Characterization of [H-3]-heparin binding in human vascular smooth muscle cells and its relationship to the inhibition of DNA synthesis

1 The glycosaminoglycan heparin inhibits vascular smooth muscle cell (VSMC) proliferation and migration, but the mechanism of its antiproliferative ac tion remains unclear. Heparin has been reported to bind to high affinity ce ll surface sites on animal VSMC before undergoing receptor mediated endocyt osis resulting in signal transduction into the cytoplasm and modulation of genes involved in proliferation. In this study, we have characterized the b inding of [H-3]-heparin to human saphenous vein-derived VSMC and examined w hether there is any relationship between the affinity of [H-3]-heparin bind ing and the inhibitory effect of heparin and its structural analogues on DN A synthesis. 2 At 4 degrees C [3H]-heparin binding to human VSMC occurred in a specific, time and concentration-dependent manner and was not influenced by the remo val of calcium ions. Binding of the ligand appeared to occur to the cell su rface and was both saturable and reversible. Kinetic and steady state data indicated a single class of binding sites. 3 The pharmacology of [H-3]-heparin binding was examined in displacement st udies using unlabelled heparin and structural analogues. A comparison of th e rank potencies of heparin, heparan sulphate fraction II, low molecular we ight heparin and trehalose octasulphate showed that there was a marked disc repancy between their estimated affinities in the binding assays and their effect on DNA synthesis. 4 In summary, we have characterized the heparin binding site on human saphe nous vein-derived VSMC. Our findings suggest that the action of heparin and its analogues on DNA synthesis does not simply reflect an interaction with the cell-associated heparin binding site defined in these studies, but may also be determined by the internalization and metabolism of the glycosamin oglycan(s).