New aspects of microvascular corrosion casting: a scanning, transmission electron, and high-resolution intravital video microscopic study.
We used intravital microscopy of small intestine and pancreas in order to show dynamic interactions between vascular wall and undiluted Mercox, because previous studies of ours have shown that Mercox diluted with monomeric methylmethacrylate penetrates cells in the vascular wall. Scanning and transmission electron microscopy were used to show three-dimensional pathways and correlating tissue structures, which cannot be identified in vivo. The microvascular diameters were not altered when the vasculature was flushed with saline/dextran solution using perfusion pressures between 70 and 140 mm Hg, but, in circumscribed areas, contraction of vascular wall was observed immediately after Mercox injection. This phenomenon was carried out by endothelial cells; pericytes were never present at the site of constrictions. Extravasation, i.e., leakage of the resin into the surrounding tissue, occurred in circumscribed areas regardless of the applied perfusion pressure. The resin also filled routes, which were not perfused with blood before casting. Scanning microscopy of corresponding specimens showed flattened cast channels, with impressions of valves and endothelial cell nuclear imprints characteristic of lymphatics. These results show that undiluted Mercox is a stimulus for vascular cellular components and that it changes the vascular wall permeability, resulting in extravasation and filling of lymphatics. Transmission electron microscopy showed that large vessels were homogeneously filled with resin and that cellular structures were not infiltrated with Mercox. Cut sections of the gold-coated surface of casts showed grooves up to 20 nm wide, suggestive of minimal deformation, while the abluminal surface of the metal film was almost smooth. Another proof of minimal deformation of undiluted Mercox casts is that the diameter of vessels was not altered during and after polymerization. Obtained casts are not fragile, as are casts of diluted Mercox, and phase separation does not occur, which would result in penetration of the cells in the vascular wall. For these reasons, the use of undiluted Mercox is recommended. Mixing 10 ml Mercox with 1 g catalyst resulted in complete polymerization within 5.5-7 min. This mixture can be used for casting biological specimens.
Aharinejad
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Department of Cell Biology, University of Massachusetts Medical Center, Worcester 01655.
MacDonald
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MacKay
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Mason-Savas
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A
D000818:Animals; D016319:Corrosion Casting; D005260:Female; D007421:Intestine, Small / Q000098:blood supply / Q000648:ultrastructure; D008208:Lymphatic System / Q000648:ultrastructure; D008297:Male; D008833:Microcirculation / Q000648:ultrastructure*; D008854:Microscopy, Electron / Q000379:methods*; D008855:Microscopy, Electron, Scanning / Q000379:methods; D010179:Pancreas / Q000098:blood supply / Q000648:ultrastructure; D051381:Rats; D017207:Rats, Sprague-Dawley