scholarly journals Plasminogen activator inhibitor-type I is a major biosynthetic product of retinal microvascular endothelial cells and pericytes in culture

1989 ◽  
Vol 259 (2) ◽  
pp. 529-535 ◽  
Author(s):  
A E Canfield ◽  
A M Schor ◽  
D J Loskutoff ◽  
S L Schor ◽  
M E Grant
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Type I ◽  
Human Skin Fibroblasts ◽  
Retinal Endothelial Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Activator Inhibitor ◽  
Pai 1

Previous studies have shown that a glycoprotein of Mr 47,000 (designated Gp47) is a major biosynthetic product of retinal endothelial cells in vitro (Canfield, Schor, West, Schor & Grant (1987) Biochem. J. 246, 121-129). We now present data indicating that (a) an identical protein is secreted by bovine retinal pericytes, (b) this protein is plasminogen activator inhibitor-type I (PAI-1), as revealed by immunoprecipitation with specific antibodies and reverse fibrin zymography, and (c) retinal endothelial cells and pericytes synthesize different species of matrix macromolecules, that is: type IV collagen is the major collagen secreted by endothelial cells, whereas pericytes produce predominantly type I collagen; fibronectin and thrombospondin are synthesized by both cell types. Our studies also indicate that PAI-1 is produced, albeit at considerably lower levels, by large vessel vascular cells (aortic endothelial and smooth muscle cells) and human skin fibroblasts. PAI-1 produced by human skin fibroblasts appears to be a distinct molecular species compared to its bovine counterpart as assessed by its slower mobility on SDS/polyacrylamide-gel electrophoresis. The potential significance of elevated PAI-1 production by retinal endothelial cells and pericytes, as well as their distinctive patterns of matrix biosynthesis, is discussed in terms of the involvement of these cells in the maintenance and remodelling of microvessel basement membrane.

scholarly journals Plasminogen activator inhibitor type I stabilizes vitronectin-dependent adhesions in HT-1080 cells.

1990 ◽  
Vol 111 (5) ◽  
pp. 2183-2195 ◽  
Author(s):  
G J Ciambrone ◽  
P J McKeown-Longo
Keyword(s):  
Binding Site ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Type I ◽  
Adherent Cells ◽  
Cell Rounding ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Activator Inhibitor ◽  
Pai 1

Polyclonal antibodies against plasminogen activator inhibitor type-I (PAI-1) caused rapid retraction and rounding of substrate-attached HT-1080 cells. The kinetics and extent of antibody-mediated cell rounding were not dependent on either urokinase or plasmin activity. Cells adherent to vitronectin-coated substrates detached within 2 h of antibody addition. Cells adherent to fibronectin were unaffected by the antibodies. Immunoblotting of substrate-attached material indicated that HT-1080 cells deposited PAI-1 into vitronectin, but not fibronectin, dependent contacts. These data suggest that the antibody-mediated cell rounding resulted from a steric disruption of vitronectin-dependent adhesions, indicating that the binding site on vitronectin for PAI-1 is near, but does not overlap, the binding site for vitronectin receptor. The accumulation of PAI-1 into vitronectin-dependent adhesion sites correlated temporally with the preferential degradation of fibronectin from the substrate. HT-1080 cells adherent to either fibronectin or vitronectin were able to activate exogenous plasminogen to plasmin. Plasmin levels were increased 200% on cells adherent to fibronectin and 100% on cells adherent to vitronectin. In the presence of a neutralizing antibody against PAI-1, vitronectin adherent cells activated plasminogen to the same extent as fibronectin adherent cells. Plasmin levels of 200% above baseline were associated with retraction of cells from the substrate. The ability of vitronectin adherent cells to activate exogenous plasmin was completely blocked in the presence of neutralizing antibodies against urokinase. These data represent the first demonstration that vitronectin-associated PAI-1 regulates urokinase in focal contact areas.

scholarly journals Hepatocyte growth factor stimulates expression of plasminogen activator inhibitor type 1 and tissue factor in HepG2 cells

Blood ◽  
1994 ◽  
Vol 84 (1) ◽  
pp. 151-157 ◽  
Author(s):  
J Wojta ◽  
T Nakamura ◽  
A Fabry ◽  
P Hufnagl ◽  
R Beckmann ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Cell Type ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Hepatocyte Growth ◽  
Activator Inhibitor ◽  
Pai 1

Abstract HGF is a powerful mitogen for both rat and human hepatocytes, epithelial cells and endothelial cells in vitro, and is angiogenic in vivo. It has considerable homology with plasminogen and has been shown to upregulate urokinase-type plasminogen activator (u-PA) in endothelial cells as well as u-PA and its receptor in kidney epithelial cells. In this study, we report that human recombinant HGF stimulates expression of plasminogen activator inhibitor type 1 (PAI-1) and tissue factor (TF) in the human hepatoma cell line HepG2. PAI-1 antigen as determined by a specific enzyme-linked immunosorbent assay increased up to threefold in conditioned media of HepG2. This increase was dose dependent with maximum stimulation achieved with a concentration of 50 ng/mL of hepatocyte growth factor (HGF). PAI-1 antigen also increased up to fourfold in the extracellular matrix in HGF treated HepG2. The production of the PAI-1 binding protein vitronectin (Vn) was not affected by HGF. In contrast, TF activity in HepG2 treated with HGF increased up to twofold. As determined by Northern blotting, PAI-1 and TF-specific mRNA were increased significantly in the presence of HGF, whereas Vn mRNA was not affected. The increase in PAI-1 and TF mRNA was also seen when HepG2 were incubated with HGF in the presence of cycloheximide, thereby indicating that de novo protein synthesis is not required to mediate the effect. u-PA could be detected neither in unstimulated or HGF-stimulated HepG2 cells on the antigen level nor on the mRNA level. In conclusion, our data give evidence that HGF, in addition to its proliferative effect for different cell types, is also involved in the local regulation of fibrinolysis and coagulation. One could speculate that HGF might modulate processes requiring matrix degradation by increasing the expression of the protease u-PA in one cell type and by upregulating the expression of the serine protease inhibitor PAI-1 in a different cell type. Because u-PA has been shown to activate latent HGF to the active form, it could furthermore be speculated that by upregulating PAI-1, which in turn could inhibit u- PA, HGF might regulate its own activation.

Apposition-Dependent Induction of Plasminogen Activator Inhibitor Type 1 Expression: A Mechanism for Balancing Pericellular Proteolysis During Angiogenesis

Blood ◽  
1998 ◽  
Vol 92 (3) ◽  
pp. 939-945 ◽  
Author(s):  
Eran Bacharach ◽  
Ahuva Itin ◽  
Eli Keshet
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
L Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Pericellular Proteolysis ◽  
Activator Inhibitor ◽  
Pai 1

Abstract Plasminogen-activator inhibitor type I (PAI-1), the primary inhibitor of urinary-type plasminogen activator, is thought to play an important role in the control of stroma invasion by both endothelial and tumor cells. Using an in vitro angiogenesis model of capillary extension through a preformed monolayer, in conjunction with in situ hybridization analysis, we showed that PAI-1 mRNA is specifically induced in cells juxtaposed next to elongating sprouts. To further establish that PAI-1 expression is induced as a consequence of a direct contact with endothelial cells, coculture experiments were performed. PAI-1 mRNA was induced exclusively in fibroblasts (L-cells) contacting endothelial cell (LE-II) colonies. Reporter gene constructs driven by a PAI-1 promoter and stably transfected into L-cells were used to establish that both mouse and rat PAI-1 promoters mediate apposition-dependent regulation. This mode of PAI-1 regulation is not mediated by plasmin, as an identical spatial pattern of expression was detected in cocultures treated with plasmin inhibitors. Because endothelial cells may establish direct contacts with fibroblasts only during angiogenesis, we propose that focal induction of PAI-1 at the site of heterotypic cell contacts provides a mechanism to negate excessive pericellular proteolysis associated with endothelial cell invasion. © 1998 by The American Society of Hematology.

Apposition-Dependent Induction of Plasminogen Activator Inhibitor Type 1 Expression: A Mechanism for Balancing Pericellular Proteolysis During Angiogenesis

Blood ◽  
1998 ◽  
Vol 92 (3) ◽  
pp. 939-945
Author(s):  
Eran Bacharach ◽  
Ahuva Itin ◽  
Eli Keshet
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
L Cells ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Pericellular Proteolysis ◽  
Activator Inhibitor ◽  
Pai 1

Plasminogen-activator inhibitor type I (PAI-1), the primary inhibitor of urinary-type plasminogen activator, is thought to play an important role in the control of stroma invasion by both endothelial and tumor cells. Using an in vitro angiogenesis model of capillary extension through a preformed monolayer, in conjunction with in situ hybridization analysis, we showed that PAI-1 mRNA is specifically induced in cells juxtaposed next to elongating sprouts. To further establish that PAI-1 expression is induced as a consequence of a direct contact with endothelial cells, coculture experiments were performed. PAI-1 mRNA was induced exclusively in fibroblasts (L-cells) contacting endothelial cell (LE-II) colonies. Reporter gene constructs driven by a PAI-1 promoter and stably transfected into L-cells were used to establish that both mouse and rat PAI-1 promoters mediate apposition-dependent regulation. This mode of PAI-1 regulation is not mediated by plasmin, as an identical spatial pattern of expression was detected in cocultures treated with plasmin inhibitors. Because endothelial cells may establish direct contacts with fibroblasts only during angiogenesis, we propose that focal induction of PAI-1 at the site of heterotypic cell contacts provides a mechanism to negate excessive pericellular proteolysis associated with endothelial cell invasion. © 1998 by The American Society of Hematology.

The human plasminogen activator inhibitor type I gene promoter targets to kidney

1998 ◽  
Vol 274 (2) ◽  
pp. F405-F412 ◽  
Author(s):  
Martin P. Emert ◽  
Christine M. Sorenson ◽  
David P. Basile ◽  
Joseph G. Rogers ◽  
Marc R. Hammerman ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Transgene Expression ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Type ◽  
Type I ◽  
Plasminogen Activator Inhibitor Type ◽  
Inhibitor Type ◽  
Activator Inhibitor Type ◽  
Activator Inhibitor ◽  
Pai 1

The plasminogen activator inhibitor type 1 (PAI-1) gene encodes the physiological inhibitor of tissue-type and urokinase-type plasminogen activators and is induced by cytokines such as transforming growth factor-β (TGF-β). Studies have identified DNA sequence elements within the first 1.3 kb of the 5′-upstream DNA that mediate cytokine responsiveness in transfected cells in vitro. However, the DNA sequences that mediate PAI-1 expression in vivo have not yet been delineated. To define these regulatory sequences, we generated transgenic mice that expressed a hybrid gene comprising sequences between −1,272 and +75 of the human PAI-1 gene ligated to a LacZ reporter gene. Transgene expression detected in two independent lines was observed only in kidney from embryonic day 13 to adult and was seen primarily in proximal tubule cells of the outer medulla. Transgene expression and activity were unchanged in response to TGF-β and remained restricted to kidney. Thus we have identified a promoter region within the PAI-1 gene that targets transgene expression to kidney but, unlike the native promoter, is unresponsive to TGF-β in the experimental protocol used.

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