Removal of heparan sulfate by heparinase treatment inhibits FGF-2-dependent smooth muscle cell proliferation in injured rat carotid arteries.

ABSTRACT

Smooth muscle cells (SMC) of the rat carotid arterial media proliferate and migrate in response to injury during the formation of a neointima. The interaction of fibroblast growth factor (FGF-2), which is released at the site of injury, with heparan sulfate proteoglycans (HSPGs) is necessary to induce signaling, which elicits an FGF-dependent mitogenic response by arterial smooth muscle cells, and also serves as a mechanism for storage of the growth factor within the extracellular matrix. However, whether these interactions are critical during neointimal formation has not been directly tested. In this study, a model of FGF-2-dependent medial SMC mitogenic response in balloon-injured rat carotid artery was used to test the effect of degradation of vessel wall heparan sulfate on subsequent SMC proliferation. Treatment of balloon-catheterized rat carotid arteries with chondroitin ABC lyase and/or heparin lyases eliminated heparan sulfates in the vessel wall, as determined by immunoperoxidase staining. In contrast, the distribution in the carotid vessel wall of the large core protein of perlecan, a major vessel wall HSPG that binds FGF-2, is not decreased. The effect of glycosaminoglycan digestion in situ on medial SMC proliferation in response to a bolus injection of FGF-2 after injury was determined by measuring the percentage of SMC nuclei that incorporated 5-bromo-2'-deoxyuridine (BrdU) 48 h after injury. Enzymatic removal of heparan sulfate reduced BrdU incorporation into medial SMC by 60-70% (P < 0.001) at 48 h after injury. Moreover, pre-incubation of FGF-2 with heparin prior to injection restored SMC replication to the levels present in injured vessels treated with buffer alone (P < 0.01). These experiments indicate that endogenous HSPGs are essential to promote FGF-2-driven medial SMC proliferation following injury, and that heparinase treatment can abrogate FGF-2-dependent responses in vivo.