Characterization of an in vitro model to study the permeability of human arterial endothelial cell monolayers

A model has been developed to study the transport of fluid and macromolecules through human arterial umbilical cord endothelial cell monolayers in vitro. Cells were cultured on fibronectin-coated polycarbonate filters and formed within a few days a tight monolayer, with an electrical resistance of 17 +/- 4 Ohm.cm2. The cells were connected by close cell contacts with tight junctions. The passage-rate of horse radish peroxidase (HRP) through these filters was 20-40 fold lower than through filters without an endothelial monolayer. The continuous presence of 10% human serum was needed to maintain the electrical resistance of the monolayer and its barrier function towards macromolecules. Chelation of extracellular calcium resulted in an increased permeability and a decreased electrical resistance of the monolayer. This process was reversible by re-addition of calcium ions to the cells. The permeation rate of dextrans of various molecular weights (9-480 kD) was inversely related to the molecular mass of the molecule. No difference was measured between the passage rate of dextran of 480 kD and dextran of 2,000 kD. Incubation of the endothelial cell monolayer with 2-deoxy-D-glucose resulted in a decreased permeability but it had no effect on electrical resistance. This suggests that the passage-process is energy-dependent. Fluid permeation through the endothelial cell monolayer on filters was measured in a perfusion chamber under 20 mmHg hydrostatic pressure. It was decreased by the presence of serum proteins and responded reversibly on the chelation and re-addition of extracellular calcium ions.