title: Visualization of the glomerular endothelial glycocalyx by electron microscopy using cationic colloidal thorium dioxide.
abstract: Biological material itself appears with poor contrast in electron microscopy (EM), due to its composition mostly of light elements. Classical staining agents such as osmium tetroxide, uranyl acetate, and lead citrate preserve and/or stain cellular structures such as membranes, cytoplasm, and organelles well for EM. However, extracellular polymeric substances (EPS) show no or only poor contrast with these staining agents. The endothelial glycocalyx in blood vessels consists mainly of proteoglycans. It can be visualized by EM only by additional staining with heavy metal ions such as copper (Alcian blue, cupromeronic blue), ruthenium (ruthenium red), or lanthanum. Best results are achieved by combined perfusion of fixative and stain. Cationic hydrous thorium dioxide colloids (named here cThO2) trace acidic groups in EPS. We describe here the use of cThO2 to visualize the glomerular endothelial glycocalyx in the mouse kidney. cThO2 shows high electron density and binds to a continuous layer of up to a few hundred nanometers thickness on the glomerular endothelium, as well as on epithelia in other blood vessels in perfused animals. The observed staining pattern gives rise to periodic densities, with a spacing varying between 50 and 200 nm, depending on the overall layer thickness, which varies between below 50 up to 300 nm. Due to high electron density of the used cThO2 particles, the introduced method allows distinct imaging and precise fine structural analysis of the endothelial glycocalyx.
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Hanover, Germany. Hegermann.Jan@mh-hannover.de.|Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Center for Lung Research (DZL), Hanover, Germany. Hegermann.Jan@mh-hannover.de.|REBIRTH Cluster of Excellence, Hanover, Germany. Hegermann.Jan@mh-hannover.de.
authors.forename: Jan
authors.initials: J
authors.lastname: Hegermann
authors.affiliation: ZEIM, Helmholtz Center for Infection Research, Braunschweig, Germany.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Lünsdorf
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Hanover, Germany.|Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), German Center for Lung Research (DZL), Hanover, Germany.|REBIRTH Cluster of Excellence, Hanover, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Ochs
authors.affiliation: Department of Nephrology, Hannover Medical School, Hanover, Germany.
authors.forename: Hermann
authors.initials: H
authors.lastname: Haller
mesh_terms: D000818:Animals; D003102:Colloids / Q000378:metabolism; D055032:Electron Microscope Tomography / Q000379:methods*; D042783:Endothelial Cells / Q000166:cytology*; D019276:Glycocalyx / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000166:cytology*; D051379:Mice; D008810:Mice, Inbred C57BL; D013194:Staining and Labeling; D013911:Thorium Dioxide / Q000378:metabolism*
chemical_list: D003102:Colloids; D013911:Thorium Dioxide
keywords: Glomerular endothelial glycocalyx; Thorium dioxide; Transmission electron microscopy