Pathogenetic implications of hyaluronan-induced modification of vascular smooth muscle cell fibrinolysis in diabetes.

Abstract

Proteolysis, modulated in part by intramural fibrinolytic system proteins and their inhibitors, appears to influence vascular smooth muscle cell (SMC) migration and proliferation and remodeling of extracellular matrix (ECM). Alterations of fibrinolysis in circulating blood and of proteolysis within vessel walls in experimental animals and patients with diabetes have been associated with accelerated vascular disease. Hyaluronan, a prominent component of ECM in normal vessels, is increased in the tunica media of macroscopically normal arterial vessels from patients with type 2 diabetes. To determine whether hyaluronan alters the expression of the fibrinolytic system protein, plasminogen activator inhibitor type-1 (PAI-1), in human vascular SMCs, thereby potentially accelerating vascular disease in patients with type 2 diabetes. Urokinase-type and tissue-type plasminogen activators (uPA and tPA) and PAI-1 were assayed in vascular SMC conditioned media and in cell lysates, using enzyme-linked immunosorbent assay and western blotting. Hyaluronan increased the 24-h release of PAI-1 into conditioned media in a concentration-dependent and time-dependent manner (1.8-fold compared with control with 1 mg/ml hyaluronan; n = 9, P < 0.01). Although the accumulation of uPA in conditioned media tended to increase also, uPA content was reduced in cell lysates (64% of control with 0.1 mg/ml hyaluronan at 24 h; n = 9, P < 0.01) without any change in PAI-1. Concentrations of tPA in conditioned media and cell lysates were unchanged. Digestion of hyaluronan with hyaluronidase (50 turbidity reducing units (TRU)/ml) or exposure of the smooth muscle cells to antihuman CD44 antibody (1 microgram/ml) that binds to the hyaluronan cell surface receptor obviated the effects of hyaluronan. Our results indicate that increases in hyaluronan increase vascular SMC expression of PAI-1, a phenomenon that may alter the balance between proteolysis and its inhibition in vessels of patients with type 2 diabetes, thereby contributing to the acceleration of macroangiopathy.