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Journal of Muscle Research & Cell Motility

, Volume 17, Issue 3, pp 313–334 | Cite as

Remodelling of adult cardiac muscle cells in culture: dynamic process of disorganization and reorganization of myofibrils

  • Asish C. Nag
  • Mei -Li Lee
  • Fazlul H. Sarkar
Papers

Summary

The myofibrils of adult rat cardiac muscle cells in long-term culture initially break down and later reassemble into mature myofibrils. The objective of this study is to examine the disorganization process of myofibrils and to determine how disorganized myofibrillar proteins, myosin, titin, actin, and α-actinin are reorganized into mature myofibrils in adult cells. Atter dismantlement of myofibrils during initial culture period (24–72 h), myofibrillar proteins became disorganized into amorphous form. These proteins later were observed in vesicular, amorphous, and nonstriated fibrillar forms. Some vesicular structures, containing mainly myosin, titin, α-actin, and α-actinin were observed on the outer surfaces of the cell and outside the cell body. Such vesicles containing F-actin were rare. Punctate structures of α-actinin emerged from the pre-existing amorphous α-actinin along with the appearance of mostly titin periodicities. The periodicities of α-actinin later became prevalent, followed by the appearance of periodicities of actin. α-actinin provided an initiation point on which titin and actin became associated, forming titin-associated I-Z-I structures. Titin traversed the I-bands on either side of the Z-line. The phalloidin-stained I-Z-I structures bound to antibodies to muscle specific sarcomeric proteins (titin, α-actin, α-actinin). The differentiation of myosin periodicities lagged behind those of titin, α-actinin, and actin although presarcomeric structures of immunolabelled titin and myosin were very closely linked in their distributions in the formative myofibrils. Variations in the temporal sequence of emergence of periodicities of α-actinin and myosin were observed among certain myocytes. Also observed was the variation of the temporal sequence of emergence of titin and actin periodicities among different myocytes and within a single myocyte. Even in the late stage of culture (30 days), when the cell body was packed with myofibrils, the myocytes contained remnants of amorphous myofibrillar proteins.

Keywords

Temporal Sequence Myofibrillar Protein Amorphous Form Initial Culture Cardiac Muscle Cell 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. Antin P. B., Forry-Schaudies S., Friedman T. M., Tapscott S. J. & Holtzer H. (1981) Taxol induces postmitotic myoblasts to assemble interdigitating microtubule-myosin arrays that exclude actin filaments. J. Cell Biol. 89, 300–8.Google Scholar
  2. Antin P. B., Tokunaka S., Nachmias V. T. & Holtzer H. (1986) Role of stress-fiber-like structures in assembling nascent myofibrils in myosheets recovering from exposure to ethyl methanesulfonate. J. Cell Biol. 102, 1464–79.Google Scholar
  3. Brooks W. H., Connell S., Cannata J., Maloney J. E. & Walker A. M. (1984) Ultrastructure of the myocardium during development from early fetal life to adult life in sheep. J. Anat. 137, 729–41.Google Scholar
  4. Clark W. A. & Samarel A. M. (1988) Evaluation of rates of protein synthesis and degradation in isolated cardiac muscle cells. In Biology of Isolated Adult Cardiac Myocytes. (edited by W. A. Clark, R. S. Decker & T. K. Borg) pp. 131–46. New York: Elsevier.Google Scholar
  5. Clark W. A. & Zak R. (1981) Assessment of fractional rates of protein synthesis in cardiac muscle cultures after equilibrium labeling. J. Biol. Chem. 256, 4863–70.Google Scholar
  6. Clark W. A., Chizzonite R. A., Everett A. W., Rabinowitz M. & Zak R. (1982) Species correlations between cardiac isomyosins. A comparison of electrophoretic and immunological properties. J. Biol. Chem. 257, 5449–54.Google Scholar
  7. Clark W. A., Rudmnick S. J., Lapres J. J., Lesch M. & Decker R. S. (1991) Cultured adult feline cardiac myocytes in long-term culture: morphological differentiation and hypertrophy induced following β-adrenergic activation of beating. Am. J. Physiol. 261, C530–42.Google Scholar
  8. Claycomb W. C., Palazzo M. S. (1980) Culture of the terminally differentiated adult cardiac muscle cells: a light and scanning electron microscopy study. Devel. Biol. 80, 446–82.Google Scholar
  9. Craig S. W. & Parodo J. V. (1979) Alpha-actinin localization in the junctional complex of intestinal epithelial cells. J. Cell Biol. 80, 203–10.Google Scholar
  10. Dlugosz A. A., Antin P. B., Nachmias V. T. & Holtzer H. (1984) The relationship between stress fiber-like structures and nascent myofibrils in cultured cardiac myocytes. J. Cell Biol. 99, 2268–78.Google Scholar
  11. Eppenberger M. E., Hauser I., Baechi T., Schaub M. C., Brunner U. T., Dechesne C. A. & Eppenberger H. M. (1988) Immunocytochemical analysis of the regeneration of myofibrils in long-term cultures of adult cardiomyocytes of the rat. Devel. Biol. 130, 1–15.Google Scholar
  12. Fürst D. O., Osborn M. & Weber K. (1989) Myogenesis in the mouse embryo: differential onset of expression of myogenic proteins and the involvement of titin in myofibril assembly. J. Cell Biol. 109, 517–27.Google Scholar
  13. Hill C. S., Duran S., Lin Z., Weber K. & Holtzer H. (1986) Titin and myosin, but not desmin, are linked during myofibrillogenesis in postmitotic mononucleated myoblasts. J. Cell Biol. 103, 2185–96.Google Scholar
  14. Horowits R., Maruyama K. & Podolsky R. J. (1989) Elastic behavior of connectin filaments during thick filament movement in activated skeletal muscle. J. Cell Biol. 109, 2169–76.Google Scholar
  15. Imanaka-Yoshinda K., Sanger J. M. & Sanger J. W. (1993) Contractile protein dynamics of myofibrils in paired adult rat cardiomyocytes. Cell Motil. Cytoskel. 26, 301–12.Google Scholar
  16. Isobe Y., Warner F. D. & Lemanski L. F. (1988) Three-dimensional immunogold localization of α-actinin within the cytoskeletal networks of cultured cardiac and nonmuscle cells. Proc. Natl Acad. Sci. USA 85, 6758–62.Google Scholar
  17. Itoh Y., Suzuki T., Kimura S., Ohasi K., Higuchi H., Sawada H., Shimuzu T., Shibata M. & Maruyama K. (1988) Extensible and less extensible domains of connectin filaments in stretched vertebrate skeletal muscle sarcomers as detected by immunofluorescence and immunoelectron microscopy using monoclonal antibodies. J. Biochem. 104, 504–8.Google Scholar
  18. Jacobson S. L. (1977) Culture of spontaneously contractin myocardial cells from adult rats. Cell Struct. Funct. 2, 1–9.Google Scholar
  19. Komiyama M., Toyota N. & Shimada Y. (1989) Morphogenesis of myofibrils in cultures of cardiac myocytes. In Mechanobiological Research on the Masticatory System (edited by K. Kubota) pp. 183–7. Berlin: VEB Verlage für Medizin und Biologie.Google Scholar
  20. Komiyama M., Maruyama K. & Shimada Y. (1990) Assembly of connectin (titin) in relation to myosin and α-actinin in cultured cardiac myocytes. J. Muscle Res. Cell Motil. 11, 419–28.Google Scholar
  21. Komiyama M., Zhen-Hua Z., Maruyama K. & Shimada Y. (1992) Spatial relationship of nebulin relative to other myofibrillar proteins during myogenesis in embryonic chick skeletal muscle cells in vitro. J. Muscle Res. Cell Motil. 13, 48–54.Google Scholar
  22. Kouchi K., Takahashi H. & Shimada Y. (1993) Incorporation of microinjected biotin-labelled actin into nascent myofibrils of cardiac myocytes: an immunoelectron microscopic study. J. Muscle Res. Cell Motil. 14, 292–301.Google Scholar
  23. Legato M. H. (1972) Ultrastructural characteristics of the rat ventricular cell grown in tissue culture, with special reference to sarcomerogenesis. J. Mol. Cell. Cardiol. 4, 299–317.Google Scholar
  24. Lessard J. L. (1988) Two monoclonal antibodies to actin, one generally reactive and the other muscle selective. Cell Motil. Cytoskel. 10, 349–62.Google Scholar
  25. Lin Z., Eshleman J., Forry-Schandies S., Duran S., Lessard J. & Holtzer H. (1987) Sequential disasembly of myofibrils induced by phorbol myristate acetate in cultured myotubes. J. Cell Biol. 105, 1365–76.Google Scholar
  26. Lin Z., Holtzer S., Schultheiss T., Murray J., Masaki T., Fischman D. A. & Holtzer H. (1989) Polygons and adhesion plaques and the disassembly and assembly of myofibrils in cardiac myocytes. J. Cell Biol. 108, 2355–67.Google Scholar
  27. Markwald R. R. (1973) Distribution and relationship of precursor Z material to organizing myofibrillar bundles in embryonic rat and hamster ventricular myocytes. J. Mol. Cell. Cardiol. 5, 341–50.Google Scholar
  28. Menko A. S., Croop J., Toyama Y., Holtzer H. & Boettiger D. (1982) The response of chicken embryo dermal fibroblasts to cytochalasin B is altered by Rous sarcoma virus-induced cell transformation. Mol. Cell. Biol. 2, 320–30.Google Scholar
  29. Moses R. L. & Claycomb W. C. (1982) Disorganization and reestablishment of cardiac muscle cell ultrastructure in cultured adult rat ventricular muscle cells. J. Ultrastruct. Res. 81, 358–74.Google Scholar
  30. Nag A. C. & Cheng M. (1981) Adult mammalian cardiac muscle cells in culture. Tissue Cell 13, 515–21.Google Scholar
  31. Nag A. C. & Lee M. L. (1992) TPA has no influence on the expression of myosin heavy chain isoforms in cultured adult cardiac muscle cells. J. Cell Biochem. 49, 399–409.Google Scholar
  32. NAG, A. C., FISCHMAN, D. A., RABINOWITZ, M. & ZAK, R. (1974) Cell culture of adult rat cardiac muscle. J. Cell Biol. 63, 238a.Google Scholar
  33. Nag A. C., Cheng M., Fischman D. A. & Zak R. (1983) Long-term culture of adult mammalian cardiac myocytes: electron microscopic and immunofluorescent analyses of myofibrillar structure. J. Mol. Cell. Cardiol. 15, 301–17.Google Scholar
  34. Nag A. C., England M. & Cheng M. (1985) Factors controlling embryonic heart cell proliferation in serumfree synthetic media. Nitro Cell. Dev. Biol. 21, 553–62.Google Scholar
  35. Nag A. C., Lee M. L. & Kosier J. (1990) Adult cardiac muscle cells in long-term serum-free culture: myofibrillar organization and expression of myosin heavy chain isoforms. Vitro Cell. Dev. Biol. 21, 464–70.Google Scholar
  36. Rhee D., Sanger J. & Sanger J. (1994) The premyofibril: evidence for its role in myofibrillogenesis. Cell Motil. Cytoskel. 28, 1–24.Google Scholar
  37. Sanger J. M., Mittal B., Pochapin M. B. & Sanger J. W. (1986) Myofibrillogenesis in living cells microinjected with fluorescently labeled alpha-actinin. J. Cell Biol. 102, 2053–66.Google Scholar
  38. Schwarzfeld T. A. & Jacobson S. L. (1981) Isolation and development in cell culture of myocardiac cells of the adult rat. J. Mol. Cell. Cardol. 13, 563–75.Google Scholar
  39. Schultheiss T., Lin Z., Lu M-H., Murray J., Fischman D. A., Weber K., Masaki T., Imamura M. & Holtzer H. (1990) Differential distribution of subsets of myofibrillar proteins in cardiac nonstriated and striated myofibrils. J. Cell Biol. 110, 1159–72.Google Scholar
  40. Shimada Y., Komiyama M., Terai M. & Maruyama K. (1990) Early phases of myofibril assembly in embryonic chick cardiac myocytes in vitro. In Etiology and Morphogenesis of Congenital Heart Disease (edited by A. Tako) pp. 67–77. New York: Futura.Google Scholar
  41. Shimizu T., Matsumura Y., Itoh T., Mannen T. & Maruyama K. (1988) An immunological homology between neurofilament and muscle elastic filament: a monoclonal antibody cross-reacts with neurofilament subunits and connectin. Biomed. Res. 9, 227–33.Google Scholar
  42. Tokuyasu K. T. & Maher P. A. (1987) Immunocytochemical studies of cardiac myofibrillogenesis in early chick embryos. I. Presence of immunofluorescent titin spots in premyofibril stages. J. Cell Biol. 105, 2781–93.Google Scholar
  43. Terai M., Komiyama M. & Shimada Y. (1989) Myofibril assembly is linked with vinculin, α-actinin, and cell substrate contacts in embryonic cardiac myocytes in vitro. Cell Motil. Cytoskel. 12, 185–94.Google Scholar
  44. Wang S.-M., Greaser M. L., Schultz E., Bulinski J. C., Lin J. J-C. & Lessard J. L. (1988) Studies on cardiac myofibrillogenesis with antibodies to titin, actin, tropomyosin, and myosin. J. Cell Biol. 107, 1075–83.Google Scholar
  45. Wulf E., Deboben A., Bautz F. A., Faulstich H. & Weiland T. (1979) Fluorescent phallotoxin, a tool for the visualization of cellular actin. Proc. Natl. Acad. Sci. USA 76, 4498–502.Google Scholar

Copyright information

© Chapman & Hall 1996

Authors and Affiliations

  • Asish C. Nag
    • 1
  • Mei -Li Lee
    • 1
  • Fazlul H. Sarkar
    • 2
  1. 1.Department of Biological SciencesOakland UniversityRochester
  2. 2.Department of PathologyWayne State University School of MedicineDetroitUSA

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