scholarly journals The Independence of Signaling Pathways Mediating Increased Expression of Plasminogen Activator Inhibitor Type1in HepG2 Cells Exposed to Free Fatty Acids or Triglycerides

2002 ◽  
Vol 3 (2) ◽  
pp. 109-118 ◽  
Author(s):  
Yabing Chen ◽  
David J. Schneider
Keyword(s):  
Fatty Acids ◽  
Free Fatty Acids ◽  
Plasminogen Activator ◽  
Hepg2 Cells ◽  
Plasminogen Activator Inhibitor ◽  
Fold Induction ◽  
Activator Inhibitor ◽  
Pai 1

We have shown that both free fatty acids (FFA) and triglycerides (TG) increase expression of plasminogen activator inhibitor type 1 (PAI-1) in vivo and in vitro. To determine signaling mechanisms responsible, HepG2 cells were exposed to FFA, emulsified TG, or the combination. The combination of FFA and TG increased PAI-1 to a greater extent than either agent alone (fold induction: 0.45mM FFA 1.7±0.2, 1000mg/dl TG 1.9±0.1, both 2.3±0.2, n=10, p<0.05 for comparison of combination with either alone). Cells transfected with PAI-1 5' flanking region containing the 4G or 5G polymorphism displayed similar activity in response to FFA, but modestly greater activity with the 4G polymorphism in response to TG (fold induction: 5G-1.28±0.14 and 4G- 1.46±0.13, n=6, p<0.05 for comparison). Deletion analyses demonstrated that FFA and TG induce PAI-1 expression through distinct regions of the promoter. Inhibition of protein kinase C inhibited the response to FFA but not TG. Accordingly, increased FFA and TG contribute to increased PAI- I through independent mechanisms.

The Roles of α2-Antiplasmin and Plasminogen Activator Inhibitor 1 (PAI-1) in the Inhibition of Clot Lysis

1993 ◽  
Vol 70 (02) ◽  
pp. 301-306 ◽  
Author(s):  
Linda A Robbie ◽  
Nuala A Booth ◽  
Alison M Croll ◽  
Bruce Bennett
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Fibrin Clot ◽  
Clot Lysis ◽  
Plasminogen Activator Inhibitor 1 ◽  
Dependent Inhibition ◽  
Activator Inhibitor ◽  
Pai 1

SummaryThe relative importance of the two major inhibitors of fibrinolysis, α2-antiplasmin (α2-AP) and plasminogen activator inhibitor (PAI-1), were investigated using a simple microtitre plate system to study fibrin clot lysis in vitro. Cross-linked fibrin clots contained plasminogen and tissue plasminogen activator (t-PA) at concentrations close to physiological. Purified α2-AP and PAI-1 caused dose-dependent inhibition. All the inhibition due to normal plasma, either platelet-rich or poor, was neutralised only by antibodies to α2-AP. Isolated platelets, at a final concentration similar to that in blood, 2.5 × 108/ml, markedly inhibited clot lysis. This inhibition was neutralised only by antibodies to PAI-1. At the normal circulating ratio of plasma to platelets, α2-AP was the dominant inhibitor. When the platelet:plasma ratio was raised some 20-fold, platelet PAI-1 provided a significant contribution. High local concentrations of PAI-1 do occur in thrombi in vivo, indicating a role for PAI-1, complementary to that of α2-AP, in such situations.

scholarly journals Tumor necrosis factor increases the production of plasminogen activator inhibitor in human endothelial cells in vitro and in rats in vivo

Blood ◽  
1988 ◽  
Vol 72 (5) ◽  
pp. 1467-1473 ◽  
Author(s):  
VW van Hinsbergh ◽  
T Kooistra ◽  
EA van den Berg ◽  
HM Princen ◽  
W Fiers ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
Blood Platelets ◽  
Interleukin 1 ◽  
Plasminogen Activator Inhibitor ◽  
Activator Inhibitor ◽  
Necrosis Factor ◽  
Pai 1

Abstract The vascular endothelium plays an important role in fibrinolysis by producing tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI). The monokine tumor necrosis factor (human recombinant TNF) increased the production of PAI by cultured human endothelial cells from umbilical vein (twofold) and from foreskin microvessles (four to eight fold). This was demonstrated by titration of endothelial cell-conditioned medium with t-PA, by reverse fibrin autography, and by immunoprecipitation of [35S]PAI-1 by anti-PAI-1 IgG. TNF also induced a marked increase of PAI-1 messenger RNA (mRNA) in the cells. The stimulation of PAI activity by TNF was seen at 4 U/mL and reached a maximum at 500 U/mL. Human recombinant lymphotoxin and interleukin-1 (alpha and beta) also stimulated the production of PAI activity, while interleukin-6 was ineffective. Separate additions of TNF or interleukin-1 (IL-1) at optimal concentrations (500 U/mL and 5 U/mL, respectively) resulted in a comparable stimulation of PAI production by endothelial cells. The simultaneous addition of both mediators resulted in an additive effect. The effect of TNF could not be prevented by the addition of polymyxin B or by anti-IL-1 antibodies. Therefore, it is unlikely that TNF acts through the induction of IL-1 secretion by endothelial cells. Two hours after a bolus injection of 250,000 U/kg TNF into rats, a fivefold increase in circulating PAI levels was found. In the next ten hours, the levels returned to normal. Blood platelets do not significantly contribute to the increase in circulating PAI, because the number of platelets did not change after TNF injection and the amount of PAI in blood platelets is not sufficient for several hours during an increase in PAI activity. The acute phase reactants, fibrinogen and alpha 2-antiplasmin in rat plasma, were altered little if any two to 24 hours after injection of 250,000 U/kg TNF. In vitro, TNF did not change PAI production by human and rat hepatocytes in primary monolayer culture. Therefore, it is most likely that vascular endothelial cells contribute to the increased amount of circulating PAI induced by TNF in vivo. This increase in PAI activity might decrease fibrinolysis.

scholarly journals Role of plasminogen activator inhibitor-1 in promoting fibrin deposition in rabbits infused with ancrod or thrombin

Blood ◽  
1993 ◽  
Vol 82 (12) ◽  
pp. 3631-3636 ◽  
Author(s):  
C Krishnamurti ◽  
C Bolan ◽  
CA Colleton ◽  
TM Reilly ◽  
BM Alving
Keyword(s):  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Fibrin Deposition ◽  
Plasminogen Activator Inhibitor 1 ◽  
Elevated Activity ◽  
Activator Inhibitor ◽  
Pai 1

The role of defective fibrinolysis caused by elevated activity of plasminogen activator inhibitor-1 (PAI-1) in promoting fibrin deposition in vivo has not been well established. The present study compared the efficacy of thrombin or ancrod, a venom-derived enzyme that clots fibrinogen, to induce fibrin formation in rabbits with elevated PAI-1 levels. One set of male New Zealand rabbits received intravenous endotoxin to increase endogenous PAI-1 activity followed by a 1-hour infusion of ancrod or thrombin; another set of normal rabbits received intravenous human recombinant PAI-1 (rPAI-1) during an infusion of ancrod or thrombin. Thirty minutes after the end of the infusion, renal fibrin deposition was assessed by histopathology. Animals receiving endotoxin, rPAI-1, ancrod, or thrombin alone did not develop renal thrombi. All endotoxin-treated rabbits developed fibrin deposition when infused with ancrod (n = 4) or thrombin (n = 6). Fibrin deposition occurred in 7 of 7 rabbits receiving both rPAI-1 and ancrod and in only 1 of 6 receiving rPAI-1 and thrombin (P “ .01). In vitro, thrombin but not ancrod was inactivated by normal rabbit plasma and by purified antithrombin III or thrombomodulin. The data indicate that elevated levels of PAI-1 promote fibrin deposition in rabbits infused with ancrod but not with thrombin. In endotoxin-treated rabbits, fibrin deposition that occurs with thrombin infusion may be caused by decreased inhibition of procoagulant activity and not increased PAI-1 activity.

scholarly journals Tumor necrosis factor increases the production of plasminogen activator inhibitor in human endothelial cells in vitro and in rats in vivo

Blood ◽  
1988 ◽  
Vol 72 (5) ◽  
pp. 1467-1473
Author(s):  
VW van Hinsbergh ◽  
T Kooistra ◽  
EA van den Berg ◽  
HM Princen ◽  
W Fiers ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Plasminogen Activator ◽  
Blood Platelets ◽  
Interleukin 1 ◽  
Plasminogen Activator Inhibitor ◽  
Activator Inhibitor ◽  
Necrosis Factor ◽  
Pai 1

The vascular endothelium plays an important role in fibrinolysis by producing tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI). The monokine tumor necrosis factor (human recombinant TNF) increased the production of PAI by cultured human endothelial cells from umbilical vein (twofold) and from foreskin microvessles (four to eight fold). This was demonstrated by titration of endothelial cell-conditioned medium with t-PA, by reverse fibrin autography, and by immunoprecipitation of [35S]PAI-1 by anti-PAI-1 IgG. TNF also induced a marked increase of PAI-1 messenger RNA (mRNA) in the cells. The stimulation of PAI activity by TNF was seen at 4 U/mL and reached a maximum at 500 U/mL. Human recombinant lymphotoxin and interleukin-1 (alpha and beta) also stimulated the production of PAI activity, while interleukin-6 was ineffective. Separate additions of TNF or interleukin-1 (IL-1) at optimal concentrations (500 U/mL and 5 U/mL, respectively) resulted in a comparable stimulation of PAI production by endothelial cells. The simultaneous addition of both mediators resulted in an additive effect. The effect of TNF could not be prevented by the addition of polymyxin B or by anti-IL-1 antibodies. Therefore, it is unlikely that TNF acts through the induction of IL-1 secretion by endothelial cells. Two hours after a bolus injection of 250,000 U/kg TNF into rats, a fivefold increase in circulating PAI levels was found. In the next ten hours, the levels returned to normal. Blood platelets do not significantly contribute to the increase in circulating PAI, because the number of platelets did not change after TNF injection and the amount of PAI in blood platelets is not sufficient for several hours during an increase in PAI activity. The acute phase reactants, fibrinogen and alpha 2-antiplasmin in rat plasma, were altered little if any two to 24 hours after injection of 250,000 U/kg TNF. In vitro, TNF did not change PAI production by human and rat hepatocytes in primary monolayer culture. Therefore, it is most likely that vascular endothelial cells contribute to the increased amount of circulating PAI induced by TNF in vivo. This increase in PAI activity might decrease fibrinolysis.

Impairment of adipose tissue development by hypoxia is not mediated by plasminogen activator inhibitor-1

2006 ◽  
Vol 95 (01) ◽  
pp. 174-181 ◽  
Author(s):  
Fabrizio Semeraro ◽  
Gabor Voros ◽  
Désiré Collen ◽  
H. Lijnen
Keyword(s):  
Adipose Tissue ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Tissue Development ◽  
Plasminogen Activator Inhibitor 1 ◽  
Adipose Tissue Development ◽  
Activator Inhibitor ◽  
Pai 1

SummaryHypoxia in rodents and humans is associated with a reduction of body fat on the one hand, and with enhanced expression of plasminogen activator inhibitor-1 (PAI-1), the main inhibitor of the fibrinolytic system, on the other hand. It was the objective of this study to investigate whether impairment of adipose tissue development by hypoxia may be mediated by PAI-1. Five week old male wild-type (WT) C57Bl/6 mice were fed a standard (SFD) or high fat (HFD) diet and kept under normoxic or hypoxic (10% O2) conditions. In addition, PAI-1 deficient mice and WT littermates were kept on HFD under normoxia or hypoxia. In vitro, the effect of hypoxia (2% O2) was investigated on differentiation of 3T3-L1 cells into adipocytes. Hypoxia induced a significant reduction of weight gain in WT mice on either SFD or HFD, accompanied by lower weights of subcutaneous (SC) and gonadal (GON) fat. Under hypoxic conditions, adipocytes in the adipose tissues were significantly smaller, whereas blood vessel size and density were larger. Serum PAI-1 levels were enhanced in hypoxic mice on SFD but not on HFD, and overall did not correlate with the observed changes in adipose tissue composition. Furthermore, the effects of hypoxia on adipose tissue in mice on HFD were not affected by deficiency of PAI-1. The inhibiting effect of hypoxia on in vitro preadipocyte differentiation was not mediated by PAI-1 activity. In conclusion, impairment of in vivo adipose tissue development and in vitro differentiation of preadipocytes by hypoxia is not mediated by PAI-1.

Synergistic Effect of Adrenal Steroids and Angiotensin II on Plasminogen Activator Inhibitor-1 Production1

2000 ◽  
Vol 85 (1) ◽  
pp. 336-344 ◽  
Author(s):  
Nancy J. Brown ◽  
Kyung-Soo Kim ◽  
Yan-Qun Chen ◽  
Lewis S. Blevins ◽  
John H. Nadeau ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Plasminogen Activator ◽  
Venous Blood ◽  
Plasminogen Activator Inhibitor ◽  
Luciferase Reporter ◽  
Ang Ii ◽  
Activator Inhibitor ◽  
Pai 1

Recent data suggest an interaction between the renin-angiotensin-aldosterone system and fibrinolysis. Although previous work has focused on the effect of angiotensin II (Ang II) on plasminogen activator inhibitor (PAI-1) expression, the present study tests the hypothesis that aldosterone contributes to the regulation of PAI-1 expression. To test this hypothesis in vitro, luciferase reporter constructs containing the human PAI-1 promoter were transfected into rat aortic smooth muscle cells. Exposure of the cells to 100 nmol/L Ang II resulted in a 3-fold increase in luciferase activity. Neither 1 μmol/L dexamethasone nor 1 μmol/L aldosterone alone increased PAI-1 expression. However, both dexamethasone and aldosterone enhanced the effect of Ang II in a dose-dependent manner. This effect was abolished by mutation in the region of a putative glucocorticoid-responsive element. A similar interactive effect of Ang II and aldosterone was observed in cultured human umbilical vein endothelial cells. The time course of the effect of aldosterone on Ang II-induced PAI-1 expression was consistent with a classical mineralocorticoid receptor mechanism, and the effect of aldosterone on PAI-1 synthesis was attenuated by spironolactone. To determine whether aldosterone affected PAI-1 expression in vivo, we measured local venous PAI-1 antigen concentrations in six patients with primary hyperaldosteronism undergoing selective adrenal vein sampling. PAI-1 antigen, but not tissue plasminogen activator antigen, concentrations were significantly higher in adrenal venous blood than in peripheral venous blood. Taken together, these data support the hypothesis that aldosterone modulates the effect of Ang II on PAI-1 expression in vitro and in vivo in humans.

scholarly journals Type 1 plasminogen activator inhibitor synthesis of endothelial cells is downregulated by smooth muscle cells

Blood ◽  
1993 ◽  
Vol 81 (5) ◽  
pp. 1277-1283 ◽  
Author(s):  
G Christ ◽  
D Seiffert ◽  
P Hufnagl ◽  
A Gessl ◽  
J Wojta ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Type Plasminogen Activator ◽  
Activator Inhibitor ◽  
Pai 1

Abstract Plasminogen activator inhibitor type 1 (PAI-1), the physiologic inhibitor of both tissue-type plasminogen activator (tPA) and urokinase- type plasminogen activator (uPA), is a major biosynthetic product of endothelial cells in vitro; endothelial cells in vivo, in contrast, do not appear to produce significant amounts of PAI-1 as made evident by in situ-hybridization studies in normal mice. This suggests that the high rate of PAI-1 synthesis of endothelial cells in vitro might be a result of the culture conditions. When human umbilical vein endothelial cells (HUVEC) were grown on human amniotic membranes, resembling the natural growth support instead of coated plastic, their morphology was changed from the cobblestone-like appearance on plastic to an in vivo like flagstone pattern. However, this morphological change had no significant effect on the synthesis and secretion of PAI-1. When smooth muscle cell (SMC) conditioned media (CM) were added to HUVEC cultures, PAI-1 antigen secretion of HUVEC was reduced by 40% to 60% as measured by enzyme-linked immunosorbent assay (ELISA). Immunoprecipitation experiments using 36S-methionine metabolically labeled HUVEC and Northern blot analysis of HUVEC PAI-1 mRNA indicate that this reduction was attributable to decreased PAI-1 synthesis and reduced steady-state levels of both the 3.2 kb and 2.2 kb form of PAI-1 mRNA. This effect was dose-dependent and observed under serum-containing as well as serum- free conditions, in the absence or presence of endothelial cell growth supplement (ECGS, 0 to 100 micrograms/mL) and attributable to a nondialyzable factor. Our data suggest that the high level of PAI-1 biosynthesis of endothelial cells in vitro may be attributable to the lack of a soluble factor produced by SMC, which controls and suppresses PAI-1 biosynthesis of endothelial cells in vivo.

scholarly journals Type 1 plasminogen activator inhibitor synthesis of endothelial cells is downregulated by smooth muscle cells

Blood ◽  
1993 ◽  
Vol 81 (5) ◽  
pp. 1277-1283
Author(s):  
G Christ ◽  
D Seiffert ◽  
P Hufnagl ◽  
A Gessl ◽  
J Wojta ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Plasminogen Activator ◽  
Plasminogen Activator Inhibitor ◽  
Type Plasminogen Activator ◽  
Activator Inhibitor ◽  
Pai 1

Plasminogen activator inhibitor type 1 (PAI-1), the physiologic inhibitor of both tissue-type plasminogen activator (tPA) and urokinase- type plasminogen activator (uPA), is a major biosynthetic product of endothelial cells in vitro; endothelial cells in vivo, in contrast, do not appear to produce significant amounts of PAI-1 as made evident by in situ-hybridization studies in normal mice. This suggests that the high rate of PAI-1 synthesis of endothelial cells in vitro might be a result of the culture conditions. When human umbilical vein endothelial cells (HUVEC) were grown on human amniotic membranes, resembling the natural growth support instead of coated plastic, their morphology was changed from the cobblestone-like appearance on plastic to an in vivo like flagstone pattern. However, this morphological change had no significant effect on the synthesis and secretion of PAI-1. When smooth muscle cell (SMC) conditioned media (CM) were added to HUVEC cultures, PAI-1 antigen secretion of HUVEC was reduced by 40% to 60% as measured by enzyme-linked immunosorbent assay (ELISA). Immunoprecipitation experiments using 36S-methionine metabolically labeled HUVEC and Northern blot analysis of HUVEC PAI-1 mRNA indicate that this reduction was attributable to decreased PAI-1 synthesis and reduced steady-state levels of both the 3.2 kb and 2.2 kb form of PAI-1 mRNA. This effect was dose-dependent and observed under serum-containing as well as serum- free conditions, in the absence or presence of endothelial cell growth supplement (ECGS, 0 to 100 micrograms/mL) and attributable to a nondialyzable factor. Our data suggest that the high level of PAI-1 biosynthesis of endothelial cells in vitro may be attributable to the lack of a soluble factor produced by SMC, which controls and suppresses PAI-1 biosynthesis of endothelial cells in vivo.

scholarly journals The pharmacokinetics of plasminogen activator inhibitor-1 in the rabbit

Blood ◽  
1990 ◽  
Vol 76 (8) ◽  
pp. 1514-1520 ◽  
Author(s):  
EJ Mayer ◽  
T Fujita ◽  
SJ Gardell ◽  
RJ Shebuski ◽  
CF Reilly
Keyword(s):  
Plasminogen Activator ◽  
Expression System ◽  
Plasminogen Activator Inhibitor ◽  
Bolus Administration ◽  
Plasminogen Activator Inhibitor 1 ◽  
Main Site ◽  
Activator Inhibitor ◽  
Pai 1

Abstract The pharmacokinetics of the activated and latent forms of plasminogen activator inhibitor-1 (PAI–1) isolated from HT1080 fibrosarcoma cells (HT1080 PAI-1) and a nonglycosylated form of human PAI-1 isolated from a yeast expression system (rPAI-1) were followed in the rabbit. As assessed by an immunologic assay specific for human PAI-1, guanidine HCI activated HT1080 PAI-1 and rPAI-1 entered the total plasma volume following intravenous bolus administration and exhibited a biphasic clearance pattern. The t1/2s of HT1080 PAI-1 for the initial and beta phases equalled 6.0 and 24.8 minutes, respectively. The t1/2s of rPAI-1 for the initial and beta phases equalled 8.8 and 34.0 minutes, respectively. Similar results were obtained by measuring PAI-1 activity in plasma and with trace amounts of 125I-rPAI-1, suggesting that the above pharmacokinetic behavior could also apply to endogenous PAI-1. The liver was the main site of rPAI-1 clearance. Unactivated, latent PAI-1 exhibited a very different pharmacokinetic profile. Over 80% of latent rPAI-1 cleared from the circulation within 10 minutes (t1/2 = 1.7 minutes). The difference in clearance behavior between activated and latent PAI-1 may be related to the ability of activated PAI-1, but not latent PAI-1, to rapidly form high-molecular-weight complexes with plasma binding factors which were observed in vitro and in vivo. Because PAI-1 could potentially tilt the fibrinolytic balance toward a prothrombotic state, its rapid clearance may represent an important control mechanism governing the circulating levels of this key component of the fibrinolytic pathway.

The Activation of Plasminogen Activator Inhibitor-1 Expression by IL-1β Is Attenuated by Estrogen in Hepatoblastoma HepG2 Cells Expressing Estrogen Receptor α

1999 ◽  
Vol 81 (03) ◽  
pp. 423-427 ◽  
Author(s):  
Marshall Scicchitano ◽  
Edward Kilbourne
Keyword(s):  
Estrogen Receptor ◽  
Plasminogen Activator ◽  
Hepg2 Cells ◽  
Plasminogen Activator Inhibitor ◽  
Estrogen Receptor Α ◽  
Plasminogen Activator Inhibitor 1 ◽  
17Β Estradiol ◽  
Activator Inhibitor ◽  
Pai 1

SummaryA low estrogen status in postmenopausal women is associated with elevated plasma levels of plasminogen activator inhibitor-1 (PAI-1). In this study, the ability of estrogen compounds to regulate PAI-1 expression was determined in a hepatocyte HepG2 cell line made to stably express estrogen receptor α (ERα). In both the wild type and ER expressing HepG2 cells, estrogen had no effect on basal PAI-1 expression. However, in the ER expressing cells the ability of IL-1β to increase PAI-1 mRNA and protein levels was attenuated by 17β-estradiol, tamoxifen and twelve estrogen components of Premarin. In contrast, the mixed agonist/antagonist raloxifene had weak agonist activity and like the pure antagonist ICI 182780, it dose dependently blocked the effect of 17β-estradiol on IL-1β stimulated PAI-1 levels. These results suggest that estrogen agonists may lower PAI-1 levels in vivo by inhibiting cytokine activated PAI-1 expression by an ER dependent mechanism.

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