJ, van Achterberg TA, van der Kruijssen CM. et al. Differentiation of aggregated murine P19 embryonal carcinoma cells is induced by a novel visceral endoderm-specific FGF-like factor and inhibited by activin A. Mech Dev 1991; 33: 157-65.
    CrossrefPubMedGoogle Scholar
  • 51 Mummery C, Ward D, van den Brink CE. et al. Cardiomyocyte differentiation of mouse and human embryonic stem cells. J Anat 2002; 200: 233-42.
    CrossrefPubMedGoogle Scholar
  • 52 Fukuda K. Development of regenerative cardiomyocytes from mesenchymal stem cells for cardiovascular tissue engineering. Artif Organs 2001; 25: 187-93.
    CrossrefPubMedGoogle Scholar
  • 53 Behfar A, Zingman LV, Hodgson D M. et al. Stem cell differentiation requires a paracrine pathway in the heart. FASEB J. 2002; 16: 1558-66.
    CrossrefPubMedGoogle Scholar
  • 54 Wobus AM, Kaomei G, Shan J. et al. Retinoic acid accelerates embryonic stem cell-derived cardiac differentiation and enhances development of ventricular cardiomyocytes. J Mol Cell Cardiol 1997; 29: 1525-39.
    CrossrefPubMedGoogle Scholar
  • 55 McBurney MW, Jones-Villeneuve EM, Edwards M K. et al. Control of muscle and neuronal differentiation in a cultured embryonal carcinoma cell line. Nature 1982; 299: 165-7.
    CrossrefPubMedGoogle Scholar
  • 56 Ventura C, Maioli M. Opioid peptide gene expression primes cardiogenesis in embryonal pluripotent stem cells. Circ.Res. 2000; 87: 189-94.
    CrossrefPubMedGoogle Scholar
  • 57 Ginis I, Luo Y, Miura T. et al. Differences between human and mouse embryonic stem 4 cells. Dev Biol 2004; 269: 360-80.
    CrossrefPubMedGoogle Scholar
  • 58 Heng BC, Haider H Kh. et al. Strategies for directing the differentiation of stem cells into the cardiomyogenic lineagein vitro . Cardiovasc Res 2004; 62: 34-42.
    CrossrefPubMedGoogle Scholar
  • 59 Sachinidis A, Fleischmann BK, Kolossov E. et al. Cardiac specific differentiation of mouse embryonic stem cells. Cardiovasc Res 2003; 58: 278-91.
    CrossrefPubMedGoogle Scholar
  • 60 Grepin C, Nemer G, Nemer M. Enhanced cardiogenesis in embryonic stem cells overexpressing the GATA-4 transcription factor. Development 1997; 124: 2387-95.
    CrossrefPubMedGoogle Scholar
  • 61 Gerecht-Nir S, Cohen S, Ziskind A. et al. Three-dimensional porous alginate scaffolds provide a conducive environment for generation of well-vascularized embryoid bodies from human embryonic stem cells. Biotechnol Bioeng 2004; 88: 313-20.
    CrossrefPubMedGoogle Scholar
  • 62 Dang SM, Gerecht-Nir S, Chen J. et al. Controlled, scalable embryonic stem cell differentiation culture. Stem Cells 2004; 22: 275-82.
    CrossrefPubMedGoogle Scholar
  • 63 Klug MG, Soonpaa MH, Koh G Y. et al. Genetically selected cardiomyocytes from differentiating embronic stem cells form stable intracardiac grafts. J Clin Invest 1996; 98: 216-24.
    CrossrefPubMedGoogle Scholar
  • 64 Meyer N, Jaconi M, Landopoulou A. et al. A fluorescent reporter gene as a marker for ventricular specification in ES-derived cardiac cells. FEBS Lett 2000; 478: 151-8.
    CrossrefPubMedGoogle Scholar
  • 65 Muller M, Fleischmann BK, Selbert S. et al. Selection of ventricular-like cardiomyocytes from ES cellsin vitro . FASEB J. 2000; 14: 2540-8.
    CrossrefPubMedGoogle Scholar
  • 66 Allegrucci C, Denning C, Steele B. et al. Human embryonic stem cells as a model for nutritional programming: An evaluation. Reprod Toxicol. 2005 In press.
    PubMedGoogle Scholar
  • 67 Vallier L, Rugg-Gunn PJ, Bouhon I A. et al. Enhancing and diminishing gene function in human embryonic stem cells. Stem Cells 2004; 22: 2-11.
    CrossrefPubMedGoogle Scholar
  • 68 Heng BC, Liu H, Cao T. Feeder cell density-a key parameter in human embryonic stem cell culture. In vitro Cell Dev Biol Anim 2004; 40: 255-7.
    CrossrefPubMedGoogle Scholar
  • 69 Xu RH, Peck R M. et al. Basic FGF and suppression of BMP signaling sustain undifferentiated proliferation of human ES cells. Nat Methods 2005; 2: 185-90.
    CrossrefPubMedGoogle Scholar
  • 70 Xu C, Inokuma MS, Denham J. et al. Feeder-free growth of undifferentiated human embryonic stem cells. Nat Biotechnol 2001; 19: 971-4.
    CrossrefPubMedGoogle Scholar
  • 71 Carpenter MK, Rosler ES, Fisk G J. et al. Properties of four human embryonic stem cell lines maintained in a feeder-free culture system. Dev Dyn 2004; 229: 243-58.
    CrossrefPubMedGoogle Scholar
  • 72 Xu C, Rosler E, Jiang J. et al. Basic fibroblast growth factor supports undifferentiated human embryonic stem cell growth without conditioned medium. Stem Cells 2005; 23: 315-23.
    CrossrefPubMedGoogle Scholar
  • 73 Amit M, Shariki C, Margulets V. et al. Feeder layerand serum-free culture of human embryonic stem cells. Biol Reprod 2004; 70: 837-45.
    CrossrefPubMedGoogle Scholar
  • 74 Mitalipova MM, Rao RR, Hoyer D M. et al. Preserving the genetic integrity of human embryonic stem cells. Nat Biotechnol 2005; 23: 19-20.
    CrossrefPubMedGoogle Scholar
  • 75 Brimble SN, Zeng X, Weiler D A. et al. Karyotypic stability, genotyping, differentiation, feeder-free maintenance, and gene expression sampling in three human embryonic stem cell lines derived prior to August 9. Stem Cells Dev 2004; 13: 585-97.
    CrossrefPubMedGoogle Scholar
  • 76 Draper JS, Smith K, Gokhale P. et al. Recurrent gain of chromosomes 17q and 12 in cultured human embryonic stem cells. Nat Biotechnol 2004; 22: 53-4.
    CrossrefPubMedGoogle Scholar
  • 77 Lakshmipathy U, Pelacho B, Sudo K. et al. Efficient transfection of embryonic and adult stem cells. Stem Cells 2004; 22: 531-43.
    CrossrefPubMedGoogle Scholar
  • 78 Menasche P, Hagege AA, Vilquin J T. et al. Autologous skeletal myoblast transplantation for severe postinfarction left ventricular dysfunction. J Am Coll Cardiol 2003; 41: 1078-83.
    CrossrefPubMedGoogle Scholar
  • 79 Hertzberg RP, Pope AJ. High-throughput screening: new technology for the 21st century. Curr Opin Chem Biol 2000; 4: 445-51.
    CrossrefPubMedGoogle Scholar
  • 80 McNeish J. Embryonic stem cells in drug discovery. Nat Rev Drug Discov 2004; 3: 70-80.
    CrossrefPubMedGoogle Scholar
  • 81 Ezashi T, Das P, Roberts RM. Low O2 tensions and the prevention of differentiation of hES cells. Proc Natl Acad Sci U S A. 2005; 102: 4783-8.
    CrossrefPubMedGoogle Scholar
  • 82 Sato N, Meijer L, Skaltsounis L. et al. Maintenance of pluripotency in human and mouse embryonic stem cells through activation of Wnt signaling by a pharmacological GSK-3-specific inhibitor. Nat Med 2004; 10: 55-63.
    CrossrefPubMedGoogle Scholar
  • 83 Zandstra PW, Bauwens C, Yin T. et al. Scalable production of embryonic stem cell-derived cardiomyocytes. Tissue Eng 2003; 9: 767-78.
    CrossrefPubMedGoogle Scholar
  • 84 Wobus AM, Boheler KR. Embryonic stem cells: prospects for developmental biology and cell therapy. Physiol Rev 2005; 85: 635-78.
    CrossrefPubMedGoogle Scholar
  • 85 Richards M, Fong CY, Chan W K. et al. Human feeders support prolonged undifferentiated growth of human inner cell masses and embryonic stem cells. Nat Biotechnol 2002; 20: 933-6.
    CrossrefPubMedGoogle Scholar
  • 86 Stojkovic P, Lako M, Stewart R. et al. An autogeneic feeder cell system that efficiently supports growth of undifferentiated human embryonic stem cells. Stem Cells 2005; 23: 306-14.
    CrossrefPubMedGoogle Scholar
  • 87 Inzunza J, Gertow K, Stromberg M A. et al. Derivation of human embryonic stem cell lines in serum replacement medium using postnatal human fibroblasts as feeder cells. Stem Cells 2005; 23: 544-9.
    CrossrefPubMedGoogle Scholar
  • 88 Martin MJ, Muotri A, Gage F. et al. Human embryonic stem cells express an immunogenic nonhuman sialic acid. Nat Med 2005; 11: 228-32.
    CrossrefPubMedGoogle Scholar
  • 89 Caspi O, Gepstein L. Potential applications of human embryonic stem cell-derived cardiomyocytes. Ann N Y Acad Sci 2004; 1015: 285-98.
    CrossrefPubMedGoogle Scholar
  • 90 Menasche P. Skeletal myoblast transplantation for cardiac repair. Expert Rev Cardiovasc Ther 2004; 2: 21-2.
    CrossrefPubMedGoogle Scholar
  • 91 Zhang M, Methot D, Poppa V. et al. Cardiomyocyte grafting for cardiac repair: graft cell death and antideath strategies. J Mol Cell Cardiol 2001; 33: 907-21.
    CrossrefPubMedGoogle Scholar
  • 92 Leor J, Aboulafia-Etzion S, Dar A. et al. Bioengineered cardiac grafts: A new approach to repair the infarcted myocardium?. Circulation 2000; 102: III56-61.
    CrossrefPubMedGoogle Scholar
  • 93 Shin M, Ishii O, Sueda T. et al. Contractile cardiac grafts using a novel nanofibrous mesh. Biomaterials 2004; 25: 3717-23.
    CrossrefPubMedGoogle Scholar
  • 94 Palmen M, Daemen M J. et al. Cardiac remodeling after myocardial infarction is impaired in IGF-1 deficient mice. Cardiovasc Res 2001; 50: 516-24.
    CrossrefPubMedGoogle Scholar
  • 95 Cheng YC, Huang ES, Lin J C. et al. Unique spectrum of activity of 9-[(1,3-dihydroxy-2-propoxy) methyl]-guanine against herpesvirusesin vitro and its mode of action against herpes simplex virus type 1. Proc Natl Acad Sci U S A. 1983; 80: 2767-70.
    CrossrefPubMedGoogle Scholar
  • 96 Schuldiner M, Yanuka O, Itskovitz-Eldor J. et al. Effects of eight growth factors on the differentiation of cells derived from human embryonic stem cells. Proc Natl Acad Sci U S A 2000; 97: 11307-12.
    CrossrefPubMedGoogle Scholar
  • 97 Markham A, Faulds D. Ganciclovir. An update of its therapeutic use in cytomegalovirus infection. Drugs 1994; 48: 455-84.
    CrossrefPubMedGoogle Scholar
  • 98 Denning C, Pitts JD. Bystander effects of different enzyme-prodrug systems for cancer gene therapy depend on different pathways for intercellular transfer of toxic metabolites, a factor that will govern clinical choice of appropriate regimes. Hum Gene Ther 1997; 8: 1825-35
    CrossrefPubMedGoogle Scholar
  • 99 Nishiyama T, Kawamura Y, Kawamoto K. et al. Antineoplastic effects in rats of 5-fluorocytosine in combination with cytosine deaminase capsules. Cancer Res 1985; 45: 1753-61.
    PubMedGoogle Scholar
  • 100 Millan MT, Shizuru J A. et al. Mixed chimerism and immunosuppressive drug withdrawal after HLA-mismatched kidney and hematopoietic progenitor transplantation. Transplantation 2002; 73: 1386-91.
    CrossrefPubMedGoogle Scholar