Three-dimensional histology: tools and application to quantitative assessment of cell-type distribution in rabbit heart

Abstract

AimsCardiac histo-anatomical organization is a major determinant of function. Changes in tissue structure are a relevant factor in normal and disease development, and form targets of therapeutic interventions. The purpose of this study was to test tools aimed to allow quantitative assessment of cell-type distribution from large histology and magnetic resonance imaging- (MRI) based datasets.Methods and resultsRabbit heart fixation during cardioplegic arrest and MRI were followed by serial sectioning of the whole heart and light-microscopic imaging of trichrome-stained tissue. Segmentation techniques developed specifically for this project were applied to segment myocardial tissue in the MRI and histology datasets. In addition, histology slices were segmented into myocytes, connective tissue, and undefined. A bounding surface, containing the whole heart, was established for both MRI and histology. Volumes contained in the bounding surface (called ‘anatomical volume’), as well as that identified as containing any of the above tissue categories (called ‘morphological volume’), were calculated. The anatomical volume was 7.8 cm3 in MRI, and this reduced to 4.9 cm3 after histological processing, representing an ‘anatomical’ shrinkage by 37.2%. The morphological volume decreased by 48% between MRI and histology, highlighting the presence of additional tissue-level shrinkage (e.g. an increase in interstitial cleft space). The ratio of pixels classified as containing myocytes to pixels identified as non-myocytes was roughly 6:1 (61.6 vs. 9.8%; the remaining fraction of 28.6% was ‘undefined’).ConclusionQualitative and quantitative differentiation between myocytes and connective tissue, using state-of-the-art high-resolution serial histology techniques, allows identification of cell-type distribution in whole-heart datasets. Comparison with MRI illustrates a pronounced reduction in anatomical and morphological volumes during histology processing.

How to improve the overall quality of cardiac morphometric data

“…For instance, we have noted an ϳ30% shrinkage of isolated myocyte volume (as assessed by Coulter Channelyzer) after hematoxylin-and-eosin processing (unpublished data). This is similar to reports by others (1). If corrections are desired, one can measure tissue section area before and after processing to determine shrinkage.…”

Myocardial interstitial remodelling in non-ischaemic dilated cardiomyopathy: insights from cardiovascular magnetic resonance

“…This contrast agent has been reported to allow in vivo molecular magnetic resonance imaging of fibrosis in a mouse model of scarred infarction [127,128], but has never entered Heart Fail Rev clinical application. Other studies investigated high-resolution 3D sequences reaching a voxel size comparable to cellular size, as a sort of magnetic resonance histology, even though they still require very high magnetic fields and very long acquisition times [129]. Finally, several studies used intravascular contrast agents [130][131][132] or adenosine stress [133] to investigate the relative contribution of the intravascular space to the ECV.…”

Energetic metabolism in cardiomyocytes: molecular basis of heart ischemia and arrhythmogenesis

“…This organ consists of various layers, the thickest of which is the myocardium. The main component of this layer is contractile cells termed cardiomyocytes [3] , with only 2% corresponding to Purkinje fibers. The Purkinje fibers are organized into an arborized structure which originates in the atrioventricular node and constitutes a specialized conduction system that allows quick and synchronic activation of the ventricles [4,5] .…”