Three-dimensional ultrastructure of the brush border glycocalyx in the mouse small intestine: a high resolution scanning electron microscopic study

Abstract

The three-dimensional ultrastructure of the filamentous glycocalyx of the brush border in the mouse small intestine was successfully demonstrated by high resolution scanning electron microscopy (SEM). The specimens were fixed with 2% glutaraldehyde in a 0.1M phosphate buffer (pH 7.4), and rinsed with buffered solutions with differently adjusted pH values (pH 3.0, 7.0 or 11.0). They were then osmicated, dried, spatter-coated with gold (1.0-1.5 nm), and observed under a high resolution SEM. The glycocalyx on the luminal surface of the intestinal villi covered the top of the microvilli of the epithelial cells and were well preserved in the specimens treated with an alkaline buffer (pH 11.0). The glycocalyx was observed as filamentous structures, 7 to 15 nm thick in diameter. These filaments repeatedly branched and anastomosed with neighboring ones to form an actual network or plexus as a whole, in contrast with superimposed images in transmission electron microscopy (TEM) which suggested that such anastomoses were pseudo-networks. The filaments thickened globularly at the sites of the filament bifurcation or branching. On the other hand, specimens rinsed with an acid or neutral buffer showed no glycocalyx on their microvilli, whose naked top had knob-like structures. Thus, the pH values of the washing buffer solutions were considered to affect the preservation of the surface coat due to molecular characteristics.

Similar Endothelial Glycocalyx Structures in Microvessels from a Range of Mammalian Tissues: Evidence for a Common Filtering Mechanism?

“…Horiuchi, avoiding tannic-acid staining, looked at the glycocalyx associated with intestinal microvilli, reporting glutaraldehyde-and osmium-fixed fibers 7-15 nm thick by scanning EM (gold 1-1.5 nm thick) (24). The online supplement to a study by van den Berg et al (25) shows fibers of 3.4 5 0.1 nm (range 2-9 nm in width) after fixation with Alcian blue/glutaraldehyde/lanthanum/osmium.…”

A Biliary HCO3 − Umbrella Constitutes A Protective Mechanism Against Bile Acid–Induced Injury in Human Cholangiocytes

“…24 This ''sugary'' layer mediates cell-cell recognition, receptor-ligand interactions, and adds to the barrier function of the plasma membrane, as best characterized for the endothelial 25 and intestinal glycocalyx. [26][27][28] This juxtamembranous layer may locally alter the pH near, but exterior, to the lipid bilayer. In melanoma cells, the pH measured in the glycocalyx was reported to be slightly higher than the pH of the medium, and the absolute number of H þ molecules differed up to 40% between different domains of the glycocalyx of a single cell.…”

The Origin of Mucosal Immunity: Lessons from the Holobiont Hydra

“…The glycocalyx represents a dense forest of highly diverse and constantly renewed transmembrane glycoproteins, proteoglycans and glycolipids (49). Beside mediation of cell-cell recognition and receptor-ligand interactions, the glycocalyx provides an important barrier function to the plasma membrane, best characterized for the intestinal glycocalyx (50–52). In mucosal tissues, such as the mammalian gastrointestinal tract, the glycocalyx is additionally coated by a mucus gel ranging in thickness from 10 µm in the eye up to 700 µm in the large intestine (53).…”