HEPARIN-INDUCED OVEREXPRESSION OF BASIC FIBROBLAST GROWTH-FACTOR, BASIC FIBROBLAST GROWTH-FACTOR RECEPTOR, AND CELL-ASSOCIATED PROTEOHEPARAN SULFATE IN CULTURED CORONARY SMOOTH-MUSCLE CELLS

Basic fibroblast growth factor (bFGF), a potent mitogen for arterial s mooth muscle cells (SMCs), plays a pivotal role in the pathogenesis of arteriosclerosis and restenosis. Heparin in nanogram quantities may p romote or even be required for binding of bFGF to its cognate receptor . Conversely, heparin in microgram doses is a strong inhibitor of arte rial SMC replication in vitro and in vivo. Bovine coronary SMCs (cSMCs ) express bFGF, bFGF receptor (FGF-R1), and cell membrane-integrated p roteoheparan sulfate (HSPG). These three molecules are known to form a trimolecular complex that promotes signal transduction and mitogenesi s. The bFGF synthesized by cSMCs is distributed to an intracellular an d a pericellular compartment. Resting cultured cells retain about 80% of their bFGF intracellularly; 20% is found in the pericellular region . During proliferation, 70% to 80% of total bFGF is expressed in the p ericellular compartment. Trypsinization generates soluble forms of the complex of bFGF with the ectodomains of the bFGF receptor and cell me mbrane-integrated HSPG in the pericellular compartment, thus allowing quantification of pericellular bFGF by a highly specific enzyme immuno assay. Standard heparin inhibits the proliferation of cSMCs by up to 8 0% in a concentration range between 10 and 100 mu g/mL medium in a dos e-dependent manner but increases the protein content of cSMCs compared with proliferating control cells. The heparin-induced increase in cel lular protein content includes a 60% to 100% increase in the expressio n of pericellular bFGF, FGF-R1, and cell membrane-integrated HSPG. Thu s, under heparin treatment, the heparan sulfate side chains of cell me mbrane-integrated HSPG incorporate more [S-35]sulfate, and the proport ion of [S-35]heparan sulfate among total glycosaminoglycans increases from 36% to 52%. Fluorescence-activated cell sorting analysis and [H-3 ]thymidine incorporation experiments provide evidence for multiple eff ects of heparin, including blocks at early and late checkpoints of the cell cycle in heparin-treated cells. These results indicate that hepa rin, despite its antiproliferative potency, stimulates the expression of all components of the bFGF system even in coronary SMCs in which gr owth is inhibited.