Backscattered Scanning Electron Microscopy Approach for Assessment of Microvessels under Conditions of Normal Microanatomy and Pathological Neovascularization.

We evaluated the efficiency of an original method for studying of the microvascular bed under conditions of normal microanatomy and pathological neovascularization. The blood vessels, tissues surrounding the stent in the pulmonary artery and subcutaneously implanted titanium nickelide plate, atherosclerotic plaque, and vascular stent with restenosis were examined. The specimens were fixed in formalin and stained in OsO4, embedded into fresh epoxy resin, grinded, polished, and counterstained with uranyl acetate and lead citrate. Numerous vasa vasorum were found in all native vessels. Around the pulmonary artery stent and metal plates, numerous newly formed vessels of small diameter were seen. The intensity of neovascularization in atherosclerosis and carotid stent restenosis differed significantly. Our technique can be successfully used for evaluation of the microvascular bed.

[Influence of unmodified and modified sutures on experimental abdominal adhesive process]. / Éksperimental'naia otsenka spaechnogo protsessa v briushnoi polosti pri ispol'zovanii nemodifitsirovannogo shovnogo materiala i modifitsirovannogo geparinom.

OBJECTIVE: Experimental assessment of the effect of modified and unmodified surgical suture material on abdominal adhesive process. MATERIAL AND METHODS: The study was performed on male rats of the Wistar subpopulation. There were 5 animals in each group. In all animals, midline abdominal incision was followed by suturing the parietal peritoneum with modified and unmodified suture material. All animals were euthanized with carbon dioxide vapors in 14 days after surgery. Macro- and microscopic assessment of severity of abdominal adhesive process was carried out. Two types of preparation of excised complexes 'peritoneum-suture material-adhesion' were applied for histological examination: paraffin sections and embedding in epoxy resin. Specimens were stained by Van Gieson and with methylene blue solution. Histological specimens were examined using Axio Imager A1 light microscope (Zeiss, Germany). RESULTS: Polypropylene filaments result extensive adhesions occupying about 75% of the area. Adhesions have a dense structure with signs of vascularization. Modification of suture material with solution of polyhydroxybutyrate/hydroxyvalerate and heparin reduce severity of adhesions. The use of modified suture material was followed by adhesions with more loose structure, no signs of vascularization. Adhesions occupied less than 25% of the area. Histological examination of excised complexes 'peritoneum-suture material-adhesion' revealed accumulation of inflammatory cells around the unmodified suture material, while there were no signs of tissue inflammatory process around the modified sutures. CONCLUSION: Application of polyhydroxybutyrate/hydroxyvalerate and heparin on the surface of surgical sutures is an effective method for prevention of abdominal adhesions.

A Novel Technique for Preparation, Staining, and Visualization of Tissue with Metal Implants and Extraskeletal Calcification Areas.

The aim of the study was to evaluate the efficacy of a novel technique for preparation, staining, and visualization of tissues containing extra-skeletal mineralization areas, all-metal implants or their prototypes for their subsequent examination using scanning electron microscopy in the backscattered electron mode. MATERIALS AND METHODS: After fixation in 10% formalin (24 h), the biomaterial (a titanium nickelide plate with the surrounding tissues after subcutaneous implantation, patented titanium alloy plates with the surrounding tissues after cranioplasty, primary and secondary calcified atherosclerotic plaques) were fixed with 1% osmium tetroxide (12 h) and then stained with 2% aqueous solution of osmium tetroxide (48 h). The samples were further stained with 2% alcoholic uranyl acetate (5 h), dehydrated with isopropanol (5 h) and acetone (1 h), impregnated with a mixture of acetone and epoxy resin Epon (1:1, 6 h) and then embedded into a fresh portion of epoxy resin (24 h), which was followed by polymerization at 60°C. After grinding and polishing, epoxy blocks were counterstained with lead citrate (7 min) and sputter-coated with carbon, then the samples were visualized by scanning electron microscopy in the backscattered electron mode. The elemental composition was studied using X-ray microanalysis. RESULTS: The developed technique allows obtaining high-quality images at five thousand-fold magnifications, provides the possibility to identify the shape and structure of intact metal and mineral inclusions, and to type the surrounding cells, distinguishing mesenchymal and immunocompetent cells by shape and cytoplasmic content. Apart from connective tissue capsule thickness and leukocyte infiltration, this technique makes it possible to estimate the number and area of newly formed small-caliber vessels representing a surrogate marker of inflammation. CONCLUSION: The proposed technique provides the possibility to investigate adequately the structure of samples when their sectioning is impossible or significantly complicated, with image quality remarkably higher than that obtained by light microscopy.