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Isolation and properties of vinculin from chicken heart muscle
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INTERSTITIAL CHANGES WITH VIRAL MYOCAROITIS. J.B. Caulfield, M. Nachtigal, Sun Ben Tao. Department of Pathology, University of South Carolina School of Medicine, Columbia, South Carolina. Encephalomyocarditis virus (ECMV) causes among other lesions an extensive necrotizing myocarditis. $hese changes were examined daily for five days after subcutaneous inoculation of 10 TCID SO per animal by light, scanning (SEM) and transmission microscopy. Necrosis of myocardial cells with calcification of mitochondria was evident at three days and occasionally apparent at two days, by light microscopy. With SEM breakdown of the basement membrane was clear at one day. Loss of basement membrane was more extensive on subsequent days. By day three loss of some collagen struts could be demonstrated. At day three fibroblasts were migrating toward and aggregating in areas of extensive loss of the basement membrane. By day five the fibroblasts had secreted collagen in the areas of loss of basement membrane. The collagen was in the form of bundles smaller than the normal struts and in greater concentrations. The arrangement of the newly secreted collagen did not reproduce the usual distribution of myocyte to myocyte struts. If sufficiently extensive this abnormal distribution of collagen could interfere with ventricular function. The ECMV infection affects the interstitium before any cellular alterations are evident suggesting that this change is an integral part of the disease.
167ISOLATION AND PROPERTIES OF VINCULIN FROM CHICKEN HEART MUSCLE. V.E. Koteliansky, G.N. Gneushev, M.A. Glukhova. Institute of Experimental Cardiology, USSR Cardiology Research Center, Moscow, USSR. Vinculin was identified and isolated from chicken heart muscle. This protein has d-helices (about an apparent molecular weight of 130 000. It has high content of 80%) and common antigenic determinants with vinculin from chicken smooth muscle. Vinculin interacts with actin and reduceslow-shear viscosity of F-actin. This effect does not depend on the presence of free calcium ions. Using filamin and &>I-actinin it was shown that vinculin increased the critical gel point as actin cross-linkers, of F-actin. In heart muscle vinculin was localized in intercalated discs, near the Z-discs of the muscle sarcomere between sarcoplasmic reticulum and myofibrils, and associated with the cell membrane. Thus, in heart vinculin appears to integrate myofibrils by linking the plasma membrane.
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ROLE OF LIPID IN THE STRUCTURE OF THE SARCOPLASMIC RETICULUM AND SARCOLEMMAL MEMBRANES. L. Herbette, R. McDaniel, T. Ashavaid and R. Calvin, Division of Cardiology, University of Connecticut Health Center, Farmington, CT 06032 The structural architecture of the skeletal and cardiac sarcoplasmic reticulum (SR) membrane differs greatly from that of the cardiac sarcolemma (SL). The average lipid to protein ratio for SR membranes was 1.5 umoles fatty acyl chains (0.9 umoles phosphorus) per mg protein in contrast to 6.1 wm~les fatty acyl chains (2.1 pmoles phosphorus) per mg SL membrane protein. Lamellar meridional x-ray diffraction from hydrated oriented SR membrane multilayers indicated that the single hydrated membrane width was 80-100 A, in contrast to 60-65 A for SL, consistent with electron microscope images of these membranes. Lipids extracted from both membranes formed bilayers with hydrated widths in the range of 55-65 A. A comparison of the electron density profiles of intact SR membranes with extracted SR lipids suggesting an asymmetric lipid distribution between both monolayers of the SR lipid bilayer was confirmed by neutron diffraction in which 10% more lipid was present in the inner monolayer of the SR lipid bilayer. In contrast, comparison of the electron density profiles of the intact SR membrane with extracted SL lipids suggests a symmetric distribution of lipid between the two monolayers of the SL lipid bilayer. These contrasting structures of these two membranes may underlie, in part, their very different functional properties. Supported by NIH HL27630, HL07420 and the American Heart Association.
INTERSTITIAL CHANGES WITH VIRAL MYOCAROITIS. J.B. Caulfield, M. Nachtigal, Sun Ben Tao. Department of Pathology, University of South Carolina School of Medicine, Columbia, South Carolina. Encephalomyocarditis virus (ECMV) causes among other lesions an extensive necrotizing myocarditis. $hese changes were examined daily for five days after subcutaneous inoculation of 10 TCID SO per animal by light, scanning (SEM) and transmission microscopy. Necrosis of myocardial cells with calcification of mitochondria was evident at three days and occasionally apparent at two days, by light microscopy. With SEM breakdown of the basement membrane was clear at one day. Loss of basement membrane was more extensive on subsequent days. By day three loss of some collagen struts could be demonstrated. At day three fibroblasts were migrating toward and aggregating in areas of extensive loss of the basement membrane. By day five the fibroblasts had secreted collagen in the areas of loss of basement membrane. The collagen was in the form of bundles smaller than the normal struts and in greater concentrations. The arrangement of the newly secreted collagen did not reproduce the usual distribution of myocyte to myocyte struts. If sufficiently extensive this abnormal distribution of collagen could interfere with ventricular function. The ECMV infection affects the interstitium before any cellular alterations are evident suggesting that this change is an integral part of the disease.
167ISOLATION AND PROPERTIES OF VINCULIN FROM CHICKEN HEART MUSCLE. V.E. Koteliansky, G.N. Gneushev, M.A. Glukhova. Institute of Experimental Cardiology, USSR Cardiology Research Center, Moscow, USSR. Vinculin was identified and isolated from chicken heart muscle. This protein has d-helices (about an apparent molecular weight of 130 000. It has high content of 80%) and common antigenic determinants with vinculin from chicken smooth muscle. Vinculin interacts with actin and reduceslow-shear viscosity of F-actin. This effect does not depend on the presence of free calcium ions. Using filamin and &>I-actinin it was shown that vinculin increased the critical gel point as actin cross-linkers, of F-actin. In heart muscle vinculin was localized in intercalated discs, near the Z-discs of the muscle sarcomere between sarcoplasmic reticulum and myofibrils, and associated with the cell membrane. Thus, in heart vinculin appears to integrate myofibrils by linking the plasma membrane.
168
ROLE OF LIPID IN THE STRUCTURE OF THE SARCOPLASMIC RETICULUM AND SARCOLEMMAL MEMBRANES. L. Herbette, R. McDaniel, T. Ashavaid and R. Calvin, Division of Cardiology, University of Connecticut Health Center, Farmington, CT 06032 The structural architecture of the skeletal and cardiac sarcoplasmic reticulum (SR) membrane differs greatly from that of the cardiac sarcolemma (SL). The average lipid to protein ratio for SR membranes was 1.5 umoles fatty acyl chains (0.9 umoles phosphorus) per mg protein in contrast to 6.1 wm~les fatty acyl chains (2.1 pmoles phosphorus) per mg SL membrane protein. Lamellar meridional x-ray diffraction from hydrated oriented SR membrane multilayers indicated that the single hydrated membrane width was 80-100 A, in contrast to 60-65 A for SL, consistent with electron microscope images of these membranes. Lipids extracted from both membranes formed bilayers with hydrated widths in the range of 55-65 A. A comparison of the electron density profiles of intact SR membranes with extracted SR lipids suggesting an asymmetric lipid distribution between both monolayers of the SR lipid bilayer was confirmed by neutron diffraction in which 10% more lipid was present in the inner monolayer of the SR lipid bilayer. In contrast, comparison of the electron density profiles of the intact SR membrane with extracted SL lipids suggests a symmetric distribution of lipid between the two monolayers of the SL lipid bilayer. These contrasting structures of these two membranes may underlie, in part, their very different functional properties. Supported by NIH HL27630, HL07420 and the American Heart Association.