Binding and endocytosis of heparin by human endothelial cells in culture.
Binding of heparin and low molecular weight heparin fragments (CY 222, Mr range 1500-8000) to human vascular endothelial cells was studied. Primary culture of human umbilical vein endothelial cells and either 125I or 3H-labeled heparin or [125I]CY 222 were used. Slow, saturable and specific binding was found. No other tested glycosaminoglycan, excepting a highly sulfated heparan fraction, was able to compete for heparin binding. Two groups of binding sites for [3H]heparin could be distinguished: one with high affinity (Kd = 0.12 microM) and another with lower affinity (Kd = 1.37 microM) and a relative large capacity of binding (1.16 X 10(7) molecules/cell) was calculated. The Kd for unlabeled heparin, as calculated from competition experiments, was 0.23 microM. Much lower affinity was calculated for unlabeled low molecular weight heparin fragments CY 222 (Kd = 4.3 microM) from competition experiments with [125I]CY 222. The binding reversibility was only partial for unfractionated heparin. Even by chasing with unlabeled compound, a fraction of 25-30% was not dissociable from endothelial cells. This fraction was much lower if incubation was carried out at 4 degrees C. The addition of basic proteins (histones) to the incubation medium greatly enhanced the undissociable binding at 37 degrees C, but not at 4 degrees C. The undissociable fraction of heparin was not available to degradation by purified microbial heparinase. These results suggest that a fraction of bound heparin is internalized by the vascular endothelium.
D001665:Binding Sites; D001667:Binding, Competitive; D002478:Cells, Cultured; D004705:Endocytosis; D004727:Endothelium / Q000187:drug effects / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism*; D019763:Heparin Lyase; D006657:Histones / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011479:Protamines / Q000494:pharmacology; D014471:Umbilical Veins / Q000378:metabolism*
D006657:Histones; D011479:Protamines; D006493:Heparin; D011133:Polysaccharide-Lyases; D019763:Heparin Lyase