Increased permeability of primary cultured brain microvessel endothelial cell monolayers following TNF-α exposure

Abstract

TNF-α is a cytokine that produces increased permeability in the peripheral vasculature; however, little is known about the effects of TNF-α on the bloodbrain barrier (BBB). Using primary cultured bovine brain microvessel endothelial cells (BBMEC) as an in vitro model of the BBB, this study shows that TNF-α produces a reversible increase in the permeability of the brain microvessel endothelial cells. The BBMEC monolayers were pre-treated with 100 ng ml of TNF-α for periods ranging from 2 to 12 hours. Permeability was assessed using three molecular weight markers, fluorescein (376 MW), fluorescein-dextran (FDX-4400; 4400 MW), and FDX-70000 (MW 70000). The permeability of BBMEC monolayers to all three fluorescent markers was increased two-fold or greater in the TNF-α treatment group compared to control monolayers receiving no TNF-α. Significant changes in permeability were also observed with TNF-α concentrations as low as 1 ng ml . These results suggest that TNF-α acts directly on the brain microvessel endothelial cells in a dynamic manner to produce a reversible increase in permeability. Exposure of either the lumenal or ablumenal side of BBMEC monolayers to TNF-α resulted in similar increases in permeability to small macromolecules, e.g. fluorescein. However, when a higher molecular weight marker was used (e.g. FDX-3000), there was a greater response following lumenal exposure to TNF-α. Together, these studies demonstrate a reversible and time dependent increase in brain microvessel endothelial cell permeability following exposure to TNF-α. Such results appear to be due to TNF's direct interaction with the brain microvessel endothelial cell.