Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.2.1.36 (hyaluronidase)
4,606 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adult rat heart was dissociated into a single cell suspension by a perfusion technique which used 0.05% collagenase and 0.1% hyaluronidase in Krebs-Ringer phosphate buffer (KRP). The non-muscle cells of the suspension were separated from the myocytes by centrifugation through 3% Ficoll solution in KRP with 0.01 mM Ca2+. An approximately 90% pure suspension of isolated single muscle cells was obtained with this method. The effects of the successive steps in the dissociation procedure on the ultrastructure of the heart were studied by scanning and transmission electron microscopy. After 30 minutes of enzyme digestion, dissociation of the inner endothelial lining of the ventricle into single cells or small groups of cells became apparent. In addition, the underlying cardiac skeleton began to disintegrate and linear arrays of cardiac muscle cells were observed. After 45 minutes of enzyme digestion the number of released single cells was higher because of the separation of intercalated discs. The majority of non-muscle cells were by now dissociated from the surfaces of muscle cells. Widening of the lateral intercellular spaces between the myocardial cells was associated with separation of desmosomes. In some regions of the heart, intact desmosomes, fasciae adherentes and gap junctions were observed even though lateral intercellular spaces had widened greatly. The majority of myocardial cells had become separated from one another after 60 minutes of enzyme digestion. Separation of gap junctional sites took place in two ways: (1) by 'unzipping' them through enzyme action; (2) by tearing them mechanically. Gap junction remnants were sometimes observed in a vesiculated state within the cell. The dissociation of the heart was ineffective when perfused with media containing 1.0 or 2 mM Ca2+. Alcian blue treatment after 60 minutes of enzyme digestion revealed that the basement membrane, and its accompanying collagen fibrils, was still present on the plasma membrane of dissociated single cells. The isolated myocardial cells retained their normal morphological characteristics. This study has enabled us to understand in detail how dismantlement of highly ordered adult cardiac tissue into a single cell suspension takes place. Cell suspensions of this type should be invaluable in the study of metabolic and synthetic activities in adult myocardial cells.
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PMID:Dissociation of adult mammalian heart into single cell suspension: an ultrastructural study. 12 Mar 52

Rat cardiac muscle was dissociated into single cells by a coronary perfusion technique with collagenase and hyaluronidase in a Ca-free medium. Retention of the cylindrical shape of isolated muscle cells could be achieved by regulation of [Ca2+]0 and temperature. Cells kept at 4 degrees C, and 0-01 mM CaCl2 remained cylindrical for more than a week and contracted spontaneously upon warming at 37 degrees C. At [Ca2+]0 between 0-1-2 mM and 37 degrees C, cells underwent contracture and rounded up. Scanning (SEM) and transmission electron microscopy were used to analyze the structure of cylindrical and rounded muscle cells. The extracellular aspect of the sarcolemma at lateral cell surfaces and intercalated disc regions were clearly revealed for SEM analysis. Both the distribution and number of T-tubule openings on the surfaces can be estimated and a three-dimensional description of the intercalated disc obtained. This study reveals that isolated adult heart cells are extremely sensitive to [Ca2+]0, but with careful control of this cation, this preparation should be helpful in the analysis of both sarcolemmal structure and the pathological changes which accompany myocardial injury.
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PMID:Studies of isolated adult rat heart cells: the surface morphology and the influence of extracellular calcium ion concentration on cellular viability. 20 Oct 46

Experiments were performed to determine the cellular associations of the molecular forms of acetylcholinesterase (AChE) in adult rat heart. For this purpose, a cardiac muscle and a non-muscle fraction were isolated from rat heart ventricles after perfusion with collagenase and hyaluronidase, extracts of these fractions were subjected to ultracentrifugation on linear density gradients of sucrose (5-20%), and fractions of these gradients were analyzed for AChE activity. The results show that only globular AChE molecular forms were present in isolated cardiac muscle cells. Globular AChE forms were also present in the non-muscle cells fraction but in different proportions. The proportions of globular AChE forms plus the high specific activity of choline acetyltransferase in the non-muscle cell fraction suggest that this fraction contains cholinergic nerve fragments. The results of this study also show that asymmetric AChE is released during the perfusion of heart with the digestive enzymes, which suggests that asymmetric AChE is bound to the extracellular matrix of heart.
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PMID:Acetylcholinesterase molecular forms in muscle and non-muscle cells of rat heart. 258 21

An 80 kDa glycoprotein was isolated from adult frog skeletal muscle by concanavalin (Con A) affinity chromatography and electrophoretic separation by molecular mass. Characteristics of the 80 kDa glycoprotein are that it: 1) binds non-covalently to gelatin-agarose or some other protein(s) bound to gelatin-agarose, 2) does not bind wheat germ agglutinin, 3) appears only at 80 kDa in both reducing and non-reducing electrophoretic separations, 4) is present in skeletal muscle but absent in smooth muscle and cardiac muscle, 5) is not collagenase or hyaluronidase-sensitive, and 6) is antigenically similar to a protein in embryonic chicken skeletal muscle. It was used to generate a polyclonal antiserum which was affinity-purified and used for immunolocalization. Indirect immunofluorescence procedures showed the antigen to be present on the surface of the skeletal muscle cells and concentrated at sites where cells are closely apposed to one another. Preparations in which adult muscle cells were depleted of basement membrane and endomysial proteins did not reduce the amount of 80 kDa protein present in skeletal muscle. These data indicate that this is a cell surface glycoprotein that may mediate attachment of the cell to extracellular proteins at sites where adjacent skeletal muscle cells are apposed.
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PMID:Identification of an 80 kDa glycoprotein located at sites of close apposition between skeletal muscle cells. 326 Aug 64

A long-term cell culture system for adult cardiomyopathic hamster cardiac muscle cells has been established. The diseased and control hearts were dissociated into single cell suspension with the modifications of our previous technique using collagenase and hyaluronidase as applied to the dissociation of the adult rat heart. The postperfusion of the diseased heart with Krebs-Ringer phosphate buffer and bovine serum albumin was very helpful in obtaining greater yield of viable diseased muscle cells; the cells were cultured for 4 wk. Approximately 60% of the myocytes from the diseased heart and 85% of the myocytes from the normal heart attached to the substrates and survived throughout the culture period. Approximately 60 to 70% of the cardiac myocytes from the diseased and control hearts were bi- or multinucleated; 30% of the diseased and 80% of the normal myocytes showed rhythmic contractility. Electron microscopy revealed the presence of two kinds of cardiac muscle cells in the diseased cell culture on the basis of their myofibril content: one with scanty myofibrils and another with abundant myofibrils. Myocytes with sparse myofibrils showed certain characteristic features that included autophagic vacuoles, amorphous matrix of fine filamentous texture, scattered strips of myofibrils, and abnormal organization of the Z-line. Cardiac muscle cells with abundant myofibrillar content contained unorganized myofibrils in certain sarcomeres. These studies demonstrate the feasibility of maintaining diseased cardiac muscle cells from adult cardiomyopathic hamsters for at least 4 wk in monolayer culture.
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PMID:Isolation, long-term culture, and ultrastructural characterization of adult cardiomyopathic cardiac muscle cells. 357 Oct 99

A method for dissociating myocardial cells from the bony fish heart is presented. Beating hearts of Pleuronectes platessa were perfused by an aerated medium containing 0.1% collagenase and 0.1% hyaluronidase. After 60 min, the hearts were quite soft and easy to dissociate. The cardiac muscle cells were then collected on Nucleopore membranes, and their surface structure was revealed by scanning electron microscopy.
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PMID:Dissociation of muscle cells from the bony fish heart. 367 58

The importance of the slow inward calcium current (Isi) in the excitation-contration coupling process of cardiac muscle is well documented. The current can be attributed mainly to a calcium translocation from the extracellular space into the cell or a subsarcolemmal compartment of it. Isi has been suggested to have its source in and to be controlled by the surface coat of the sarcolemma (glycocalyx). The glycocalyx is destroyed in myocytes dissociated from adult heart tissue with solutions containing low calcium, collagenase and hyaluronidase (Fig. 1). By comparing the Isi data obtained in isolated myocytes with those reported for trabeculae or papillary muscles, we have now obtained evidence suggesting that the glycocalyx is not important in the genesis of Isi.
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PMID:Glycocalyx is not required for show inward calcium current in isolated rat heart myocytes. 624 5

Isolated cardiac muscle cells enzymatically digested with collagenase and hyaluronidase from whole rat myocardium demonstrate the characteristics of an intact membrane in that they tolerate millimolar concentrations of free Ca2+ and exhibit phasic contractions with electrical excitation. These isolated cells maintain their characteristic A-I band striation patterns when at rest or during contraction. An apparatus has been developed to directly image these cells with phase-contrast micrography onto a 1,728-element charge-coupled device photodiode array for rapid data storage in a digital computer. The digitized striation pattern profile was analyzed for individual and average sarcomere spacing. In isotonic media the average resting sarcomere length ranged from 1.77 to 1.91 micrometers in 13 cells, with a mean length of 1.83 +/- 0.12 micrometers. Electrically stimulated phasic contractions in three cells demonstrated a synchronous 20% decrease in sarcomere spacing to a mean of 1.51 micrometers. Striation spacing decreased under hypertonic stress but elongated only up to 1.93 micrometers in hypotonic solutions, suggesting that some internal elastic constraint exists that limits elongation of the cell.
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PMID:Direct measurement of sarcomere length from isolated cardiac cells. 627 4

Adult rat heart was dissociated into a single-cell suspension by a retrograde perfusion technique with collagenase and hyaluronidase in Krebs-Ringer phosphate buffer. Long-term culture of these isolated single cardiac muscle cells was established for up to 45 days. Transmission electron microscopy and immunofluorescence analysis with monoclonal antibodies to cardiac myosin were used to examine sequentially the external and internal structural organization of the cardiac myocytes. Most of the cardiac myocytes exhibited prominent alterations in their external and internal structural organization during the first two weeks of culture. As they attached to the substrate and spread out, the myocytes assumed various shapes and sizes, with the exception of a few which maintained their original cylindrical shape. Electron microscopy of 2 to 4-day cultures revealed that most of the muscle cells contained disorganized myofibrils and surface blebs with enclosed mitochondria and myofilaments, which were eventually extruded from the cytoplasm. With progressive culture, the cardiac myocytes appeared to lose myofibrillar material; fewer myofilaments or sacromere fragments with interfibrillar mitochondria were observed in the sarcoplasm. Such cells resembled cultured embryonic or neonatal cardiac myocytes. However, some muscle cells retained closely packed, well organized myofibrils characteristic of freshly dissociated or in vivo cardiac myocytes. Immunofluorescence microscopy demonstrated that the cultured cardiac myocytes were strongly myosin positive throughout their morphological changes and subsequent maintenance in culture. Two patterns of fluorescence were observed in these cells in correlation with the fine structural evidence for myofibrillar distribution. One pattern exhibited bright fluorescence near the central region of the cell with a more weakly diffuse fluorescence throughout the cytoplasm; the other pattern was characterized by bright fluorescence throughout the sarcoplasm. Most of the myocytes retained their contractility throughout the culture period excepting the initial 24 to 48 h of cell attachment and flattening. These studies demonstrate the feasibility of maintaining contractile cardiac muscle cells from adult rats for at least 1 1/2 months in monolayer culture, although some variability in myofibrillar organization has been observed.
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PMID:Long-term cell culture of adult mammalian cardiac myocytes: electron microscopic and immunofluorescent analyses of myofibrillar structure. 635 Jun 10

Antigens extracted from cells of Streptococcus pyogenes T6 and Streptococcus mutans strains AHT, BHT, 10449, OMZ175, and K1R adsorbed to the sarcolemmal sheath of cardiac muscle cells in vitro. Similar preparations from S. salivarius, S. sanguis, Staphylococcus aureus, and Lactobacillus casei had weak or negligible tissue-binding activity. Tissue-bound bacterial antigens were detected with homologous rabbit antisera with both indirect immunofluorescence tests and an indirect radioimmunoassay. Serological cross-reactivity was observed between the tissue-binding factors of S. pyogenes and S. mutans cells but not between the bacteria and muscle tissue. In a comparative study of extraction procedures, the greatest yield of tissue-binding factors was obtained from group A streptococci by cell disruption in buffer at 4 degrees C. Hot aqueous phenol and hot water extracts were inactive. Antibodies specific for the tissue-binding factor(s) were readily adsorbed from rabbit anti-S. pyogenes serum by a preparation of isolated cytoplasmic membranes but not by a suspension of cell wall fragments. The heart-binding component of S. pyogenes cell extracts was inactivated by protease digestion and heat treatment and to a lesser extent by periodic acid oxidation. The capacity of heart cell components to adsorb streptococcal antigens was reduced by protease treatment but not by the action of neuraminidase, hyaluronidase, organic solvents, or detergents.
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PMID:Binding of streptococcal antigens to muscle tissue in vitro. 699 20


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