Connective Tissue Skeleton of the Human Heart
A Demonstration by Cell-Maceration Scanning Electron Microscope Method
The stroma of the heart maintains the structure of the myocardium, determining tissue tensile strength and stiffness.1 In addition, it contributes to ventricular function through the transmission of myocyte-generated force to the atrial and ventricular chambers and to the relengthening of myocytes in diastole.2 The three-dimensional configuration of cardiac collagen has been determined by scanning electron microscopy3 4 5 : the epimysium envelops the entire cardiac muscle; the perimysium, which is an extension of the epimysium, serves to enwrap groups of myocytes; and the endomysium, as final arborization of the perimysium, supports and connects individual cells. The endomysial weave envelops each individual myocyte and is connected to adjacent myocytes by lateral struts.
Because this knowledge was obtained through studies on whole fixed myocardial tissue without removal of its nonfibrous elements, we attempted to dissolve the cellular elements and leave behind a noncollapsed matrix, aiming for a better three-dimensional view. For this, we used a modification of the NaOH maceration technique reported by Ohtani.6 This method was reported to be able to remove cellular elements much more effectively than any other method. Small fragments, 10×5×3 mm in size, of the anterior wall at the midventricular region were obtained from three human hearts, weighing between 300 and 350 g, without any pathological changes. All samples were fixed in 10% neutral formalin. After being rinsed in distilled water, the specimens were immersed in a 10% NaOH solution for 4 to 6 days at room temperature and then rinsed in distilled water until they became transparent. Then they were immersed in 1% tannic acid for 4 hours. Subsequently, the specimens were rinsed in distilled water overnight, rinsed, post-fixed in 1% osmium tetroxide for 2 hours, dehydrated in graded concentrations of ethanol, sectioned transversely or longitudinally with a very sharp, clean blade under a dissecting microscope, freeze-dried, coated with gold, and observed under a Zeiss 940-A scanning electron microscope.
The figure clearly shows, for the first time, the three-dimensional architecture of collagen fibrils in human myocardium after digestion of the cellular elements. This is expected to contribute further to the understanding of the morphology of the connective tissue skeleton of the heart.
Professor Rossi is Senior Investigator of the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq).
The editor of Images in Cardiovascular Medicine is Hugh A. McAllister, Jr, MD, Chief, Department of Pathology, St Luke’s Episcopal Hospital and Texas Heart Institute, and Clinical Professor of Pathology, University of Texas Medical School and Baylor College of Medicine.
Circulation encourages readers to submit cardiovascular images to Dr Hugh A. McAllister, Jr, St Luke’s Episcopal Hospital and Texas Heart Institute, 6720 Bertner Ave, MC1-267, Houston, TX 77030.
- Copyright © 1998 by American Heart Association