Binding of Urokinase to Low Density Lipoprotein-related Receptor (LRP) Regulates Vascular Smooth Muscle Cell Contraction

Abstract

Urokinase plasminogen activator (uPA) is a multifunctional protein that has been implicated in several physiological and pathological processes involving cell adhesion and migration in addition to fibrinolysis. In a previous study we found that two-chain urokinase plasminogen activator (tcuPA) stimulates phenylephrine-induced vasoconstriction of isolated rat aortic rings. In the present paper we report that uPA ؊/؊ mice have a significantly lower mean arterial blood pressure than do wild type mice and that aortic rings from uPA ؊/؊ mice show an attenuated contractile response to phenylephrine. In contrast, the blood pressure of urokinase receptor knockout (uPAR ؊/؊ ) mice and the response of their isolated aortic rings to phenylephrine were normal, indicating that the effect of uPA on vascular contraction is independent of uPAR. Addition of mouse and human uPA almost completely reversed both the impaired vascular contractility and the lower arterial blood pressure in vivo. The in vitro and in vivo effects of infused uPA on aortic contractility and the restoration of normal blood pressure in uPA ؊/؊ mice were prevented by antibody to low-density lipoprotein receptor-related protein/␣ 2 -macroglobulin receptor (LRP). A modified form of uPA that lacks the kringle failed to restore the blood pressure in uPA ؊/؊ mice, notwithstanding having a longer half-life in the circulation. Ligands that regulate the interaction of uPA with LRP, such as PAI-1 or the PAI-1-derived peptide (EEIIMD), abolished the vasoactivity of tcuPA in vitro and in vivo. These studies identify a novel signal transducing cellular receptor pathway involved in the regulation of vascular contractility.Urokinase plasminogen activator (uPA) 1 is a multifunctional protein that has been implicated in several physiological and pathological processes, including fibrinolysis. Transgenic mice with a targeted disruption in the uPA gene (uPA Ϫ/Ϫ ) are prone to form thrombi when exposed to endotoxin (1) or hypoxia (2, 3) or when the uPA gene is disrupted in otherwise healthy tPA Ϫ/Ϫ mice (1, 4). We recently reported that clearance of pulmonary microemboli in uPA Ϫ/Ϫ mice is delayed, despite the presence of an intact tPA system (5).uPA has also been implicated in other pathophysiological processes, such as pulmonary inflammation and repair, in which the relationship to fibrinolysis is less clear. For example, uPA Ϫ/Ϫ mice are more susceptible to lethal pulmonary infection (6) and to the development of pulmonary fibrosis (7), endpoints that might reflect contribution of uPA in cell adhesion (8) and migration to these phenotypes. However, the mechanism by which uPA is involved in these processes has not been established.In a previous study we found that uPA enhances phenylephrine and endothelin-induced vasoconstriction of aortic rings isolated from rats (9). The possibility that uPA contributes to the regulation of vascular tone may help to explain some of the phenotypic changes described in uPA Ϫ/Ϫ mice and perhaps provide a broader understanding of the role pl...

Urokinase Plasminogen Activator Regulates Pulmonary Arterial Contractility and Vascular Permeability in Mice

“…The procontractile effect of uPA is independent of uPAR, and is mediated through LRP (16). LRP antagonists abolish the vasoactivity of uPA in vitro and in vivo, as do ligands that regulate the interaction of uPA with LRP, such as PAI-1 or a PAI-1-derived peptide (EEIIMD) that binds to the "docking site" in uPA for PAI-1 (16). Similar effects of uPA on vascular contractility were observed within the cerebral circulation (17,20,21).…”

“…The protein was synthesized and characterized as we reported previously (6), and was co-immunoprecipitated as previously described (6,16,29) (see the online supplement for additional details).…”

“…Isometric tension in aortic rings from rats and mice and pulmonic rings from rats were measured as described previously (6,16) (see the online supplement for additional details).…”

“…uPA is a multifunctional protein that was implicated in tracheal smooth muscle cell (SMC) contractility (6), the activation of leukocytes (7), and vascular remodeling (8)(9)(10), among other activities relevant to lung injury (11)(12)(13)(14). Some of these phenotypic changes require the proteolytic activity of uPA, whereas others involve intracellular signaling through cellular receptors, including the uPA receptor (uPAR) (10,15), the low-density lipoproteinrelated receptor (LRP) (16,17), specific integrins (7), and N-methyl-D-aspartate receptors (NMDA-Rs) (6).…”

“…We previously reported that uPA regulates the vascular contractility of isolated aortic rings (16,19), and that uPA 2/2 mice exhibit significantly lower mean arterial blood pressure than do wild-type mice, indicating a predominant procontractile effect of uPA in the systemic arterial circulation under physiological conditions (16). The procontractile effect of uPA is independent of uPAR, and is mediated through LRP (16). LRP antagonists abolish the vasoactivity of uPA in vitro and in vivo, as do ligands that regulate the interaction of uPA with LRP, such as PAI-1 or a PAI-1-derived peptide (EEIIMD) that binds to the "docking site" in uPA for PAI-1 (16).…”