Skip to main content
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1989 Mar 1;108(3):1067–1078. doi: 10.1083/jcb.108.3.1067

Growth and partial differentiation of presumptive human cardiac myoblasts in culture

PMCID: PMC2115380  PMID: 2522096

Abstract

A cell culture model for human cardiac myogenesis is introduced. Human fetal myocardial cells were dissociated enzymatically, and cultured in a mitogen-rich medium that promoted the growth of presumptive cardiac myoblasts. Strains of human cardiac myoblasts were generated from different anatomical regions of the fetal heart. The cells could be cultured for at least 30 generations, or frozen and recovered for later use. Differentiation was induced by culturing the cardiac myoblasts in a mitogen-poor medium. Differentiation of cardiac myoblasts was marked primarily by transcriptional activation of the atrial natriuretic factor (ANF) gene. Evidence is presented that posttranscriptional processing of ANF transcripts is affected by the anatomical origin of the cardiac myoblasts and the presence of cocultured neuronal cells. Cardiac myoblasts induced to differentiate in culture synthesized only low levels of sarcomeric myosin and cardiac alpha-actin, suggesting that differentiation of these cells progresses through two phases: an initial, noncontractile phase that is represented by the differentiating cultured cells; and a later contractile phase, in which myofibrillar assembly is accentuated and modulated by secondary signals from the cardiac milieu.

Full Text

The Full Text of this article is available as a PDF (3.1 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bader D., Masaki T., Fischman D. A. Immunochemical analysis of myosin heavy chain during avian myogenesis in vivo and in vitro. J Cell Biol. 1982 Dec;95(3):763–770. doi: 10.1083/jcb.95.3.763. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bains W., Ponte P., Blau H., Kedes L. Cardiac actin is the major actin gene product in skeletal muscle cell differentiation in vitro. Mol Cell Biol. 1984 Aug;4(8):1449–1453. doi: 10.1128/mcb.4.8.1449. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bloch K. D., Seidman J. G., Naftilan J. D., Fallon J. T., Seidman C. E. Neonatal atria and ventricles secrete atrial natriuretic factor via tissue-specific secretory pathways. Cell. 1986 Dec 5;47(5):695–702. doi: 10.1016/0092-8674(86)90512-x. [DOI] [PubMed] [Google Scholar]
  4. Clark W. A., Jr, Chizzonite R. A., Everett A. W., Rabinowitz M., Zak R. Species correlations between cardiac isomyosins. A comparison of electrophoretic and immunological properties. J Biol Chem. 1982 May 25;257(10):5449–5454. [PubMed] [Google Scholar]
  5. Claycomb W. C. Biochemical aspects of cardiac muscle differentiation. Possible control of deoxyribonucleic acid synthesis and cell differentiation by adrenergic innervation and cyclic adenosine 3':5'-monophosphate. J Biol Chem. 1976 Oct 10;251(19):6082–6089. [PubMed] [Google Scholar]
  6. Clegg C. H., Linkhart T. A., Olwin B. B., Hauschka S. D. Growth factor control of skeletal muscle differentiation: commitment to terminal differentiation occurs in G1 phase and is repressed by fibroblast growth factor. J Cell Biol. 1987 Aug;105(2):949–956. doi: 10.1083/jcb.105.2.949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Cummins P., Lambert S. J. Myosin transitions in the bovine and human heart. A developmental and anatomical study of heavy and light chain subunits in the atrium and ventricle. Circ Res. 1986 Jun;58(6):846–858. doi: 10.1161/01.res.58.6.846. [DOI] [PubMed] [Google Scholar]
  8. Dulak N. C., Temin H. M. A partially purified polypeptide fraction from rat liver cell conditioned medium with multiplication-stimulating activity for embryo fibroblasts. J Cell Physiol. 1973 Apr;81(2):153–160. doi: 10.1002/jcp.1040810204. [DOI] [PubMed] [Google Scholar]
  9. Etlinger J. D., Zak R., Fischman D. A. Compositional studies of myofibrils from rabbit striated muscle. J Cell Biol. 1976 Jan;68(1):123–141. doi: 10.1083/jcb.68.1.123. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Ewton D. Z., Florini J. R. Relative effects of the somatomedins, multiplication-stimulating activity, and growth hormone on myoblasts and myotubes in culture. Endocrinology. 1980 Feb;106(2):577–583. doi: 10.1210/endo-106-2-577. [DOI] [PubMed] [Google Scholar]
  11. Flink I. L., Rader J. H., Banerjee S. K., Morkin E. Atrial and ventricular cardiac myosins contain different heavy chain species. FEBS Lett. 1978 Oct 1;94(1):125–130. doi: 10.1016/0014-5793(78)80921-1. [DOI] [PubMed] [Google Scholar]
  12. Garcia R., Cantin M., Thibault G. Role of right and left atria in natriuresis and atrial natriuretic factor release during blood volume changes in the conscious rat. Circ Res. 1987 Jul;61(1):99–106. doi: 10.1161/01.res.61.1.99. [DOI] [PubMed] [Google Scholar]
  13. Gospodarowicz D. Purification of a fibroblast growth factor from bovine pituitary. J Biol Chem. 1975 Apr 10;250(7):2515–2520. [PubMed] [Google Scholar]
  14. Gospodarowicz D., Weseman J., Moran J. S., Lindstrom J. Effect of fibroblast growth factor on the division and fusion of bovine myoblasts. J Cell Biol. 1976 Aug;70(2 Pt 1):395–405. doi: 10.1083/jcb.70.2.395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Greenberg M. E., Ziff E. B. Stimulation of 3T3 cells induces transcription of the c-fos proto-oncogene. Nature. 1984 Oct 4;311(5985):433–438. doi: 10.1038/311433a0. [DOI] [PubMed] [Google Scholar]
  16. Gunning P., Ponte P., Okayama H., Engel J., Blau H., Kedes L. Isolation and characterization of full-length cDNA clones for human alpha-, beta-, and gamma-actin mRNAs: skeletal but not cytoplasmic actins have an amino-terminal cysteine that is subsequently removed. Mol Cell Biol. 1983 May;3(5):787–795. doi: 10.1128/mcb.3.5.787. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hamid Q., Wharton J., Terenghi G., Hassall C. J., Aimi J., Taylor K. M., Nakazato H., Dixon J. E., Burnstock G., Polak J. M. Localization of atrial natriuretic peptide mRNA and immunoreactivity in the rat heart and human atrial appendage. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6760–6764. doi: 10.1073/pnas.84.19.6760. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Heintz N., Sive H. L., Roeder R. G. Regulation of human histone gene expression: kinetics of accumulation and changes in the rate of synthesis and in the half-lives of individual histone mRNAs during the HeLa cell cycle. Mol Cell Biol. 1983 Apr;3(4):539–550. doi: 10.1128/mcb.3.4.539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hoh J. F., McGrath P. A., Hale P. T. Electrophoretic analysis of multiple forms of rat cardiac myosin: effects of hypophysectomy and thyroxine replacement. J Mol Cell Cardiol. 1978 Nov;10(11):1053–1076. doi: 10.1016/0022-2828(78)90401-7. [DOI] [PubMed] [Google Scholar]
  20. Izumo S., Nadal-Ginard B., Mahdavi V. All members of the MHC multigene family respond to thyroid hormone in a highly tissue-specific manner. Science. 1986 Feb 7;231(4738):597–600. doi: 10.1126/science.3945800. [DOI] [PubMed] [Google Scholar]
  21. Kikuchi K., Nakao K., Hayashi K., Morii N., Sugawara A., Sakamoto M., Imura H., Mikawa H. Ontogeny of atrial natriuretic polypeptide in the human heart. Acta Endocrinol (Copenh) 1987 Jun;115(2):211–217. doi: 10.1530/acta.0.1150211. [DOI] [PubMed] [Google Scholar]
  22. Kirby M. L., Gale T. F., Stewart D. E. Neural crest cells contribute to normal aorticopulmonary septation. Science. 1983 Jun 3;220(4601):1059–1061. doi: 10.1126/science.6844926. [DOI] [PubMed] [Google Scholar]
  23. Kohtz D. S., Georgieva-Hanson V., Kohtz J. D., Schook W. J., Puszkin S. Mapping two functional domains of clathrin light chains with monoclonal antibodies. J Cell Biol. 1987 Apr;104(4):897–903. doi: 10.1083/jcb.104.4.897. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lemanski L. F., Mooseker M. S., Peachey L. D., Iyengar M. R. Studies of muscle proteins in embryonic myocardial cells of cardiac lethal mutant mexican axolotls (Ambystoma mexicanum) by use of heavy meromyosin binding and sodium dodecyl sulfate polyacrylamide gel electrophoresis. J Cell Biol. 1976 Feb;68(2):375–388. doi: 10.1083/jcb.68.2.375. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lompré A. M., Nadal-Ginard B., Mahdavi V. Expression of the cardiac ventricular alpha- and beta-myosin heavy chain genes is developmentally and hormonally regulated. J Biol Chem. 1984 May 25;259(10):6437–6446. [PubMed] [Google Scholar]
  26. Mahdavi V., Chambers A. P., Nadal-Ginard B. Cardiac alpha- and beta-myosin heavy chain genes are organized in tandem. Proc Natl Acad Sci U S A. 1984 May;81(9):2626–2630. doi: 10.1073/pnas.81.9.2626. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mahdavi V., Periasamy M., Nadal-Ginard B. Molecular characterization of two myosin heavy chain genes expressed in the adult heart. Nature. 1982 Jun 24;297(5868):659–664. doi: 10.1038/297659a0. [DOI] [PubMed] [Google Scholar]
  28. Maki M., Takayanagi R., Misono K. S., Pandey K. N., Tibbetts C., Inagami T. Structure of rat atrial natriuretic factor precursor deduced from cDNA sequence. Nature. 1984 Jun 21;309(5970):722–724. doi: 10.1038/309722a0. [DOI] [PubMed] [Google Scholar]
  29. Manasek F. J. Histogenesis of the embryonic myocardium. Am J Cardiol. 1970 Feb;25(2):149–168. doi: 10.1016/0002-9149(70)90576-x. [DOI] [PubMed] [Google Scholar]
  30. Perez-Polo J. R., Werbach-Perez K., Tiffany-Castiglioni E. A human clonal cell line model of differentiating neurons. Dev Biol. 1979 Aug;71(2):341–355. doi: 10.1016/0012-1606(79)90174-x. [DOI] [PubMed] [Google Scholar]
  31. Powell T., Twist V. W. A rapid technique for the isolation and purification of adult cardiac muscle cells having respiratory control and a tolerance to calcium. Biochem Biophys Res Commun. 1976 Sep 7;72(1):327–333. doi: 10.1016/0006-291x(76)90997-9. [DOI] [PubMed] [Google Scholar]
  32. Price K. M., Littler W. A., Cummins P. Human atrial and ventricular myosin light-chains subunits in the adult and during development. Biochem J. 1980 Nov 1;191(2):571–580. doi: 10.1042/bj1910571. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  34. Samuel J. L., Rappaport L., Mercadier J. J., Lompre A. M., Sartore S., Triban C., Schiaffino S., Schwartz K. Distribution of myosin isozymes within single cardiac cells. An immunohistochemical study. Circ Res. 1983 Feb;52(2):200–209. doi: 10.1161/01.res.52.2.200. [DOI] [PubMed] [Google Scholar]
  35. Sartore S., Pierobon-Bormioli S., Schiaffino S. Immunohistochemical evidence for myosin polymorphism in the chicken heart. Nature. 1978 Jul 6;274(5666):82–83. doi: 10.1038/274082a0. [DOI] [PubMed] [Google Scholar]
  36. Seidler F. J., Slotkin T. A. Presynaptic and postsynaptic contributions to ontogeny of sympathetic control of heart rate in the pre-weanling rat. Br J Pharmacol. 1979 Mar;65(3):431–434. doi: 10.1111/j.1476-5381.1979.tb07847.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Sonnenberg H., Veress A. T. Cellular mechanism of release of atrial natriuretic factor. Biochem Biophys Res Commun. 1984 Oct 30;124(2):443–449. doi: 10.1016/0006-291x(84)91573-0. [DOI] [PubMed] [Google Scholar]
  38. Spizz G., Roman D., Strauss A., Olson E. N. Serum and fibroblast growth factor inhibit myogenic differentiation through a mechanism dependent on protein synthesis and independent of cell proliferation. J Biol Chem. 1986 Jul 15;261(20):9483–9488. [PubMed] [Google Scholar]
  39. Sréter F. A., Bálint M., Gergely J. Structural and functional changes of myosin during development: comparison with adult fast, slow and cardiac myosin. Dev Biol. 1975 Oct;46(2):317–325. doi: 10.1016/0012-1606(75)90108-6. [DOI] [PubMed] [Google Scholar]
  40. Stewart D. E., Kirby M. L., Sulik K. K. Hemodynamic changes in chick embryos precede heart defects after cardiac neural crest ablation. Circ Res. 1986 Nov;59(5):545–550. doi: 10.1161/01.res.59.5.545. [DOI] [PubMed] [Google Scholar]
  41. Tiemeier D. C., Tilghman S. M., Leder P. Purification and cloning of a mouse ribosomal gene fragment in coliphage lambda. Gene. 1977;2(3-4):173–191. doi: 10.1016/0378-1119(77)90016-6. [DOI] [PubMed] [Google Scholar]
  42. Veress A. T., Sonnenberg H. Right atrial appendectomy reduces the renal response to acute hypervolemia in the rat. Am J Physiol. 1984 Sep;247(3 Pt 2):R610–R613. doi: 10.1152/ajpregu.1984.247.3.R610. [DOI] [PubMed] [Google Scholar]
  43. Yazaki Y., Raben M. S. Cardiac myosin adenosinetriphosphatase of rat and mouse. Distinctive enzymatic properties compared with rabbit and dog cardiac myosin. Circ Res. 1974 Jul;35(1):15–23. doi: 10.1161/01.res.35.1.15. [DOI] [PubMed] [Google Scholar]
  44. Zhong R., Roeder R. G., Heintz N. The primary structure and expression of four cloned human histone genes. Nucleic Acids Res. 1983 Nov 11;11(21):7409–7425. doi: 10.1093/nar/11.21.7409. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES