Ultrastructural evidence of differential solubility in Triton X-100 of endothelial vesicles and plasma membrane

Abstract

Endothelial plasmalemmal vesicles (EV) are distinct membrane-bound structures characteristic for all vascular endothelia, being involved in transcytosis of plasma macromolecules. EV are considered to be similar to the caveolae (characterized by a specific peptide called caveolin) found in other cell types. Caveolin-rich membrane domains were recently isolated from whole lung and chicken gizzard as a Triton X-100 (TX)-insoluble membrane fraction. However, ultrastructural data on the localization of these domains within cells have not yet been reported. We have examined whether EV are TX-insoluble structures. Cultured bovine aortic endothelial cells (BAEC) briefly fixed in paraformaldehyde (10 min, 37 degrees C) were exposed to 0.1% TX for 5 min at 22 degrees C and further subjected to standard electron microscopy procedure. The results showed an extensive solubilization of endothelial plasmalemma as well as other intracellular membranes. Individual or clusters of EV were not affected by TX extraction, retaining their trilaminar unit membrane appearance and dimensions. Moreover, a crude membrane fraction prepared from unfixed BAEC was also extracted with 1% TX for 20 min at 4 degrees C and the insoluble material was examined by electron microscopy. In this fraction clusters of about 10 membranous vesicles were found. These data suggest that EV and plasma membrane have a different lipid composition; the low TX solubility is a characteristic common to caveolin-rich domains (caveolae) of other cells types and EV, whereas the ultrastructural complexity and intracellular localization of the latter are specific for endothelia.