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TEM and SEM study of rat endocardium following myocardial necrosis elicited by isoproterenol
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SURFACE MORPHOLOGY OF ISOLATED RAT CARDIAC MYOCYTES. S. P. Bishop and J . L . Drummond. Department of Pathology, University of Alabama in Birmingham, Birmingham, Alabama. We studied the morphologic features of cardiac myocytes isolated from 200 g rats using a variety of media and perfusion conditions. Up to 80% isolated myocytes were elongated and excluded trypan blue when obtained by perfusion on a modified Langendorf apparatus with Ca ++ free Hank's solution, 3% fetal calf serum, 95% 02 , 5% CO 2, pH 7.4 for 12 min, the same media With 0.1% collagenase for 12 min~ and ffnaliy with 0.5 mM EOTA for 12 min. Initial flow rate was l0 ml/min and reduced to 3-4 ml/min when enzyme was added. Some hearts were perfusion fixed with 2% glutaraldehyde for light (LM) and electron microscopy (EM) of in situ cells. Isolated myocytes were examined as wet mounts or by scanning electron microscopy (SEM). Poor preparations were obtained if perfusion pressure exceeded i00 mm Hg, if Ca was included, if pH, perfusion flow rates or times were not carefully ccntrollcd. Poor ~rc;araticns contained ~any contracted c=lls wlLh vdcuolated cytoplasm and ruptured plasma membrane~ by LM and EM, trypan blue uptake in wet mounts, and ruptured membrane blebs by SEM. By SEM, elongated cells excluding trypan blue had a uniformly corrugated surface with depressions at sarcomere ~nds; irregularly located holes in the Z-line ridge within the depressions apparent]y were T-tubules. Cells had irregular elongated shapes due to many transverse intercalated disc areas not only at cell ends but also along lateral edges. Isolated cardiac myocytes are a useful model to study cell function and structure, provided consideration is given to morphologically damaged cells in the preparation.
TEM AND SEM STUDY OF RAT ENDOCARDIUM FOLLOWING MYOCARDIAL NECROSIS ELICITED BY ISOPROTERENOL. M. Boutet, O. Rona, H. Turcotte and P.E. Roy. Laval and McGill Universities, Quebec, Canada. Samples from control and isoproterenol (ISO0 45 mg/kg) treated male Wistar rat hearts (24e 48, 72 hours and 1 week post [SO) were processed for transmission - (TEM) and scanning electron microscopic (SEM) studies. Endocardial surface of controls showed a continuous endothelial cell layer with a polygonal pattem corresponding to intercellular boundaries. Scattered mlcrovilli were observed only on some cell surfaces. Studies on ISO induced subendocardial necrosis revealed a progressive increase in the number of mlcrobilli on the surface of endocardial endothelial cells up to one week. By 24 hours, monocytic cells showed phyllopodic attachments to endothellum. At 48 hours in addition to fibrin mesh containing red blood cells and platelets, a very thine smooth and transparent layer was formed. Parallel TEM studies confirmed a close relationship of this vell with endothelial microvilll suggesting that this material represents an external cell coat-llke layer secreted by the endothelium. By electron microscopy the endothelial cells showed signs of increased protein synthesls. The ISO induced myocardial necrosis may serve as a model for studying the reaction of endocardial endothelium to injury. (Supported by the Medical Research Council of Canada (Grant MT-3635), and the Canadian Heart Foundation (Grant 7003).
SURFACE MORPHOLOGY OF ISOLATED RAT CARDIAC MYOCYTES. S. P. Bishop and J . L . Drummond. Department of Pathology, University of Alabama in Birmingham, Birmingham, Alabama. We studied the morphologic features of cardiac myocytes isolated from 200 g rats using a variety of media and perfusion conditions. Up to 80% isolated myocytes were elongated and excluded trypan blue when obtained by perfusion on a modified Langendorf apparatus with Ca ++ free Hank's solution, 3% fetal calf serum, 95% 02 , 5% CO 2, pH 7.4 for 12 min, the same media With 0.1% collagenase for 12 min~ and ffnaliy with 0.5 mM EOTA for 12 min. Initial flow rate was l0 ml/min and reduced to 3-4 ml/min when enzyme was added. Some hearts were perfusion fixed with 2% glutaraldehyde for light (LM) and electron microscopy (EM) of in situ cells. Isolated myocytes were examined as wet mounts or by scanning electron microscopy (SEM). Poor preparations were obtained if perfusion pressure exceeded i00 mm Hg, if Ca was included, if pH, perfusion flow rates or times were not carefully ccntrollcd. Poor ~rc;araticns contained ~any contracted c=lls wlLh vdcuolated cytoplasm and ruptured plasma membrane~ by LM and EM, trypan blue uptake in wet mounts, and ruptured membrane blebs by SEM. By SEM, elongated cells excluding trypan blue had a uniformly corrugated surface with depressions at sarcomere ~nds; irregularly located holes in the Z-line ridge within the depressions apparent]y were T-tubules. Cells had irregular elongated shapes due to many transverse intercalated disc areas not only at cell ends but also along lateral edges. Isolated cardiac myocytes are a useful model to study cell function and structure, provided consideration is given to morphologically damaged cells in the preparation.
TEM AND SEM STUDY OF RAT ENDOCARDIUM FOLLOWING MYOCARDIAL NECROSIS ELICITED BY ISOPROTERENOL. M. Boutet, O. Rona, H. Turcotte and P.E. Roy. Laval and McGill Universities, Quebec, Canada. Samples from control and isoproterenol (ISO0 45 mg/kg) treated male Wistar rat hearts (24e 48, 72 hours and 1 week post [SO) were processed for transmission - (TEM) and scanning electron microscopic (SEM) studies. Endocardial surface of controls showed a continuous endothelial cell layer with a polygonal pattem corresponding to intercellular boundaries. Scattered mlcrovilli were observed only on some cell surfaces. Studies on ISO induced subendocardial necrosis revealed a progressive increase in the number of mlcrobilli on the surface of endocardial endothelial cells up to one week. By 24 hours, monocytic cells showed phyllopodic attachments to endothellum. At 48 hours in addition to fibrin mesh containing red blood cells and platelets, a very thine smooth and transparent layer was formed. Parallel TEM studies confirmed a close relationship of this vell with endothelial microvilll suggesting that this material represents an external cell coat-llke layer secreted by the endothelium. By electron microscopy the endothelial cells showed signs of increased protein synthesls. The ISO induced myocardial necrosis may serve as a model for studying the reaction of endocardial endothelium to injury. (Supported by the Medical Research Council of Canada (Grant MT-3635), and the Canadian Heart Foundation (Grant 7003).