THE EFFECT OF THROMBIN ON LOW-DENSITY-LIPOPROTEIN PERMEABILITY AND UPTAKE BY AN ARTERIAL ENDOTHELIAL SMOOTH-MUSCLE CELL BILAYER

Thrombin, a mediator of thrombosis, has been shown to directly alter the function of vascular cells. We studied the effect of thrombin on low-density lipoprotein permeability and uptake by an arterial endothelial cell-smooth muscle cell bilayer to determine its potential role in atherogenesis. Confluent cell bilayers were incubated in media containing thrombin (10 or 50 units/ml) for a period of 24 hours to 9 days. Iodine 125 (I-125)-LDL (10-mu-g protein/ml) was then added to the media, and after a 3-hour incubation, I-125-LDL transit through the endothelial cell layer as well as membrane binding and uptake by each cell type were measured. The lower concentration of thrombin caused a delayed increase in both the permeability (p < 0.0001) and uptake (p < 0.05) of LDL, but had no effect on membrane binding of the lipoprotein. The higher thrombin concentration led to an immediate increase in endothelial cell permeability to LDL (p < 10(-7)) and a significant reduction in both cellular uptake (p < 10(-7)) and membrane binding (p < 0.0005). In contrast, smooth muscle cell binding and uptake were unaffected at the lower concentration of thrombin. At the higher concentration, smooth muscle cell uptake of LDL was increased (p < 10(-7)) disproportionately to a more limited increase in membrane binding (p < 0.05). Endothelial DNA content, reflecting cell number, was increased at 10 units/ml thrombin (p < 0.001) but markedly reduced at 50 units/ml thrombin (p < 0.0005), whereas smooth muscle cell DNA content remained unchanged. H-3-adenine and chromium 51 release assays indicated a greater susceptibility of endothelial cells to thrombin-induced injury. These results suggest that endothelial cells and smooth muscle cells are affected differently by thrombin. Thrombin-induced endothelial toxicity, in combination with enhanced LDL uptake by underlying smooth muscle cells, may promote the formation of atherosclerotic plaque.