Long-term cell culture of adult mammalian cardiac myocytes: Electron microscopic and immunofluorescent analyses of myofibrillar structure

Nag, Asish C.

doi:10.1016/0022-2828(83)91342-1

Cited by 68 publications

(31 citation statements)

References 8 publications

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“…This once again demonstrates the heterogeneity of myocytes in culture. The general pattern of changes can be described as starting with an initial bright apparently nonspecific staining similar to that reported by Nag et al 29 This takes place during massive internal reorganization in cells converting from a cylindrical to a round shape. This is followed by more diffuse indistinct staining as new sarcoplasm is expressed.…”

Section: Myosin Expression In Culturementioning

confidence: 74%

Differentiation of adult rat cardiac myocytes in cell culture.

Bugaisky

Zak

1989

Circulation Research

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Cardiac myocytes isolated from adult rat hearts were grown on laminln coated culture dishes for more than a month. During this tune, the cells underwent a morphological transformation which has also been referred to by others as cell remodeling (Guo J-X, Jacobson SL, Brown DL: Cell Mot Cytoskeleton 1986;6:291-304). This results in a change in myocyte morphology from its typical in vivo cylindrical shape to one which is more pleiomorphic. Despite the long-term change in morphology, myocytes expressed for differing lengths of time several aspects of the adult phenotype as evidenced by the following: 1) maintenance of cylindrical shape and/or evident cross-striations for the first 24-48 hours in culture, 2) reappearance of cross-striations during the second week in culture, 3) little or no spontaneous contractility for the first 4 days in culture, 4) expression of only the V, isoform of myosin for at least 7 days, and 5) altered myosin isoform expression in response to changes in environmental conditions. These factors taken together suggest that in culture the adult cardiac myocyte remains a highly differentiated cell (as opposed to possible dedifferentiation) and maintains many of its previous in vivo characteristics. Such highly differentiated adult cells should be suitable as an in vitro system for studying the direct cellular effects of factors which regulate growth and differentiation of the in vivo heart. (Circulation Research 1989;64:493-500) T he adult cardiac myocyte of the rat is a binucleated, nondividing cell which participates in organ growth solely by cellular hypertrophy. While this cell type constitutes only about 25% of the total cellular population in the heart, it represents approximately 75% of the organ's total mass 1 and thus forms the major functional and structural unit of the heart. Little is known about those factors that at the cellular level directly regulate growth, differentiation, and the response to physiological stress in this cell type. To investigate these processes, it is necessary to have a well-characterized cell culture system in which the properties of the cardiac myocyte can be studied devoid of systemic influences. Investigators began to approach this problem more than 20 years ago using

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Section: Myosin Expression In Culturementioning

confidence: 74%

Differentiation of adult rat cardiac myocytes in cell culture.

Bugaisky

Zak

1989

Circulation Research

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“…The ultrastructure of cultured neonatal and adult rat ventricular cardiac myocytes grown on polystyrene surfaces has been described in previous studies (Moses and Kasten, 1979;Moses and Claycomb, 1982a,b;Moses and Delcarpio, 1983;Nag et al, 1983). The ultrastructure of the myocytes cultured on laminin was similar to their counterparts grown on plastic.…”

Section: Morphological Observationsmentioning

confidence: 83%

Ultrastructural morphometric analysis of cultured neonatal and adult rat ventricular cardiac muscle cells

1989

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Neonatal and adult rat ventricular cardiac muscle cells cultured on laminin differed from similar myocytes grown on plastic in the amount and distribution of their mitochondria and transverse tubules. Point-count morphometry was used at the electron microscopic level to quantify these differences. Adult myocytes grown on laminin contained more mitochondria per unit volume than adult myocytes grown on plastic. No significant differences were observed in the volume percent of myofibrils in either adult or neonatal ventricular myocytes when grown on laminin and compared to those grown on plastic. The transverse tubule system in neonatal and adult myocytes was reduced significantly when both groups were cultured on laminin. Furthermore, neonatal and adult myocytes cultured on laminin were flatter than those cultured on plastic. This may indicate a relationship between the surface/volume ratio and transverse tubule development in cultured myocytes. These studies establish that point-count morphometry can be used to quantify changes in the organelle volume densities of cultured cardiac muscle cells.

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“…The construct was delivered to myocytes of WT and SERCA mice in vivo via adenoviral gene therapy. In vivo transfection (as opposed to infection of isolated myocytes in cell culture) was employed to allow expression in a working heart in its physiological milieu to avoid myocyte remodeling and the potential decrease in SR activity that accompanies quiescent myocytes kept in culture (32,48). Using the ratiometric pericam construct, we were able to demonstrate that basal levels of calcium were similar in both groups and that electrical stimulation of cardiac myocytes results in a greater increase of mitochondrial calcium in SERCA than in WT myocytes.…”

Section: Discussionmentioning

confidence: 99%

Increased expression of SERCA in the hearts of transgenic mice results in increased oxidation of glucose

Belke¹,

Swanson²,

Suárez³

et al. 2007

American Journal of Physiology-Heart and Circulatory Physiology

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Dillmann WH. Increased expression of SERCA in the hearts of transgenic mice results in increased oxidation of glucose. Am J Physiol Heart Circ Physiol 292: H1755-H1763, 2007. First published December 1, 2006; doi:10.1152/ajpheart.00884.2006.-While several transgenic mouse models exhibit improved contractile characteristics in the heart, less is known about how these changes influence energy metabolism, specifically the balance between carbohydrate and fatty acid oxidation. In the present study we examine glucose and fatty acid oxidation in transgenic mice, generated to overexpress sarco(endo) plasmic reticulum calcium-ATPase (SERCA), which have an enhanced contractile phenotype. Energy substrate metabolism was measured in isolated working hearts using radiolabeled glucose and palmitate. We also examined oxygen consumption to see whether SERCA overexpression is associated with increased oxygen utilization. Since SERCA is important in calcium handling within the cardiac myocyte, we examined cytosolic calcium transients in isolated myocytes using indo-1, and mitochondrial calcium levels using pericam, an adenovirally expressed, mitochondrially targeted ratiometric calcium indicator. Oxygen consumption did not differ between wildtype and SERCA groups; however, we were able to show an increased utilization of glucose for oxidative metabolism and a corresponding decreased utilization of fatty acids in the SERCA group. Cytosolic calcium transients were increased in myocytes isolated from SERCA mice, and they show a faster rate of decay of the calcium transient. With these observations we noted increased levels of mitochondrial calcium in the SERCA group, which was associated with an increase in the active form of the pyruvate dehydrogenase complex. Since an increase in mitochondrial calcium levels leads to activation of the pyruvate dehydrogenase complex (the rate-limiting step for carbohydrate oxidation), the increased glucose utilization observed in isolated perfused hearts in the SERCA group may reflect a higher level of mitochondrial calcium. mitochondrial calcium; pyruvate dehydrogenase; cardiac energetics; sarco(endo)plasmic reticulum calcium-adenosine 5Ј-triphosphatase IN THE HEART, OXIDATIVE METABOLISM involving ATP production and workload are tightly linked. An increase in cardiac output is associated with an increase in oxygen consumption and an increase in energy substrate metabolism (carbohydrates and fatty acids) (36). An increase in cardiac performance occurs in vivo as a result of increased stimulation by the sympathetic nervous system and can be mimicked in ex vivo isolated heart preparations (Langendorff, working heart) through the use of positive inotropic agents (dobutamine and isoproterenol), an increase in heart rate, an increase in perfusate calcium concentrations, and/or an increase in preload and afterload perfusion pressures (19,23,35,42). The increased cardiac performance is associated with an increased calcium load within the myocyte, affecting the increase in force generation (23). In an att...

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Long-term cell culture of adult mammalian cardiac myocytes: Electron microscopic and immunofluorescent analyses of myofibrillar structure

Cited by 68 publications

References 8 publications

Differentiation of adult rat cardiac myocytes in cell culture.

Differentiation of adult rat cardiac myocytes in cell culture.

Ultrastructural morphometric analysis of cultured neonatal and adult rat ventricular cardiac muscle cells

Increased expression of SERCA in the hearts of transgenic mice results in increased oxidation of glucose

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