Characterization of the blood-brain barrier: protein composition of the capillary endothelial cell membrane.
Microvessels were isolated from canine cerebral cortex, and the composition of the endothelial cell membrane was investigated. Endothelial cell membranes were separated from the surrounding basement membrane, solubilized, and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis in 12% gels. Staining with Coomassie Blue revealed a characteristic banding pattern of at least 12 major proteins with apparent molecular weights between 14,000 and 250,000. When proteins from red blood cell ghosts were run simultaneously, no similarities were observed, except for proteins at apparent molecular weights of 43,000 (band 3) and 35,000 (band 4). These two proteins migrated exactly to the positions of the erythrocyte proteins actin and glyceraldehyde 3-phosphate dehydrogenase, respectively. Membrane glycoproteins in gels were also examined by the use of fluorescent lectins. Of the fluorescein isothiocyanate-conjugated (FITC) lectins tested, only FITC-concanavalin A had an affinity for any membrane components. Diazotized [125I]iodosulfanilic acid, a membrane-impermeable reagent, was used to label the internal (lumen) cell surface and the external (antilumen) cell surface. Autoradiography and determination of radioactivity levels in gel slices showed that several proteins were specifically labeled, and that major differences in radioactivity of proteins existed in internal and external labeling experiments. It is concluded that the protein composition of the luminal membrane is different from that of the antiluminal membrane.
D000818:Animals; D001812:Blood-Brain Barrier; D002196:Capillaries / Q000032:analysis*; D002462:Cell Membrane / Q000032:analysis; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004727:Endothelium / Q000032:analysis; D004910:Erythrocyte Membrane / Q000032:analysis; D008565:Membrane Proteins / Q000032:analysis*; D008854:Microscopy, Electron; D008970:Molecular Weight
D008565:Membrane Proteins