Histological evaluation of surgical experiments in animal models

Introduction: The dissertation is based on six studies that focus on the application of quantitative histology in animal model experiments. It includes a presentation of virtual microscopy procedures and image field sampling strategies, mapping changes in the microscopic structure of ovine and porcine carotid segments and their comparison with human coronary arteries and internal thoracic arteries, vascularization assessment in a mouse model of lymphoma xenografts (PDX), the effect of hyperbaric oxygen therapy on type III collagen production and on vascularization in a skin wound in a Zucker Diabetic Fatty rat. Methods: The review article about virtual microscopy was focused on an example of sampling images from various areas of quantitative histology. In other studies, histologically processed sections were stained with a variety of methods for vascular wall construction, cell infiltration (orcein, picrosirius red, Verhoeff's hematoxylin and green trichrome, Gill's hematoxylin, alcian blue) and immunohistochemical antigen detection (α-smooth muscle actin, neurofilament protein, CD-31, von Willebrand factor). Using unbiased sampling and stereological methods, we quantified the area fraction of components (elastin, collagen, smooth muscle actin and chondroitin sulfate) using a stereological grid...Úvod: Dizertační práce je založena na šesti studiích, které se zaměřují na uplatnění kvantitativní histologie v hodnocení experimentů u zvířecích modelů. Zahrnuje představení postupů virtuální mikroskopie a strategií vzorkování obrazových polí, mapování změn mikroskopické struktury segmentů ovčích a prasečích krkavic a jejich porovnání s lidskými koronárními cévami a arteria thoracica interna, hodnocení vaskularizace u myšího modelu s xenografty lymfomů (PDX), vliv hyperbarické oxygenoterapie na tvorbu kolagenu typu III a na vaskularizaci v kožní ráně u Zucker Diabetic Fatty potkana. Metody: Přehledový článek o virtuální mikroskopii byl zaměřen na ukázku příkladu vzorkování snímků z různých oblastí kvantitativní histologie. V ostatních studiích byly histologicky zpracované řezy barvené škálou metod zaměřených na stavbu cévní stěny, a buněčné osídlení (orcein, pikrosiriová červeň, Verhoeffův hematoxylin a zelený trichrom, Gillův hematoxylin, alcianová modř) a imunohistochemickým průkazem antigenů (α-hladký svalový aktin, neurofilamentový protein, CD-31, von Willebrandův faktor). Pomocí nevychýleného vzorkování a stereologických metod jsme kvantifikovali plošné podíly složek (elastin, kolagen, hladkosvalový aktin a chondroitinsulfát) použitím stereologické bodové mřížky; dvourozměrnou hustotu...Ústav histologie a embryologieLékařská fakulta v PlzniFaculty of Medicine in Pilse

Histological evaluation of surgical experiments in animal models

Introduction: The dissertation is based on six studies that focus on the application of quantitative histology in animal model experiments. It includes a presentation of virtual microscopy procedures and image field sampling strategies, mapping changes in the microscopic structure of ovine and porcine carotid segments and their comparison with human coronary arteries and internal thoracic arteries, vascularization assessment in a mouse model of lymphoma xenografts (PDX), the effect of hyperbaric oxygen therapy on type III collagen production and on vascularization in a skin wound in a Zucker Diabetic Fatty rat. Methods: The review article about virtual microscopy was focused on an example of sampling images from various areas of quantitative histology. In other studies, histologically processed sections were stained with a variety of methods for vascular wall construction, cell infiltration (orcein, picrosirius red, Verhoeff's hematoxylin and green trichrome, Gill's hematoxylin, alcian blue) and immunohistochemical antigen detection (α-smooth muscle actin, neurofilament protein, CD-31, von Willebrand factor). Using unbiased sampling and stereological methods, we quantified the area fraction of components (elastin, collagen, smooth muscle actin and chondroitin sulfate) using a stereological grid..

Histological mapping of porcine carotid arteries - An animal model for the assessment of artificial conduits suitable for coronary bypass grafting in humans

Background: Using animal models in experimental medicine requires mapping of their anatomical variability. Porcine common carotid arteries (CCA) are often preferred for the preclinical testing of vascular grafts due to their anatomical and physiological similarity to human small-diameter arteries. Comparing the microscopic structure of animal model organs to their human counterparts reveals the benefits and limitations of translational medicine. Methods: Using quantitative histology and stereology, we performed an extensive mapping of the regional proximodistal differences in the fractions of elastin, collagen, and smooth muscle actin as well as the intima-media and wall thicknesses among 404 segments (every 1 cm) of porcine CCAs collected from male and female pigs (n = 21). We also compared the microscopic structure of porcine CCAs with segments of human coronary arteries and one of the preferred arterial conduits used for the coronary artery bypass grafting (CABG), namely, the internal thoracic artery (ITA) (n = 21 human cadavers). Results: The results showed that the histological structure of left and right porcine CCA can be considered equivalent, provided that gross anatomical variations of the regular branching patterns are excluded. The proximal elastic carotid (51.2% elastin, 4.2% collagen, and 37.2% actin) transitioned to more muscular middle segments (23.5% elastin, 4.9% collagen, 54.3% actin) at the range of 2-3 centimeters and then to even more muscular distal segments (17.2% elastin, 4.9% collagen, 64.0% actin). The resulting morphometric data set shows the biological variability of the artery and is made available for biomechanical modeling and for performing a power analysis and calculating the minimum number of samples per group when planning further experiments with this widely used large animal model. Conclusions: Comparison of porcine carotids with human coronary arteries and ITA revealed the benefits and the limitations of using porcine CCAs as a valid model for testing bioengineered small-diameter CABG vascular conduits. Morphometry of human coronary arteries and ITA provided more realistic data for tailoring multilayered artificial vascular prostheses and the ranges of values within which the conduits should be tested in the future. Despite their limitations, porcine CCAs remain a widely used and well-characterized large animal model that is available for a variety of experiments in vascular surgery