The mechanisms for the transendothelial transport of macromolecules are poorly understood. Much of this lack of understanding is due to the complexity of studying transendothelial transport in situ. We have developed an in vitro model of the endothelium which facilitates precise control and monitoring of the forces on both sides of the endothelium. The model consists of monolayers of endothelial cells cultured on a subsrate impregnated micropore filter. Tight junctions develop between the endothelial cells, the endothelial monolayer develops an electrical resistance comparable to in situ endothelia, and the albumin permeability of the monolayers is also similar to in situ endothelia. Using this model, we have recently observed that the endothelium actively transports albumin from interstitium to lumen. The interstitial-to-luminal flux is favored over the luminal-to-interstitial flux by a factor of 10-40 depending on interstitial albumin concentration. The positive interstitial-to-luminal flux ratio is preserved against a concentration gradient and is abolished by depleting endothelial cell energy stores. In more recent experiments, we have found that ouabain inhibits this active interstitial-to-luminal albumin transport. The proposed studies are designed to more completely characterize this active transport process in terms of its kinetics, its energy dependence, its coupling to Na-K-ATPase activity and its ability to be regulated by messenger molecules. The active transport of albumin may be important to the normal function of the endothelium in controlling the egress of water and solutes from the vascular space. It may also be important in controlling the entrance of macromolecules into the vessel wall and hence in the pathogenesis of atherosclerosis.
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