Scanning Electron Microscopy and 3D Morphometry of Vascular Corrosion Casts: Techniques and Current Applications in Biomedical Research

Abstract

Background:  Vascular corrosion casting is an old method that has found application by anatomists for a long time. Initially, the viscosity of casting media was very high and thus the technique failed to cast the entire circulatory system from the arterial injection site through small arteries, arterioles, capillaries, venules, small veins and caval veins. Casting media soon were improved and thus enabled casting of the entire circulatory system. Examination of these vascular casts under the light microscope does not allow a thorough analysis with high resolution. Only when in 1971 Murakami applied Scanning Electron Microscopy SEM. to study vascular corrosion casts a breakthrough was achieved and for the first time, microvascular networks, too, were able to be analysed with a spatial resolution high enough to study the surface of the casts also termed injection replicas. and a depth of focus high enough to examine a large field of view enabling to reliably trace individual vessels either over very short or over extremely long distances.

Scanning electron microscopy SEM. is a tool with great potential for morphological research. The images generated via SEM have great resolution and a high depth of focus, which gives SEM micrographs pseudo three-dimensionality. Adversely, the high depth of focus prevents accurate dimensional or spatial measurements of imaged microstructures from either the SEM video-display, printed micrographs or from photonegatives. Macroscopic objects are viewed close up using binocular vision. Binocular vision is also used in microscopy where stereophotogrammetry and related techniques applying stereo paired images, and a variety of hardware tools calculate the third dimension z-coordinate. using the parallax.

 Morphometric SEM 3D analysis is currently used to analyse i. the geometry of microvascular trees in terms of vascular parameters i.e., diameters, interbranching distances, branching angles and intervascular distances. and ii. to determine bifurcation indices, asymmetry and area ratios given in arterial bifurcations respectively in venous mergings. Moreover, it allows to generate anaglyphic 3D images and to calculate optimality principles underlying the construction and maintenance of such delicate vascular networks i.e. principles of minimal lumen volume, minimal pumping power, minimal lumen surface and minimal endothelial shear force.

Aim: The main goal of the method is to characterize microvascular networks in human and animal tissues in order to demonstrate their anatomical situations and to reflect onto their physiological conditions. These methods enable not only to qualitatively describe but also to quantitatively explore the microvascularisation by means of three-dimensional morphometry and thus to compare different developmental stages and progress of disease.

Conclusions: The combination of the techniques described above allows a thorough study of microvascular networks in healthy- and pathologic tissues and organs. This enables i. to enhance the knowledge of vascular- / anatomical situations and development, ii. to compare the physiological conditions in health and different pathologies and iii. to gain insights and to develop therapeutical strategies in various clinical pictures, which are critically dependent on a proper functioning vascularization.