Vascular Plasminogen Activator Inhibitor-1 Research Articles

PAI-1 and TGF-β

Atherosclerosis is an extremely complex disease process in which genetic, lipid, cellular, and immunologic factors combine to determine the location, severity, and timing of lesion development and clinical events.1 With the current epidemic of obesity and the metabolic syndrome, additional factors are now recognized as contributors to the vascular disease equation, including plasminogen activator inhibitor-1 (PAI-1),2 which is one of the critical physiological regulators of plasminogen activation. PAI-1 accumulates in atherosclerotic lesions3 and contributes to a variety of vascular pathologies including coronary artery thrombosis4 and perivascular fibrosis.5,6 Numerous factors are known to regulate vascular PAI-1 production, including nitric oxide (NO), which directly suppresses PAI-1 expression.7 Simply inhibiting vascular NO production stimulates arterial PAI-1 accumulation and promotes the development of PAI-1–dependent perivascular fibrosis.8 Other factors promote vascular pathology and arteriosclerosis through mechanisms that likely involve PAI-1, including Angiotensin II9 and …

Chronic blockade of the angiotensin II receptor has a differential effect on adipose and vascular PAI-1 in OLETF rats

Angiotensinogen (AGT) and plasminogen activator inhibitor-1 (PAI-1) are expressed in both vascular and adipose tissues. Angiotensin II (AG II) has an adipogenic effect and increases PAI-1 expression. To evaluate the chronic effects of AG II type 1 receptor (AT 1R) antagonism on adipose mass and PAI-1 expression in vascular and adipose tissues, losartan (30 mg/kg/day) was administered to Otsuka Long-Evans Tokushima Fatty (OLETF) rats, a model of type 2 diabetes, for 20 weeks. Adipose mass and regional fat distribution in the abdomen did not change after chronic AT 1R antagonism in OLETF rats. AGT and PAI-1 mRNA expressions in adipose tissue of OLETF rats were significantly increased compared with Long-Evans Tokushima Otsuka (LETO) rats, the normal control. Chronic losartan therapy further increased the level of adipose AGT in OLETF rats, but did not affect the level of adipose PAI-1 mRNA. In contrast, aortic PAI-1 expression in OLETF rats was attenuated by chronic losartan therapy. Our results have two implications. First, adipose tissue may be an important source of AG II in metabolic syndrome even after chronic losartan therapy. Second, chronic AT 1R antagonism with losartan causes differential effects on vascular and adipose PAI-1 expression.

PAI-1 and atherothrombosis.

Plasminogen activator inhibitor-1 (PAI-1) is the major physiologic inhibitor of tissue-type plasminogen activator in plasma, and is elevated in a variety of clinical situations that are associated with increased risk of ischemic cardiovascular events. Recent insights into the biology of PAI-1 suggest that it is more than just an innocent bystander in the pathogenesis of ischemic heart disease. Elevated PAI-1 levels appear to increase the risk of atherothrombotic events and may also promote the progression of vascular disease. The development and testing of specific PAI-1 antagonists will enable basic and clinical investigators the opportunity to test the hypothesis that vascular PAI-1 excess promotes the development of intravascular thrombosis and atherosclerosis.

Plasminogen activator inhibitor-1 deficiency prevents hypertension and vascular fibrosis in response to long-term nitric oxide synthase inhibition.

Long-term inhibition of nitric oxide synthase (NOS) is known to induce hypertension and perivascular fibrosis. Recent evidence also suggests that long-term NOS inhibition induces expression of plasminogen activator inhibitor-1 (PAI-1) in vascular tissues and that PAI-1 may contribute to the development of fibrosis after chemical or ionizing injury. On the basis of these observations, we hypothesized that PAI-1 may influence the vascular response to long-term NOS inhibition by N(omega)-nitro-L-arginine methyl ester (L-NAME). We compared the temporal changes in systolic blood pressure and coronary perivascular fibrosis in PAI-1-deficient (PAI-1(-/-)) and wild-type (WT) male mice (N=6 per group). At baseline, there were no significant differences in blood pressure between groups. After initiation of L-NAME, systolic blood pressure increased in both groups at 2 weeks. Over an 8-week study period, systolic blood pressure increased to 141+/-3 mm Hg in WT animals versus 112+/-4 mm Hg in PAI-1(-/-) mice (P<0.0001). The extent of coronary perivascular fibrosis increased significantly in L-NAME-treated WT mice (P<0.01 versus PAI-1(-/-) mice). Cardiac type I collagen mRNA expression was greater in control (P<0.01) and L-NAME-treated PAI-1(-/-) (P<0.05) groups than in control WT mice, indicating that PAI-1 deficiency prevents the increase of collagen deposition by promoting matrix degradation. These findings suggest that PAI-1 deficiency alone is sufficient to protect against the structural vascular changes that accompany hypertension in the setting of long-term NOS inhibition. Direct inhibition of vascular PAI-1 activity may provide a new therapeutic strategy for the prevention of arteriosclerotic cardiovascular disease.

Vascular release of plasminogen activator inhibitor-1 impairs fibrinolysis during acute arterial thrombosis in mice

The role of plasminogen activator inhibitor-1 (PAI-1) in the plasma, blood platelets, and vessel wall during acute arterial thrombus formation was investigated in gene-deficient mice. Photochemically induced thrombosis in the carotid artery was analyzed via transillumination. In comparison to thrombosis in C57BL/6J wild-type (wt) mice (113 +/- 19 x 10(6) arbitrary light units [AU] n = 15, mean +/- SEM), thrombosis in PAI-1(-/-) mice (40 +/- 10 x 10(6) AU, n = 13) was inhibited (P <.01), indicating that PAI-1 controls fibrinolysis during thrombus formation. Systemic administration of murine PAI-1 into PAI-1(-/-) mice led to a full recovery of thrombotic response. Occurrence of fibrinolytic activity was confirmed in alpha(2)-antiplasmin (alpha(2)-AP)-deficient mice. The sizes of thrombi developing in wt mice, in alpha(2)-AP(+/-) and alpha(2)-AP(-/-) mice were 102 +/- 35, 65 +/- 8.1, and 13 +/- 6.1 x 10(6) AU, respectively (n = 6 each) (P <.05), compatible with functional plasmin inhibition by alpha(2)-AP. In contrast, thrombi in wt mice, t-PA(-/-) and u-PA(-/-) mice were comparable, substantiating efficient inhibition of fibrinolysis by the combined PAI-1/alpha(2)-AP action. Platelet depletion and reconstitution confirmed a normal thrombotic response in wt mice, reconstituted with PAI-1(-/-) platelets, but weak thrombosis in PAI-1(-/-) mice reconstituted with wt platelets. Accordingly, murine (wt) PAI-1 levels in platelet lysates and releasates were 0.43 +/- 0.09 ng/10(9) platelets and plasma concentrations equaled 0.73 +/- 0.13 ng/mL. After photochemical injury, plasma PAI-1 rose to 2.9 +/- 0.7 ng/mL (n = 9, P <.01). The plasma rise was prevented by ligating the carotid artery. Hence, during acute thrombosis, fibrinolysis is efficiently prevented by plasma alpha(2)-AP, but also by vascular PAI-1, locally released into the circulation after endothelial injury.

Vascular release of plasminogen activator inhibitor-1 impairs fibrinolysis during acute arterial thrombosis in mice

AbstractThe role of plasminogen activator inhibitor-1 (PAI-1) in the plasma, blood platelets, and vessel wall during acute arterial thrombus formation was investigated in gene-deficient mice. Photochemically induced thrombosis in the carotid artery was analyzed via transillumination. In comparison to thrombosis in C57BL/6J wild-type (wt) mice (113 ± 19 × 106 arbitrary light units [AU] n = 15, mean ± SEM), thrombosis in PAI-1−/− mice (40 ± 10 × 106 AU, n = 13) was inhibited (P < .01), indicating that PAI-1 controls fibrinolysis during thrombus formation. Systemic administration of murine PAI-1 into PAI-1−/− mice led to a full recovery of thrombotic response. Occurrence of fibrinolytic activity was confirmed in α2-antiplasmin (α2-AP)–deficient mice. The sizes of thrombi developing in wt mice, in α2-AP+/− and α2-AP−/− mice were 102 ± 35, 65 ± 8.1, and 13 ± 6.1 × 106 AU, respectively (n = 6 each) (P < .05), compatible with functional plasmin inhibition by α2-AP. In contrast, thrombi in wt mice, t-PA−/− and u-PA−/−mice were comparable, substantiating efficient inhibition of fibrinolysis by the combined PAI-1/α2-AP action. Platelet depletion and reconstitution confirmed a normal thrombotic response in wt mice, reconstituted with PAI-1−/− platelets, but weak thrombosis in PAI-1−/− mice reconstituted with wt platelets. Accordingly, murine (wt) PAI-1 levels in platelet lysates and releasates were 0.43 ± 0.09 ng/109 platelets and plasma concentrations equaled 0.73 ± 0.13 ng/mL. After photochemical injury, plasma PAI-1 rose to 2.9 ± 0.7 ng/mL (n = 9, P < .01). The plasma rise was prevented by ligating the carotid artery. Hence, during acute thrombosis, fibrinolysis is efficiently prevented by plasma α2-AP, but also by vascular PAI-1, locally released into the circulation after endothelial injury.

P2Y receptor regulation of PAI-1 expression in vascular smooth muscle cells.

P2Y-type purine and pyrimidine nucleotide receptors play important roles in the regulation of vascular hemostasis. In this article, the regulation of plasminogen activator inhibitor-1 (PAI-1) expression in rat aortic smooth muscle cells (RASMCs) by adenine and uridine nucleotides was examined and compared. Northern analysis revealed that RASMCs express multiple P2Y receptor subtypes, including P2Y(1), P2Y(2), and P2Y(6). Treatment of RASMCs with UTP increased PAI-1 mRNA expression and extracellular PAI-1 protein levels by 21-fold (P<0.001) and 7-fold (P<0.001), respectively. The ED(50) for the effect of UTP on PAI-1 expression was approximately 1 micromol/L, and its maximal effect occurred at 3 hours. UDP stimulated a 5-fold increase (P<0.005) in PAI-1 expression. In contrast to these potent stimulatory effects of uridine nucleotides, ATP and 2-methylthioadenosine triphosphate (2-MeSATP) caused a small and transient increase in PAI-1 mRNA at 1 hour, followed by a rapid decrease to baseline levels. ADP produced only an inhibitory effect, reducing PAI-1 mRNA levels by 63% (P<0.05) at 3 hours. The relative nucleotide potency in stimulating PAI-1 expression is UTP>UDP>ATP=2-MeSATP, consistent with a predominant role of the P2Y(6) receptor. Further studies revealed that exposure of RASMCs to either ATP or ADP for 3 hours inhibited both UTP- and angiotensin II-stimulated PAI-1 expression by up to 90% (P<0.001). Thus, ATP induced a small and transient upregulation of PAI-1 that was followed by a strong inhibition of PAI-1 expression. These results show that extracellular adenine and uridine nucleotides exert potent and opposing effects on vascular PAI-1 expression.

In Vivo Stimulation of Vascular Plasminogen Activator Inhibitor-1 Production by very Low-Density Lipoprotein Involves Transcription Factor Binding to a VLDL-Responsive Element

High plasma levels of plasminogen activator inhibitor-1 (PAI-1) are associated with an increased risk of cardiovascular disease. There is also a close relation between high plasma levels of PAI-1 and hypertriglyceridemia. Cell culture studies have shown that very low density lipoprotein (VLDL) increases the production and secretion of PAI-1 in endothelial cells and hepatocytes, suggesting a possible mechanism for this association. To determine whether VLDL stimulates PAI-1 production in vascular cells also in vivo, Sprague-Dawley rats were injected intravenously with 6 mg/kg of VLDL (derived from human subjects with type IV hyperlipidemia). Previous studies have demonstrated that this results in an accumulation of human VLDL in the aorta and other arteries followed by increased nuclear factor-kappa B (NF-kappaB) activation. Endothelial, but not smooth muscle cells, showed a basal PAI-1 mRNA and protein expression as assessed by in situ hybridization and immunohistochemistry, respectively. Six to twenty-four hours after the VLDL injection, lipoprotein particle accumulation was seen in the aortic wall, which was accompanied by increasing PAI-1 mRNA and protein expression in endothelial and smooth muscle cells. Within the rat PAI-1 promoter we identified a sequence located at -589 to -571 with 74% homology with the recently described VLDL responsive element in the human PAI-1 promoter and located adjacent to a 4-guanosine motif presumably corresponding to the human 4G/5G polymorphism. Transient transfection studies showed that VLDL exerts its stimulatory effects on rat PAI-1 gene expression in vascular cells by interaction with promoter sequences located within bp -656 and -505. Electrophoretic mobility shift assays showed that VLDL increases the binding of as yet incompletely characterized factors to this response element. Taken together these observations support a direct influence of VLDL on vascular PAI-1 gene expression ill vivo. This stimulation is exerted on the level of PAI-1 gene transcription, and involves transcription factor binding to a VLDL responsive element adjacent to a 4G motif within the PAI-1 promoter.

Natriuretic factors and nitric oxide suppress plasminogen activator inhibitor-1 expression in vascular smooth muscle cells. Role of cGMP in the regulation of the plasminogen system.

Increased expression of plasminogen activator inhibitor-1 (PAI-1) has been reported in atherosclerotic and balloon-injured vessels. Little is known regarding the factors and mechanisms that may negatively regulate PAI-1 expression. In this report, the effect of cGMP-coupled vasoactive hormones, including natriuretic factors and nitric oxide, on the regulation of PAI-1 expression in vascular smooth muscle cells was examined. Atrial natriuretic factor 1-28 (ANF) and C-type natriuretic factor-22 (CNP) reduced angiotensin II (Ang II)- and platelet-derived growth factor-stimulated PAI-1 mRNA expression in rat aortic smooth muscle cells by 50% to 70%, with corresponding reductions in PAI-1 protein release. Treatment of human aortic smooth muscle cells with CNP similarly inhibited both platelet-derived growth factor-induced PAI-1 mRNA expression and PAI-1 protein release by 50%. Dose-response studies revealed that the inhibitory effects of CNP and ANF on PAI-1 expression were concentration dependent, with IC50s of approximately 1 nmol/L for both natriuretic peptides. Ang II-stimulated PAI-1 expression was also inhibited by the nitric oxide donor S-nitroso-N-acetylpenicillamine. The membrane-permeant cGMP analogue 8-Br-cGMP reduced Ang II-stimulated PAI-1 expression by 60%, and an inhibitor of soluble guanylyl cyclase (1H-[1,2,4]oxadiazolo[4, 3-a]quinoxalin-1-one) significantly impaired the inhibitory effects of S-nitroso-N-acetylpenicillamine on Ang II-stimulated PAI-1 expression. Studies of PAI-1 mRNA stability in cells treated with actinomycin D showed that ANF did not alter PAI-1 mRNA half-life, suggesting that natriuretic factors reduce PAI-1 transcription. These data show that natriuretic factors and nitric oxide, via a cGMP-dependent mechanism, inhibit PAI-1 synthesis in vascular smooth muscle cells. Thus, cGMP-coupled vasoactive hormones may play an important role in suppressing vascular PAI-1 expression.