Abstract
In adult regenerating cardiomyocytes in culture, in contrast to fetal cells, mitochondrial creatine kinase (Mi-CK) was expressed. In the same cell, two populations of mitochondria, differing in shape, in distribution within the cell and in content of Mi-CK, could be distinguished. Immunofluorescence studies using antibodies against Mi- CK revealed a characteristic staining pattern for the two types of mitochondria: giant, mostly cylindrically shaped, and, as shown by confocal laser light microscopy, randomly distributed mitochondria exhibited a strong signal for Mi-CK, whereas small, "normal" mitochondria, localized in rows between myofibrils, gave a much weaker signal. Transmission EM of the giant mitochondria demonstrated paracrystalline inclusions located between cristae membranes. Immunogold labeling with anti-Mi-CK antibodies revealed a specific decoration of these inclusions for Mi-CK. Addition of 20 mM creatine, the substrate of Mi-CK, to the essentially creatine-free culture medium caused the disappearance of the giant cylindrically shaped mitochondria as well as of the paracrystalline inclusions, accompanied by an increase of the intracellular level of total creatine. Replacement of creatine in the medium by the creatine analogue and competitor beta- guanidinopropionic acid caused the reappearance of the enlarged mitochondria. It is believed that the accumulation of Mi-CK within the paracrystalline inclusions, similar to those observed in certain myopathies, represents a compensatory effect of the cardiomyocytes to cope with a metabolic stress situation caused by low intracellular total creatine levels.
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- Adams V., Bosch W., Schlegel J., Wallimann T., Brdiczka D. Further characterization of contact sites from mitochondria of different tissues: topology of peripheral kinases. Biochim Biophys Acta. 1989 Jun 6;981(2):213–225. doi: 10.1016/0005-2736(89)90031-x. [DOI] [PubMed] [Google Scholar]
- Borg T. K., Rubin K., Lundgren E., Borg K., Obrink B. Recognition of extracellular matrix components by neonatal and adult cardiac myocytes. Dev Biol. 1984 Jul;104(1):86–96. doi: 10.1016/0012-1606(84)90038-1. [DOI] [PubMed] [Google Scholar]
- Bähler M., Moser H., Eppenberger H. M., Wallimann T. Heart C-protein is transiently expressed during skeletal muscle development in the embryo, but persists in cultured myogenic cells. Dev Biol. 1985 Dec;112(2):345–352. doi: 10.1016/0012-1606(85)90405-1. [DOI] [PubMed] [Google Scholar]
- Claycomb W. C., Lanson N., Jr Isolation and culture of the terminally differentiated adult mammalian ventricular cardiac muscle cell. In Vitro. 1984 Aug;20(8):647–651. doi: 10.1007/BF02619615. [DOI] [PubMed] [Google Scholar]
- Claycomb W. C., Palazzo M. C. Culture of the terminally differentiated adult cardiac muscle cell: a light and scanning electron microscope study. Dev Biol. 1980 Dec;80(2):466–482. doi: 10.1016/0012-1606(80)90419-4. [DOI] [PubMed] [Google Scholar]
- Duan J. M., Karmazyn M. Acute effects of hypoxia and phosphate on two populations of heart mitochondria. Mol Cell Biochem. 1989 Oct 5;90(1):47–56. doi: 10.1007/BF00225220. [DOI] [PubMed] [Google Scholar]
- Eggleton P., Elsden S. R., Gough N. The estimation of creatine and of diacetyl. Biochem J. 1943;37(5):526–529. doi: 10.1042/bj0370526. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Eppenberger-Eberhardt M., Flamme I., Kurer V., Eppenberger H. M. Reexpression of alpha-smooth muscle actin isoform in cultured adult rat cardiomyocytes. Dev Biol. 1990 Jun;139(2):269–278. doi: 10.1016/0012-1606(90)90296-u. [DOI] [PubMed] [Google Scholar]
- Eppenberger M. E., Hauser I., Baechi T., Schaub M. C., Brunner U. T., Dechesne C. A., Eppenberger H. M. Immunocytochemical analysis of the regeneration of myofibrils in long-term cultures of adult cardiomyocytes of the rat. Dev Biol. 1988 Nov;130(1):1–15. doi: 10.1016/0012-1606(88)90408-3. [DOI] [PubMed] [Google Scholar]
- Eppenberger M. E., Schoenenberger R., Eppenberger H. M. Myofibrillar M-line structure in normal and dystrophic hamster muscle. Muscle Nerve. 1984 May;7(4):304–311. doi: 10.1002/mus.880070409. [DOI] [PubMed] [Google Scholar]
- Farrants G. W., Hovmöller S., Stadhouders A. M. Two types of mitochondrial crystals in diseased human skeletal muscle fibers. Muscle Nerve. 1988 Jan;11(1):45–55. doi: 10.1002/mus.880110109. [DOI] [PubMed] [Google Scholar]
- Fitch C. D., Shields R. P., Payne W. F., Dacus J. M. Creatine metabolism in skeletal muscle. 3. Specificity of the creatine entry process. J Biol Chem. 1968 Apr 25;243(8):2024–2027. [PubMed] [Google Scholar]
- Hanzlíková V., Schiaffino S. Mitochondrial changes in ischemic skeletal muscle. J Ultrastruct Res. 1977 Jul;60(1):121–133. doi: 10.1016/s0022-5320(77)80048-8. [DOI] [PubMed] [Google Scholar]
- Kamieniecka Z., Schmalbruch H. Neuromuscular disorders with abnormal muscle mitochondria. Int Rev Cytol. 1980;65:321–357. doi: 10.1016/s0074-7696(08)61964-6. [DOI] [PubMed] [Google Scholar]
- Karpati G., Carpenter S., Melmed C., Eisen A. A. Experimental ischemic myopathy. J Neurol Sci. 1974 Sep;23(1):129–161. doi: 10.1016/0022-510x(74)90148-8. [DOI] [PubMed] [Google Scholar]
- Kuhlmann W. D., Peschke P. Advances in ultrastructural postembedment localization of antigens in Epon sections with peroxidase labelled antibodies. Histochemistry. 1982;75(2):151–161. doi: 10.1007/BF00496006. [DOI] [PubMed] [Google Scholar]
- LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
- LUFT J. H. Improvements in epoxy resin embedding methods. J Biophys Biochem Cytol. 1961 Feb;9:409–414. doi: 10.1083/jcb.9.2.409. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LUFT R., IKKOS D., PALMIERI G., ERNSTER L., AFZELIUS B. A case of severe hypermetabolism of nonthyroid origin with a defect in the maintenance of mitochondrial respiratory control: a correlated clinical, biochemical, and morphological study. J Clin Invest. 1962 Sep;41:1776–1804. doi: 10.1172/JCI104637. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Meyer R. A., Brown T. R., Krilowicz B. L., Kushmerick M. J. Phosphagen and intracellular pH changes during contraction of creatine-depleted rat muscle. Am J Physiol. 1986 Feb;250(2 Pt 1):C264–C274. doi: 10.1152/ajpcell.1986.250.2.C264. [DOI] [PubMed] [Google Scholar]
- Moses R. L., Claycomb W. C. Disorganization and reestablishment of cardiac muscle cell ultrastructure in cultured adult rat ventricular muscle cells. J Ultrastruct Res. 1982 Dec;81(3):358–374. doi: 10.1016/s0022-5320(82)90064-8. [DOI] [PubMed] [Google Scholar]
- Mukherjee T. M., Dixon B. R., Blumbergs P. C., Swift J. G., Hallpike J. F. The fine structure of the intramitochondrial crystalloids in mitochondrial myopathy. J Submicrosc Cytol. 1986 Jul;18(3):595–604. [PubMed] [Google Scholar]
- Nag A. C., Cheng M. Adult mammalian cardiac muscle cells in culture. Tissue Cell. 1981;13(3):515–523. doi: 10.1016/0040-8166(81)90023-9. [DOI] [PubMed] [Google Scholar]
- Nag A. C., Cheng M., Fischman D. A., Zak R. Long-term cell culture of adult mammalian cardiac myocytes: electron microscopic and immunofluorescent analyses of myofibrillar structure. J Mol Cell Cardiol. 1983 May;15(5):301–317. doi: 10.1016/0022-2828(83)91342-1. [DOI] [PubMed] [Google Scholar]
- Ohira Y., Kanzaki M., Chen C. S. Intramitochondrial inclusions caused by depletion of creatine in rat skeletal muscles. Jpn J Physiol. 1988;38(2):159–166. doi: 10.2170/jjphysiol.38.159. [DOI] [PubMed] [Google Scholar]
- Page E., McCallister L. P. Quantitative electron microscopic description of heart muscle cells. Application to normal, hypertrophied and thyroxin-stimulated hearts. Am J Cardiol. 1973 Feb;31(2):172–181. doi: 10.1016/0002-9149(73)91030-8. [DOI] [PubMed] [Google Scholar]
- Palmer J. W., Tandler B., Hoppel C. L. Biochemical properties of subsarcolemmal and interfibrillar mitochondria isolated from rat cardiac muscle. J Biol Chem. 1977 Dec 10;252(23):8731–8739. [PubMed] [Google Scholar]
- Papadimitriou J. M., Mastaglia F. L. Ultrastructural changes in human muscle fibres in disease. J Submicrosc Cytol. 1982 Jul;14(3):525–551. [PubMed] [Google Scholar]
- Perry S. B., McAuliffe J., Balschi J. A., Hickey P. R., Ingwall J. S. Velocity of the creatine kinase reaction in the neonatal rabbit heart: role of mitochondrial creatine kinase. Biochemistry. 1988 Mar 22;27(6):2165–2172. doi: 10.1021/bi00406a052. [DOI] [PubMed] [Google Scholar]
- REYNOLDS E. S. The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J Cell Biol. 1963 Apr;17:208–212. doi: 10.1083/jcb.17.1.208. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Reznik M., Hansen J. L. Mitochondria in degenerating and regenerating skeletal muscle. Arch Pathol. 1969 Jun;87(6):601–608. [PubMed] [Google Scholar]
- Rossi A. M., Eppenberger H. M., Volpe P., Cotrufo R., Wallimann T. Muscle-type MM creatine kinase is specifically bound to sarcoplasmic reticulum and can support Ca2+ uptake and regulate local ATP/ADP ratios. J Biol Chem. 1990 Mar 25;265(9):5258–5266. [PubMed] [Google Scholar]
- SHY G. M., GONATAS N. K. HUMAN MYOPATHY WITH GIANT ABNORMAL MITOCHONDRIA. Science. 1964 Jul 31;145(3631):493–496. doi: 10.1126/science.145.3631.493. [DOI] [PubMed] [Google Scholar]
- Schlegel J., Wyss M., Eppenberger H. M., Wallimann T. Functional studies with the octameric and dimeric form of mitochondrial creatine kinase. Differential pH-dependent association of the two oligomeric forms with the inner mitochondrial membrane. J Biol Chem. 1990 Jun 5;265(16):9221–9227. [PubMed] [Google Scholar]
- Schlegel J., Zurbriggen B., Wegmann G., Wyss M., Eppenberger H. M., Wallimann T. Native mitochondrial creatine kinase forms octameric structures. I. Isolation of two interconvertible mitochondrial creatine kinase forms, dimeric and octameric mitochondrial creatine kinase: characterization, localization, and structure-function relationships. J Biol Chem. 1988 Nov 15;263(32):16942–16953. [PubMed] [Google Scholar]
- Schnyder T., Gross H., Winkler H., Eppenberger H. M., Wallimann T. Structure of the mitochondrial creatine kinase octamer: high-resolution shadowing and image averaging of single molecules and formation of linear filaments under specific staining conditions. J Cell Biol. 1991 Jan;112(1):95–101. doi: 10.1083/jcb.112.1.95. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schnyder T., Sargent D. F., Richmond T. J., Eppenberger H. M., Wallimann T. Crystallization and preliminary X-ray analysis of two different forms of mitochondrial creatine kinase from chicken cardiac muscle. J Mol Biol. 1990 Dec 20;216(4):809–812. doi: 10.1016/S0022-2836(99)80002-3. [DOI] [PubMed] [Google Scholar]
- Scholte H. R., Weijers P. J., Wit-Peeters E. M. The localization of mitochondrial creatine kinase, and its use for the determination of the sidedness of submitochondrial particles. Biochim Biophys Acta. 1973 Feb 16;291(3):764–773. doi: 10.1016/0005-2736(73)90479-3. [DOI] [PubMed] [Google Scholar]
- Sengers R. C., Stadhouders A. M., Trijbels J. M. Mitochondrial myopathies. Clinical, morphological and biochemical aspects. Eur J Pediatr. 1984 Feb;141(4):192–207. doi: 10.1007/BF00572761. [DOI] [PubMed] [Google Scholar]
- Shah A. J., Sahgal V., Muschler G., Subramani V., Singh H. Morphogenesis of the mitochondrial alterations in muscle diseases. J Neurol Sci. 1982 Jul;55(1):25–37. doi: 10.1016/0022-510x(82)90167-8. [DOI] [PubMed] [Google Scholar]
- Shields R. P., Whitehair C. K. Muscle creatine: in vivo depletion by feeding beta-guanidinopropionic acid. Can J Biochem. 1973 Jul;51(7):1046–1049. doi: 10.1139/o73-136. [DOI] [PubMed] [Google Scholar]
- Shoubridge E. A., Jeffry F. M., Keogh J. M., Radda G. K., Seymour A. M. Creatine kinase kinetics, ATP turnover, and cardiac performance in hearts depleted of creatine with the substrate analogue beta-guanidinopropionic acid. Biochim Biophys Acta. 1985 Oct 30;847(1):25–32. doi: 10.1016/0167-4889(85)90148-x. [DOI] [PubMed] [Google Scholar]
- Stadhouders A. M., Sengers R. C. Morphological observations in skeletal muscle from patients with a mitochondrial myopathy. J Inherit Metab Dis. 1987;10 (Suppl 1):62–80. doi: 10.1007/BF01812848. [DOI] [PubMed] [Google Scholar]
- Studer D., Michel M., Müller M. High pressure freezing comes of age. Scanning Microsc Suppl. 1989;3:253–269. [PubMed] [Google Scholar]
- Tandler B., Erlandson R. A., Wynder E. L. Riboflavin and mouse hepatic cell structure and function. I. Ultrastructural alterations in simple deficiency. Am J Pathol. 1968 Jan;52(1):69–96. [PMC free article] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wallimann T., Eppenberger H. M. Localization and function of M-line-bound creatine kinase. M-band model and creatine phosphate shuttle. Cell Muscle Motil. 1985;6:239–285. doi: 10.1007/978-1-4757-4723-2_8. [DOI] [PubMed] [Google Scholar]
- Wallimann T., Eppenberger H. M. The subcellular compartmentation of creatine kinase isozymes as a precondition for a proposed phosphoryl-creatine circuit. Prog Clin Biol Res. 1990;344:877–889. [PubMed] [Google Scholar]
- Wallimann T., Schnyder T., Schlegel J., Wyss M., Wegmann G., Rossi A. M., Hemmer W., Eppenberger H. M., Quest A. F. Subcellular compartmentation of creatine kinase isoenzymes, regulation of CK and octameric structure of mitochondrial CK: important aspects of the phosphoryl-creatine circuit. Prog Clin Biol Res. 1989;315:159–176. [PubMed] [Google Scholar]
- van der Voort H. T., Brakenhoff G. J., Baarslag M. W. Three-dimensional visualization methods for confocal microscopy. J Microsc. 1989 Feb;153(Pt 2):123–132. doi: 10.1111/j.1365-2818.1989.tb00553.x. [DOI] [PubMed] [Google Scholar]