Abstract:

Electrophysiological modeling of cardiac tissue is commonly based on functional and structural properties measured in experiments. Our knowledge of these properties is incomplete, in particular their remodeling in disease. Here, we introduce a methodology for quantitative tissue characterization based on fluorescent labeling, three-dimensional scanning confocal microscopy, image processing and reconstruction of tissue micro-structure at sub-micrometer resolution. We applied this methodology to normal rabbit ventricular tissue and tissue from hearts with myocardial infarction. Our analysis revealed that the volume fraction of fibroblasts increased from 4.830.42% (meanstandard deviation) in normal tissue up to 6.510.38% in myocardium from infarcted hearts. The myocyte volume fraction decreased from 76.209.89% in normal to 73.488.02% adjacent to the infarct. Numerical field calculations on three-dimensional reconstructions of the extracellular space yielded an extracellular longitudinal conductivity of 0.2640.082 S/m with an anisotropy ratio of 2.0951.11 in normal tissue. Adjacent to the infarct, the longitudinal conductivity increased up to 0.4000.051 S/m, but the anisotropy ratio decreased to 1.2950.09. Our study indicates an increased density of gap junctions proximal to both fibroblasts and myocytes in infarcted versus normal tissue, supporting previous hypotheses of electrical coupling of fibroblasts and myocytes in infarcted hearts. We suggest that the presented methodology provides an important contribution to modeling normal and diseased tissue. Applications of the methodology include the clinical characterization of disease-associated remodeling. 1.

Abstract:

This work covers several fields of studies. Biological techniques, knowledge of imaging systems and their features as well as a background in system theory and digital image processing were combined. Biopsy punches from six rat hearts and one rabbit heart were obtained providing seven biopsies from rat and three from rabbit. The tissue was sectioned and quad-labelled in a three day procedure to make extracellular space, intracellular space of fibroblasts, cellular nuclei and gap junctions visible. Afterwards three to four three-dimensional image stacks per biopsy were produced by confocal microscopy. The image stacks were preprocessed including depth-dependent attenuation correction, noise reduction, and deconvolution. The resulting data after this final step is used for analysis or further processing (segmentation). My exemplary analysis surveyed the orientation of Cx43 covered membrane surface.Image stacks of rat hearts samples provided good results. The different channels were distinguish- able and allowed analysis of ventricular histology. In contrast the quad-labelling protocol did not work as well for rabbit but crosstalk was visible. Confer Figs. 4.4 and 4.5. Analysis of the orien- tation of the membrane surface covered with Cx43 showed that the relatively highest percentage of Cx43 is found in orientation of the principal axis.